Podcasts about transcriptional

Play Episode Listen Later Oct 19, 2023 45:35

In this episode of the Epigenetics Podcast, we caught up with Sheila Teves from the University of British Columbia to talk about her work on the inheritance of transcriptional memory by mitotic bookmarking. Early in her research career, Sheila Teves focused on the impact of nucleosomes on torsional stress and gene regulation. She also highlights the development of a genome-wide approach to measure torsional stress and its relationship to nucleosome dynamics and RNA polymerase regulation. The conversation then shifts to her focus on transcriptional memory and mitotic bookmarking during her postdoc in the Tijan lab. She explores the concept of mitotic bookmarking, whereby certain transcription factors remain bound to their target sites during mitosis, facilitating efficient reactivation of transcription after cell division. She discusses her findings on the behavior of transcription factors on mitotic chromosomes, challenging the notion that they are excluded during mitosis. She also discusses the differences in binding behavior between the general transcription factor TBP and other transcription factors. Finally, the effect of formaldehyde fixation on the potential to find transcription factors bound to mitotic chromosomes is discussed. References Teves, S., Henikoff, S. Transcription-generated torsional stress destabilizes nucleosomes. Nat Struct Mol Biol 21, 88–94 (2014). https://doi.org/10.1038/nsmb.2723 Sheila S Teves, Luye An, Anders S Hansen, Liangqi Xie, Xavier Darzacq, Robert Tjian (2016) A dynamic mode of mitotic bookmarking by transcription factors eLife 5:e22280. https://doi.org/10.7554/eLife.22280 Sheila S Teves, Luye An, Aarohi Bhargava-Shah, Liangqi Xie, Xavier Darzacq, Robert Tjian (2018) A stable mode of bookmarking by TBP recruits RNA polymerase II to mitotic chromosomes eLife 7:e35621. https://doi.org/10.7554/eLife.35621 Kwan, J. Z. J., Nguyen, T. F., Uzozie, A. C., Budzynski, M. A., Cui, J., Lee, J. M. C., Van Petegem, F., Lange, P. F., & Teves, S. S. (2023). RNA Polymerase II transcription independent of TBP in murine embryonic stem cells. eLife, 12, e83810. https://doi.org/10.7554/eLife.83810 Price, R. M., Budzyński, M. A., Shen, J., Mitchell, J. E., Kwan, J. Z. J., & Teves, S. S. (2023). Heat shock transcription factors demonstrate a distinct mode of interaction with mitotic chromosomes. Nucleic acids research, 51(10), 5040–5055. https://doi.org/10.1093/nar/gkad304 Related Episodes In Vivo Nucleosome Structure and Dynamics (Srinivas Ramachandran) From Nucleosome Structure to Function (Karolin Luger) Structural Analysis of Nucleosomes During Transcription (Lucas Farnung) Contact Epigenetics Podcast on Twitter Epigenetics Podcast on Instagram Epigenetics Podcast on Mastodon Active Motif on Twitter Active Motif on LinkedIn Email: podcast@activemotif.com

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Control your DNA! Transcriptional and Translational Control I Jack Westin MCAT Podcast

Jack Westin MCAT Podcast

Play Episode Listen Later Oct 12, 2023 45:14

How do we control what DNA is expressed? Azaii and Phil talk about all the ways that our body controls the rate of protein production via regulating transcription and translation. About Jack Westin - The team at Jack Westin is dedicated to a single goal: giving students the highest quality learning resources. Jack Westin understands that students can't crush the MCAT without the perfect blend of critical thinking and fundamental science knowledge. To this end, Jack Westin is dedicated to providing students with cutting edge comprehensive tools, courses, and practice materials. The Jack Westin MCAT science and CARS courses, taught by the world's best and most engaging MCAT instructors, are designed to do more than just teach students the MCAT—it supercharges studying and encourages lifelong learning. Want to learn more? Shoot us a text at 415-855-4435 or email us at podcast@jackwestin.com!

