Podcasts about CD36

What Causes Cancer to Metastasize and How to Potentially Help Control it with Diet A type of saturated fat concentrated in meat and dairy seems to cause cancer to metastasize. Check out today's episode to find out more. Written by Dr. Michael Greger at @NutritionFacts.org #vegan #plantbased #plantbasedbriefing #cancer #saturatedfat #metastaticcancer #metastasis #cd36 ============================ Original post: https://nutritionfacts.org/video/friday-favorites-what-causes-cancer-to-metastasize-and-how-to-potentially-help-control-it-with-diet    Related Episodes: Use search feature at https://www.plantbasedbriefing.com/episodes-search  Search for CANCER   ============================ Dr. Michael Greger is a physician, New York Times bestselling author, and internationally recognized speaker on nutrition, food safety, and public health issues. A founding member and Fellow of the American College of Lifestyle Medicine, Dr. Greger is licensed as a general practitioner specializing in clinical nutrition. He is a graduate of the Cornell University School of Agriculture and Tufts University School of Medicine. He founded NUTRITIONFACTS.ORG is a non-profit, non-commercial, science-based public service provided by Dr. Michael Greger, providing free updates on the latest in nutrition research via bite-sized videos. There are more than a thousand videos on nearly every aspect of healthy eating, with new videos and articles uploaded every day.   His latest books —How Not to Age, How Not to Die, the How Not to Die Cookbook, and How Not to Diet — became instant New York Times Best Sellers. His two latest books, How to Survive a Pandemic and the How Not to Diet Cookbook were released in 2020.  100% of all proceeds he has ever received from his books, DVDs, and speaking engagements have always and will always be donated to charity. ============================== FOLLOW THE SHOW ON: YouTube: https://www.youtube.com/@plantbasedbriefing     Spotify: https://open.spotify.com/show/2GONW0q2EDJMzqhuwuxdCF?si=2a20c247461d4ad7 Apple Podcasts: https://podcasts.apple.com/us/podcast/plant-based-briefing/id1562925866 Your podcast app of choice: https://pod.link/1562925866 Facebook: https://www.facebook.com/PlantBasedBriefing   LinkedIn: https://www.linkedin.com/company/plant-based-briefing/   Instagram: https://www.instagram.com/plantbasedbriefing/     

References Biosci Rep. 2022 Jun 30; 42(6): BSR20211854 Nature Reviews Endocrinology 2018. volume 14:452–463 Bone. 2022-11-01, Volume 164, Article 116539 Beethoven LV. 1806. Violin Concerto in D Major OP 61 https://youtu.be/rhdXr9l5gPM?si=0WcFfHzKajylaUBA Hunter-Garcia. 1973 Row Jimmy on [Wake of the Flood;lp] https://youtu.be/B4k9XfHBVEE?si=T3mXTER0Nvh2cccG --- Support this podcast: https://podcasters.spotify.com/pod/show/dr-daniel-j-guerra/support

Palmitic acid, a saturated fat concentrated in meat and dairy, can boost the metastatic potential of cancer cells through the fat receptor CD36. Randomized controlled trials show that lowering saturated fat intake can lead to improved breast cancer survival.

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.23.533959v1?rss=1 Authors: King, S. D., Cai, D., Fraunfelder, M. M., Chen, S.-Y. Abstract: BACKGROUND: Atherosclerosis is a progressive inflammatory disease where macrophage foam cells play a central role in the pathogenesis. Surfactant protein A (SPA) is a lipid-associating protein involved with regulating macrophage function in various inflammatory diseases. However, the role of SPA in atherosclerosis and macrophage foam cell formation has not been investigated. METHODS: Primary resident peritoneal macrophages were extracted from wild-type (WT) and SPA deficient (SPA-/-) mice to determine the functional effects of SPA in macrophage foam cell formation. SPA expression was assessed in healthy vessels and atherosclerotic aortic tissue from the human coronary artery and WT or apolipoprotein e-deficient (ApoE-/-) mice brachiocephalic arteries fed high fat diets (HFD) for 4 weeks. Hypercholesteremic WT and SPA-/- mice fed a HFD for 6 weeks were investigated for atherosclerotic lesions in vivo. RESULTS: In vitro experiments revealed that global SPA deficiency reduced intracellular cholesterol accumulation and macrophage foam cell formation. Mechanistically, SPA-/- dramatically decreased CD36 cellular and mRNA expression. SPA expression was increased in atherosclerotic lesions in humans and ApoE-/- mice. In vivo SPA deficiency attenuated atherosclerosis and reduced the number of lesion-associated macrophage foam cells. CONCLUSIONS: Our results elucidate that SPA is a novel factor for atherosclerosis development. SPA enhances macrophage foam cell formation and atherosclerosis through increasing scavenger receptor cluster of differentiation antigen 36 (CD36) expression. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.12.527706v1?rss=1 Authors: Barrachina, M. N., Pernes, G., Becker, I. C., Allaeys, I., Hirsch, T. I., Groeneveld, D. J., Khan, A. O., Freire, D., Guo, K., Carminita, E., Morgan, P. K., Collins, T. J., Mellett, N., Wei, Z., Almazni, I., Italiano, J. E., Luyendyk, J., Meikle, P. J., Puder, M., Morgan, N. V., Boilard, E., Murphy, A. J., Machlus, K. R. Abstract: Lipids contribute to hematopoiesis and membrane properties and dynamics, however, little is known about the role of lipids in megakaryopoiesis. Here, a lipidomic analysis of megakaryocyte progenitors, megakaryocytes, and platelets revealed a unique lipidome progressively enriched in polyunsaturated fatty acid (PUFA)-containing phospholipids. In vitro, inhibition of both exogenous fatty acid functionalization and uptake and de novo lipogenesis impaired megakaryocyte differentiation and proplatelet production. In vivo, mice on a high saturated fatty acid diet had significantly lower platelet counts, which was prevented by eating a PUFA-enriched diet. Fatty acid uptake was largely dependent on CD36, and its deletion in mice resulted in thrombocytopenia. Moreover, patients with a CD36 loss-of-function mutation exhibited thrombocytopenia and increased bleeding. Our results suggest that fatty acid uptake and regulation is essential for megakaryocyte maturation and platelet production, and that changes in dietary fatty acids may be a novel and viable target to modulate platelet counts. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2022.12.06.519076v1?rss=1 Authors: Hara, Y., Hirano, K.-i. Abstract: 123I-15-(p-iodophenyl)-(R,S)-methyl pentadecanoic acid (BMIPP) is a long-chain fatty acid (LCFA) analog developed to examine myocardial LCFA metabolism and has been used as a tracer for nuclear cardiology. However, its use is limited because of the specialized features of cardiac scintigraphy. In this study, a novel BMIPP-based probe was utilized, in which iodine-123 was replaced with a fluorescent compound, to extend the use of 123I-BMIPP to a wider variety of cells ex vivo. To confirm that this fluorescent LCFA analog (fluorescent BMPP) was imported into cells, fluorescence-activated cell sorting (FACS) analysis and fluorescent cell imaging were performed using cultured cells. The analysis showed that the import of fluorescent BMPP into the cells occurred in a concentration-dependent manner. This import into cells was inhibited by Sulfosuccinimidyl Oleate in a dose-dependent manner, which is an inhibitor of CD36, a well-known LCFA transporter, suggesting that fluorescent BMPP could be imported into cells via the same pathway as LCFA. FACS and cell imaging intensities of the cells importing fluorescent BMPP were attenuated after incubation in the non-Alexa680-BMPP medium. These results suggest that fluorescent BMPP can be transported into and from cells, reflecting the metabolism of LCFA. Fluorescently-labeled BMPP has the potential to be used as a probe for studying LCFA metabolism in various cells. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

The discussion over what aspect of our diet has been driving the escalation of metabolic disease over the last 100 years has been a confusing one full of contradictory information. It would be logical to look for a change in our diet associated with such a dramatic change in the health of a population There are both correlated evidence and dietary trials to support the theory that the increase in linoleic acid, provided by the introduction of PUFA processed oils into our diet, is the driver. In this episode of Hunger Hunt Feast, I want to share a collection of them to help clarify some of the confusion. -- Episode Specific Links:  Linoleic acid concentrations in fats and oils http://www.distributionkatrina.com/english/comparison-of-dietary-fats.html Increase in Adipose Tissue Linoleic Acid of US Adults in the Last Half-Century https://www.ncbi.nlm.nih.gov/labs/pmc/articles/PMC4642429/ Composition of adipose tissue and marrow fat in humans by 1H NMR at 7 Tesla https://www.sciencedirect.com/science/article/pii/S0022227520346733 Diets could prevent many diseases https://www.researchgate.net/publication/10673140_Diets_could_prevent_many_diseases Corn Oil in Treatment of Ischemic Heart Disease https://ncbi.nlm.nih.gov/pmc/articles/PMC2166702/ Linoleic acid causes greater weight gain than saturated fat without hypothalamic inflammation in the male mouse https://pubmed.ncbi.nlm.nih.gov/27886622/#:~:text=the%20male%20mouse-,Linoleic%20acid%20causes%20greater%20weight%20gain%20than%20saturated%20fat%20without,doi%3A%2010.1016%2Fj Effects of fatty acids on mitochondria: implications for cell death https://pubmed.ncbi.nlm.nih.gov/12206909/ Can linoleic acid contribute to coronary artery disease? https://pubmed.ncbi.nlm.nih.gov/8192728/ Effects of linoleate-enriched and oleate-enriched diets in combination with alpha-tocopherol on the susceptibility of LDL and LDL subfractions to oxidative modification in humans https://pubmed.ncbi.nlm.nih.gov/8148354/ Acrolein is a product of lipid peroxidation reaction https://www.jbc.org/article/S0021-9258(18)80708-6/fulltext#seccestitle90 The role of dietary oxidized cholesterol and oxidized fatty acids in the development of atherosclerosis https://pubmed.ncbi.nlm.nih.gov/16270280/ Rapeseed oil and sunflower oil diets enhance platelet in vitro aggregation and thromboxane production in healthy men when compared with milk fat or habitual diets https://pubmed.ncbi.nlm.nih.gov/1641826/ Stearoyl-CoA Desaturase-1 Is Associated with Insulin Resistance in Morbidly Obese Subjects https://link.springer.com/article/10.2119/molmed.2010.00078 Circulating levels of linoleic acid and HDL-cholesterol are major determinants of 4-hydroxynonenal protein adducts in patients with heart failure https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3909262/ Circulating levels of linoleic acid and HDL-cholesterol are major determinants of 4-hydroxynonenal protein adducts in patients with heart failure https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3909262/ Role of Physiological Levels of 4-Hydroxynonenal on Adipocyte Biology: Implications for Obesity and Metabolic Syndrome https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4038367/ “The Hateful Eight: Enemy Fats That Destroy Your Health - Dr. Cate.” Dr. Cate, 22 May 2020,  https://drcate.com/the-hateful-eight-enemy-fats-that-destroy-your-health/ Effects of diets enriched in linoleic acid and its peroxidation products on brain fatty acids, oxylipins, and aldehydes in mice https://www.ncbi.nlm.nih.gov/labs/pmc/articles/PMC6180905/ Strong increase in hydroxy fatty acids derived from linoleic acid in human low-density lipoproteins of atherosclerotic patients https://pubmed.ncbi.nlm.nih.gov/9488997/ Brief episode of STZ-induced hyperglycemia produces cardiac abnormalities in rats fed a diet rich in n-6 PUFA https://journals.physiology.org/doi/full/10.1152/ajpheart.00480.2004?rfr_dat=cr_pub++0pubmed&url_ver=Z39.88-2003&rfr_id=ori%3Arid%3Acrossref.org Changes in Dietary Fat Intake Alter Plasma Levels of Oxidized Low-Density Lipoprotein and Lipoprotein(a) https://pubmed.ncbi.nlm.nih.gov/9844997/ “Dr. Knobbe Presents, ‘Macular Degeneration - Preventable & Treatable - With an Ancestral Diet?' at Weston A. Price Foundation's Annual Conference - Wise Traditions - 2017 - Cure AMD Foundation.” Cure AMD Foundation, https://www.cureamd.org/dr-knobbe-presents-macular-degeneration-preventable-treatable-with-an-ancestral-diet-at-weston-a-price-foundations-annual-conference-wise-traditions-2017/ Unsaturated fatty acids and their oxidation products stimulate CD36 expression in human macrophages https://www.researchgate.net/publication/11260045_Unsaturated_fatty_acids_and_their_oxidation_products_stimulate_CD36_expression_in_human_macrophages Lowering dietary linoleic acid (LA) reduces bioactive oxidized linoleic acid metabolites in humans https://www.ncbi.nlm.nih.gov/labs/pmc/articles/PMC3467319/ Can linoleic acid contribute to coronary artery disease? https://www.researchgate.net/publication/15005994_Can_linoleic_acid_contribute_to_coronary_artery_disease A high linoleic acid diet increases oxidative stress in vivo and affects nitric oxide metabolism in humans https://pubmed.ncbi.nlm.nih.gov/9844997/ Dietary Linoleic Acid Elevates Endogenous 2-AG and Anandamide and Induces Obesity https://onlinelibrary.wiley.com/doi/full/10.1038/oby.2012.38 Use of dietary linoleic acid for secondary prevention of coronary heart disease and death: evaluation of recovered data from the Sydney Diet Heart Study and updated meta-analysis https://www.bmj.com/content/346/bmj.e8707 Connect with Zane: ReLyte Electrolytes by Redmond Real Salt: https://shop.redmond.life?afmc=Zane Follow me on Instagram: https://www.instagram.com/zanegriggsfitness Follow me on YouTube: https://www.youtube.com/c/ZaneGriggs QUICK EPISODE SUMMARY What causes modern disease The hateful eight oils The correlation between vegetable oils and obesity The oxidation effects of LDL The truth about the dairy data How much cholesterol is in your brain  Where you can find a copy of today's mentioned study What we know about Linoleic acid