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Long-Range Transcriptional Control by 3D Chromosome Structure (Luca Giorgetti)

Play Episode Listen Later Aug 24, 2023 40:12

In this episode of the Epigenetics Podcast, we caught up with Luca Giorgetti from the Friedrich Miescher Institute to hear about his work on long-range transcriptional control by 3D chromosome structure. Luca Giorgetti's research focuses on chromosomal interactions, transcriptional output, and the dynamics of enhancer-promoter relationships. His lab investigated the causal relationship between chromosome interactions and transcriptional events. They've found that by manipulating the contact probabilities between an enhancer and a promoter by changing their distance, these changes had a substantial effect on transcription levels. This project was an experiment that Luca Giorgetti was eager to do, and it allowed him to establish a smooth functional relationship between contact probabilities and changes in transcription levels. References Giorgetti, L., Galupa, R., Nora, E. P., Piolot, T., Lam, F., Dekker, J., Tiana, G., & Heard, E. (2014). Predictive polymer modeling reveals coupled fluctuations in chromosome conformation and transcription. Cell, 157(4), 950–963. https://doi.org/10.1016/j.cell.2014.03.025 Redolfi, J., Zhan, Y., Valdes-Quezada, C., Kryzhanovska, M., Guerreiro, I., Iesmantavicius, V., Pollex, T., Grand, R. S., Mulugeta, E., Kind, J., Tiana, G., Smallwood, S. A., de Laat, W., & Giorgetti, L. (2019). DamC reveals principles of chromatin folding in vivo without crosslinking and ligation. Nature structural & molecular biology, 26(6), 471–480. https://doi.org/10.1038/s41594-019-0231-0 Zuin, J., Roth, G., Zhan, Y., Cramard, J., Redolfi, J., Piskadlo, E., Mach, P., Kryzhanovska, M., Tihanyi, G., Kohler, H., Eder, M., Leemans, C., van Steensel, B., Meister, P., Smallwood, S., & Giorgetti, L. (2022). Nonlinear control of transcription through enhancer-promoter interactions. Nature, 604(7906), 571–577. https://doi.org/10.1038/s41586-022-04570-y Mach, P., Kos, P. I., Zhan, Y., Cramard, J., Gaudin, S., Tünnermann, J., Marchi, E., Eglinger, J., Zuin, J., Kryzhanovska, M., Smallwood, S., Gelman, L., Roth, G., Nora, E. P., Tiana, G., & Giorgetti, L. (2022). Cohesin and CTCF control the dynamics of chromosome folding. Nature genetics, 54(12), 1907–1918. https://doi.org/10.1038/s41588-022-01232-7 Related Episodes scDamID, EpiDamID and Lamina Associated Domains (Jop Kind) Epigenetics and X-Inactivation (Edith Heard) Spatial Organization of the Human Genome (Wendy Bickmore) Contact Epigenetics Podcast on Twitter Epigenetics Podcast on Instagram Epigenetics Podcast on Mastodon Active Motif on Twitter Active Motif on LinkedIn Email: podcast@activemotif.com

RBL2 represses the transcriptional activity of Multicilin to inhibit multiciliogenesis

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Play Episode Listen Later Aug 4, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.08.04.551992v1?rss=1 Authors: Quiroz, E. J., Kim, S., Gautam, L. K., Borok, Z., Kintner, C., Ryan, A. L. Abstract: A core pathophysiologic feature underlying many respiratory diseases is multiciliated cell dysfunction, leading to inadequate mucociliary clearance. Due to the prevalence and highly variable etiology of mucociliary dysfunction in respiratory diseases, it is critical to understand the mechanisms controlling multiciliogenesis that may be targeted to restore functional mucociliary clearance. Multicilin, in a complex with E2F4, is necessary and sufficient to drive multiciliogenesis in airway epithelia, however this does not apply to all cell types, nor does it occur evenly across all cells in the same cell population. In this study we further investigated how co-factors regulate the ability of Multicilin to drive multiciliogenesis. Combining data in mouse embryonic fibroblasts and human bronchial epithelial cells, we identify RBL2 as a repressor of the transcriptional activity of Multicilin. Knockdown of RBL2 in submerged cultures or phosphorylation of RBL2 in response to apical air exposure, in the presence of Multicilin, allows multiciliogenesis to progress. These data demonstrate a dynamic interaction between RBL2 and Multicilin that regulates the capacity of cells to differentiate and multiciliate. Identification of this mechanism has important implications for facilitating MCC differentiation in diseases with impaired mucociliary clearance. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