On Episode 17 of the Stroke Alert Podcast, host Dr. Negar Asdaghi highlights two articles from the June 2022 issue of Stroke: “Vitamin D Enhances Hematoma Clearance and Neurologic Recovery in Intracerebral Hemorrhage” and “Acute Ischemic Stroke, Depressed Left Ventricular Ejection Fraction, and Sinus Rhythm: Prevalence and Practice Patterns.” She also interviews Dr. Bruce Campbell on his article “Role of Intravenous Thrombolytics Prior to Endovascular Thrombectomy.” Dr. Negar Asdaghi:         Let's start with some questions. 1) Is vitamin D that golden key to recovery from intracerebral hemorrhage? 2) Endovascular therapies seem to have prevailed where thrombolytics have failed. In the era of fast and furious thrombectomy, what is the role of pre-thrombectomy thrombolysis? 3) And finally, 20 years of clinical research has failed to demonstrate the superiority of anticoagulation over antiplatelet therapies for treatment of patients in sinus rhythm with low left ventricular ejection fraction, and yet, our practice patterns have not changed. Why do we remain resolute in prescribing anticoagulation despite the lack of evidence? We're back here to tackle the toughest questions with our Stroke Alert Podcast because this is the latest in Stroke. Stay with us. Dr. Negar Asdaghi:         Welcome back to another extremely motivating Stroke Alert Podcast. My name is Negar Asdaghi. I'm an Associate Professor of Neurology at the University of Miami Miller School of Medicine and your host for the monthly Stroke Alert Podcast. The June 2022 issue of Stroke contains a number of interesting articles. As part of our Advances in Stroke, we have two articles, one on the topic of cost-effectiveness of stroke care to inform health policy and the second on the current state and the future of emerging stroke therapies. As part of our Original Contributions category, we have an interesting study by Dr. [Ben] Assayag and colleagues from the Department of Neurology at Tel Aviv Sourasky Medical Center, where we learned that just over 10% of patients with TIA and stroke developed post-traumatic stress disorder, or PTSD. Higher presenting stroke severity, preexisting white matter disease, and having anxious coping styles are risk factors for development of post-stroke PTSD. Dr. Negar Asdaghi:         In another Original Contribution, by Dr. Daehoon Kim and colleagues from Yonsei University College of Medicine in Seoul, South Korea, we read with interest on the topic of whether or not we should be anticoagulating frail patients with atrial fibrillation. In this large population-based cohort, which included patients with atrial fibrillation older than 65 years of age with frailty as defined by a score of equal or greater than five on Hospital Frailty Risk Score, we learned that despite their frailty, patients with atrial fibrillation still significantly benefit from oral anticoagulation therapy. In this study, those treated with anticoagulation had lower net adverse clinical events as compared to those untreated. We also learned that direct oral anticoagulants provided lower incidence of stroke, bleeding, and mortality over Coumadin. This paper really provided practical information on treatment of frail patients with atrial fibrillation. So, I encourage you to review these papers in addition to listening to our podcast today. Later in the podcast, I have the great pleasure of interviewing Dr. Bruce Campbell from University of Melbourne in Australia on an especially timely topic, that is the role of intravenous thrombolytics prior to endovascular therapy. Dr. Campbell is a leading authority on the topic, and his interview does not disappoint. But first, with these two articles. Dr. Negar Asdaghi:         In the setting of intracerebral hemorrhage, or ICH, aside from the primary brain insult that occurs at the time of hemorrhage, secondary brain injuries continue for days and sometimes to months mostly due to the pathological response of the brain to byproducts of hematoma lysis or RBC degradation products. Today, the majority of spontaneous ICH cases are not surgically evacuated, so we rely on the body's own ability to clear blood for hematoma clearance, and obviously the faster the clearance, the better the outcome. Erythrophagocytosis by monocyte-derived macrophages contributes to hematoma clearance and ultimately to the functional recovery from ICH. So, it's conceivable that therapeutic approaches to enhance the endogenous erythrophagocytosis can potentially improve ICH outcomes. Vitamin D has been known to have variety of functions within the central nervous system, and it turns out that it may also be one such therapeutic option to improve the much needed erythrophagocytosis in intracerebral hemorrhage. Dr. Negar Asdaghi:         In the current issue of the journal, in the study titled "Vitamin D Enhances Hematoma Clearance and Neurologic Recovery in Intracerebral Hemorrhage," a group of researchers led by Dr. Jiaxin Liu from the Department of Surgery at Queen Mary Hospital at the University of Hong Kong studied the effects of oral vitamin D administered two hours after the induction of hematoma in a rodent model of ICH using direct collagenase injection into the striatum of the mouse. Eighty-nine young mice and 78 middle-aged mice were included in the study and randomly divided into three groups. Group one were sham-operated mice; group two, ICH mice treated with vehicle, which was corn oil; and group three, vitamin D-treated ICH mice. In the third group, 1000 international unit per kg of vitamin D diluted in corn oil was administered orally using a pipette two hours after the induction of ICH to mice, and then daily afterwards. And here are their top three findings of this study. Dr. Negar Asdaghi:         Number one, vitamin D-treated mice did better than vehicle on two neurobehavioral tests that were completed in the study. On the cylinder test, treatment with vitamin D significantly alleviated the asymmetric usage of four limbs at day seven, and vitamin D elongated the duration that the mice could run on the accelerated rod at day 10 on the rotarod test. Dr. Negar Asdaghi:         Number two, in terms of hematoma resolution and perihematoma edema, it's an issue that we deal with, with ICH, they used MRI imaging for edema measurement on T2-weighted images, and then sacrificed the mice and used digital quantification of hematoma volume with fresh brain specimens. And they found that treatment with vitamin D significantly alleviated both the ICH-associated brain swelling on MR and resulted in significant reduction in hematoma volume on the fresh brain specimens when compared with the vehicle-treated group at day three and day five. Dr. Negar Asdaghi:         And finally, their third main finding is in terms of erythrophagocytosis. So, the pathway that is mediated by the monocyte-derived macrophages is an endogenous pathway, that is, PPAR-γ (which stands for peroxisome proliferator-activated receptor γ) and its downstream scavenger receptor CD36 mediated. This pathway is essential for directing the endogenous erythrophagocytosis. Using flow cytometry, they found that vitamin D-treated mice had more mature macrophages expressing the scavenger receptor CD36, which was not expressed by the undifferentiated monocytes. Dr. Negar Asdaghi:         Western blot analysis confirmed that vitamin D treatment increased the tissue levels of CD36 and the upstream PPAR-γ levels in the brain at day five after collagenase model. Locally, vitamin D-enriched phagocytes that were positive for PPAR-γ and CD36 in the perihematoma regions. So, in summary, vitamin D increased the number of mature macrophages rather than undifferentiated monocytes in the perihematoma region and accelerated the differentiation of reparative macrophages from bone marrow-derived monocytes. So, bottom line is that in vitamin D, we have a simple, accessible, and well-tolerated agent to improve both the ICH outcomes and enhance hematoma resolution, but this we all observed in rodents. So, we stay tuned with interest to find out whether the same success will be seen in humans treated with vitamin D after intracerebral hemorrhage. Dr. Negar Asdaghi:         Patients with depressed left ventricular ejection fraction, or low EF, are at risk of development of ischemic stroke even if they remain in sinus rhythm. The optimal antithrombotic treatment for these patients is still unknown. Over the past two decades, we have a number of randomized trials studying the efficacy of oral anticoagulation, predominantly Coumadin, over aspirin therapy in prevention of all forms of stroke, that is ischemic and hemorrhagic, and death in patients with a low EF in sinus rhythm. Dr. Negar Asdaghi:         The meta-analysis of WASH, HELAS, WATCH, and WARCEF trials showed that treatment of low ejection fraction patients in sinus rhythm with Coumadin does reduce the subsequent risk of stroke, but it comes at the cost of a higher major bleeding risk in this population. The COMMANDER HF clinical trial published in New England Journal of Medicine in October 2018 studied whether low-dose rivaroxaban at 2.5 milligram BID was superior to placebo in patients with recent worsening of chronic heart failure, reduced ejection fraction, coronary artery disease, but no atrial fibrillation, and very similar to its prior counterparts, it did not show that rivaroxaban was associated with a lower rate of combined death, myocardial infarction, or stroke as compared to placebo. But very similar to prior studies, it also showed that rivaroxaban-treated patients had a lower risk of subsequent ischemic stroke. This poses a conundrum for stroke neurologists treating patients with this condition, especially after they present with an embolic-appearing stroke. So, the question is, how often do we encounter this situation, and what do we do in routine practice? We know that when there is equipoise, there's practice variation. Dr. Negar Asdaghi: In the current issue of the journal, in the study titled "Acute Ischemic Stroke, Depressed Left Ventricular Ejection Fraction, and Sinus Rhythm," Dr. Richa Sharma from the Department of Neurology at Yale School of Medicine and colleagues examined the prevalence of heart failure with sinus rhythm among hospitalized patients with acute ischemic stroke and the physician's practice patterns with regard to the choice of antithrombotics in this population. Dr. Negar Asdaghi:         So, let's look at their study. The study was comprised of five separate study cohorts of hospitalized acute ischemic stroke patients in the Greater Cincinnati Northern Kentucky Stroke Study for the year 2005, 2010, and 2015, and then four additional academic hospital-based cohorts in the United States during different timeframes. These were the Massachusetts General Hospital from 2002 to 2016, Rhode Island Hospital from 2016 to 2018, Yale-New Haven Hospital 2015 to 2017, and Cornell Acute Stroke Academic Registry from 2011 to 2018. All of these cohorts combined contributed to the 19,155 total number of patients in this study, which included over 14,000 patients that had documented left ventricular ejection fraction. Amongst those, 1,426 had a depressed EF and were included in this study. The investigator obviously excluded those with documented atrial fibrillation and flutter. And so the sample size for this analysis was 805 patients. And here are their main results. Dr. Negar Asdaghi:         The overall prevalence of this condition, that is low ejection fraction and sinus rhythm, among hospitalized acute ischemic stroke patients was 5%. It varied slightly between the different cohorts in this study from 4 to 6%. In terms of the antithrombotic treatment patterns, this information was available in close to 500 patients in the cohort. Overall, 59% of patients were discharged on an antiplatelet treatment alone, and 41% on anticoagulation. But these percentages significantly varied between the different institutions and was as low as 22% in one of the cohorts and as high as 45% in another cohort. Dr. Negar Asdaghi:         So, what were the factors that were associated with the use of anticoagulation at discharge? They found that the absolute percentage of left ventricular ejection fraction and the presenting NIH Stroke Scales were associated with anticoagulation use. That is, the lower the percentage of EF and the higher the presenting NIH Stroke Scale, the more likely physicians were to discharge the patients on an anticoagulation in univariate analysis, but in multivariate analysis, only the study site and presenting NIH Stroke Scale over eight were independently associated with anticoagulation use. Dr. Negar Asdaghi:         Now, interestingly, 2002 to 2018, which was their overall study period, was a time during which some of the largest and neutral randomized trials on the topic of anticoagulation versus antiplatelet were published, including the WATCH and the WARCEF trial. But the authors found no temporal variation in anticoagulation practice patterns before and after the publication of the results of these trials. So, it appears that we didn't change our minds. So, overall, we have some important takeaway messages from this study. We learned that 5% of hospitalized acute ischemic stroke patients have low left ventricular ejection fraction and remain in sinus rhythm without atrial fibrillation. Today, over 40% of patients with this condition are anticoagulated at discharge despite the results of the randomized trials, but the practice is widely variable among different institutions, and a higher presenting NIH Stroke Scale is a significant predictor of anticoagulation use at discharge in this population. Dr. Negar Asdaghi:         Almost 20 years after the approval of intravenous thrombolysis for treatment of patients with acute ischemic stroke, endovascular therapy was approved for treatment of select ischemic stroke patients with a large vessel occlusion. The two treatments are, therefore, entangled, as one was the standard of care while the second one was being tested. Therefore, all endovascularly treated patients enrolled in randomized trials would've received intravenous thrombolysis if eligible. Now, with the overwhelming success of endovascular therapy in achieving reperfusion in areas where IV thrombolysis has drastically failed, there're still critical questions regarding the added value of IV thrombolysis to endovascularly treated patients. The critical question remains as to whether eligible ischemic stroke patients who have immediate access to endovascular thrombectomy should receive prior IV thrombolysis, or should we skip the thrombolysis step altogether and just move to the angio suite as fast as possible. And there are, of course, arguments for and against each approach. Dr. Negar Asdaghi:         In this issue of the journal, in an invited topical review titled "The Role of Intravenous Thrombolytics Prior to Endovascular Thrombectomy," we learn about these arguments as the authors go through a comprehensive review of the current literature on this issue. I'm joined today by the first author of this review, Dr. Bruce Campbell, to discuss this paper. Dr. Campbell absolutely needs no introduction to our Stroke listeners. He's a professor of neurology and head of neurology and stroke at Royal Melbourne Hospital, University of Melbourne, in Australia. He's a pioneer in the field of acute stroke therapies and acute neuroimaging. He has served as the lead investigator of multiple landmark randomized trials, including EXTEND-IA and EXTEND-IA TNK, and holds multiple leadership roles. He's the clinical director of the Stroke Foundation and co-chairs the Australian Stroke Guidelines Working Party and the coordinator of the National Brain School Training Program for Neurologists in Training. And, of course, last but not least, he's my friend. So, I'm delighted to welcome him to our podcast today. Top of the morning to you, Bruce, 6:00 a.m. in Melbourne. That's quite some dedication. Thank you for being here. Dr. Bruce Campbell:       It's great to be with you. Thanks for the invitation. Dr. Negar Asdaghi:         Congrats on the paper, really exciting topic. So, let's just start with this question as part of a case. We have a patient with an M1 occlusion, a large clinical syndrome presenting two hours out from their symptom onset, and we are at a hospital where the angio suite is ready. What are some of the benefits of basically spending time in giving IV thrombolytics first rather than quickly going to the angio suite? Dr. Bruce Campbell:       I think a key element of this case is that the patient has presented directly to a hospital with immediate access to thrombectomy. Thrombolytic used in drip-and-ship transfer patients really isn't controversial, and the recent randomized trials excluded them. So, the debate's all about this context of bridging thrombolytics in patients presenting directly to a comprehensive stroke center. And you mentioned spending time giving lytics, but in fact, if you do things in parallel, that shouldn't be the case. It shouldn't delay thrombectomy if you go and give thrombolysis. Dr. Bruce Campbell:       So, the general principle is that getting the artery open faster by any means is better, and IV thrombolytic certainly has the potential to open the artery before thrombectomy in a proportion of patients, perhaps not that many, but it may also facilitate the thrombectomy. So, in the randomized trials, reperfusion after the thrombectomy was significantly better when patients had had bridging thrombolytic despite a low rate of pre-endovascular reperfusion. Other reasons for giving the lytics are the potential safety net it provides if the thrombectomy procedure is unexpectedly delayed or fails to get the artery open, and there's also this potential for lytics to dissolve distal embolic fragments and perhaps improve microvascular reperfusion. Dr. Negar Asdaghi:         So, great. So, let me summarize for our listeners what you mentioned. First off, so these are arguments in favor of giving lytics. As you mentioned, we're not really wasting time. These processes occur in parallel, so it's not like we're wasting time in giving a therapy that is potentially not as efficacious as thrombectomy is. And number two, we have improved the possibility of early reperfusion, perhaps, with the lytics. And if there are some fragments or distal clots that thrombectomy wouldn't have reached, then the lytics would. And then also there is also the chance that the thrombectomy might have failed in difficult access, and so on and so forth, and at least the patient has some chance of revascularization with the lytics. So, if these are the arguments for giving lytics, what are the arguments against giving lytics in this scenario? Dr. Bruce Campbell:       The main argument is the potential to reduce both the intracerebral and systemic hemorrhagic complications. There's also potential cost saving by skipping thrombolytics. That's probably more relevant in low-resource settings, particularly when relatives may have to pay for the thrombolytic before treatment is initiated, and that can be burdensome and also potentially delay the thrombectomy. There's a theoretical concern about thrombus fragmentation with lytics and potential migration of the clot out of reach of the thrombectomy or to new territories. But final reperfusion, as I mentioned, was, on average, better with the patient having a lytic on board in the randomized trials. Dr. Negar Asdaghi:         Perfect. And I want to highlight this issue of thrombus fragmentation because I think our readers will read more and more about this idea of, as you mentioned, fragmentation will potentially make an accessible clot for thrombectomy inaccessible. But I see that later in our questions, we're going to address that as part of the findings of randomized trials as well. So, these are some of the arguments for and against. And before we go to the randomized trials, I'd like to get an overview of what we knew as part of observational studies and non-randomized studies prior to more recent randomized trials on this topic. Dr. Bruce Campbell:       There've been a couple of nice systematic reviews and meta-analyses of the observational data, and notably in most of these studies, the direct thrombectomy patients had contraindications to lytics, and that introduces confounding factors that are difficult to adjust for. For what that's worth, the functional independence, mortality outcomes were better in the bridging patients. Hemorrhage rates weren't always higher with the lytic, and one study by Jonathan Coutinho in JAMA Neurology for the SWIFT and STAR studies showed the opposite despite them having really careful adjustment for all the confounders they could think of. And the meta-analysis by Eva Mistry in Stroke did not detect a difference in symptomatic ICH between the direct and bridging strategies. One thing that should be less affected by the patient characteristics would be the technical efficacy outcomes, and it was interesting that in the observational data, the patients who'd had bridging lytic had higher mTICI 2b-3 rates and also fewer device passes. Dr. Negar Asdaghi:         Okay. And now we do have further information with all of these new randomized trials. So, why don't we start with some of the earlier studies, the three, SKIP, DEVT, and DIRECT-MT, and start with those studies first before we move to some more recent European trials. Dr. Bruce Campbell:       SKIP was performed in Japan, and it used the lower 0.6 milligram per kilogram dose of alteplase that's standard there, and DEVT and DIRECT-MT were performed in China. All three of them showed numerically similar functional outcomes with slight trends favoring direct thrombectomy. SKIP had a smaller sample size and did not meet its non-inferiority criteria, and the other two trials did meet their specified non-inferiority margin, but it could be argued those margins were overly generous. If you think about non-inferiority trials, we generally try to set a margin for non-inferiority such as lower 95% confidence interval for the trial intervention would sacrifice up to 50% of the reference treatment effect. And it's difficult to estimate the effect of alteplase in this specific population. But if you think of the Emberson meta-analysis of alteplase, overall zero to three hours alteplase versus placebo has a 10% effect size and mRS 0-1, three to four and a half hours of 5% effect size. And we regard that as clinically important. So, half of 5%, 2.5%, is a lot tighter margin than any of the direct randomized trials employed. Dr. Negar Asdaghi:         So, Bruce, let me recap what you just mentioned. Two out of the three earlier trials seem to suggest that perhaps skipping IV therapy is the way to go rather than bridging as these two trials met the non-inferiority criteria if we believe that non-inferiority margins you mentioned. And now we have a couple of more trials, more recent trials. Can you tell us about these trials please? Dr. Bruce Campbell:       MR CLEAN-NO IV in a European population did not demonstrate non-inferiority, and the point estimate slightly favored bridging. Interestingly, in that trial, the symptomatic intracerebral hemorrhage risk, which was one of the main drivers for trying this strategy, was 5.9% in the direct and 5.3 in the bridging group. So, there's no hint of benefit from dropping the lytic on that metric. SWIFT-DIRECT was more selective in only enrolling internal carotid and M1 occlusions, which had a lower chance of early recanalization with lytic. But the protocol also specified giving the full dose of lytic. In the other trials, it seems the alteplase infusion was often stopped once the patient was in the angio suite, so the full dose may not have been delivered. And despite very low pre-endovascular recanalization in that selected group in SWIFT-DIRECT, the end of procedure reperfusion was significantly better in the bridging group, which is a consistent finding across the trials and suggests that the lytic may improve the thrombectomy outcome. Dr. Bruce Campbell:       DIRECT-SAFE, the final of those trials, was interesting in that the patients were enrolled roughly 50:50 from Australia, New Zealand, versus Asia. And in contrast to the original three randomized trials in Asian patients, DIRECT-SAFE found a significant benefit of bridging lytic in Asian patients. So, it'd be very interesting to see the results of the IRIS individual patient data meta-analysis, but we may not find a difference in Asian versus Caucasian patients despite those initial trials and despite substantial differences in the prevalence of intracranial atherosclerosis, which has often been proposed as something that would increase the risk of having bridging thrombolytic on board. Dr. Bruce Campbell:       The original study level estimate of symptomatic hemorrhage had a borderline significant 1.8% absolute reduction in the direct group. Whether those data were not all core lab adjudicated and the final analysis may show a smaller difference than that. Notably, given that trend with symptomatic intracerebral hemorrhage, mortality did not differ significantly, and, in fact, the trend favored bridging patients. So, the symptomatic hemorrhage slight trend into increase did not translate into any hint of increased mortality. Dr. Negar Asdaghi:         So, Bruce, a lot of information, and I need a recap for me. So, let me try to recap some of the things you said, and please jump in. So, so far, the newer data really basically don't show us any convincing evidence that skipping is the way to go, and direct endovascular we really don't have data in favor of going directly to the angio suite. And the jury is still out regarding an increase in the symptomatic intracerebral hemorrhage rate amongst those that actually are pre-treated with IV therapy. Is that correct? Dr. Bruce Campbell:       That's correct. So, none of the three recent trials met their non-inferiority margins. And again, we had this issue of relatively generous non-inferiority margins, and the symptomatic hemorrhage, it would make sense that there's a small difference, but it's not really been borne out in the data to be statistically significant at this stage. And again, this individual patient data meta-analysis is keenly awaited to get the most accurate estimate on that. Dr. Negar Asdaghi:         So, while we wait that, I'm going to digress a little bit and ask you a question that's not addressed in the paper that you have in this issue of the journal, and that's the CHOICE trial. So, by now, we have the results of CHOICE trial. Do you mind first give us a brief overview of what CHOICE was and how you feel that the results of CHOICE would affect this field of direct versus bridging in general? Dr. Bruce Campbell:       CHOICE is a very interesting study in that it tested giving the intra-arterial lytic at the end of a thrombectomy procedure that had achieved an mTICI 2b or better, which is what we traditionally regarded as angiographic success. The idea was to improve microvascular flow, and that may be the case. The trial was terminated early due to logistic reasons and showed a very large effect size that requires replication. The subgroup analyses are interesting in that the benefits seem to mostly accrue in patients who'd not already had intravenous lytic. Dr. Bruce Campbell:       So, perhaps giving the IV lytic before thrombectomy can still benefit patients after the thrombectomy, as well as achieving early recanalization in a proportion of patients and perhaps facilitating the thrombectomy. The other issue to address with the DIRECT trials is that with the exception of a few patients in DIRECT-SAFE, the comparator was alteplase and not tenecteplase. And we have data from EXTEND-IA TNK that tenecteplase bridging is not just non-inferior, but superior to alteplase bridging. There's an ongoing Brazilian trial of exactly that, tenecteplase versus the direct approach, which will be very interesting. Dr. Negar Asdaghi:         So, great, Bruce. I just want to repeat this segment again for our listeners. So, CHOICE is a very interesting study, looked at giving intraarterial alteplase to patients after endovascular therapy was completed and after they'd already achieved the complete and successful revascularization, and the trial was terminated early because of logistic reasons. So, we have to keep in mind, this was a smaller study, early termination, but the effect size was pretty large in favor of giving lytics. Dr. Negar Asdaghi:         So, what you mentioned is interesting, and I think that it's really worth paying attention to, that the majority of the benefits seem to have occurred from intraarterial thrombolytics in patients that have not been given intravenous lytics prior to endovascular therapy. So, in other words, you need some sort of lytics either before or after the endovascular thrombectomy to achieve that ultimate improved outcome. So, moving forward now from the randomized trials that we have on bridging versus direct thrombectomy, you have mentioned in the paper some interesting subgroups that may benefit or not benefit as much from bridging versus direct thrombectomy. Do you want to elaborate a little more about those subgroup analyses? Dr. Bruce Campbell:       The idea of precision selection or individualized treatment is being talked about a lot given there didn't seem to be much overall difference between strategies in the randomized trials, but it's important to note that the randomized trial actually disadvantages the bridging group by delaying lytic until the patient was firstly confirmed eligible for thrombectomy and then consented and randomized. Putting that aside, if we could identify a subgroup who clearly benefit from skipping lytic and, importantly, identify them without delaying lytic for those who likely benefit, that's clearly attractive. Dr. Bruce Campbell:       Currently, I'd say we have not identified that kind of subgroup, and the planned IRIS individual patient data meta-analysis will be critical for that. Patients with a large ischemic core are one potential group where there's a high risk of bleeding hypothesized. To date, there is no definitive data to indicate the risk is lower with the direct approach. Patients who need stents certainly may benefit from not having a lytic on board because they often need adjuvant antithrombotics that could increase the bleeding risk. But the question there is whether we can confidently identify those patients before the procedure, and I think that's unclear at this stage. Patients with really large clot burdens and proximal occlusions have sometimes been said not to benefit from IV lytic based on the low rates of pre-endovascular reperfusion, but the randomized trials really hinted other benefits like this potential facilitative thrombectomy. So, that hypothesis may be insecure as well. Dr. Negar Asdaghi:         And how about age? Have you come across and has there been any signal towards an impact or interaction between age and benefit from pre-endovascular thrombectomy and thrombolytics? Dr. Bruce Campbell:       It's an interesting question because age has not generally been a treatment effect modifier in previous stroke studies with thrombolytics and thrombectomy, and the individual direct thrombectomy trials that have reported subgroups haven't shown any convincing heterogeneity by age. There's certainly no indication that older patients are at risk from bridging in what I've seen so far. Dr. Negar Asdaghi:         So, this question comes up in clinical practice all the time, that a person's older, perhaps more atrophy, more vascular risk factors and white matter disease, and they're more prone, so to speak, of having a symptomatic intracerebral hemorrhage. So, what you're saying is, from the data we have, there's really no signal in favor of withholding pre-thrombectomy lytics in this population. So, it's important to know this. Bruce, what should be our final takeaway message from this study? Dr. Bruce Campbell:       I tend to agree with the recent European Stroke Organization and ESMINT guideline that for now, patients should receive lytic as early as possible and in parallel with the decision to perform thrombectomy such that neither treatment delays the other. I think if we can identify a subgroup that benefits from direct thrombectomy, and that's confirmed in the individual patient data and meta-analysis, and we can identify them without disadvantaging the majority of patients, and also that the ongoing improvements in IV lytic strategies don't render the existing trial data obsolete, then we may, in future, skip lytic for some patients, but we are not there yet. Dr. Negar Asdaghi:         So, that's amazing, Bruce. We look forward to reviewing the paper and individual data meta-analysis and interviewing you, hopefully at a better hour your time, on that. Thank you very much for joining us on the podcast today. Dr. Bruce Campbell:       Thanks again for the invitation. It's been great talking to you. Dr. Negar Asdaghi:         Thank you. Dr. Negar Asdaghi:         And this concludes our podcast for the June 2022 issue of Stroke. Please be sure to check out this month's table of contents for the full list of publications, including three very interesting images that are presented as part of a new article type, Stroke Images, and a special report in Comments and Opinions section on "Bias in Stroke Evaluation: Rethinking the Cookie Theft Picture." June is the month of Pride, and in spirit of equality, we hope to do our part to reduce all biases in stroke processes of care, diagnosis, and outcomes as we continue to stay alert with Stroke Alert. Dr. Negar Asdaghi:         This program is copyright of the American Heart Association, 2022. The opinions expressed by speakers in this podcast are their own and not necessarily those of the editors or of the American Heart Association. For more, visit AHAjournals.org.

This month on Episode 34 of Discover CircRes, host Cynthia St. Hilaire highlights four original research articles featured in the March 4 and March 18th issues of Circulation Research. This episode also features a conversation with Dr Mireille Ouimet and Sabrina Robichaud from the University of Ottawa Heart Institute to discuss their study, Autophagy is Differentially Regulated in Leukocyte and Non-Leukocyte Foam Cells During Atherosclerosis.   Article highlights:   Pauza, et al. GLP1R in CB Suppress Chemoreflex-Mediated SNA   Lim, et al. IL11 in Marfan Syndrome   Hohl, et al. Renal Denervation Prevents Atrial Remodeling in CKD   Liu, et al. Smooth Muscle Cell YAP Promotes Arterial Stiffness   Cindy St. Hilaire:        Hi and welcome to Discover CircRes, the podcast of the American Heart Association's journal, Circulation Research. I'm your host, Cindy St. Hilaire from the Vascular Medicine Institute at the University of Pittsburgh, and today I'm going to be highlighting articles from our March issues of Circulation Research. I'm also going to speak with Dr Mireille Ouimet and Sabrina Robichaud from the University of Ottawa Heart Institute, and they're with me to discuss their study, Autophagy is Differentially Regulated in Leukocyte and Non-Leukocyte Foam Cells During Atherosclerosis.   The first article I want to share is titled GLP1R Attenuates Sympathetic Response to High Glucose via Carotid Body Inhibition. The first author is Audrys Pauza, and the corresponding authors are Julian Paton and David Murphy at the University of Bristol.   Cindy St. Hilaire:        Hypertension and diabetes are risk factors for cardiovascular disease. And yet, for many patients with these two conditions, lowering blood pressure and blood sugar is insufficient for eliminating the risk. The carotid body is a cluster of sensory cells in the carotid artery, and it regulates sympathetic nerve activity. Because hypertension and diabetes are linked to increased sympathetic nerve activation, this group investigated the role of the carotid body in these disease states. They performed a transcriptome analysis of crowded body tissue, from rats with and without spontaneous hypertension. And they found among many differentially-expressed genes that the transcript encoding glucagon-like peptide-1 receptor or GLP1R, was considerably less abundant in hypertensive animals.   Cindy St. Hilaire:        This was of particular interest because the gut hormone GLP-1 promotes insulin secretion and tends to be suppressed in Type 2 diabetes. Moreover, GLP1R agonists are already used as diabetic treatments. This group showed that treating rat carotid body with GLP1R agonist suppresses sympathetic nerve activation and arterial blood pressure, suggesting that these drugs may provide benefits in more than one way. Perhaps the carotid body could be a novel target for lowering cardiovascular disease risk in metabolic syndrome.   Cindy St. Hilaire:        The second article I want to share is titled Inhibition of IL11 Signaling Reduces Aortic Pathology in Murine Marfan syndrome. The first author is Wei-Wen Lim, and the corresponding author is Stuart Cook and they're from the National Heart Center in Singapore. People with the genetic connective tissue disorder Marfan syndrome, are typically tall and thin with long limbs and are prone to skeletal, eye and cardiovascular problems, including a life-threatening weakening of the aorta. While Marfan syndrome patients commonly take blood pressure-lowering treatments to minimize risk of aortic aneurysm and dissection, there's currently no cure for Marfan syndrome or targeted therapy.   Cindy St. Hilaire:        The cytokine IL11 is strongly induced in vascular smooth muscle cells upon treatment with the growth factor TGF-beta, which is over activated in Marfan syndrome patients. And TGF-beta is also considered a key feature of the syndrome's molecular pathology. This study found that IL11 is strongly upregulated in the aortas of Marfan syndrome model mouse, and that genetically eliminating IL11 in these animals protected them against aortic dilation, fibrosis, inflammation, elastin degradation and loss of smooth muscle cells. Treating Marfan syndrome mice with anti-IL11 neutralizing antibodies exhibited the same beneficial effects. These results suggest that perhaps inhibiting IL11's activity could be a novel approach for protecting the aortas of Marfan syndrome patients.   Cindy St. Hilaire:        The next article I want to mention is titled Renal Denervation Prevents Atrial Arrhythmogenic Substrate Development in Chronic Kidney Disease. The first authors are, Mathias Hohl, Simina-Ramona Selejan and Jan Wintrich, and the corresponding authors also Mathias Hohl, and they're from Saarland University. People with chronic kidney disease have a two to three fold higher risk than the general population of developing atrial fibrillation, which is a common form of arrhythmia that can be life-threatening. Chronic kidney disease is associated with activation of the sympathetic nervous system, which can be damaging to the heart. Thus, this group examined myocardial tissues from atrial fibrillation patients with and without chronic kidney disease to see how they differ. They found that atrial fibrosis was more pronounced in patients with both conditions than in patients with atrial fibrillation alone, suggesting that chronic kidney disease perhaps exacerbates or even drives arterial remodeling.   Cindy St. Hilaire:        Sure enough, induction of chronic kidney disease in rats led to greater atrial fibrosis and incidence of atrial fibrillation than seen in the control animals. Renal denervation is a treatment in which the sympathetic nerves are ablated, and it's a medical procedure that's used for treating uncontrolled hypertension, and it has also been shown in animals to reduce atrial fibrillation. Performing renal denervation in the rats with chronic kidney disease reduced atrial fibrosis and atrial fibrillation susceptibility. This study not only shows that chronic kidney disease induces atrial fibrosis and in turn atrial fibrillation, but also suggests that renal denervation may be used in chronic kidney disease patients to break this pathological link and prevent potentially deadly arrhythmias.   Cindy St. Hilaire:        The last article I want to highlight is titled YAP Targets the TGFβ Pathway to Mediate High-Fat/High-Sucrose Diet-Induced Arterial Stiffness. First author is Yanan Liu and the corresponding author is Ding Ai from Tianjin Medical University. Metabolic syndrome is characterized as a collection of conditions that increase the risk of cardiovascular diseases, such as obesity, hypertension and diabetes. Among the tissue pathologies associated with metabolic syndrome is arterial stiffness, which itself is a predictor of cardiovascular disease incidence and mortality. To specifically investigate how arterial stiffness develops in metabolic syndrome, this group fed mice a high-fat, high-sugar diet, which is known to induce metabolic syndrome and concomitant arterial stiffness.   Cindy St. Hilaire:        After two weeks on the diet, the animals' aorta has exhibited significant upregulation of TGF-beta signaling, which is a pathway known for its role in tissue fibrosis, and the aorta has also exhibited increased levels of yes-associated protein, or YAP, which has previously been implicated in vascular remodeling, collagen deposition and inflammation. YAP gain and loss of function experiments in transgenic mice revealed that while knockdown of protein in the animals' smooth muscle cells attenuated arterial stiffness, increased expression exacerbated the condition.   Cindy St. Hilaire:        The team went on to show that YAP interacted with and prevented the activation of PPM-1 B, which is a phosphatase that normally inhibits TGF-beta signaling and thus fibrosis. Together the results suggest that targeting the YAP, PPM-1 B pathway, could be a strategy for reducing arterial stiffness and associated cardiovascular disease risk in metabolic syndrome.   Cindy St. Hilaire:        Today, Sabrina Robichaud and Dr Mireille Ouimet from University of Ottawa Heart Institute are with me to discuss their study Autophagy is Differentially Regulated in Leukocyte and Non-Leukocyte Foam Cells During Atherosclerosis, which is in our March 18 issue of Circulation Research. So thank you both for joining me today.   Sabrina Robichaud:    Thank you so much for having us. It's a pleasure.   Mireille Ouimet:         Thank you for having us.   Cindy St. Hilaire:        Yeah, and congrats on the study. So we know that LDL particles contain cholesterol and fats, and these are the initiating factors in atherosclerosis. And it's also really now appreciated that inflammation in the vessel wall is a secondary consequence to this lipid accumulation. Macrophages are an immune cell that, in the context of the plaque, gobble up this cholesterol to the point that they become laden with lipids and exhibit this foamy appearance, which we now call foam cells. And these foam cells can exhibit atheroprotective properties, one of them called reverse cholesterol transport, and that's really one of the focuses of your paper. So before we dig into what your paper is all about, could you give us a little bit of background about what reverse cholesterol transport is in the context of the atherosclerotic plaque? And maybe introduce how it links to this cellular recycling program, autophagy, which is also a big feature of your study.   Mireille Ouimet:         Yes, so the reverse cholesterol transport pathway is a pathway that's very highly anti-atherogenic. It's linked to HDL function and the HDL protective effects, in that HDL can serve as a cholesterol acceptor for any excess cholesterol from arterial cells or other cells of the body and return this excess cholesterol to the liver for excretion into the feces. There is also trans-intestinal cholesterol efflux that can help eliminate any excess bodily cholesterol. Mireille Ouimet:         So reverse cholesterol transport is a way that we can eliminate excess cholesterol from foam cells in the vascular wall, and that's why we're really interested in the process. But the rate-limiting step of cholesterol efflux out of foam cells in plaques is actually, they have to be mobilized in the form of free cholesterol to be pumped out of the cells through the action of the ATP-binding cassette transporters. And so the rate-limiting step of the process is the hydrolysis of the cholesterol esters and the lipid droplets, because that's where the excess cholesterol is stored in foam cells.   Mireille Ouimet:         And so for years, people investigated the actions of cytosol like lipases in mobilizing free cholesterol from lipid droplets, although the identity of those lipases are not well-known and in macrophage themselves, but our recent work showed a role for autophagy in the catabolism of lipid droplets. And in fact, in macrophage foam cells, 50% of lipid droplet hydrolysis is attributable to autophagy while the other half is mediated by neutral lipases, which makes it really important to investigate the mechanisms of autophagy-mediated lipid droplet catabolism.   Cindy St. Hilaire:        That is so interesting. I guess I didn't realize it was that significant a component in that kind of rate-limiting step. That's so cool. So really, a lot of the cholesterol efflux studies, and maybe this is just limited to my knowledge of a lot of these cholesterol efflux studies, but to my knowledge, it's been really focused on the foam cell itself, the macrophage foam cell. However, there's been a lot of recent work that has now implicated vascular smooth muscle cells in this process. So could you share some of the research specific to smooth muscle cells and smooth muscle-derived foam cells that led you to want to investigate the contributions of smooth muscle cell-derived foam cells in cholesterol efflux?   Mireille Ouimet:         Yeah, so you're right in the sense that macrophages have always been the culprit foam cells in the atherosclerotic plaques but pioneering work from several groups, including Edward Fisher and Gordon Francis, they've shown that the smooth muscle cells can actually acquire a macrophage-like phenotype becoming lipid-loaded and foamy. And there's been work specifically looking at the ABC transporters, and their ability to efflux cholesterol from these vascular smooth muscle cell-derived foam cells, because as they trans-differentiate into macrophage-like cells, they acquire the expression of ABCA1, but this is to a lower extent, as compared to their macrophage counterparts.   Mireille Ouimet:         And the efflux is defective because there's an impairment in liposomal cholesterol processing of the lipoproteins that's really important to activate a like cell, and the expression of the ABC transporters, so vascular smooth muscle cell-derived foam cells are very poor effluxes.   Sabrina Robichaud:    There's very few studies that look at the vascular smooth muscle cell foam cells, and the very few that did look at it mostly focused on the ABCA1 transporters, and did show that they were poor effluxes. And as we all know, ABC1 is not the only cholesterol transporters that can transport cholesterol out of cells, there's also ABCG1 which is also one of our major findings in our paper.   Cindy St. Hilaire:        Can you tell us a little bit about the models you chose in the study and why you picked them? And also maybe a step back in terms of, what are the pros and cons of using mouse models in atherosclerotic studies?   Sabrina Robichaud:    So we chose to use the GFP-LC3 reporter mouse model because it allows us to track in lifestyle the movement of LC3, which is the main component of the autophagosome which is involved in pathology. So by using this reporter model, we could infer whether or not the cells had high autophagy or low autophagy. And to induce atherosclerosis in these mice, instead of backcrossing them to either an LDLR knockout or an ApoE knockout, we chose to do the adeno-associated virus that encode the gain of function PCSK9 instead to kind of minimize the time for breeding. It did have the effect that we needed in terms of raising plasma cholesterol to induce the atherosclerosis. So that was one of the models that we used in our paper.   Mireille Ouimet:          There's not very many good mouse models to study autophagy flux in vivo and GFP-LC3 is kind of the main one currently. We're working on developing some other tools to track lipophagy in vivo, but these things take time to put in place. So in the future, we hope to have some better tools to track lipophagy in real-time in vivo.   Cindy St. Hilaire:        How difficult is it to measure autophagy flux in vivo? I know there's certain part like LC3 or P62, a lot of people use a western blot and it's like, oh, it's high, it must be active, but it's a flux. So it's a little bit more... There's more subtleties to that, dynamic than that. So how difficult is it to really measure this flux in in vivo tissues?   Mireille Ouimet:         Yes, so now there are more recent mouse models that have been developed more recently to replace kind of the GFP-LC3 is the Rosella LC3. So it has both a red and a green tag, and so two LC3, so when autophagosomes are fused to lysosomes and are degraded, then there's preferential quenching of the GFP first, and then you have the red appearance that predominates so we know that then it's kind of like it a live flux measurements. Because we use the GFP-LC3 mouse, Sabrina treated her cells ex vivo. When we dissected out the aortic arches, digested the cells then we divided those into two components and added bafilomycin so that we can inhibit lysosome acidification to see the changes in the flux. And that's really to get the differences in untreated versus bafilomycin-treated.   Mireille Ouimet:         When we inhibit the lysosome, then we're sure that it is a functional flux or not. But it's kind of an indirect way of measuring it, and it reads very complex when we're talking about P62 and LC3 degradation with or without lysosome inhibition, but you really need that lysosomal inhibition, to show that if you block the degradation of the autophagosomes that fuse in with a lysosome, then you get an increase in the LC3 and the P62, and that's when you know that the flux is you intact.   Mireille Ouimet:         Because you could get an increase in LC3, that's just related to a defect in the breakdown of the autophagosome. But in our study, we've used phosphorylated ATG16L1, which is a now better marker of active autophagy. And I would recommend researchers to begin to use that rather than the combination of P62 and LC3 together with or without a lysosome inhibitors such as- Cindy St. Hilaire:        Oh, interesting. So let's repeat that, phosphorylated ATG-   Mireille Ouimet:         16L1, yes. So there's been an antibody that was developed by a colleague at the University of Ottawa, Dr Ryan Russell, and it's commercially available through cell signaling now, and it really has been a great tool to track active autophagy.   Cindy St. Hilaire:        That's great. I remember my lab was looking at that at one point, and I was trying to explain the flux as... I don't know if people are going to remember this, but there's this amazing, I Love Lucy skit, where her and Ethel are working on a chocolate factory conveyor belt, and it picks up speed. And because she can't get it all done quick, she starts stuffing them in her mouth. And it's like, if you just took a snapshot of that, you would not know whether it's going too fast, or not functioning properly. And so I equate the flux experiments to that. Which are probably aging myself a lot on so.   Cindy St. Hilaire:        All right, so sticking to kind of the autophagy angle, what were the differences you found in autophagy in early and late atherosclerotic plaques? Because I know you looked at those two time points, but also, importantly, between the macrophage foam cells and the smooth muscle cell-derived foam cells?   Sabrina Robichaud:    So surprisingly, there weren't that big of a difference between each time point when we were looking at the individual cell type by themselves. Surprisingly, we did find that the macrophages did have a functional autophagy flux, even at the later stages of atherosclerosis, which was kind of interesting in itself. But when we looked at the vascular smooth muscle cell foam cells, though, that was a whole other story, and we found that these were actually defective at a very early stage and stayed defective up until the very late stage of atherosclerosis.   Cindy St. Hilaire:        And what is the very early stage like? What's that definition with the smooth muscle cell?   Sabrina Robichaud:    So we did a six-week time points in terms of our atherosclerosis study, and then a 25-week time point. So there are far apart, which shows like the very early, early stage and what would be considered the most effective autophagy at that point with the necrotic core and everything. So surprisingly, the two phenotype were quite similar at early and both late stages for both cell types, but were functional in the macrophages but dysfunctional in the smooth muscle cells.   Cindy St. Hilaire:        So you mentioned at one point in the discussion that you observed inconsistent lipid loading of the smooth muscle cells, and you mentioned that a lipase, which is excreted from the foam cells can then be internalized by, I assume kind of neighboring or in the vicinity, smooth muscle cells. And so the question I had it's kind of one of those chicken-and-egg question, and it's, is the smooth muscle cell-derived foam cell an independent process? Does it happen alone or de novo as a function of a smooth muscle-mediated process? Or is it really dependent first on this macrophage foam cell providing this lipid that is efflux that is then internalized by a smooth muscle cell that kind of goes on to become a foam cells. It's kind of a question of like the continuum of an atherosclerotic plaque and what do you think is happening, either based on your data or just kind of a hunch?   Mireille Ouimet:         That's an excellent question. And there's no doubt that macrophages really drive the initiating events of atherosclerosis. So I don't think that without the macrophage there would ever be a vascular smooth muscle cell, or there would be minimal vascular smooth muscle cell-derived foam cells. Definitely the inconsistencies that we observed in our study, were if we added like aggregated LDL on its own to a primary mouse vascular smooth muscle cell, we would get poor lipid loading and a very low percentage of those cells that would become foamy, relative to treating them with cyclodextrin complex cholesterol, for instance.   Mireille Ouimet:         So free cholesterol, that's cell permeable, will go into the vascular smooth muscle cell, no problem, and generate the foaminess and then allow that cell to acquire the macrophage-like phenotype. But aggregated LDL on its own in our hands, just gave very poor loading. And when we treated the vascular smooth muscle cells with aggregated LDL along with macrophage-derived condition media, we got some improvements, but it was still kind of inconsistent. But then we thought if we treat the vascular smooth muscle cells with aggregated LDL in the presence of conditioned media from macrophage foam cells that were preloaded with the aggregated LDL, would that promote their foaminess to a greater extent? And it did.   Mireille Ouimet:         So, there have been studies from Gordon Francis's lab that showed that adding recombinant lysosomal acid lipase to vascular smooth muscle cells that contained aggregated LDL, promoted the lysosomal hydrolysis of the aggregated LDL and to generate the foamy macrophages and allow the lysosomal processing. So we know that that vascular smooth muscle cells take up lysosomal acid lipase, and we know that macrophages undergo lysosome exocytosis and they can secrete lysosome acid lipase and acidify the extracellular milieu.   Mireille Ouimet:         So work from Fred Maxfield group has shown the presence of these cell surface connected compartments that are acidified, containing macrophage-derived lysosomal acid lipase, that even hydrolyze extra cellularly-aggregated LDL for macrophages. So we're not sure whether there's probably a local production of free cholesterol in the plaque by macrophages, this free cholesterol could be taken up by the vascular smooth muscle cell. And also the vascular smooth muscle cells do express some scavenger receptors, whether the expression of these scavenger receptors like LRP or CD36 even goes up when they've taken up a little bit of the free cholesterol. And then that allows the aggregated LDL to come in and then there would be some lysosomal acid lipase secreted by the macrophage foam cells that would promote the lysosomal processing of this aggregated LDL. All of those are very complex questions that will require some addressing in vivo models.     Cindy St. Hilaire:        You also mentioned in the paper that studies... There's a handful of them now. Studies have shown that between 30% and 70% of the cells that are staining positively for macrophage markers, meaning they're foam cells, are of the smooth muscle cell lineage. And so I believe people have seen that in mouse plaques with lineage tracing, but they've also used newer techniques to really see this also in human atherosclerotic plaques. So we know it's not just from a mouse, we know that smooth muscle cells can turn into a macrophage-like foam cell, and it's 30% to 70%, which is a huge range. Cindy St. Hilaire:        So do we know the factors that dictate whether a specific plaque is going to have more or less smooth muscle cell derived foam cells? And I guess more important to what you found in your paper is, how important would it be to know whether a plaque is on the 30% end or on the 70% end in terms of therapeutic strategies?   Sabrina Robichaud:    Yeah, most of these studies, the range can be attributed to the different time points at which these studies have been collected early on will be a little bit more macrophage understanding would be at a later time point. Now of course in terms of therapeutics, as we saw in our paper, metformin actually will positively increase cholesterol efflux in the vascular smooth muscle cell foam cells, but not in the macrophages. So obviously, being able to know at which point there's a majority of macrophages versus vascular smooth muscle cells, definitely going to determine which therapeutic we're going to be able to use.   Sabrina Robichaud:    Ideally, we would be able to find a therapeutic that would work in both foam cell, but from what we've seen, the mechanistic behind the autophagy dysfunction between both cell types are so different, that I'm not entirely sure that that would be possible, we would need some sort of combination therapy. But again, we need to be a little bit more targeted depending on the percentage of the foam cells that are comprising the plaque at that particular moment in time.   Cindy St. Hilaire:        Yeah, so you mentioned there's a function of time there. If you look earlier, there's more macrophage, if you look later, the percent of smooth muscle cell-derived foam cell increases. Is there a point in a very advanced atherosclerotic plaque where it's just mostly smooth muscle cells? Or do those macrophage foam cells stay, and it's just the increasing number of smooth muscle cell-derived foam cells? Do we know?   Mireille Ouimet:         This is an excellent question, and I was going to bring up the topic of clonal expansion of the vascular smooth muscle cells. So it's a very heterogeneous population and understanding that might be some of the differences that we see in different studies. It could be the model has one type of a smooth muscle cell that's expanding more than another, what are the factors that govern that? Does one clone take over at the later stages versus the earlier stages? We don't know.   Mireille Ouimet:         But we were surprised in our studies to see that the macrophages that are present at least on the lumen of the plaques were very active in autophagy. They had the highest staining for the phospho-ATG16L1 in that late stage. So we're not sure if it's newly-recruited macrophages that come in, that are more active and in autophagy, and then have good lysosomal capacity that keeps degrading the lipid present in the plaque and tries to ingest it, but also as a consequence keeps releasing some of the degraded cholesterol into the milieu where the smooth muscle cells that are proliferating are internalizing it and becoming more foamy. So these are really great open questions that need to be addressed in the field.   Cindy St. Hilaire:        So drug-eluting stents are coated with rapamycin or the various chemical compositions that are derived from rapamycin. And rapamycin itself induces autophagy. So while the thought behind using this coating on stents was to prevent smooth muscle cell proliferation, and thus restenosis or ingrowing of the stent, your study suggests that this could also help to promote autophagy in the cells underlying the stent. So has anyone gone in and looked at plaques that have been stented and either failed or not, and investigated the foam cell content or markers for autophagy activity?   Mireille Ouimet:         Not to my knowledge, and this has been something we've definitely... We think that this is what's happening. Some of the protective effects of these drug-eluting stents that have everolimus or sirolimus or the rapamycin or rapamycin analogs, we do believe that some of their protective effect can be attributed to autophagy activation, but this remains to be demonstrated. We think that autophagy activation locally would promote reverse cholesterol transport and would be one of the processes that prevents restenosis because we can promote the efflux of cholesterol out.   Cindy St. Hilaire:        Great. So I guess stemming from my question on the stents, what are the other translational implications of the findings of your study? And what would you like to see come out of this?   Mireille Ouimet:         So one of the things is, as Sabrina mentioned, would be to target both foam cell populations because it seems as though the vascular smooth muscle cell foam cells are very much defective in their autophagy capacity, and they're very poor effluxes, but we could potentially restore autophagy in the cell population to promote reverse cholesterol transport.   And looking at prevention of atherosclerosis is a bit different than looking at regression, because regression is at a later stage where the plaques are more advanced. And if they're mostly vascular smooth muscle cell-derived, maybe then those drugs that we're considering that protect against the development of atherosclerosis are effective on the macrophage themselves early on, but might not be mimicking what we would see in the clinic where the patients that present are older.   Cindy St. Hilaire:        Yeah, it's kind of really reminiscent of like the CANTOS trial and like, where do we want to target the therapy? It's going to be very different if it's an early smaller plaque, versus a late-stage possibly pro close to rupturing type of plaque. Well, Sabrina Robichaud and Dr Ouimet, thank you so much for joining me today. Congratulations again on a wonderful study, and I'm really looking forward to hearing more about this from your group.   Sabrina Robichaud:    Thank you.   Mireille Ouimet:         Thank you very much. And we also want to thank all the co-authors on the study, specifically also Adil Rasheed, who is co-first author on the work and Katey Rayner's group for all the support and involvement in this study.   Cindy St. Hilaire:        That's it for the highlights from the March issues of Circulation Research. Thank you for listening. Please check out the CircRes Facebook page and follow us on Twitter and Instagram with the handle @circres and #DiscoverCircRes. Thank you to our guests, Sabrina Robichaud and Dr Mireille Ouimet Sabrina. This podcast is produced by Ashara Ratnayaka, edited by Melissa Stoner and supported by the editorial team of Circulation Research. Some of the copy text for highlighted articles was provided by Ruth Williams. I'm your host, Dr Cindy St. Hilaire and this is Discover CircRes, you're on-the-go source for the most up-to-date and exciting discoveries in basic cardiovascular research. This program is copyright of the American Heart Association 2022, The opinions expressed by speakers in this podcast are their own and not necessarily those of the editors or of the American Heart Association. For more information, visit ahajournals.org.

In this week's episode, we'll review a research article showing beneficial effects of prenatal immunotherapy in a mouse model of anti-CD36-mediated fetal and neonatal alloimmune thrombocytopenia. Next, we'll look at results of a simulation analysis suggesting that gene therapy for hemophilia B is more cost-effective than on-demand or prophylactic factor treatment. We'll conclude with a report which provides important new insights into regulation of terminal erythroid maturation at the transcriptional level that may help improve our understanding of normal and abnormal erythropoiesis.

I discuss the complex but potent interactions of TLR signaling with plasma membrane protein complexes including semaphorin cytoplasmic kinase interactions leading to PPAR gamma transcription that fires off CD36 and beta oxidation gene transcription to switch M1 to M2 macrophage polarity thus feedback regulating the pro-inflammatory glycolytic mToRC system dynamics. Aging is associated with a corruption in this fine-tuning that presents in the older sedentary human individual as a simple dysregulation of inflammation that ultimately impairs the ability to ward off otherwise relatively innocuous pathogens while simultaneously punctuating the equilibrium otherwise exerted to control tissue repair and the potential for cellular proliferation and oncogenesis. References Nat Immunol. 2018 Jun;19(6):561-570. FEBS Letters 2011 Volume 585, Issue 23, Pages 3829-3835 Scientific Reports 2015 volume 5, Article number: 11789 Ann Allergy Asthma Immunol. 2000 Jul;85(1):9-18 --- Send in a voice message: https://anchor.fm/dr-daniel-j-guerra/message Support this podcast: https://anchor.fm/dr-daniel-j-guerra/support

Palmitic acid, a saturated fat concentrated in meat and dairy, can boost the metastatic potential of cancer cells through the fat receptor CD36.