An integrated single-cell RNA-seq atlas of the mouse hypothalamic paraventricular nucleus links transcriptional and functional types

Play Episode Listen Later Jul 19, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.07.19.549666v1?rss=1 Authors: Berkhout, J., Poormoghadam, D., Yi, C.-X., Kalsbeek, A., Meijer, O., Mahfouz, A. Abstract: The hypothalamic paraventricular nucleus (PVN) is a highly complex brain region that is crucial for homeostatic regulation through neuroendocrine signalling, outflow of the autonomic nervous system, and projections to other brain areas. The past years, single-cell datasets of the hypothalamus have contributed immensely to the current understanding of the diverse hypothalamic cellular composition. While the PVN has been adequately classified functionally, its molecular classification is currently still insufficient. To address this, we created a detailed atlas of PVN transcriptional cell types by integrating various PVN single-cell datasets into a recently published hypothalamus single-cell transcriptome atlas. Furthermore, we functionally profiled transcriptional cell types, based on relevant literature, existing retrograde tracing data and existing single-cell data of a PVN-projection target region. In our PVN atlas dataset, we identify the well-known different neuropeptide types, each composed of multiple novel subtypes. We identify Avp-Tac1, Avp-Th, Oxt-Foxp1, Crh-Nr3c1 and Trh-Nfib as the most important neuroendocrine subtypes based on markers described in literature. To characterize the pre-autonomic functional population, we integrated a single-cell retrograde tracing study of spinally-projecting pre-autonomic neurons into our PVN atlas. We identify these (pre-sympathetic) neurons to co-cluster with the Adarb2+ clusters in our dataset. Finally, we identify expression of receptors for Crh, Oxt, Penk, Sst, and Trh in the dorsal motor nucleus of the vagus, a key region that pre-parasympathetic PVN neurons project to. Concluding, our study present a detailed overview of the transcriptional cell types of the murine PVN, and provides a first attempt to resolve functionality for the identified populations. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Pseudospatial transcriptional gradient analysis of hypothalamic ependymal cells: towards a new tanycyte classification

Play Episode Listen Later Jul 7, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.07.06.547914v1?rss=1 Authors: Brunner, M., Lopez-Rodriguez, D., Messina, A., Thorens, B., Santoni, F., Langlet, F. Abstract: The ependyma lining the third ventricle (3V) in the mediobasal hypothalamus is recognized as a critical player in controlling energy balance and glucose homeostasis. Its molecularly distinct cell types, including diverse tanycyte subpopulations and typical ependymal cells, confer a high functional heterogeneity. The study of gene expression profiles and dynamics of ependymal cells has the potential to uncover fundamental mechanisms and pathways involved in metabolic regulation. Here, we cataloged 5481 hypothalamic ependymocytes using FACS-assisted single-cell RNA sequencing from fed, 12h-fasted, and 24h-fasted adult male mice. First, standard clustering analysis revealed the limitation of the current characterization regarding the different ependymal cell subpopulations along the 3V. Indeed, while typical ependymal cells and {beta}2-tanycytes are sharply defined at the molecular level, other subpopulations (i.e., {beta}1-, 2-, and 1 tanycytes) display fuzzy boundaries and very few specific markers. Moreover, we observed that 12h- and 24h-fasting dynamically modulate gene expression, increasing tanycyte subgroup heterogeneity. Secondly, pseudospatial trajectory analysis based on peculiar UMAP neuroanatomical distribution improved the identification of tanycyte markers, distinguishing specific versus overlapping features and better segregating tanycyte specific versus standard functions. Intriguingly, we discovered numerous functions related to tanycyte-neuron and tanycyte-synapse interactions with modulation by energy balance. Finally, combining pseudospatial analysis and gene regulatory network inference, we observed that fasting dynamically shifts patterns in gene expression and transcription activity along the 3v, creating a metabolic and functional switch for some subpopulations. Altogether, this data provides a mechanism through which energy status leads to distinct cell type-specific responses along the 3V and gives new insights into molecular diversity underlying tanycyte classification. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Single neuron analysis of aging associated changes in learning reveals progressive impairmentsin transcriptional plasticity