In this episode we speak with Elliott Watkins aka Muselk the Executive Director at Click Management. Muselk stops by to break down what it takes to find long term success on YouTube. Creative Disruption Podcast with Ricky Ray Butler and Derral EvesInsights and stories from leading creators, writers, producers and marketers on how the worlds of advertising, entertainment, and data science are converging. Subscribe To YouTube

Dr. Knobbe began his practice of ophthalmology in 1994, after completing his residency training at the University of Colorado Health Sciences Center, in Denver, Colorado, USA. He was certified by the American Board of Ophthalmology in 1997 and has remained board-certified since then. Eye physician and surgeon, Chris A. Knobbe, MD, had been in practice nearly 20 years when, in 2013, he asked himself the question, “Could macular degeneration be a ‘Westernized disease’? Could AMD be a disease that is the result of a Westernized diet?” That question would forever change his life. The hypothesis that Dr. Knobbe would proffer holds that macular degeneration, which is the leading cause of irreversible vision loss and blindness in developed nations, is not only preventable but treatable in the early to moderate stages, with an ancestral diet.   Time Stamps:  0:09:24 Podcast Begins 0:12:39 200 years of nutritional research 0:23:09 Needs a title 60-sec audiogram 0:30:09 Timeline for Processed Food Introductions 0:36:44 Non-negotiable: Animal fats are essential 0:44:17 150 years of Vegetable Oil Consumption 0:49:07 Who's the real culprit? 0:51:46 Heart Disease Deaths vs Sturated fat and Vegetable Oil 1:05:32 Can linoleic acid contribute to coronary artery disease? https://www.researchgate.net/publication/15005994_Can_linoleic_acid_contribute_to_coronary_artery_disease 1:08:12 Why do we blame the carbs? 1:11:35 Sweden vs US adipose Linoleic Acid and obesity 1:14:02 The Case Of Japan 1:27:20 The problem is neither calories nor carbs 1:33:49 Is processed sugar harmful in and of itself? | What carbs to eat 1:43:36 The Case Of Okinawa 1:55:41 Diets could prevent many diseases Https://www.researchgate.net/publication/10673140_Diets_could_prevent_many_diseases 2:02:50 Lowering dietary linoleic acid reduces bioactive oxidized linoleic acid metabolites in humans https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3467319/ 2:03:29 Unsaturated fatty acids and their oxidation products stimulate CD36 expression in human macrophages https://www.researchgate.net/publication/11260045_Unsaturated_fatty_acids_and_their_oxidation_products_stimulate_CD36_expression_in_human_macrophages 2:04:13 Low-density lipoprotein rich in oleic acid is protected against oxidative modification: implications for dietary prevention of atherosclerosis https://www.ncbi.nlm.nih.gov/pmc/articles/PMC54010/ 2:06:00 A high linoleic acid diet increases oxidative stress in vivo and affects nitric oxide metabolism in humans https://www.researchgate.net/publication/12880728_A_high_linoleic_acid_diet_increases_oxidative_stress_in_vivo_and_affects_nitric_oxide_metabolism_in_humans 2:10:50 Changes in Dietary Fat Intake Alter Plasma Levels of Oxidized Low-Density Lipoprotein and Lipoprotein(a) https://www.ahajournals.org/doi/10.1161/01.atv.0000118012.64932.f4 2:11:39 Where to find Chris Knobbe -- Heart & Soil: www.heartandsoil.co  White Oak Pastures: www.whiteoakpastures.com code CarnivoreMD for 10% off first order Belcampo: www.belcampo.com code CarnivoreMD for 20% off BluBlox: www.blublox.com, use code CarnivoreMd for 15% off Sacred Hunting: www.sacredhunting.com/paul for $250 off of your hunt

New in 2021, we will feature 2 Feature Discussions every other week. For this week, we start with author Michael Gold and editorialist Sana Al-Khatib as they discuss the article "Primary Results from the Understanding Outcomes with the S-ICD in Primary Prevention Patients with Low Ejection Fraction (UNTOUCHED) Trial." Then, we switch to an important discussion about children and COVID-19 as author Israel Valverde and Associate Editor Gerald Greil discuss "Acute Cardiovascular Manifestations in 286 Children with Multisystem Inflammatory Syndrome Associated with COVID-19 Infection in Europe." TRANSCRIPT BELOW: Dr. Carolyn Lam: Welcome to Circulation on the Run. Your weekly podcast summary and backstage pass to the journal and its editors. We're your co-hosts. I'm Dr. Carolyn Lam, associate editor from the National Heart Center and Duke National University of Singapore. Greg Hundley: And I'm Greg Hundley, director of the Pauley Heart Center at VCU Health in Richmond, Virginia. Well, Carolyn, we have two features in this issue. Dr. Carolyn Lam: I know. Greg Hundley: Yeah. And I've got the first one. It's going to be evaluating the acute cardiovascular manifestations in 286 children with multi-system inflammatory syndrome. Dr. Carolyn Lam: Wow. That is so important in the current pandemic. Well, the other is also so important. It's really, really critical results from the subcutaneous ICD trial called UNTOUCHED. I think that one might change clinical practice. So, do you want to tell us about other papers first? Greg Hundley: Yes. Carolyn, I'm going to grab my cup of coffee and we'll get started and my first paper comes from China. Professor Junbo Ge. So Carolyn PCSK9, mainly secreted by the liver and released into the blood elevates plasma load density lipoprotein cholesterol by degrading LDL receptors. Pleiotropic effects of PCSK9 beyond lipid metabolism have been shown. However, the direct effects of PCSK9 on platelet activation and thrombosis, as well as the underlying mechanisms still remain unclear. Greg Hundley: So this group detected the direct effects of PCSK9 on agonist-induced platelet aggregation, dense granule ATP release, integrin a2b beta-3 activation, alpha granule release, spreading and clot retraction. They also investigated the underlying mechanisms. Using myocardial infarct models, they explored the effects of PCSK9 on microvascular obstruction and infarct expansion, post myocardial infarction. Dr. Carolyn Lam: Oh, nice. And what did they find? Greg Hundley: Well, Carolyn, PCSK9 in plasma directly enhances platelet activation and in vivo thrombosis, as well as myocardial infarct expansion post MI by binding to platelet CD36 and thus activating the downstream signaling pathways. PCSK9 inhibitors, or aspirin, abolished the enhancing effects of PCSK9 supporting the use of aspirin in patients with high plasma PCSK9 levels in addition to PCSK9 inhibitors to prevent thrombotic complications. Dr. Carolyn Lam: Wow. Very interesting. Indeed, the pleiotropic effects. Well guess what? My paper talks about the first non-LDL lowering treatment that has been shown to reduce CABG in a blinded randomized trial. Greg, can you guess what that treatment was? Well, here's a hint, here's a hint. It's the REDUCE-IT trial. Greg Hundley: Well, Carolyn, thanks for the hint. It must be icosapent ethyl. Dr. Carolyn Lam: Indeed, indeed. As a reminder, REDUCE-IT was a multicenter double-blind placebo controlled trial, which randomized statin treated patients with elevated triglycerides, controlled LDL and either established cardiovascular disease or diabetes plus other risk factors to receive icosapent ethyl at four grams daily or placebo. The primary and secondary composite end points were significantly reduced. The current paper examined all coronary revascularizations, recurrent revascularizations and revascularization subtypes. First revascularizations were reduced by icosapent ethyl versus placebo with a hazard ratio of 0.66, which is a number needed to treat of only 24. Similar reductions were observed in total revascularizations and across elective, urgent and emergent revascularizations. Icosapent ethyl significantly reduced PCI and CABG with a hazard ratio of 0.61. So, icosapent ethyl reduce first and total coronary revascularization, including PCI and CABG in patients with elevated triglycerides and high cardiovascular risk despite well controlled LDL. Isn't that cool? Greg Hundley: Very nice Carolyn. Well, my next paper comes from Professor Kari Alitalo, from the University of Helsinki. So Carolyn recent discoveries have indicated that in the developing heart, sinus venosus and endocardium provide major sources of endothelium for coronary vessel growth that supports the expanding myocardium. The author set out to study the origin of the coronary vessels that develop in response to vascular endothelial growth factor B or VEGF-B in the heart and the effect of VEGF-B on recovery from myocardial infarction. Dr. Carolyn Lam: So what were their results? Greg Hundley: Well Carolyn, the myocardial VEGF-B trans-gene promotes the formation of endocardium-derived coronary vessels during development, endothelial proliferation in sub-endocardial myocardium in adult mice and structural and functional rescue of cardiac tissue after myocardial infarction. So VEGF-B could provide a new therapeutic strategy for cardiac neovascularization after coronary occlusion to rescue the most vulnerable myocardial tissue. Well, Carolyn that's all of the main articles. How about we turn to some of the other articles and letters in the issue? Dr. Carolyn Lam: Yeah. Why not? And Greg, let me start by talking about an exchange of letters between Dr. Wei and Dr. Fox on interpreting the net clinical benefit from rivaroxaban plus aspirin versus aspirin for chronic vascular disease. There's also an ECG challenge by Dr. Patel elusively entitled, A Rainy Day. Here's a hint, it's about hypothermia. In cardiology news by Bridget Kuhn. She talks about how the pandemic throws cardiovascular trials off course, there is an On My Mind paper by Dr. Most entitled, The Striking Similarities of Multi-system Inflammatory Syndrome in Children and a Myocarditis-like Syndrome in Adults: The overlapping manifestations of COVID-19. As a couple of Research Letters, one by Dr. Malhotra on Defining the Normal Spectrum of Electrocardiographic and Left Ventricular Adaptations in Mixed Race, Male, Adolescent Soccer Players, as well as by Dr. Qi on adherence to a healthy sleep pattern and incident heart failure, a prospective study of more than 400,000 UK Biobank participants. Dr. Greg Hundley: Very nice Carolyn. Well, I've got a primmer review of Cardiac Involvement in Multi-system Inflammatory Syndrome in Children with the corresponding author being Dr. Kevin Freedman. Well, how about we get off quickly to those next two feature discussions? Dr. Carolyn Lam: Yay! Let's go, Greg. Dr. Greg Hundley: Well, listeners, we are to the first of our double feature for the new year 2021. And with me is Israel Valverde from King's College in London and our own associate editor, Dr. Gerald Greil from UT Southwestern in Dallas. Welcome gentlemen. And as we get started Israel, could you tell us a little bit about the background that framed this study and the hypothesis that you wanted to address? Dr. Israel Valverde: Thank you. I mean, the main problem we had in Europe around April 2020, was that both the Center for Disease Control and Prevention in the U.S. simulated a clinical ALIT about a newly described immune disease and inflammatory syndrome in children associated with a COVID infection. And there were similarities with all the well-known syndromes, such as viral myocarditis, Kawasaki disease, Kawasaki shock syndrome, and toxic shock syndrome, that we were a bit of confused, because of the overlap in clinical presentation, because it was a true diagnostics challenge. So the hypothesis in this study is we wanted to describe the cardiovascular implications in this newly described sinus syndrome. Dr. Greg Hundley: Very nice. So tell us a little bit about your study design and what study population did you assemble to address this question? Dr. Israel Valverde: I think that was the most difficult thing in the study, because thanks to the Association For European Pediatric Cardiology, the APC, we coordinated a multicenter study involving all they would appear on centers that we were 55 centers from 17 countries all over Europe. And we were able to recruit 286 children with this new newly described syndrome and cardiovascular manifestations. Dr. Greg Hundley: And tell us, what did you find? Dr. Israel Valverde: I think we can summarize that in three main findings. First is that cardiac involvement is very common in children with multi-system inflammatory syndrome associated with coronavirus disease, 2019 infection. Second, that inflammatory markers were significantly raised in most children, particularly their C-reactive protein, ferritin, pro-calcitonin, N-terminal pro BNP natriuretic peptide, interleukin 6, and D-dimer level. And finally, that 65% of patient with MIS-C had evidence of previous infection with severe acute respiratory syndrome coronavirus too, either by PCR, immunoglobulin M or immunoglobulin G. Dr. Greg Hundley: Were there any particular aspects of your results that may have segregated to the boys versus the girls? Dr. Israel Valverde: Not really. We couldn't find any differences between boys and girls, because our study population was quite similar, but we couldn't find any differences between them. What we found is that there is a huge difference between children and adults. Dr. Greg Hundley: And what was the age range of these children? Dr. Israel Valverde: From a couple of weeks, until 18 years old. Dr. Greg Hundley: And no differences in the scope of the syndrome that they experienced in the younger children versus the- Dr. Israel Valverde: Initially we found two peaks. Similarly to the peaks of viral myocarditises, which is affecting more of the children below two years, and also adolescence. Dr. Greg Hundley: Well Gerald, multi-system inflammatory syndrome in children. How to results from this study compare with perhaps other inflammatory disease processes that have been observed in children, such as Kawasaki disease, et cetera. Dr. Gerald Greil: So we and Circulation were very lucky to get a bunch of submissions regarding a MIS-C and also have a radio article regarding this and I would like to point our readers towards that. I think the key issue is it's a new disease entity. The community is pretty clear about it, that other diseases like Kawasaki disease have similarities, but it's not the same thing. And we wanted to be very clear about this. Obviously, more things need to be investigated, but the key findings where Dr. Valverde pointed out in other study groups within the U.S. and within Europe, all point to the same direction, that is a new entity, we need to further investigate. Dr. Greg Hundley: How about the cardiovascular aspects, what differences in cardiovascular disease with this syndrome, again, relative to other perhaps inflammatory diseases that might affect the heart in children? Dr. Gerald Greil: As far as we know. And once again, when we're talking about a very preliminary data, is that the outcome in children having MIS-C usually good. Dr. Valverde pointed out in his study series of more of 286 children, there was only one death and recovery was pretty high rate. We have similar findings from U.S. groups, other findings from a group in Paris and Europe. So, I would like to point our readers to other summaries where it's pretty clear that it's a new disease entity. Overall, it's actually rare in children. So we want to make a pretty clear point that it's a rare disease, which at this point in time seems to have a good outcome. Dr. Greg Hundley: Well Israel, I'm going to come back to you. What do you see is the next study that should be performed perhaps in this general area of cardiovascular disease for children? Dr. Israel Valverde: So I think what we have learned is that the way to move forward is the collaboration between centers. So now that we have a large study multicenter group, our idea is continue describing the long-term outcomes of the study population, because most of them recovered, but a few of them keep, for example, coronary artery dilatation. So do they recovering when you have time until you have time, do they do the ejection fraction, the function of the heart go back to normality soon? Or do we have to wait? So I think that's the next step forward to probably the long-term outcome of the study population. Dr. Greg Hundley: Longitudinal follow-up. And Gerald, do you have anything to add to that? Dr. Gerald Greil: I would just like to reiterate what Israel said, that we need to focus on multicenter studies. I hope Circulation can be a platform to reunite different groups around the world. Because as I mentioned before, I think we have a new disease entity in front of us. We don't know the long-term outcomes. And in the interest of our children confronted with new disease, we need to be very, very careful to learn how we need to follow up these children and how we potentially need to treat them in the long-term. Dr. Greg Hundley: Well, listeners, this has been a fantastic discussion with Dr. Israel Valverde from King's College in London and our own Dr. Gerald Greil from UT Southwestern, revealing some aspects of this multi-system inflammatory syndrome that's associated with COVID-19 in children. And so, on behalf of Carolyn and myself, well, we've got to get to the next feature. So, I'm not going to let you go just yet. Well, listeners, we are in the double feature year 2021. In our second feature discussion today, we have Dr. Michael Gold from Medical University of South Carolina and our own associate editor, Dr. Sana Al-Khatib, who has written an editorial on this paper. And she is from Duke University. Welcome to you both. Michael, we'll start with you. Could you describe for us a little bit about the background related to this paper and what hypothesis did you want to address? Dr. Michael Gold: Well, thank you. And thank you for this opportunity. The implantable defibrillator is a fundamental aspect of the treatment and prevention of sudden cardiac death. It's been around for almost 40 years now and is commonly used in the system has evolved from being a surgical procedure, requiring a thoracotomy to a transvenous procedure in which leads could be placed into the heart. While it's very effective, the major limitation of this in many people's minds were, major limitations were both of complications associated with the transvenous lead. Those include infections and lead failures, as well as unnecessary or inappropriate shock to patients. And based on that, the subcutaneous ICD was developed as the latest iteration of this technology in which a lead is placed under the skin and tunneled up along the sternum, so that one could sense and shock the heart when necessary without being subjected to an intravascular lead. Dr. Michael Gold: And the device has been around for 10 years or so. It's proven to be effective, but primarily used in niche populations of younger patients, patients with poor vascular access and those considered at relatively low risk with few comorbidities, such as patients with Brugada syndrome or long QT syndrome or possibly hypertrophic cardiomyopathy. We felt it was important both to establish the role of the subcutaneous ICD in a more typical group of defibrillator patients, as well as with the multiple evolutions now of programming, as well as technology within the device of seeing if the more modern contemporary devices were as effective in these sicker populations, and also could reduce some of the issues seen earlier in terms of higher rates of inappropriate shock therapy. Dr. Greg Hundley: So Michael we're working with subcutaneous ICDs. What was your study population and how did you design this study to really test the efficacy of the subcutaneous ICDs relative to the more conventional transvenous lead systems in these high-risk patients? Dr. Michael Gold: So what we did was to first identify a population and the most common population price of the implantation in the United States have for sure, in many of the countries is primary prevention patients and low ejection fraction. So we restricted the study to patients with an ejection fraction less than 35% and primary prevention, meaning have never had an episode of sustained ventricular tachycardia or cardiac arrest. So starting with that population, this was a prospective large registry. The bandwidth was out there to do a very large randomized trial, which was just done, although not as in a sicker patient population that study called PRAETORIAN was going on simultaneously. And what we decided was to be aggressive if you will. And we defined our endpoints for the study by looking at identified studies previously with transvenous ICDs that have the most contemporary programming and the lowest risk of inappropriate shock. Dr. Michael Gold: So we used the MADEIT-RIT study, which most people probably would consider it to be the ultimate, if not one of the two contemporary studies for that. And we wanted to essentially compare head-to-head results with that, obviously without those specific patients, but using that as in some ways a historical control or benchmark. So, we set what we wanted to use as our performance goal based on the MADEIT-RNT study for inappropriate shocks, as well as looked at complication rates, using performance goals that have been well-established by the FDA. And we use an 18-month time window, because that's a time when most inappropriate shocks as well as most complications will show up with this device. Dr. Greg Hundley: So what did you find? Dr. Michael Gold: So, what we found was first that we accomplished the goal of having patients who were had much more comorbidities or sicker, if you will. A majority of patients of the over 1,000 patients in the study had ischemic heart disease. I mean, ejection fraction, it was down 26%. In perspective, the early studies of the S-ICD, I mean, ejection fractions are 40% higher, almost 90% of patients have heart failure rather than a third of patients in previous studies and the incidents of diabetes, kidney disease, hypertension and other things with two to three times higher than what would have been done or shown in any early registries of the S-ICD. So we're very pleased about that. Again, there were over 1,100 patients in this study. And what we showed was that at 18 months, the inappropriate shock rate for the whole population was only 4%, hopefully 2.7% annually, which is quite low compared to any other S-ICD study and was very favorably compared with the MADEIT-RIT study easily met the performance goal for that. Dr. Michael Gold: And if we looked at that subgroup of patients who had generation three of the most modern of the S-ICD devices that have a newer discrimination algorithm, that number even became lower than that. So again, very, very reassuring to see what we found there and for the generation three device, it was only 2.9% of patients in 18 months had any inappropriate shock for that. And if we then looked at the other aspects of it, the all shock rate for appropriate and inappropriate shock was only 10% of 18 months, which meant the performance goal for that as well. And what's important about the all shock rate is that it's nice to show that the device can defibrillate a patient in the lab when you're testing them. And this was successful in 98, 99% of the time, which is typical for transvenous or subcutaneous devices. But when we looked at spontaneous arrhythmic events, it was highly effective, 100% of VT storms, and all but one case of a VTVF episode was converted with defibrillator, that one case actually spontaneously converted, but was officially listed as a failure, because it didn't convert right away. Dr. Michael Gold: But no patients had a cardiac due to ventricular fibrillation that could not be successfully treated or ended up requiring external defibrillation. And finally, despite a much sicker population, again, if you will, we showed that at 18 months, the overall complication rate from the device was only about 7% in total, which again is very low compared to what we'd expect to see with other devices. And no patient developed a bloodborne infection or a true lead failure, which is one of the major complications that, and dreaded complications of transvenous devices. Dr. Greg Hundley: Well, Sana as our editorialist, what stood out to you as being really important findings in this study? Dr. Sana Al-Khatib: First of all, I want to start by congratulating Michael and his team on the completion of this important study. And I agree that this has been a really important addition to the armamentarium of studies related to subQ-ICDs. We have had registries in the past. In fact, Michael alluded to the publication of the PRAETORIAN trial, which was really the first trial to compare the S-ICD with the transvenous ICD in a randomized control trial design. However, despite the important contributions of the PRAETORIAN trial, there were several things that remain unaddressed or unanswered in terms of the rate of the end points, especially in appropriate shocks with the newer S-ICD models. So where we had an abundance of those in the UNTOUCHED study, and then really the important question of do the results of that trial, which was largely not done in the United States, apply to the average patient, seen in the United States. Dr. Sana Al-Khatib: And then could those results be extrapolated to sicker patients, because as Michael very nicely highlighted that those patients were not as sick as patients that we see in routine clinical practice. So from my perspective, those are the findings of the UNTOUCHED study are really important, because we now have confirmation that the S-ICD is actually effective and safe in a patient population representative of the average patient seen in clinical practice in the U.S. with a primary prevention indication for the ICD, no really sicker patients, good representation of women, black patients. So overall, I think this trial really gives me a reassurance that the subcutaneous ICD is safe and effective, that the rate of inappropriate shocks with the newer generation of ICDs is really low. That said, I just wanted to highlight a couple of points that I'm hoping to see more data on as we go forward. Dr. Sana Al-Khatib: The study was only 18 months long. It would be good really to have a longer studies. Again, the average age of patients enrolled in the trial was on the younger side in the 50s, and I would love really to see more data on the S-ICD in older patients. And also as Michael said that these results were obtained in patients who had a primary prevention indication for the ICD on the basis of systolic heart failure. Well, we have other patient populations. And especially with, for example, patients with hypertrophic cardiomyopathy, where the rate of inappropriate shocks might be higher, and it would be great to see even more data as we go forward on that patient population and other subgroups of patients that were not included or not well-represented in the trial, but overall a really important trial. Dr. Greg Hundley: Very nice Sana. And Michael, just swinging back to you, what do you think are some of the next studies that might be performed in this space? Dr. Michael Gold: No, I think that Sana made some nice points. I should point out that, in its, I think appropriate vision, the FDA require as long-term evaluation of devices and studies. So the S-ICD post-approval study, which I'm fortunate enough to be involved with as well, is a five-year followup study. We've already published a little bit on two and three-year data, but we will have five-year data on this device in a patient that is closer to the S-ICD than the early registries. It was not restricted to just low EF patients, but it was a U.S.-only study of over 1,500 patients. So, a larger study that has the typical distribution of patients that we see in a U.S. practice who would be eligible for the S-ICD, those without pacing indications and both primary and secondary prevention patients. So I think that will be very important. Dr. Michael Gold: Where the technology is going is largely, there will still be iterations of longer battery life, smaller devices and those things, but there's been a real push to be able to also provide pacing therapy without the need for intravenous leads. So there are several models and systems approaches being used, including adding a lead-less pacemaker that will communicate with a subcutaneous ICD or placing a lead under the sternum, still extra vascular with the capabilities of pacing. So I think the next sort of group of trials will likely be seeing on this device other than simply looking at other populations in a little more depth will be further expansion of the technology to allow for a greater use of a device for those patients who may require a pacing. Dr. Greg Hundley: Well listeners, we want to thank Dr. Michael Gold from Medical University of South Carolina and Dr. Sana Al-Khatib from Duke University, our editorialists for describing this study. And in regards to those, or in regards to subcutaneous ICDs, providing confirmation that these devices are both effective and safe, particularly in a U.S. population for primary prevention of sudden death and really a high-risk patient population with low ejection fraction. On behalf of Carolyn and myself, we want to wish you a great week and we will catch you next week on the Run. This program is copyright of the American Heart Association, 2021.  

Petro Dobromylskyj is a veterinarian, trained at the RVC, London University. He was fortunate enough to intercalate a BSc degree in physiology into his veterinary degree. He was even more fortunate to study under Patrick Wall at UCH, who set me on course to become a veterinary anaesthetist, mostly working on acute pain control. That led to the Certificate then Diploma in Veterinary Anaesthesia and enough publications to allow him to enter the European College of Veterinary Anaesthesia and Analgesia as a de facto founding member. Anaesthesia teaches you a lot. Basic science is combined with the occasional need to act rapidly. Wrong decisions can reward you with catastrophe in seconds. Thinking is mandatory. He stumbled onto nutrition completely by accident. Once you have been taught to think, it's hard to stop.   Time Stamps:   0:08:42  Podcast Begins 0:09:52  Peter's Background 0:16:36  The Electron Transport Chain 0:45:30  Insulin resistance is a cellular antioxidant defense mechanism   https://www.pnas.org/content/106/42/17787 0:47:50 High rates of superoxide production in skeletal-muscle mitochondria respiring on both complex I- and complex II-linked substrates   https://www.researchgate.net/publication/5929069_High_rates_of_superoxide_production_in_skeletal-muscle_mitochondria_respiring_on_both_complex_I-_and_complex_II-linked_substrates 0:49:50  We need insulin resistance 0:54:53  All about a1a 1:00:37  How PUFAs affect mitochondria 1:02:40  PUFAs and cellular lipid accumulation 1:05:34  Pathological insulin sensitivity   1:07:52  Insulin-induced translocation of CD36 to the plasma membrane is reversible and shows similarity to that of GLUT4  https://www.sciencedirect.com/science/article/abs/pii/S1388198107002296?via%3Dihub 1:12:34  The center of metabolic health: the adipocytes  1:14:08  PUFAs and tumor growth 1:15:52  Eating the perfect ratio of fatty acids 1:20:18  Differential Metabolic Effects of Saturated Versus Polyunsaturated Fats in Ketogenic Diets  https://academic.oup.com/jcem/article/89/4/1641/2844241 1:20:18  How PUFAs break your metabolism 1:23:20  Premature Atherosclerosis Associated With Monogenic Insulin Resistance  https://www.ahajournals.org/doi/10.1161/01.cir.103.18.2225?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.org&rfr_dat=cr_pub%20%200pubmed 1:25:21  Role of Physiological Levels of 4-Hydroxynonenal on Adipocyte Biology: Implications for Obesity and Metabolic Syndrome  https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4038367/ 1:30:16  Lowering dietary linoleic acid reduces bioactive oxidized linoleic acid metabolites in humans  https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3467319/ 1:30:53  Context matters in interpreting lipids 1:36:49  You might want to rethink your avocado and olive oil 1:40:17  Plant are not benevolent 1:47:55  How to find Peter Dobromylskyj 1:51:01  Peter's radical running habits 1:53:54  Green tea extract only affects markers of oxidative status postprandially: lasting antioxidant effect of flavonoid-free diet   https://www.cambridge.org/core/product/identifier/S0007114502000673/type/journal_article 1:55:42  The roots of the chronic disease epidemic 1:58:23  Diet Fat Composition Alters Membrane Phospholipid Composition, Insulin Binding, and Glucose Metabolism in Adipocytes from Control and Diabetic Animals  jbc.org/content/265/19/11143.long 1:59:29 Insulin-sensitive obesity   https://journals.physiology.org/doi/full/10.1152/ajpendo.00586.2009?rfr_dat=cr_pub++0pubmed&url_ver=Z39.88-2003&rfr_id=ori%3Arid%3Acrossref.org Heart & Soil is my passion based company founded to help a few million more people reclaim their ancestral birthright to radical health through nose to tail nutrition. We are making grass fed, grass finished desiccated organ capsules from regenerative farms in New Zealand and developing a US based supply chain. Check us out at: www.heartandsoil.co, @heartandsoilsupplements on Instagram  Belcampo: www.belcampo.com Use the code “carnivoremd” for 20% off your order!  Nutrisense (Continuous Glucose Monitor- CGM): www.Nutrisense.io    BluBlox: www.blublox.com use the code CarnivoreMD for 15% off your order White Oak Pastures: Use the code CARNIVOREMD at www.whiteoakpastures.com for 10% off your first order!

Hello there, Today it's a pleasure to bring you "Smoothing Out the Wrinkles" a poem by Samantha Terrell. Samantha Terrell is an American poet raised in the Midwest, whose work emphasizes emotional integrity and social justice. Her work has been published in Dove Tales by Writing for Peace, Ebola chapbook (West Chester University), Poetry Quarterly (Pennsylvania), and many other online and in-print journals. She and her family reside in Upstate NY. You can find her on Twitter @honestypoetry. Check out Samantha's website for more of her work! https://poetrybysamantha.weebly.com/ Stop by idlewy.blog to submit your work, and to discover more stuff by and for working class creatives! Much love, j Produced by J. Alejandro @idlewy Sound effects obtained from https://www.zapsplat.com Special thanks to Dust Jones @ Speer Productions

Wes is the co-author of the successful initiative to make the complete Abolition of Abortion onto the Texas Republican Party platform. Jason and Wes discuss politics, interposition, uniting under the banner of Christ with other abolitionists and much more.

This month on Episode 7 of the Discover CircRes podcast, host Cindy St. Hilaire highlights two featured articles from the December 6, 2019 issue of Circulation Research and talks with Roy Silverstein and Yiliang Chen about their article, Mitochondrial Metabolic Reprogramming by CD36 Signaling Drives Macrophage Inflammatory Responses. Article highlights: McArdle, et al, et al. Migratory and Dancing Atherosclerotic Macrophages Skaria, et al. Cardioprotection with Endogenous αCGRP Transcript Dr Cindy St. Hilaire: Hi, welcome to Discover CircRes the monthly podcast of the American Heart Association journal, Circulation Research. I'm your host, Dr Cindy St. Hilaire, and I'm an Assistant Professor at the University of Pittsburgh. In this episode I'm going to share with you highlights from recent articles published in the December 6 issue of Circulation Research. We're also going to have an in-depth conversation with Drs Roy Silverstein and Yiliang Chen about their recent article on how macrophage CD36 modulates immunometabolism. Also, the American Heart Association Scientific Sessions were recently held in Philadelphia, PA and in this edition of Discover CircRes, we're going to feature a conversation with the editors in chief of Circulation Research and Circulation, Drs Jane Friedman and Joe Hill. The first article I'd like to highlight is titled Migratory Dancing Atherosclerotic Macrophages. The first author is Sarah McCardell and the corresponding author is Klaus Ley and the work was conducted at the La Jolla Institute of Immunology in La Jolla, California. A major component of atherosclerosis is the inflammatory response and atherosclerotic plaques contain a mix of macrophages. Some macrophages arise from proliferation of resident cells, while other macrophages can infiltrate in from the blood. And a few studies have shown that smooth muscle cells can acquire some macrophage-like markers. Some macrophages are anti-inflammatory while others are more pro-inflammatory. These variations have largely been determined using techniques that examine the cell surface marker expression, the transcription profiles, or by mass spectrometry. But how all these different types of macrophagia cells look and function in vivo has not been clearly defined nor visualized. McCardell and colleagues have now observed fluorescently-labeled macrophages in the atherosclerotic plaques of live mice. First, using single cell RNA sequencing, they identified key markers of macrophage subsets. These markers are Cx3cr1 and CD11c. They then generated Apoe knockout mice that could then express green fluorescent protein under the direction of the Cx3cr1 promoter and yellow fluorescent protein under the direction of CD11c. These fluorescent proteins could be expressed individually, they could be expressed together, or they could be expressed not at all. And then in these mice they used intravital microscopy to look at the carotid artery plaques and they found while green cells and double positive cells, so that is, cells expressing Cx3cr1or both Cx3cr1 and CD11c--these cells tended to stay in one place, but they could extrude these protrusions akin to dancing, while the yellow cells or the cells that were expressing CD11c alone were more spherical and migratory. RNA analysis revealed that migratory genes were indeed upregulated in the yellow cells as compared to the green cells. The work provides preliminary insights into plaque macrophage dynamics and presents a technical resource for investigating how such behaviors may influence disease progression and I highly recommend you check this article out online. They have included several videos in the supplementary data and they're really beautiful. You can actually see the macrophages moving around and dancing and moving through the tissue and it's really neat to think about maybe how people are going to use this in the future to study the role of macrophages and maybe even other inflammatory cells in atherosclerotic disease progression. The next paper I want to highlight is titled Blood Pressure Normalization-Independent Cardioprotective Effects of Endogenous, Physical Activity-Induced Alpha Calcitonin Gene-Related Peptide (αCGRP) in Chronic Hypertensive Mice. The first author is Tom Skaria and the corresponding author is Johannes Vogel and they are from the University of Zurich in Zurich, Switzerland. So chronic hypertension affects a ton of people, over a billion worldwide, and it is a main driver of cardiovascular mortality and morbidity and it's a leading risk for heart failure. The way chronic hypertension can contribute to heart failure is by increasing the sarcomere gene expression in cardiomyocytes. And this gene expression helps to promote cellular hypertrophy or the swelling of cells, the enlarging of cells. High blood pressure can also promote interstitial fibrosis. And this fibrosis, which is happening in between the cardiomyocytes, impairs the contractile function of those cardiomyocytes. And while there are some medications available to help treat hypertension, many patients are unresponsive to these anti-hypertensive therapies. Interestingly, we all know exercise is good for us, and we all know exercise is good for specifically our heart, but exercise itself also induces cardiac hypertrophy, but it does so without impairing cardiac contractility. So how does this do this? How does exercise cause hypertrophy, but do it without impairing contractility? One of the proteins thought to be involved is called alpha calcitonin gene related peptide or alpha CGRP and mice that have been deleted of alpha CGRP, when they exercise, they exhibit hearts that look like hypertensive hearts. So this group hypothesized that exercise-induced endogenous alpha CGRP suppresses hypertension induced pathological cardiac remodeling and they tested this hypothesis in a murine model of chronic hypertension. What they found was interesting--they found that endogenous alpha CGRP suppresses pathological cardiac remodeling and it helps to preserve the heart function and it also mediates the cardioprotective effects of regular exercise in the setting of chronic hypertension. A really interesting thing that this article highlights is that alpha CGRP is currently approved for the treatment of migraines. So what might that mean? That might mean that someone who is taking this long-term for migraines may actually carry the risk of cardiac impairment if they have chronic hypertension. Mid-November is when the annual American Heart Association Scientific Sessions are held. This year's scientific sessions were in Philadelphia, PA and the Editors-In-Chief of Circulation and Circulation Research, Drs Joe Hill and Jane Friedman, had the chance to sit down for a chat and I'm going to share with you what they discussed, and I hope you enjoy it. Here they are. Amit Khera: I'm Amit Khera. I'm Digital Strategies Editor for Circulation and I'm standing in this week for Carolyn Lam and Greg Hunley. And I'm also doing the Circ on the Run Podcast as well as Discover CircRes podcast with our two Editors-In-Chief. This is Jane Friedman, who recently took over as editor-in-chief of Circulation Research and Joseph Hill, who is the editor-in-chief of Circulation. So welcome to you both. We’re excited to do this. Dr Joseph Hill:Thank you. Dr Jane Freedman: Thank you. Amit Khera: The idea behind this, there's a session here at Sessions where we're running a little bit about Circulation Research and Circulation, pulling back the cover, if you will, and seeing behind the cloak as to what happens in the journal. So Dr Freedman, I'll start with you. Tell me a little bit about as the incoming editor of Circulation Research, some of your vision for the journal, what you're excited about. Dr Jane Freedman: Well, I'm thrilled to be the new editor of Circulation Research and I've assembled a fabulous team of associate editors, deputy editors and other staff and support that are going to continue to grow what's already a wonderful journal, to be the preeminent and primary journal for basic and translational cardiovascular sciences and also support and interact with the other AHA family of journals. Amit Khera: So obviously that starts with a great team and it sounds like you've assembled that. Anything new that you're thinking about and the redesign of Circ Research in your term? Dr Jane Freedman: So we're hoping to expand the original scientific content, so we can have a larger number of articles in original science and we can have the pages to be able to handle other areas of basic cardiovascular science to include new areas, emerging areas, things like that. We're also increasing some of our early career initiatives, so that's very important to us as well. Amit Khera: Fantastic. And you talk about expanding for science and Joe, that that leads to you. In the session tomorrow, one of the goals is when people submit their science, it really goes into a black box and people don't know what happens on the Editorial level. Can you maybe enlighten us a little what happens? Dr Joseph Hill: Jane and I had been friends for 20 or more years and we now have established a bidirectional mutually synergistic collaboration where we send papers each way. We have distinct missions but yet with significant overlap, and I think it's an incredibly exciting time for the entire portfolio of AHA journals. So as you say, most people that you hit send and you wait four to six weeks and either get a happy note or an unhappy note. And what happens at both our journals is we have a strategy of multiple touches on every paper. The paper that first comes in is first touched by a senior editor, either myself or James de Lemos and two or three others. And we will reject without review about 50% of the papers at that point. We publish six papers a week, but we get 110 a week, so we don't need to review 50 of them to pick the top six. Out of respect to our authors to save them time, out of respect to our reviewers who devote tremendous effort to reviewing papers, we don't send them papers that we don't think have a shot. That said, if a paper makes it past that first stage, there's about a 50% chance it'll get published either in our journal or in one of the subspecialty journals. Probably a 50/50 chance it'll be published somewhere in an AHA family journal. So if it makes it past that stage, we send it to an Associate Editor, of which you are one, and we have about 50 of them. A third are in Dallas, another third is in the US outside of Dallas, and another third are in countries around the world, 17 different countries. And that person will probably reject without review another 5% or 10% maybe, but he or she will dig into that paper and, in parallel send it out to two or sometimes three reviewers, who are trusted and valued advisors. They help that associate editor make a strong recommendation. He or she makes a decision to bring to the larger group that is informed by those reviewers. So already that paper has been touched by five different investigators. Typically, that associate editor will reach out electronically within his or her affinity group. We have an affinity group in epidemiology, heart failure intervention, basic science… asking other AEs, "Could you take a look at this paper? One reviewer said this, one said that, I'm sort of thinking this…" And then we'll have a conversation on our weekly video conference and then a decision goes out to the authors. So every paper is touched by at least five and sometimes 10 different editors and reviewers, which we have found has been a powerful way to really dig into and identify things that one or two people might've missed. Amit Khera: You know, one thing I note here is how many people touch these articles, yet how efficient and how fast this process is. And that's a testament to the goals of the journal to be really responsive and rapid for our authors. One big part of that, and I'll come back to Dr Freedman, is peer review, right? So associate editors have a lot of work and we're affinity groups and so forth, but really critical is these peer reviewers, and in the modern era we're all so busy. Tell us a little bit about the value of peer review and how we enhance the value to the peer reviewers themselves. Dr Jane Freedman: Just as you said, the peer reviewers are absolutely central, valued, and vital parts of making the journal run correctly and we, like Circulation, our associate editors send them out to three different peer reviewers and they have a very fixed amount of time to review the articles and they provide these wonderful comments. We also very heavily rely on our Editorial Board. They know the drill that it needs to be back within a fixed amount of time and for the most part they do it. It's an interesting question. What's the value to them? I've been a reviewer too. It's part of your payback. It's part of educating yourself about what's new and interesting. There's a lot of reasons for doing it. People enjoy being on the Editorial Board and interacting with the journal. But fundamentally, as an editor, you're incredibly grateful to your reviewers. They are the unsung heroes of making a journal work. Amit Khera: And you mentioned sending out to three. When you have disparate reviews, it's amazing when some people love it and some people hate it. Dr Jane Freedman: Yeah. Amit Khera: How do you handle that? Dr Jane Freedman: Yeah. Well, sometimes it's apparent from the reviews why that happened. Someone may have focused on something that the editorial group thinks is less important or they focused on something that's addressable. The other thing we do similar to Joe, is we have a video conference call every single week on Wednesdays, and that's a period where people can vet any concerns or questions. And then my editors, my associate and deputy editors, know we have an open communication at all times. So I very frequently, when they have questions about reviews and how to reconcile disparate reviews, we'll have an ongoing conversation about that. Amit Khera: It sounds like of course you're actively engaged in how this is a dynamic process. I mentioned one thing as digital strategies editor, and I know both at Circ Research and Circulation, I was thinking how do we bring these articles to life? How do we have the most people read them or engage with them? And one is traditional social media, so Twitter and Facebook, which is incredibly important. Podcast, we have a monthly podcast, we have a weekly podcast, and really hope that people listen to them because they're really full of important information. And finally, I think what people don't appreciate is the media. So we work with AHA media. Some of our top stories get over a million media impressions, go all around the world and there's Professional Heart Daily. So there's so many ways that we're bringing articles to life. Joe, I'm going to finish with you. This is a Circ family. The value of having a family of journals and how we keep cohesion and for authors when they're submitting to serve a family of journals. What's the value and how does that add? Dr Joseph Hill: Well, there has been complete turnover of all the Editors-In-Chief in the entire family of journals, of which there are 12. And we are all quite similar in our personalities, in our perspectives on the importance, the ultimate importance of validity. The first question we ask is this true? If it's not, it's gone. It doesn't get referred. We reject it. Even if it's going to be on the front page of the New York Times and cited 10,000 times. And all of us hold ourselves to that same standard. So our vectors are all pointed in the same direction. We also care about impact, not impact factor, but does it change the way you think? Does it matter? Is it incremental or does it really move the needle? So we are now in a situation, I think, a wonderful situation where we all sink or swim together. We send papers all around, as you know very well. We send papers to the sub-specialty journals, we send 20 or 30 a week on an extraordinarily regular basis, and we send papers horizontally to Circ Research or Hypertension or Stroke and so forth. So it is a syncytium now I would say of a family of journals where we are all looking out for each other. Jane cares about our journal and we care about her journal and that's a really a wonderful situation to be in. Amit Khera: Well thanks. That family and how this fluidity of articles and thought and exchanges is really part of the value and ultimately the goal is for a great paper to find a great home and I think in the Circ family we do that. Dr Cindy St. Hilaire: Great. So I'm here with Drs Roy Silverstein and Yiliang Chen and today we're going to be discussing their paper titled Mitochondrial Metabolic Reprogramming by CD36 Signaling Drives Macrophage Inflammatory Responses. And this article is in the December 6th, 2019 edition of Circulation Research. So thank you both for joining me today. I'm really looking forward to learning more about the study, but before we really dig into it, could you please introduce yourselves and maybe give us a little bit about your background? Dr Roy Silverstein: Hi, I'm Roy Silverstein. I am a physician scientist, chair of the department of medicine at Medical College of Wisconsin in Milwaukee and also a senior investigator at the Blood Research Institute, which is part of what is now called Versiti Blood Center of Wisconsin. I'm a hematologist. Dr Yiliang Chen: Hi, my name is a Yiliang Chen. I graduate a PhD from University of Toledo, Ohio State. Then I chose to join Roy Silverstein's lab because I'm fascinated with this macrophage biology and immune functions in a disease called atherosclerosis, which is well-known inflammatory diseases. Dr Roy Silverstein: Can I make a little note that Dr Chen is currently supported by a scientist development grant from the American Heart Association, which is I think a nice tie-in? Dr Cindy St. Hilaire: Yeah. Dr Yiliang Chen: Yeah. I want to take this opportunity really saying American Heart Association to support our research. Dr Cindy St. Hilaire: Well that's wonderful. And now we get to publish this beautiful story. So it's come full circle. So you stated the objective of this paper was to investigate the mechanisms by which dyslipidemia, oxidative stress, and macrophage activation are linked in athero. And you focused on immunometabolism and you also focused on a protein called CD36. So before we get too deep in the weeds, can you give us a short little primer on what is immunometabolism in the context of athero, and also maybe a little bit about the molecule CD36? Dr Roy Silverstein: Well let me take the CD36 piece and then I'll let Dr Chen take the immunometabolism piece. So CD36 is a protein that's expressed on quite a few different cell types. We think that on muscle and fat its main purpose is to translocate free fatty acids from the external environment into the cell. In the case of fat, for storage, and in the case of muscle, for beta oxidation and energy, but in macrophages and immune cells and platelets, it has a different role. It serves as a scavenger receptor, part of the innate immune system, and it recognizes structures that we call DAMPs-danger associated molecular patterns. And the specific DAMPs that are recognized include oxidized low-density lipoprotein or what we call Ox-LDL. Dr Cindy St. Hilaire: Okay, so now could you give us a little bit about immunometabolism? Dr Yiliang Chen: Sure. So for metabolism, especially the process related to ATP or energy production, normally we call it bioenergetics and it is import. For so many years, people understand for the immune cell to get activated, they may produce proteins or somehow sometimes they need to proliferate. So there's a lot of energy is required during this whole process, right? But the old dogma is that the metabolism is only activated just to support the production of energy, especially the ATP. Right. But the emerging evidence has shown that, actually it's not that simple. For example, if you use LPS or bacterial product to activate to M1 status, the cells mainly use the glycolysis. They don't use the mitochondria Ox-LDL so TCA cycle to produce ADP. While the M2 activation is total different story. They switch it to the mitochondria ATP production and not using the glycolysis and it seems the metabolism is the underlying mechanism that driving these immune activations of the macrophages. So we want to ask, what kind of metabolism is going on in those ox-LDL-stimulated macrophages and is it related to atherosclerosis? So finally we figured out, okay, everything is focusing on a mitochondria function, which is interesting. But in our situation, very interesting, we find when the cells treat with oxidized LDL, actually they largely shut down the mitochondria OXPHOS. Then the Mito can switch ROS production of reactive oxygen species in shall we call it ROS. So that makes things quite interesting because it is well known oxidative stress is commonly observed during atherosclerosis. And also the mitochondria dysfunction actually, they are also commonly observed in the human patients with cardiovascular diseases. That kind of thing, everything together. Dr Cindy St. Hilaire: So you found that fatty acid metabolism, which is induced by this oxidized LDL, leads to the metabolic shift in the mitochondria, this switch you just described. And that shift leads to an accumulation of long chain fatty acids, but you also noticed independent of the metabolism in the mitochondria, you notice dysfunctions in what you call the mitochondrial network, and I'm wondering is it the accumulation of these long chain fatty acids that drives alterations in the mitochondrial network, or is it the other way around? I guess what I'm curious about is the interplay between that metabolic shift and just the baseline function of the mitochondria. Is one causing the other? Is it bi-directional? Dr Yiliang Chen: It sounds like a chicken and egg question. Dr Cindy St. Hilaire: Exactly. Dr Yiliang Chen: I think it's not that simple. For me, I was saying initially the cell try to adapt to this oxo LDL microenvironment. They try to stimulate the fatty acid trafficking into the mitochondria. But a side effect, what we think is, when you shut down a fatty acid oxidation while you're trafficking them there, that naturally will lead to accumulation of fatty acid. Those lipids may very well insert into the inner membrane of mitochondria then leads to the defects and that pretty much explained the EN images we show in our paper. Yeah. Exactly. Dr Cindy St. Hilaire: So the fact that the cell can't break down these long chain fatty acids, they're accumulating and potentially disrupting the mitochondrial membrane integrity. Dr Roy Silverstein: It's a form of lipodystrophy, not lipodystrophy, but lipotoxicity. Dr Yiliang Chen: Essentially, I think this is a defect in metabolism that leads to chronic inflammation. Dr Cindy St. Hilaire: Yeah. So your study focused on macrophages and atherosclerotic plaques and there's a huge body of evidence that shows inflammation and macrophage contribute to atherosclerotic disease progression pretty much throughout the whole plaque development. But there's also a body of evidence that shows smooth muscle cells can transdifferentiate and acquire macrophage-like phenotypes and they can also express CD36. It's one of the markers that people look for in that. And so I'm wondering, do you think this metabolic shift is operative in the smooth muscle or the macrophage-like smooth muscle cells? I guess that's the better thing to call them. Do you think that metabolic shift is operative and is contributing more so to the plaque or do you think this is innate to the macrophage from the immune system? Dr Roy Silverstein: I think that's a very provocative question. Thank you for it. Our in vitro experiments would not answer that question yet. We'd start with macrophages in those experiments, but it would be very interesting to look at smooth muscle cells that have been pushed towards that phenotype. We do, however, have some in vivo data that suggests that the cells that we call macrophages are behaving this way. And we can't say for certain that those are haematopoietically-derived cells versus smooth muscle derived cells. Dr Cindy St. Hilaire: So your study focused on the role of CD36 on macrophages, however, you mentioned in your introduction at the beginning that CD36 has also, it's in the GI track, it's on the muscle cells, it's on adipose tissue. It's also on the skin. And I'm wondering if you think these findings are specific only to the resident macrophages in the plaques or is this a broader function in macrophages? And I guess I'm thinking of this in the context of your study because it used the Apoe CD36 double knockout. So these are full body knockouts missing Apoe and functional CD36. And so I'm wondering, I guess, what would happen if CD36 was only removed from the macrophage cells itself? And I guess I'm thinking about this in context of something like metabolic syndrome. Could this be operative in adipose cells expressing CD36 or muscle cells or something like that? Kind of a more speculative question. Dr Roy Silverstein: That's great. You're helping us write our next grant. Dr Yiliang Chen: Yeah. Great question. Dr Cindy St. Hilaire: Give me 10%. Give me a little bit. Dr Roy Silverstein: I think one of the things that we've found over the years is that CD36 signaling in response to DAMPs, and even in response to fatty acid, involves a generalizable pattern that involves recruitment of what you might call a signalosome inside the cell. That signalosome typically would include members of the SARC family kinases, specific map kinases, a guanine nucleotide exchange factor called VAV, and other downstream signaling complexes. So we believe that that creates some opportunity for context specific signaling, but it does seem that a common theme is the generation of intracellular reactive oxygen species. Dr Cindy St. Hilaire: So I guess the bigger question after all of this, after your great findings, is what is the potential to leverage these findings in terms of developing therapies? Is there a novel pathway we can start to target or to think about targeting and how would you kind of go about that? Dr Yiliang Chen: Yeah. For that, that is our ongoing investigation. So based on the story in this paper, we are saying the mitochondria dysfunction and ROS production will activate and be pathway and drive this chronic inflammation, right? So if this is true, the particular question we are asking now is can we find a way to suppress mitochondrial ROS production or find a way to correct this fatty acid defect? Dr Cindy St. Hilaire: Do you think your findings on the metabolic shift of macrophage and how those contribute to atherosclerosis, how do you think those findings inform what the Cantos trial showed and the Cantos trial, which people may not be familiar with, used immunomodulation. Essentially, it was an antibody to block IO1 beta signaling and it had mediocre affects. The MI numbers were down, but the death rates were the same. Do you think that targeting the metabolism of the immune cells specifically as opposed to targeting the inflammation pathway outside of the cells is a more targeted and therefore maybe more precise approach? Dr Roy Silverstein: Yeah. I think that's a good observation. In my mind, what the Cantos study really showed us is that blocking inflammation in the broad sense, in a very upstream sense, can have an impact on human atherosclerotic heart disease. And I think that's really important observation and it validates the concept of inflammation as a target for atherosclerosis. Dr Cindy St. Hilaire: All of us breathed a sigh of relief once we're okay. It is an inflammation disease. Dr Roy Silverstein: Yeah. And it also decreased cancer, right? So it's double victory. Dr Cindy St. Hilaire: Yes. It did. Exactly. Yeah. Dr Roy Silverstein: But my thought is that we could maybe get a little bit upstream of that in a specific way. And perhaps the most translatable discovery here is the importance of mitochondrial reactive oxygen species as the source. Most people have looked at the NOX pathways, the NADP H oxidase pathways or broad spectrum. Dr Cindy St. Hilaire: I looked at that in my graduate studies. Dr Roy Silverstein: Yeah. So, you know that literature, and using broad heavy-handed approaches to create a “antioxidant effect” and most of those clinical trials have been extremely disappointing. But they haven't really targeted specific reactive oxygen species or specific sources. And we have this inhibitor that Dr Chen used in his experiment called mito-TEMPO which targets the mitochondria through a molecular mechanism and had a significant impact on the downstream production of pro-inflammatory product. So we think, or I think at least, that that is potentially an interesting target to basically prevent that reversal of mitochondrial function. Dr Cindy St. Hilaire: Thank you so much for taking the time to speak with me today. It's been wonderful, and I look forward to reading more of your papers in the future. Dr Roy Silverstein: Thanks. Dr Yiliang Chen: Thank you. Dr Cindy St. Hilaire: That's it for highlights from the December 6th issue of Circulation Research. Thank you so much for listening. This podcast is produced by Rebecca McTavish, edited by Melissa Stoner, and supported by the editorial team of Circulation Research. Some of the copy text for the highlighted articles is provided by Ruth Williams. Thank you to our guests, Dr Roy Silverstein and Dr Yiliang Chen and Drs Jane Friedman and Joe Hill for sharing their discussion with us. I'm your host, Dr Cindy St Hilaire, and this is Discover CircRes, your source for the most up-to-date and exciting discoveries in basic cardiovascular research.  

Sometimes, Free Speech requires a price to be paid. As a result, in today's polarizing world climate we see violence against political activists, government censorship, self-censorship, "Cancel Culture", etc. Join the Carolina Desis this week as they discuss their observations on these costs, their effects on social media, and why it can still be worth saying what's on your mind in the present day. As always, thank you Yash Mistry for editing this month’s episode!

Previously seen as an older man’s disease, increased rates of prostate cancer in younger men has been associated with an increased incidence of obesity. “There are a number of changes that happen in the body when you have obesity, that could lead to the progression of cancer,” says Professor Watt, Head of the Department of Physiology, School of Biomedical Sciences at the University of Melbourne. “Essentially what we’ve found is that, unlike other cancer types, which rely heavily on glucose to fuel both their growth and their proliferation, prostate cancer cells are very highly reliant on fatty acids.” Working with researchers at Monash University, Professor Watt’s group have shown that they can block the capacity of these prostate cells to take up fatty acids which slows their growth dramatically. “We think this has very important implications in understanding both the progression of the disease, but also ways in which we might be able to therapeutically target prostate cancer in the future.” Episode recorded: May 6, 2019. Interviewer: Dr Andi Horvath. Producer, audio engineer and editor: Chris Hatzis. Co-producers: Silvi Vann-Wall and Dr Andi Horvath. Banner image: Shutterstock.

Dr Carolyn Lam:                Welcome to Circulation on the Run, your weekly podcast summary and backstage pass to the journal and its editors. I'm Dr Carolyn Lam, Associate Editor from the National Heart Center and Duke National University of Singapore.                                                 Is there a unique lipoprotein profile for incident peripheral artery disease as opposed to coronary or cerebral vascular disease? Well, you're just gonna have to wait for our feature discussion to find out. That's coming right up after these summaries.                                                 Our first original paper this week tells us that gene variance known to be associated with idiopathic and peripartum cardiomyopathy are also associated with preeclampsia. First and corresponding author Dr Gammill from University of Washington and colleagues studied 181 participants with confirmed preeclampsia from the Preeclampsia Registry in BioBank. Saliva samples were collected for DNA isolation and whole exome sequencing was performed to detect rare variants in 43 genes known to be associated with cardiomyopathy.                                                 Results were compared with data from two controlled groups, unrelated women with a gynecological disorder, sequence using the same methods and instruments, as well as published variant data from 33,000 subjects in the Exome Aggregation Consortium.                                                 The results showed that women who developed preeclampsia are more likely to carry protein altering mutations in genes associated with cardiomyopathy, particularly, the TTN gene which encodes the sarcomeric protein titin. Thus, detecting these gene variants may allow more specific diagnosis, classification, counseling and management of women at risk.                                                 Prior trials have shown that nonsteroidal anti-inflammatory drugs or NSAIDS confer cardiovascular risk. Now this has been postulated to be due to enhanced formation of methyl arginines in the kidney that would limit the action of nitric oxide throughout the vasculature. However, the next original paper in this week's journal suggests that this may not be correct. First author, Dr Ricciotti, corresponding author, Dr FitzGerald from University of Pennsylvania Perelman School of Medicine and colleagues, used multiple genetic and pharmacological approaches to disrupt the COX 2 pathway in mice and analyze plasma from patients taking NSAIDS.                                                 However, they did not observe an increase in methyl arginines. In contrast, they did observe an increase in plasma asymmetric dimethylarginine or EDMA in mice-rendered hypertensive by infusion of angiotensin II at a dose that also caused renal impairment. After a four week washout period following the infusion of angiotensin II, blood pressure, creatinine, and ADMA levels all fell back to normal levels.                                                 Celecoxib-treated mice also exhibited increased ADMA and plasma creatinine in response to infusion of angiotensin II and their levels also returned to normal thereafter. Thus, it seems likely that the previous reported elevations in ADMA reflected renal dysfunction rather than a direct consequence of COX 2 deletion or inhibition. The authors end by suggesting that the most plausible mechanism by which NSAIDS confer a cardiovascular risk, is by suppression of COX 2 derived cardioprotective prostaglandins such as Prostacyclin rather than by enhanced formation of methyl arginines.                                                 The next original paper identifies new targets with the potential to prevent vascular malformations in patients with hereditary hemorrhagic telangiectasia. Co-corresponding authors, Dr Ola and Eichmann from Yale University School of Medicine and colleagues looked at SMAD4, which is a downstream effector of transforming growth factor-beta/bone morphogenetic protein family ligands that signal via activin-like kinase receptors.                                                 The authors generated a tamoxifen inducible postnatal endo-fetal specific SMAD for a mutant mouse and showed that SMAD4 prevented flow-induced arterial venous malformations by inhibiting casein kinase II. The uncovered pathways provided novel targets for the treatment of vascular lesions in hereditary hemorrhagic telangiectasia related juvenile polyposis patients carrying SMAD4 mutations.                                                 The next original paper provides important data for the accurate diagnosis of long QT syndrome. Long QT syndrome can be a challenging diagnosis partly because the optimal method for QT assessment is not unequivocally established. QT experts advocate manual measurements with a tangent or threshold method.                                                 In today's paper, first and corresponding author, Dr Vink from Academic Medical Center University of Amsterdam and colleagues, aimed to assess similarities and differences between these two methods of QT interval analysis among 1,484 patients with a confirmed pathogenic variant in either KCNQ1, KCNH2 or SNC5A genes from 265 families. Both QT measurement methods yielded a high inter and intra reader validity and a high diagnostic accuracy.                                                 Using the same current guideline cutoff of QTC interval 480 milliseconds, both methods had similar specificity but yielded a different sensitivity. QTC interval cutoff values for the QT measured by the tangent method was lower compared to that measured by the threshold method. Plus, values were different depending on the correction for heart rate, age, and sex.                                                 The authors provided an adjusted cutoff values specified for method, correction formula, age, and sex. In addition, a freely accessible online probability calculator for long QT syndrome at www.QTcalculator.org has been made available as an aid in the interpretation of the QT interval.                                                 The next original paper demonstrates for the first time that thrombin mediated signaling may play a role in diet-induced atherogenesis. Co-first authors, Dr Raghavan and Singh, corresponding author Dr Rao from University of Tennessee Health Science Center and colleagues, used a mouse model of diet-induced atherosclerosis and molecular biological approaches and explored the role of thrombin and its G protein coupled receptor signaling in diet-induced atherosclerosis.                                                 They found that thrombin-induced CD36 expression and foam cell formation required protease activated receptor 1, G alpha 12, Pyk2, GAB 1, and protein kinase C theta dependent activating transcription factor 2 activation. Thus, inhibition of thrombin G protein coupled receptor signaling could be a promising target for the development of new drugs in reducing the risk of diet-induced atherogenesis.                                                 The next study provides insights into the long- term association of LDL cholesterol with coronary heart disease mortality in individuals at low tenure risks of atherosclerotic cardiovascular disease. First and corresponding author, Dr Abdullah, from VA North Texas Medical Center and UT Southwestern Medical Center and colleagues studied more than 36,000 subjects in the Cooper Clinic Longitudinal Study cohort who are at low tenure estimated risk of atherosclerotic cardiovascular disease. In other words, a low tenure risk of less than 7.5%. They've followed these patients for more than two decades.                                                 Results showed that LDL cholesterol and non-HDL cholesterol at or above 160 milligrams per deciliter were independently associated with a 50 to 80% increased relative risk of cardiovascular disease mortality. The associations between LDL cholesterol and cardiovascular disease mortality were more robust when follow up was extended beyond the traditional 10 year estimated risk period.                                                 The associations remain significant in those with an estimated tenure atherosclerotic cardiovascular disease risk of less than 5%. These data suggests that LDL cholesterol levels at or above 160 milligrams per deciliter in individuals deemed to be at low tenure atherosclerotic cardiovascular risk are associated with worse long term cardiovascular disease mortality. These findings, along with other observational data and data extrapolated from clinical trials, support further consideration of appropriate LDL cholesterol thresholds for lipid lowering interventions in individuals categorized as low short-term risk.                                                 The final paper this week uncovers a novel therapeutic target for the prevention and treatment of thoracic aortic aneurysms. First author, Dr Nogi, corresponding author Dr Shimokawa from Tohoku University Graduate School of Medicine and colleagues, used genetically modified mice to show a pathogenic role of the small GTP binding protein, GDP dissociation stimulator in the development of angiotensin 2 induced thoracic aortic aneurysms and dissection. Down regulation of this protein contributed to dysfunction of aortic smooth muscle cells and hence oxidative stress, and matrix metalloproteinase activities in the pathogenesis of thoracic aortic aneurysms and dissection.                                                 Local over expression of this small GTB binding protein GDP dissociation stimulator around the thoracic aorta inhibited aortic dilatation and rupture in deficient mice. And that wraps it up for this week's summaries. Now for our feature discussion.                                                 Atherosclerosis has been considered a systemic process, meaning that when we see a disease in one vascular bed, we assume that that's a risk marker for disease in other vascular territories, and that they share pathophysiology, they share risk factors. However, if we think about it, the prior studies have all been sort of focusing on coronary and cerebral vascular disease, but today's feature paper changes that a bit because it addresses a key knowledge gap in peripheral artery disease risk, and interestingly suggests that there may be a unique lipid profile that's related to peripheral artery disease.                                                 This is gonna be an exciting discussion and I have the first author, Dr Aaron Aday from Vanderbilt University Medical Center currently. We have our editorialist, Dr Parag Joshi from UT Southwestern, and our associate editor, Dr Anand Rohatgi from UT Southwestern. Welcome gentlemen and Aaron, could we start with you sharing about your study? Dr Aaron Aday:                 So, as you mentioned, a lot of the previous epidemiologic data on atherosclerosis have been primarily in coronary artery disease and stroke, and when we looked at peripheral artery disease or PAD, there seemed to be some subtle differences. So for instance, total cholesterol on HTL cholesterol seemed to be the strongest risk factors for future peripheral artery disease and in terms of LDL cholesterol, the data are somewhat mixed. Some have found a weak association, some have actually found no association. And so building on that, we wanted to see if using nuclear magnetic resonance spectroscopy, we could elucidate more details about the litho protein pathways associated with peripheral artery disease.                                                 And we did this in the women's health study which is a prospective cohort study of women free of cardiovascular disease, the baseline, they were aged 45 and older. And what we've found in terms of the standards with their profiles, we again found that there was no association between LDL cholesterol and future peripheral artery disease, whereas certain standard lipid measures like HDL cholesterol were strongly associated with PAD, and then using the Endemol spectroscopy tool, we found that actually, small LDL particles and total LDL particles were concentrations of both of those markers, were strong risk factors for future PAD and other measures like total HDL particle concentration were even more strongly associated with future PAD than coronary artery disease.                                                 So essentially the signature associated with future peripheral artery disease, had some important differences than that for a composite of coronary artery disease and stroke. Dr Carolyn Lam:                Aaron thanks for that. That's beautifully described and just so intriguing. Parag, could you tell us how should we be thinking about results like this? Dr Parag Joshi:                   It's a great paper and it really highlights a new and unique approach in that we ... Peripheral artery disease as an isolated incident event is fairly understudied I guess we could say and so, this is a really nice paper to start choosing out some of the risk factors for that. I think overall, when we think of peripheral arterial disease in general, I think historically, we've thought of it as similar pathophysiology, you know LDL particles and perhaps other particles depositing in the arterial space. But this does highlight some important differences that might exist and I think one of those seems to be that maybe this is more a signature of elevated remnant lipoproteins or triglyceride rich remnant lipoproteins, small dent LDL particles, low HDL, that sort of metabolic syndrome type patterns that we look at as a high risk factor that may be more contributory to peripheral artery disease than coronary disease, or at least more specific to peripheral artery disease.                                                 I guess one of my main questions about that from your work Aaron is, how can we be sure this isn't just a pre-clinical marker of diabetic patients which we know have this type of pattern? Dr Aaron Aday:                 Sure, it's certainly a possibility. I think what's notable in the cohort, at least a time enrollment. And there was a very little diabetes and actually there was a much greater prevalent of metabolic syndrome. So in my mind, it may be more of a metabolic syndrome specific marker rather than necessarily down the diabetes pathway, but it's certainly something that needs to be explored further. Dr Parag Joshi:                   I wonder whether women's health studies such a healthy cohort that I wonder if this is picking up some signal before the answer to diabetes or as you said, metabolic syndrome, you know which certainly suggests an insulin resistance pattern and we know the association of diabetes with peripheral artery disease is stronger and so I wonder if this may be a sort of earlier way of picking that up. Dr Aaron Aday:                 It may be. I think one thing to notice is the outcome of peripheral artery disease that we're using. So it is symptomatic disease. So, we're not picking up a lot of ulcers that are developing in the future, it's more the claudication and then people who've undergone revascularization. Certainly diabetics have both of those as well but I think that may suggest it's not fully unexplained by developing diabetes than peripheral artery disease further down the line. Dr Parag Joshi:                   Yeah that's a great point. Dr Carolyn Lam:                Yeah great questions, great thoughts. Anand, what about you? Did you have questions too? Dr Anand Rohatgi:            I think from my perspective and thinking about it for circulation and its readership, we found this really interesting for several reasons. Number one, I think is, as you all have discussed, peripheral arterial disease just is not as well characterized and you can see that here in over 25,000 people, add about a 100 a bed, so I think in younger folk, it takes a lot of people to study, to be able to really understand kind of the pathophysiology of peripheral arterial disease.                                                 The other thing that they think they really shed some light on is how this is happening in women in particular and in women, of course as we know have been understudied in all cardiovascular diseases, but in particular, diseases like this which are less common. It's really insightful to see that these lipid abnormalities in women are contributing to peripheral arterial disease more so than your typical LDL cholesterol management and interestingly enough, most of the women who had PAD events in this study, did not have other cardiovascular events.                                                 They really just had PAD events exclusively and I thought that was really intriguing, and the use of this advanced lipoprotein testing, this NMR modality has been very useful in terms of biology and research, and I think that's the case here where we really go under the hood Carolyn, as you said, and get kind of deep dive, the lipid metalobles on abnormalities. And I think Parag and Aaron hit the nail in the head that this is really capturing an insulin resistance of phenotype and what I really liked about this is, instead of studying people who are 70, 80 years old and a lot of things are sort of clustering, a lot of diseases are clustering and they're manifesting all at the same time, it's very hard to tease apart the effective age.                                                 Here, we captured women in their 50s and middle aged, just as they have kind of gone through menopause and this adverse metabolite's phenotype starts to rise in women. And then we could follow them over time and see what the natural history of that is, and the women who have this phenotype go on to have this devastating consequence, this peripheral arterial disease. One of the questions I had then, Aaron for you is, what do you think the implications are from these findings? Does it mean that in terms of diagnostics, we should be doing more advanced testings looking at LDL and HDL type particles with NMR or some other mortality? Does it change therapies with new therapies beings studies right now? What do you think the implications are from your work? Dr Aaron Aday:                 That's important right. I think you mentioned this and I see the inter marked tool in this study, is really a way to try to dig further into the biology of peripheral artery disease as a form of atherosclerosis. I think that we already know patients who are extremely high risk or PAD, those are patients with diabetes, smoking history, metabolic syndrome et cetera., and as you can see in a patient population in 28,000 middle aged women who are pretty healthy, we only had just over a 100 PAD events.                                                 So, I think even if you were to scale this up in terms of cost, I'm not sure that that would necessarily be a viable option for patients, but I think it does suggest that truly focusing on LDL in a very high-risk patient population, meaning patients with PAD, or we may not be fully addressing their risk. And so I think this is a need to highlight that important gap, think about other therapeutic options and we'll soon have ongoing trials, triglyceride low in therapy that may be particularly beneficial in this patient population and so that's how I see this being used. Dr Anand Rohatgi:            That makes a lot of sense and particular because in middle aged women like this, your standard risk score algorithms will not really capture that they're at increased risk, even if they smoke, just because they're women and they're younger and so, I think this really is a call to arms to more refined risk assessment in these women. Dr Parag Joshi:                   Aaron, do you think there's actually a difference in the biology in the peripheral arteries compared to the coronary and cerebral vascular beds, or is there data to kind of look at that or maybe histopathological data to look at that? Dr Aaron Aday:                 We know there's a lot of overlaps, so I don't wanna suggest that PAD is not a former atherosclerosis. I think one limitation is that the primary animal model for PAD is the hyperCKemia model. That doesn't fully recapitulate what's happening in a limb with PAD and so I think that has been one limitation in understanding the biology. But I think what we're starting to see in some clinical trials that have come out in the last couple of years or starting to see a somewhat different signal for therapies in patients with PAD so for instance, in 48, we actually saw that there was a greater benefit to LDL lower [inaudible 00:21:00] inhibitors than for coronary disease. We now have the compass trial results, again, more events, higher risk among these patients but for their benefit, add on River Oxodine therapy, we've seen lymph events or lymph signals in the SGLP2 inhibitor trials. So, I think we're starting to get a sense that there may be something else on top of the traditional ascariasis biology that may be a potential target on down the road. Dr Parag Joshi:                   I think it's really a fascinating biological question of how these different territories might actually differ in their pathophysiology. I think it's a really a nice time to look at this. Also I think, Anand and Aaron both mentioned ongoing trials. The omega 3 fatty acid trials I think reduce it, will be soon to be presented and hopefully published in the next month or so. It would be nice to see if they evaluate peripheral events in that group, I'm sure they will. Dr Carolyn Lam:                Indeed, these have been just such great thoughts and discussion. Nothing really much to add there. I suppose I could say something cheeky like for the first time, and I never thought I'd say it on the podcast, I feel kind of bad that there are no men included in this trial but anyway, I just learnt so much from this. I just wanna thank you gentlemen for a great discussion.                                                 Thank you, listeners, for joining us today and don't forget to tune in again next week to Circulation on the Run.

Carolyn:                               Welcome to Circulation on the Run, your weekly podcast summary and backstage pass to The Journal and its editor's. I'm Dr. Carolyn Lam, associate editor from the National Heart Center and Duke National University of Singapore. Our Journal this week features important new data telling us that a common genetic variant risk score is associated with risk of drug induced QT prolongation and torsades de pointes.                                                 First, let's give you your summary of this week's journal. The first paper provides both clinical and experimental data to show that the adipokine, retinal binding protein four promotes atherosclerosis. First author, Dr. Liu, corresponding author, Dr. Xia and colleagues from Sun Yat Sen University in Guangzhou, China first evaluated the association between serum retinal binding four levels and the incidents of adverse cardiovascular events in a community based prospective cohort and then examined the effects of retinal protein four gain or loss of function on macrophage foam cell formation and atherogenesis in an apple lipase protein E deficient mouse model. They found, in the clinical cohort study, that base line serum retinal binding protein four level was an independent predictor of incidents of adverse cardiovascular events after adjustment for traditional risk factors.                                                 In the experimental study's, they showed that retinal binding protein four promoted macrophage derived foam cell formation through the activation of scavenger receptor CD36 mediated cholesterol uptake. In turn dependent on June and terminal kinase and signal transducer and activator of transcription one, as well as upstream regulation by the tracing kinase CSRC. These findings, therefore, support the use of retinal binding protein four as a novel biomarker for the prediction of cardiovascular risk. The data also provide insight into the mechanism of action of retinal binding protein four in the path of physiology of atherosclerosis.                                                 The next paper is the first clinical trial, looking at remote ischemic pre conditioning prior to carotid artery stinting in patients with severe carotid artery stenosis. Remote ischemic pre conditioning is a protective, systemic strategy by which cycles of bilateral limb ischemia are applied briefly to confer protection from subsequent severe ischemia and distant organs. First author, Dr. Zhao, corresponding authors, Dr. Ji, and colleagues from Xuanwu Hospital, Capital Medical University in Beijing, China performed a proof of concept, single center, prospective, randomized control trial to assess whether remote ischemic preconditioning was safe and effective in attenuating ischemic injury related to carotid artery stinting in 189 patients with severe carotid artery stenosis. Results show that daily remote ischemic pre conditioning for two weeks, prior to carotid artery stenting, was feasible, safe, well tolerated, and may effectively attenuate secondary brain injury as evidence by a decreased incidence and reduced volumes of new ischemic legions on magnetic residence imaging performed within 48 hours post operation. The clinical implications are that if results are confirmed by future, larger studies, remote ischemic preconditioning may evolve into a nonpharmacological, neuro protective method for inhibiting carotid artery stenosis related cerebral ischemic events.                                                 This potential for clinical translation in discussed in an accompanying editorial by Doctors Bell and Yellen, from University College, London.                                                 The final paper discusses firefighting and the heart. What's the link? Well, cardiovascular events are the leading cause of death amongst firefighters and the risk is known to be substantially increased during fire suppression duties. In the current study, first author Dr. Hunter, corresponding author, Dr. Mills, and colleagues from University of Edinburgh in United Kingdom sought to understand this link better by assessing the effects of simulated fire suppression on measures of cardiovascular health in an open label, randomized cross over study of 19 healthy firefighters. These firefighters performed a standardized training exercise in a fire simulation facility or like duties for 20 minutes. Following each exposure, ex vivo thrombus formation, fibrinolysis, platelet activation and for armed blood flow in response to intra-arterial infusions of endothelium dependent and independent vasodilators were all measured. The authors found that exposure to extreme heat and physical exertion during fire suppression activated platelets, increased thrombus formation, impaired vascular function, and promoted myocardial ischemia and injury in healthy fire fighters. These finding provided pathogenic mechanisms to explain the association between fire suppression activity and acute myocardial infarction in fire fighters.                                                 The implications of these findings for prevention are discussed in an accompanying editorial from Dr. Kales, of Harvard school of Public Health and Dr. Smith from Skidmore College and University of Illinois fire service institute.                                                 Well, those were your summaries. Let's welcome our guests for our feature discussion.                                                 Today's feature paper describes a pilot study that shows that a common genetic variant risk score, is associated with drug induced QT prolongation and torsades de pointes. This paper is so interesting to me because I found that the learning points, at least for me, really extended well beyond the trial itself. I'm so delighted to have with me the co corresponding authors, Dr. David Strauss from the US FDA, as well as Dr. Christopher Newton-Cheh from Massachusetts General Hospital. Welcome, gentlemen. David:                                   Thanks very much, glad to be here. Christopher:                       Thank you, Carolyn. Carolyn:                               So, I've always thought that common genetic variants identified via GWAS, for example, are individually very weak effects on medical traits. For example, systolic blood pressure or in this case, QT interval. But what I'm so impressed with this study is that you show, I think for the first time, that even these small effects can add up to clinically meaningful results that are testable or demonstrable in a trial. David, could you begin by telling us a little bit about this trial and what the primary results were. David:                                   In the study, we tested the hypothesis that a weighted combination of common genetic variants, contributing to the QT interval at base line, identified through prior GWAS studies, can predict individual response to multiple QT prolonging drugs. We performed a genetic analysis of 22 subjects and a secondary analysis of a randomized, double blind, placebo controlled cross over trial, that included three QT prolonging drugs, with 15 tie matched QT and plasma drug concentration measurements. This allowed us to carefully control for the inter individual differences in pharmacokinetics and just focus on the pharmacodynamics so the direct effect of the drug on the heart.                                                 What we found was, there was a significant correlation between the weighted combination of common genetic variants, which we call the genetic QT score, and drug induced QT prolongation. More specifically, we found that the genetic QT score explained 30 percent of the variability in response to dofetilide, 23 percent in response to quinidine, and 27 in response to ranolazine.                                                 We also investigated how response to one QT prolonging drug predicted the response to other QT prolonging drugs. There were significant correlations between all the drug/drug relationships with response to each drug explaining 24 to 29 percent of the variability in response to each of the other drugs. It's important to note that QT prolongation, by itself, is not harmful. The real concern is torsades de pointes, which can degenerate into ventricular fibrillation and cause sudden death. So, the test, irrelevant to the common genetic variants in predicting drug induced torsades, we then went on to examine a previously published, genome wide association study that included 215 patients with drug induced torsades, compared to 771 ancestry match controls and that prior study that was previously published had found that each individual common genetic variant did not reach genome wide significance, as you suggested, Carolyn. However, when we applied the weighted combination of common genetic variants, we found that the genetic QT risk score was associated with significantly increased risk of drug induced torsade, explaining 12 percent of the variation in risk. Carolyn:                               So, my simplistic understanding was more or less there. That these genetic risks of these common variants kind of add up. I'm just curious ... Chris, do you think that this has implications for even other diseases? That's one question. And then secondly, I really appreciated your comment about using an intermediate trait, if you may, of QT interval versus looking at the disease itself of torsade de pointes. Could you give me comments on both these things? Christopher:                       The study of intermediate traits, such as, quantitative traits like QT variability on the EKG are, I think very tractable for the study of genetic bases of underlying physiologic processes because we can study so many people. So the original genome wide association study that detected these individually weak genetic effects could only find them because we studied about 75,000 people who had had genome wide genome typing and QT intervals measured. It requires such large sample sizes to reach p values that are able to distinguish true positive associations from false positive associations, due to the multiple testing burden.                                                 I think a challenge of what to do with these genetic effects once they've been reliably detected is that they do have weak effects and they influence intermediate traits. Nobody really cares whether their QT interval is three milliseconds longer, or three milliseconds shorter. What they care about is hard outcomes, or the likelihood that they'll have a toxic drug response. So, it was a natural follow on to that work to try to test these variants, and we knew that based on their weak effects individually on QT interval in the general population, that it was unlikely that they would individually explain a significant portion of either drug response or torsade. Which is why we aggregated the facts into the weighted score.                                                 I think we tried to examine what we thought were the most proximal, clinically relevant outcomes. Specifically, drug response. QT drug response to drugs that are established to cause QT prolongation and arrhythmias. Whether the QT score will have meaningful or detectable impact on drugs that have much weaker effects on re polarization and risk of torsade, I think, would remain to be seen. Carolyn:                               That's really remarkable.                                                 David, how about your perspective of the implications of this? It's so unique that you're actually from the FDA so, why is this important to the FDA? David:                                   As Chris mentioned, the specific application we studied here, a drug induced QT prolongation and torsade have resulted in the withdrawal of several drugs from the market both in the US and worldwide. Many critical drugs remain on the market that are associated with QT prolongation and torsade…over 100 drugs, likely. What some people may not be familiar with is that at FDA we perform research to move new science into the drug review process and close the gap between scientific innovation and drug review. Like practicing clinicians, we seek to understand inter patient variabilities and we conduct research to better evaluate, benefit, and risk of medications. This is in line with the broader initiative ... the precision medicine initiative, which seeks to move away from the traditional “one size fits all” approach for medical therapy and instead, take into account specific characteristics of individual patients.                                                 People are most familiar with this being applied in oncology and advances in pharmacogenomics have been more limited in other areas with the exception of the genetic bases of metabolism and pharmacokinetics  where the traits are often controlled by one or a few genetic mechanisms, rather than the many mechanisms responsible for complex traits and diseases, as Chris discussed. As I mentioned earlier, what was relatively unique about this study is that we were able to control for the difference in pharmacokinetics and investigate the inter individual differences in the direct effect of drugs on the heart, the pharmacodynamics. We think it's very exciting that a combination of common genetic variants and aggregate can explain a significant portion of the inter individual variability and, as Chris mentioned, this is also important because the incidence of torsade is quite low. Only a small number of patients will develop drug induced torsade. It's possible that in the future analysis of a large number of common genetic variants that can be identified through genome wide association studies as in this case, may help to better define the personalized benefit risk profiles for individual patients. Carolyn:                               You've really articulated that remarkably. That's exactly the excitement I think the entire editorial team shared when we read your paper. Thank you so much for it. Maybe just one last question thrown out to both of you, what's the next step? What's in the future. Christopher:                       I think one next step, based on this proof of principle study, will be to try to test the impact of these genetic risk scores in real world clinical settings where individual patients with the diversity of different comorbidities and different drug exposures are also receiving QT prolonging drugs. Because that will have the biggest relevance for our patients who faced increased risk of drug toxicity. David:                                   The issue of cardiac safety of drugs is something that is very important to us at the FDA and we have some parallel initiatives that, in collaboration with other global drugs ... regulatory agencies and industry and academic collaborators ... we are working to develop new cardiac safety evaluation paradigms for new drugs, or existing drugs, that could even be applied in the preclinical setting and really focus on the mechanistic base, pro arrhythmic risk. So, we should have more exciting work coming forward in the near future for better prediction and individualized prediction of benefit and risk of medication. Carolyn:                               Thank you, listeners, for joining us. You've been listening to Circulation on the Run. Join us next week.  

Two new biotech companies launched, TO Health! launches a new online health hub, and Government flexes its financial muscle backing science initiatives across the nation. We have this and more on this week’s show! Welcome to Biotechnology Focus Podcast. I’m your host Shawn Lawrence. Story 1 We kick things of this week in Toronto, where TO Health!, an industry led cluster organization has launched a new online networking site, the Toronto Health Hub (engage.tohealth.ca).  According to the organization, the new Hub will showcase news and events in the burgeoning Human Health & Sciences (HHS) cluster in Toronto, as well as provide a centralized place for government, business and academia to engage around critical topics pertaining to the region.  The new platform coincides with organization’s mandate to make Toronto a Top 5 global health science cluster within the next 10 years. Toronto Health Hub is powered by the content sharing publishing platform Pressly, which was developed by a Toronto based start-up Pressly Inc..  The platform has helped companies like Deloitte and The Economist to foster more meaningful conversations and content sharing amongst a broad community of stakeholders and innovators. The Toronto region’s Human Health Science cluster is home to the largest geographic concentration of health sciences assets in Canada, with more than 38,000 jobs and includes private and public sector organizations and ventures including biopharma, medical and digital health companies, universities, hospitals, research institutes, innovation organizations, service providers and government agencies.  All parties are invited and encouraged to engage on this new community building site. For more information about the Toronto Health Hub and to join the community, visit http://engage.tohealth.ca. Story 2 Heading to the prairies, The University of Saskatchewan recently marked the official launch of its unique Plant Phenotyping and Imaging Research Centre (P2IRC) with an international symposium and demonstration of new drone technology to be used in novel crop development approaches. The creation of the P2IRC stems from a $37.2-million award over seven years from the Canada First Research Excellence Fund (CFREF) for the research program “Designing Crops for Global Food Security”—one of only five CFREF grants awarded across Canada last year in the inaugural competition. The new centre will not only enhance the U of S biosciences cluster—but also capitalize on one of the largest clusters of food-related researchers in the world.” The P2IRC is led by Maurice Moloney, executive-director of the university’s Global Institute for Food Security (GIFS), who has been building a multi-disciplinary team of researchers from across the U of S campus and from other Canadian universities and centres. He hopes to create by the year 2022 a unique global resource for plant breeders seeking to develop new crop varieties at unprecedented speed and scale at the centre. At the same time, hopes are the centre will sustain Canada’s positon global powerhouse in agricultural research and lead to commercial spin-offs involving field and aerial sensors, satellite imaging, robotics, and big data analytics. Moloney noted the P2IRC is unique in that it combines plant genomics with crop phenotyping (the identification of useful traits), high-performance computing, and digital imaging technology, as well as undertakes research to address societal and developing world impacts. Research projects in four theme areas have already undergone rigorous international peer review involving an eight-member International Scientific Advisory Committee that included experts from Australia, Germany, France, and the U.K., several of whom participated in the symposium signaling the launch of the centre. The new centre also involves partnerships with four Canadian universities, three international institutes, and more than 15 private and public organizations, including the National Research Council and Agriculture and Agri-Food Canada. Story 3 In funding news, The National Research Council’s-Industry Research Assistance Program (NRC-IRAP) has awarded Vancouver Aspect Biosystems Ltd. $450,000 in grant funding to help the company further advance the development of its Lab-on- a-Printer™ platform technology and associated 3D bioprinted tissue applications. According to Tamer Mohamed, president and CEO of Aspect, the Canadian government-sponsored program has been very supportive of the company’s ongoing research and development efforts and the new funding will assist Aspect Biosystem in strategically deploying its unique platform to world-class research labs. Founded in 2013 as a collaboration between two research groups based in the departments of Engineering and Medicine at UBC, Aspect Biosystems’ platform enables advances in fundamental biological research, drug development through novel pre-clinical models, and regenerative medicine through the use of its 3D bioprinting and tissue engineering technology. The project funding arrives as the company prepares to launch its Early Platform Access Program and expand its partnerships. Story 4 A few months ago we mentioned on this show that AmorChem, an early-stage life sciences fund managed by Montréal-based venture capital firm GeneChem, was planning on launching a new company. Well, this past week the fund made good on its promise spinning out Mperia Therapeutics, focused on the development a novel immunotherapy approach based on CD36 cell receptor drug-ligands. As part of the company launch, AmorChem has transferred all rights to a CD36 technology to Mperia, and backed the company with a Series A investment of $1M. The capital will allow Mperia to pursue the early-stage development of lead candidates for the treatment of dry Age-related Macular Degeneration (AMD). The technology itself is the result of almost 20 years of research conducted by Dr. Huy Ong and his collaborators at the Université de Montréal, where the CD36 scavenger receptor has been shown to play a key role in the clearance of oxidized lipids, and more recently, in chronic inflammation. Promising CD36 drug-ligands have been tested in several dry AMD animal models and demonstrated significant efficacy to preserve and restore cell layers at both the retinal and subretinal levels. Dry AMD represents almost 90 per cent of all AMD cases, with five million patients affected by the disease in North America and Europe. It remains an unmet medical need with no approved treatments or cure for this disease indication. With the spinning off of this technology into Mperia and a first seed financing, AmorChem believes the company is poised to raise more capital and bring the first CD36 drug-ligand to the clinic in a near future. Additionally, ipon creation of Mperia Therapeutics, AmorChem appointed Dr. Maxime Ranger as the company’s new president and CEO, and Dr. Huy Ong as its chief scientific officer. A serial entrepreneur, Dr. Ranger has been entrepreneur-in- residence at Univalor for several months, in addition to ensuring the supervision of Dr Ong’s CD36 research project. In such context, Dr Ranger will thus lead Mperia in steps towards clinical proof-of- concept of its first product and its next fundraising. Likewise As CSO of the company, Dr Ong adds significant scientific value as a prominent investigator in the field of CD36 receptor biology. Story 5 Across the country, in British Columbia, to mark World Sepsis Day on September 13, the Centre for Drug Research and Development (CDRD) also has launched a new spin-off company, called Sepset Biosciences Inc. The choice to do it on World Sepsis day was no accident, as the new company is developing a novel rapid diagnostic test that will aid healthcare professionals in providing earlier and more targeted treatment of sepsis – a global healthcare problem that is more common than heart attack and claims more lives than any cancer. Sepset’s technology, which is based on extensive  work led by renowned University of British Columbia (UBC) researcher Dr. Robert Hancock, hopes to meet this dire clinical need as current methods to diagnose sepsis take more than 24 hours after a patient enters the emergency ward – by which time, the patient may already be well on their way towards tissue damage, organ failure, and death. For every three-hour delay in diagnosis, the rate of mortality and morbidity grows by almost 25 per cent. Dr. Hancock, a professor of Microbiology and Immunology and the director of the Centre for Microbial Diseases and Immunity Research at UBC, explains, the new company hopes to change the way scientists and health care professionals look at sepsis. Sepset’s blood-based test works by detecting, at the time a patient enters the hospital, a unique biomarker signature based on the body’s immune response rather than the presence of a pathogen. Dr. Hancock adds that the results of initial clinical studies show it to be a very promising approach and he and his team are now in the process of advancing to larger multi-centre, multi-country trials. Story 6 In a major funding commitment, Kirsty Duncan, Canada’s Science Minister announced that the federal government will invest $900 million towards research efforts at 13 postsecondary institutions across Canada through the Canada First Research Excellence Fund. Among the biggest recipients are the Université de Montréal, McGill University, Polytechnique Montréal, and HEC Montréal who together received $213 million to be split between three projects related to research in artificial intelligence, brain health, and medical technologies. According to a release, Polytechnique Montréal will use its hospital’s infrastructure to shape future medical technologies for complex diseases, while McGill’s Healthy Brains for Healthy Lives project will establish a central hub to work on projects to improve brain health in Canada. Two of the other more larger grant recipients included Dalhousie University, which is getting $93 million for its Safe and Sustainable Development of the Ocean Frontier to develop Canada’s oceans, and the University of Waterloo which is receiving $72 million for Transform Quantum Technologies, an initiative for advancing its work in developing quantum technologies. Additionally, the University of Saskatchewen is receiving $77.84 million for World Water Futures: Solutions to Water Threats in an Era of Global Change, while the University of Guelph is receiving approximately $76.6 million for Food from Thought: Agricultural Systems for a Healthy Planet. Rounding out the list is the University of Alberta, getting $75 million for the Future Energy Systems Research Institute, the University of Calgary getting $75 million for the Global Research Initiative in Sustainable Low Carbon Unconventional Resources, Western University getting $66 million for BrainsCAN: Brain health for life, $63.744 million to Queen’s University for the Canadian Particle Astrophysics Research Centre, $49.2 million going to Laurentian Universtiy for Metal Earth and York University receiving $33.3 million for Vision: Science to Applications (VISTA). An interesting side note, the Canada First Research Excellence Fund was actually established by the Harper Conservatives in the 2014 budget to help support research on big ideas and big themes. The funding is allocated on a competitive basis with researchers applying to an arms-length selection board. Story 7 Our final story of the week is also a funding announcement. Backed by the Federal Government, Genome Canada says it is investing $4 million in 16 new bioinformatics and computational biology (B/CB) research projects to be conducted at academic institutions across Canada. The investments are being made through Genome Canada’s 2015 Bioinformatics and Computational Biology Competition, a partnership with the Canadian Institutes of Health Research (CIHR). Parliamentary secretary to the Minister of International Development, Karina Gould  made the funding announcement on behalf of Science Minister Kristy Duncan at McMaster University. At the press conference, Gould highlighted the work of McMaster University’s Dr. Andrew McArthur, who is a co-leader on two B/CB projects looking to develop new software and database tools that will empower public health agencies and the agri-food sector to more rapidly respond to threats posed by infectious disease outbreaks such as, food-borne illnesses or the growing crisis of microbes resistant to antimicrobials. Additionally Marc LePage, president and CEO of Genome Canada highlighted the importance of the 16 projects and harnessing the power of genomics and the big data it provides researchers across multiple sectors. He cited Bioinformatics and computation biology as a priority area for Genome Canada given the growing need for enhanced tools and methodologies to make sense of the huge and growing influx of data stemming from genomics research. Some of the other projects in this funding round will enhance diagnosis and treatment for patients, improve crops of importance to Canada and strengthen environmental monitoring. Each project will receive approximately $250,000.  A full backgrounder of the 16 projects is available at www.genomecanada.ca/sites/genomecanada/files/2015_bcb-backgrounder- en.pdf. With that we’ve come to the end of this week’s program. We hope you enjoyed it. A big thanks to our production manager Laskey Hart and the rest of the Biotechnology Focus team. You can find past episodes online at www.biotechnologyfocus.ca and we’re always looking for your feedback, story ideas and suggestions so we’d love to hear from you. Simply reach out to us on twitter: @BiotechFocus or by email biotechnology_focus@promotive.net. For all of us here at Biotechnology Focus, thank you for listening.

Roy Silverstein explains how the Na+/K+-ATPase helps turn macrophages into foam cells at atherosclerotic plaques.

The cell type-, organ-, and species-specific expression of the Toll-like receptors (TLRs) are well described, but little is known about the respective expression profiles of their accessory molecules. We therefore determined the mRNA expression levels of LBP, MD2, CD36, CD14, granulin, HMGB1, LL37, GRP94, UNC93b1, TRIL, PRAT4A, AP3B1, AEP and the respective TLRs in human and mouse solid organs. Humans and mice displayed significant differences between their respective mRNA expression patterns of these factors. In addition, the expression profiles in transient tissue inflammation upon renal ischemia-reperfusion injury, in spleens and kidneys from mice with lupus-like systemic autoimmunity, and in progressive tissue fibrosis upon unilateral ureteral obstruction were studied. Several TLR co-factors were specifically regulated during the different phases of these disease entities, suggesting a functional involvement in the disease process. Thus, the organ-and species-specific expression patterns need to be considered in the design and interpretation of studies related to TLR-mediated innate immunity, which seems to be involved in the tissue injury phase, in the phase of tissue regeneration, and in progressive tissue remodelling.

Unter in vivo Bedingungen kann es in Zellen, die shear stress ausgesetzt sind (wie z.B. Endothel und Epithelzellen) ständig zu Rupturen der Plasmamembran kommen. Das sogenannte Membrane Resealing stellt die Fähigkeit von Zellen dar, auf eine solche Schädigung zu reagieren und die Membranintegrität durch schnelle Fusionsprozesse intrazellulärer Vesikel an Verletzungsstellen wiederherzustellen. Vielfach belegt ist hierbei die Beteiligung lysosomaler Vesikel sowie Enlargeosomen. In der vorliegenden Arbeit konnte erstmals eine Beteiligung ER-generierter Vesikel an diesen Reparaturprozessen der Zellmembran nachgewiesen werden. In verschiedenen experimentellen Ansätzen wurde eine Translokation von ER-Membranen an die geschädigten Areale der Zellmembran gezeigt. Eine Fusion der ER-Membranen mit der Zellmembran wurde durch den Nachweis luminaler Domänen transmembranöser ER-Proteine (CNX) sowie luminaler (löslicher) ER-Proteine (ERp57) an den Verletzungsstellen von Axonen des Rückenmarkes in vivo bestätigt. Durch die Blockade der am ERES freigesetzten COPII-Vesikel (Sar1) wurde der frühe sekretorische Transportweg vom ER zum Cis-Golgi-Netzwerk unterbunden. Damit einhergehend kam es zu einem verminderten Resealing der geschädigten Areale in der Zellmembran. Die Ergebnisse zeigen, dass die schnelle Freisetzung von ER-Vesikeln nach mechanischer Verletzung bzw. Schädigung der Plasmamembran durch bakterielle Toxine entscheidend an der Reparatur und Regenerierung geschädigter Zellen beteiligt ist. Nach mechanischer Schädigung kommt es auch zur Freisetzung von exozytotischen Vesikeln, sogenannten Mikropartikeln (MP), in den extrazellulären Raum. Bisher ist weitgehend unbekannt, wie die Homöostase der externalisierten MP koordiniert wird. In der vorliegenden Arbeit wurde unter in vitro- und in vivo-Bedingungen gezeigt, dass die extrazelluläre Konzentration der MP über die Clearance mittels verschiedener Endozytose-/Phagozytoseprozesse reguliert wird. An der Internalisierung dieser Vesikel ist der Class B Scavenger-Rezeptor CD36 beteiligt. Eine Blockade dieses Rezeptors in vitro zeigte eine deutliche Reduktion der Aufnahme von MP in phagozytosefähige Zellen. In vivo konnte eine CD36-abhängige Reduktion der MP-Aufnahme in verschiedenen Organen (vor allem Niere, Milz) in CD36-defizienten Tieren im Vergleich zu Kontrolltieren nachgewiesen werden. Des Weiteren wurden unter in vitro-Bedingungen Unterschiede bei der Internalisierung normaler und karzinomatöser MP nachgewiesen. Im Gegensatz zur zellulären Aufnahme von MP aus nicht-transformierten Zellen, wurden MP aus karzinomatösen Zellen nicht über Endozytose/Phagozytose internalisiert. Hingegen kam es hierbei zu einer Fusion von karzinomatösen MP mit der Membran der Akzeptorzelle, einem Mechanismus, der an der Transformation normaler Zellen in karzinomatöse Zellen beteiligt sein könnte. Insgesamt gesehen wurde hierdurch gezeigt, dass MP über Endozytose/Phagozytose in Zellen internalisiert werden, und dass dies organspezifisch über den Scavenger Rezeptor CD36 vermittelt wird.

Ein bedeutender Mechanismus zur Prävention und Regression von atherosklerotischen Läsionen ist die Abräumung von akkumulierten extrazellulären Lipiden in der Gefäßwand und deren Einschleusung in den reversen Cholesterintransport durch Makrophagen. Wichtigste molekulare Effektoren sind dabei Scavenger Rezeptoren wie CD36 und Cholesterin-Exporter wie ABCA1 und ABCG1. Deren Expression wird durch spezifische oxidierte Sterole, die die nukleären Transkriptionsfaktoren wie PPARgamma und LXRalpha aktivieren, induziert. Da hochdosierte lipidlösliche Antioxidantien diese regulatorischen Oxylipide beeinflussen könnten, war es Ziel dieser Arbeit am Makrophagen-Modell die Wirkung von hochdosiertem alpha-Tocopherol auf Signalwege und Schlüsselrezeptoren der Cholesterin-Homöostase zu untersuchen. Der Einfluss von alpha-Tocopherol und teilweise auch von gamma-Tocopherol wurde auf regulatorischer, transkriptioneller, translationeller und funktioneller Ebene mittels Realtime RT-PCR, Reportergen-Assays, FACS, Immunoblot und Lipidaufnahme- und Lipidefflux-Assays analysiert. Der LDL-R wurde durch hochdosiertes alpha-Tocopherol nicht beeinflusst, während die Expression des Scavenger Rezeptors CD36, konzentrationsabhängig sowohl auf mRNA-Ebene als auch auf Protein-Ebene durch alpha-Tocopherol beeinträchtigt wurde. Auf funktioneller Ebene verringerte alpha-Tocopherol die Aufnahme von [H³]-Cholesterin markiertem oxLDL durch Makrophagen. Der Effekt konnte ebenso mikroskopisch dargestellt werden. Die verminderte Expression von CD36 durch alpha-Tocopherol konnte zumindest teilweise durch eine dosisabhängige Verminderung der mRNA-Transkription von PPARγ und eine verminderte Aktivierung von PPARgamma im PPRE-Luziferase-Assay auch durch exogene Stimuli erklärt werden. gamma-Tocopherol hatte keinen vergleichbaren Effekt auf die CD36- und PPARgamma-spezifische mRNA, weswegen bereits auch ein direkter transkriptioneller Effekt von alpha-Tocopherol postuliert wurde. Die vermehrte zelluläre Aufnahme von oxidiertem LDL über Scavenger Rezeptoren wie CD36 induziert normalerweise auch eine vermehrte Einschleusung von Cholesterin in den reversen Cholesterintransport durch ABC-Exporter wie ABCA1 und ABCG1, wodurch die Schaumzellbildung zumindest verzögern werden kann. Diese Induktion der Cholesterin-Exporter wird durch oxidierte Sterole vermittelt, die LXRalpha aktivieren. Deshalb wurde ebenfalls eine mögliche Interferenz von hochdosiertem alpha-Tocopherol mit dem zellulären Cholesterin-Export untersucht. In der Tat wurde der Cholesterin-Efflux von Makrophagen auf delipidiertes HDL durch alpha-Tocopherol beeinträchtigt, wodurch der zelluläre Cholesterin-Bestand anstieg. Dieser Effekt zeigte auch mikroskopisch vermehrte Lipidgranula. Die Aktivierung des LXR-Response Elements im Luziferase-Assay durch exogene Stimuli wie 22-OHC oder oxidiertes LDL wurde durch alpha-Tocopherol ebenfalls negativ beeinflusst. Dadurch könnte die Reduktion der Expression von ABCA1 und ABCG1 auf mRNA-Ebene und von ABCA1 auf Proteinebene zumindest teilweise erklärt werden. Mit gamma-Tocopherol konnte nur eine geringe Reduktion auf mRNA Ebene, sowohl für ABCA1 als auch LXRalpha festgestellt werden. Bei der verminderten Expression von ABCA1 und ABCG1 durch hochdosiertes alpha-Tocopherol handelt es sich also wahrscheinlich um einen spezifischen, teilweise durch LXRalpha vermittelten Prozess. Es scheinen aber weitere Signalwege beteiligt zu sein: Unerwarteterweise wurde die Transkription und die Aktivierung von LXRalpha auch durch delipidiertes HDL stimuliert, was durch hochdosiertes alpha-Tocopherol ebenfalls dosisabhängig reduziert werden konnte. Nichtsdestotrotz war ABCA1 in Makrophagen nach Cholesterinverarmung durch delipidiertes HDL supprimiert. Die gefundenen Effekte von alpha-Tocopherol auf Schlüsselrezeptoren der Cholesterin-Homöostase in Makrophagen können zur Erklärung der enttäuschenden Resultate der Preventionsstudien mit hochdosiertem alpha-Tocopherol beitragen: Durch Hemmung des Scavenger Rezeptors CD36 reduziert alpha-Tocopherol zwar einerseits den ersten Schritt zur Schaumzellbildung um den Preis einer verzögerten Abräumung extrazellulärer Lipiddepots, alpha-Tocopherol verlangsamt aber auch durch Hemmung von ABCA1 und ABCG1, den endgültigen Abtransport von Cholesterin aus der Gefäßwand durch den reversen Cholesterin-Transport.

Phospholipids form the matrix of plasmamebranes (PM) consisting of an asymmetric lipid bilayer. The high amounts of sphingolipids and cholesterol in the PM are furthermore the basis to form liquid-ordered domains / rafts by saturated long chain fatty acid interactions. Hence, the lipid trafficking pathways enabling the membrane biogenesis and lateral segregation into membrane compounds remain largely undefined. Here we could demonstrate that the scavenger receptor CD36 selectively mediates the lipid uptake of sphingomyelin (SM) and phosphatidylcholine (PC) into PM in an endocytosis independent way. In human monocytes we could further show that the selective phospholipid uptake for SM, PC and phosphatidylethanolamine was almost exclusively promoted by CD36 and SR-BI. Whereas CD36 was mainly localized in rafts, we found about 2/3rds of SR-BI located in non-rafts. Thereby, the main cell entrance for phospholipids and cholesterol was mediated by SR-BI into non-raft compartments. SR-BI was then internalized by clathrin-coated pits and promoted the formation of intracellular microdomains in early endosomes by recruitment of the newly acquired raft lipids. Thus, we propose a leading role for SR-BI in generating new rafts by the incorporation of the raft building lipids SM and cholesterol. These may provide a pool for the regeneration of raft domains within the PM.

In der vorliegenden Arbeit wurden Effekte von IL-10 auf monocytäre Lipidrezeptoren und -Transporter untersucht, die zusätzlich zu den anti-inflammatorischen Wirkungen von IL-10 für dessen anti-atherosklerotische Wirkung von Bedeutung sein könnten. Es konnte gezeigt werden, dass IL-10 die Expression des quantitativ wichtigsten Scavenger Rezeptors, CD36, in Monocyten/Makrophagen sowohl auf RNA- als auch auf Proteinebene hemmt. Diese Hemmung ging einher mit einer verringerten zellulären Aufnahme von oxidiertem LDL. Durch PPARgamma-FACS und -Western Blot in Cytosol-Kern-Fraktionen sowie Koinkubationen mit IL-10 und den PPARgamma-Agonisten 15d-PGJ2 (15-deoxy-Δ12,14-Prostaglandin J2) und Indomethacin konnte gezeigt werden, dass die IL-10 Hemmung von CD36 über die Hemmung des Transkriptionsfaktors PPARgamma vermittelt wird. Im Gegensatz dazu stimulierte IL-10 den Cholesterinexporter ABCA1 und dessen wichtigsten Transkriptionsfaktor LXRalpha auf RNA- und Proteinebene. Die Induktion von ABCA1 hatte funktionell einen verstärkten zellulären Cholesterinefflux zur Folge, welcher die Monocyten und Makrophagen vor Lipidakkumulationen schützte und das vermehrte Einschleusen von Cholesterin in den reversen Cholesterintransport ermöglicht. IL-10 stimulierte ABCA1 dabei trotz der Hemmung des Transkriptionsfaktor PPARgamma. Die IL-10 Stimulation von ABCA1 war durch Piceatannol, einem Inhibitor der proximalen, STAT3-vermittelten Signalübertragung des IL-10 Rezeptors, hemmbar. Mittels LXRalpha "knock down" Zellen konnte weiter gezeigt werden, dass für die IL-10-stimulierte ABCA1 Expression ein intakter LXRalpha-Signaltransduktionsweg nötig ist. Koinkubationen mit Fenofibrat, 9-cis RA (9-cis retinoic acid) und 22-OHC (22-Hydroxycholesterol) ergaben, dass IL-10 auch die Induktion von ABCA1 durch die beiden Transkriptionsfaktoren, PPARalpha und LXRalpha, verstärkte. Durch Hemmung der Proteinkinase A (PKA) und Messung der zellulären cAMP-Spiegel konnte des Weiteren ein distaler cross-talk des IL-10-Signalweges mit dem cAMP/PKA-Weg für die ABCA1 Stimulation nachgewiesen werden. Dagegen hatte IL-10 keinen anhaltenden Einfluss auf die Transkription von SR-BI. Der Scavenger Rezeptor BI (SR-BI) kann je nach Konzentrationsgradient sowohl die zelluläre Cholesterinaufnahme wie die Cholesterinabgabe vermitteln. Des Weiteren reduzierte IL-10 die LPS-induzierte ICAM-1 Expression, was auf die Attenuierung der NFkappaB- und PPARgamma-vermittelten ICAM-1 Expression zurückgeführt werden konnte. Insgesamt befördert IL-10 somit den ABCA1-initiierten peripheren Cholesterinabtransport aus Monocyten/Makrophagen durch HDL. Die gezeigten Effekte von IL-10 erklären tierexperimentelle Befunde eines deutlich reduzierten Lipidgehalts in atherosklerotischen Plaques unter IL-10 Behandlung. Sie erklären auch die niedrigen HDL-Spiegel bei IL-10 knock out (IL-10-/-) Mäusen, die durch exogene IL-10 Substitution korrigiert werden können. Demnach sollte IL-10 nicht nur durch seine anti-inflammatorischen Mechanismen, sondern auch durch seine Effekte auf das Cholesterinhandling von Monocyten/Makrophagen in der Gefäßwand die Entstehung und Progression atherosklerotischer Plaques reduzieren.

Das natürliche Vitamin Nikotinsäure wird seit 1955 in pharmakologischer Dosierung als Medikament zur Behandlung von Dyslipidämien und bei arteriosklerotischen Gefäßveränderungen verwendet. Von Nikotinsäure konnte als erstem Medikament bereits 1975 im Coronary Drug Project nachgewiesen werden, dass es die Mortalität nach Myokardinfarkt signifikant und anhaltend reduziert. Nikotinsäure senkt den LDL-Plasmaspiegel und erhöht den HDL-Spiegel. Während der Nikotinsäureeffekt auf LDL vielfach untersucht wurde, ist über den Mechanismus der HDL-Erhöhung bisher wenig bekannt. Nikotinsäure stimuliert massiv die PGD2-Synthese in vivo. Der Hauptmetabolit von PGD2, das 15-deoxy-Δ12,14-Prostaglandin J2, wurde kürzlich als wichtigster endogener Aktivator des nukleären Transkriptionsfaktors PPAR erkannt. PPAR ist entscheidend an der Regulation des Scavenger Rezeptors CD36 und des zellulären Cholesterinexporters ABCA-1 beteiligt. Diese Rezeptoren dominieren die zelluläre Aufnahme modifizierter LDL-Partikel und die Ausschleusung zellulären Cholesterins auf HDL-Partikel und damit die Cholesterinhomöostase in Monozyten/Makrophagen in der Gefäßwand. Deshalb war es Ziel der Arbeit an einem Makrophagenmodell zu untersuchen, ob Nikotinsäure Scavenger-Rezeptoren und zelluläre Cholesterin-Transporter tatsächlich beeinflusst und so über einen gesteigerten reversen Cholesterintransport aus der Peripherie zur Leber seinen klinischen Nutzen vermitteln könnte. Als Modelle wurden die differenzierte humane Monozytenlinie MM6, die humane hepatische Linie HepG2 und frisch präparierte humane Monozyten verwendet. Die Expression der Scavenger Rezeptoren CD36, SR-BI, LOX-1, des LDL-R, des Cholesterinexporters ABCA-1, des Transkriptionsfaktors PPAR und von -Aktin wurden durch reverse Transkription der spezifischen mRNAs, nachfolgende PCR und Quantifizierung der Amplifikate über HPLC bestimmt. Die Proteinexpression von CD36 und PPAR wurden mittels spezifischer Antikörper nach Fluoreszenzmarkierung im FACS gemessen. Die Änderung des zellulären Cholesteringehalts durch Inkubationen mit Nikotinsäure, oxLDL und delipidiertem HDL wurde nach zellulärer Lipidextraktion in einem adaptierten enzymatischen Assay gemessen. Im Makrophagenmodell stimulierte die Inkubation der Zellen mit Nikotinsäure schon nach 3 h und mindestens bis 48 h anhaltend die Transkription von PPAR, des PPAR abhängigen Scavenger-Rezeptors CD36 und des zellulären Cholesterinexporters ABCA-1. Dagegen blieb die Transkription des ApoB-spezifischen LDL-R und des Scavenger-Rezeptors LOX-1 unverändert. Vergleichbare Effekte waren auch am Hepatozytenmodell nachweisbar. Die Effekte auf die PPAR und CD36 Expression waren tendenziell auch auf Proteinebene nachweisbar. Die Stimulation von CD36 und ABCA-1 durch Nikotinsäure konnte auf RNA-Ebene auch an frisch präparierten peripheren Monozyten von Normalpersonen nachgewiesen werden. Die funktionelle Bedeutung der Nikotinsäureeffekte wurde in einem Cholesterin-Aufnahme und Efflux-Assay überprüft. Dabei reduzierte die Inkubation mit Nikotinsäure den zellulären Cholesteringehalt basal und unter oxLDL-Exposition und steigerte den zellulären Cholesterin-Efflux auf delipidiertes HDL. Diese neuen Effekte der Nikotinsäure auf mehrere Lipid-Rezeptoren und -Transporter können Lipidablagerungen in der Gefäßintima reduzieren, der Schaumzellbildung entgegenwirken und durch vermehrte Einschleusung von zellulärem Cholesterin in den reversen Cholesterintransport zurück zur Leber die HDL-Spiegel erhöhen. Diese peripheren Effekte der Nikotinsäure ergänzen die Effekte von Statinen und liefern ein Rational für einen potentiell überadditiven klinischen Nutzen durch die Kombinationstherapie, die gegenwärtig klinisch geprüft wird.

Die Differentialdiagnose entzündlicher Hauterkrankungen setzt seitens des Dermatologen ein hohes Maß an klinischer Erfahrung voraus. Labortechnische und histologische Untersuchungsverfahren sind dabei nur eingeschränkt hilfreich. Ausgehend von der Annahme, dass erstens das atopische Ekzem ein krankheitsspezifisches Mikromilieu in der Epidermis und Dermis aufweist und dass sich zweitens dieses spezifische Mikromilieu im Phänotyp der immunkompetenten CD1a positiven epidermalen dendritischen Zellpopulation widerspiegelt, wurde seit 1995 ein Verfahren zur Immunphänotypisierung von Langerhans Zellen und IDEC entwickelt. Es konnte gezeigt werden, dass das atopische Ekzem durch eine hohe Expression des hochaffinen IgE-Rezeptors (FceRI) bei niedriger Expression des IgG-Rezeptors FcgRII auf den dentritischen Zellen der Epidermis hoch sensitiv und spezifisch von anderen Dermatosen abgegrenzt werden kann. Ziel der vorgelegten Arbeit war es, die standardisierte Phänotypisierung CD1a positiver epidermaler Zellen so zu erweitern, dass neben dem atopischen Ekzem auch die Diagnose anderer entzündlicher Hauterkrankungen möglich wird. Es wurden 296 Patienten mittels standardisierter Phänotypisierung epidermaler dendritischer Zellen untersucht. Bei 192 Patienten konnte nach Abschluss aller klinisch diagnostischen Untersuchungen eine eindeutige Diagnose gestellt werden, so dass die an diesen Patienten erhobenen Phänotypisierungsdaten für die Analyse krankheitsspezifischer Veränderungen im Oberflächenmarkerprofil von Langerhans Zellen und IDEC herangezogen wurden. Die Patienten verteilten sich auf 8 Gruppen: Normale Haut (n=31); Atopisches Ekzem (n=59); Psoriasis (n=41); Allergisches Kontaktekzem (n=23); Kontaktekzem bei atopischer Diathese (n=14); Lichen ruber (n=6); Mycosis fungoides (n=10) und Nummuläres Ekzem (n=8). Es konnte gezeigt werden, dass der prozentuale Anteil epidermaler dendritischer Zellen in Abhängigkeit von der Diagnose variiert und dass die bei allen entzündlichen Dermatosen nachweisbare Vermehrung epidermaler dendritischer Zellen vorrangig auf eine de novo Einwanderung von IDEC in die Epidermis zurückzuführen ist. Das wesentliche Ergebnis der vorgelegten Arbeit ist die Erweiterung der diagnostischen Phänotypisierung auf die Psoriasis und das allergische Kontaktekzem. So konnte die Psoriasis durch eine Ratio von CD64/CD11b auf IDEC sinnvoll abgegrenzt werden. Das Kontaktekzem war mittels einer Ratio aus E-Cadherin/CD86 auf Langerhans Zellen zu identifizieren. Die Gruppe der Patienten mit Kontaktekzem bei atopischer Diathese wurde durch die Ratio E-Cadherin/CD80 auf IDEC erkannt. Die bereits für das atopische Ekzem publizierte Ratio FcRI/CD32 auf allen CD1a positiven Zellen wurde mit der gleichen Ratio, ermittelt nur für IDEC, verglichen. Eine Verbesserung der Sensitivität und Spezifität für die Diagnosestellung des atopischen Ekzems konnte hierdurch nicht erreicht werden. Die starke Expression von FceRI auf LC und IDEC kennzeichnet das atopische Ekzem. Typisch für das atopische Ekzem ist weiterhin die Expression von CD36 auf LC und IDEC. Die Psoriasis ist durch eine besonders starke Expression der Fcg-Rezeptoren auf IDEC gekennzeichnet. Darüber hinaus findet sich eine erhöhte passive Anlagerung von sCD23 an IDEC bei Psoriasis. Gegenüber anderen Diagnosen ist die Expression von CD11b und CD11c auf IDEC vermindert. Für das Kontaktekzem ist die im Vergleich mit anderen entzündlichen Dermatosen niedrige CD49d Expression auf LC kennzeichnend. Die Einführung neuer diagnostischer Algorithmen in die Phänotypisierung epidermaler dendritischer Zellen konnte das diagnostische Spektrum dieser Methode deutlich erweitern. Neben atopischem Ekzem und normaler Haut wurden auch Algorithmen für die Psoriasis und das allergische Kontaktekzem beschrieben. Die systematische Analyse weiterer Dermatosen mit epidermaler Beteiligung dürfte auch für diese Erkrankungen diagnostische Algorithmen aufzeigen, die das Mikromilieu der entsprechenden Krankheiten widerspiegeln. Die Phänotypisierung epidermaler dendritischer Zellen dürfte insbesondere im Rahmen klinisch-therapeutischer Studien vermehrt zur Anwendung kommen, da mit dieser Methode eine objektive, quantitative Untersuchung einzelner behandelter oder unbehandelter Hautareale im zeitlichen Verlauf möglich ist.

Thu, 15 Jan 2004 12:00:00 +0100 https://edoc.ub.uni-muenchen.de/1728/ https://edoc.ub.uni-muenchen.de/1728/1/Jostarndt_Kristina.pdf Jostarndt, Kristina

Topical tacrolimus (FK506) leads to profound phenotypic and functional alterations of epidermal antigen-presenting dendritic cells in atopic dermatitis. BACKGROUND: Atopic dermatitis (AD) is a chronic inflammatory skin disease in which antigen-presenting epidermal dendritic cells (DCs), ie, Langerhans cells and the so-called inflammatory dendritic epidermal cells (IDECs) expressing the high-affinity receptor for IgE (FcepsilonRI) may play a significant pathophysiologic role. Therapeutic efficacy of the immunosuppressive macrolide tacrolimus (FK506) in AD has been demonstrated in clinical trials, but little is known of its mode of action. OBJECTIVE: The present study focused on the effects of topical tacrolimus treatment on epidermal CD1a+/FcepsilonRI+ DC populations in lesional AD. METHODS: Immunohistological analysis, epidermal DC phenotyping, and functional studies were performed on skin biopsy specimens from treated and untreated lesional skin of 10 patients with AD participating in a clinical trial with tacrolimus. RESULTS: Untreated lesional skin was characterized by a high proportion of CD1a+ cells, which was largely due to a high proportion of IDECs strongly expressing FcepsilonRI. Epidermal DCs isolated from untreated lesional skin exhibited high stimulatory activity toward autologous T cells, which was strongly reduced while clinical improvement was seen during application of tacrolimus. Concomitantly, a decreased FcepsilonRI expression was observed in both Langerhans cells and IDECs. Finally, topical tacrolimus led to a progressive decrease in the IDEC population within the pool of CD1a+ epidermal DCs and also to a decrease in their CD36 expression, which is indicative of lower local inflammation. CONCLUSION: Epidermal CD1a+ DCs may represent a target for topical tacrolimus in the treatment of AD.