Play Episode Listen Later Jun 25, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.06.23.546336v1?rss=1 Authors: Badal, K. K., Sadhu, A., McCracken, C., Raveendra, B. L., Lozano-Villada, S., Shetty, A. C., Gillette, P., Zhao, Y., Stommes, D., Fieber, L. A., Schmale, M. C., Mahurkar, A., Hawkins, R. D., Puthanveettil, S. V. Abstract: Molecular mechanisms underlying aging associated impairments in learning and long-term memory storage are poorly understood. Here we leveraged the single identified motor neuron L7 in Aplysia, which mediates a form of non-associative learning, sensitization of the siphon-withdraw reflex, to assess the transcriptomic correlates of aging associated changes in learning. RNAseq analysis of the single L7 motor neuron isolated following short-term or long-term sensitization training of 8,10 and 12 months old Aplysia, corresponding to mature, late mature and senescent stages, has revealed progressive impairments in transcriptional plasticity during aging. Specifically, we observed modulation of the expression of multiple lncRNAs and mRNAs encoding transcription factors, regulators of translation, RNA methylation, and cytoskeletal rearrangements during learning and their deficits during aging. Our comparative gene expression analysis also revealed the recruitment of specific transcriptional changes in two other neurons, the motor neuron L11 and the giant cholinergic neuron R2 whose roles in long-term sensitization were previously not known. Taken together, our analyses establish cell type specific progressive impairments in the expression of learning- and memory-related components of the transcriptome during aging. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Phenotypic Switching of Vascular Smooth Muscle Cells in Duchenne Muscular Dystrophy

Play Episode Listen Later Jun 25, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.06.23.546309v1?rss=1 Authors: Xuan, W., Cheng, F., Han, X., Tipparaju, S., Ashraf, M. Abstract: Background: Extensive studies have been conducted in skeletal muscle and myocardium affected by Duchenne Muscular Dystrophy (DMD) disease but there is a significant gap of research in the role of vascular smooth muscle cells (VSMCs) in DMD. Here, we investigated the role of dystrophin deficiency in the maintenance of VSMCs contractile phenotype. Methods: 12-14 months old mdx mice and DMD induced pluripotent stem cells (iPSC) derived VSMCs were used as disease models. Morphological and immunohistochemistry analyses were performed to determine histological changes and the expression of contractile markers. Transmission Electron Microscopy (TEM) was used to assess ultrastructural changes in the VSMCs. Mito-tracker staining and TUNEL staining were performed to determine mitochondria fission-fusion and apoptosis respectively. mRNA Sequencing for normal iPSC derived VSMCs (WT-VSMCs) and DMD iPSC derived VSMCs (DMD-VSMCs) with or without oxidative stress was performed. KEGG signaling pathway enrichment, Go function enrichment and Gene set enrichment analysis (GESA) were conducted to explore the potential mechanism responsible for these changes. In addition, transcription factor enrichment analysis was performed to unravel mechanistic pathways of regulatory networks. Results: Spontaneous abnormal VSMCs proliferation, loss of vascular structure and degenerative changes occurred in VSMCs in aorta from 12-14 months old mdx mice. The DMD-VSMCs showed maturation defect, loss of mitochondrial hemostasis, and increased vulnerability to oxidative stress compared with WT-VSMCs. Transcriptome analysis revealed dysregulation of smooth muscle proliferation, differentiation, and vascular development in DMD-VSMCs. Transcriptional factor, target, and motif discovery analysis of the dysregulated gene set suggested potential contributions of transcriptional factors GADD45A, SOX9, TIA1, RBBP9 and FOXM to the phenotypes of DMD-VSMCs. Under oxidative stress, initiation of apoptotic process was significantly enhanced in DMD-VSMCs while their response to hypoxia and oxidative stress was downregulated. Conclusions: Dystrophin deficiency induced VSMCs phenotype switching and disrupted mitochondrial metabolism. The findings in this study underscore the importance of vascular dysfunction in DMD disease and therapeutic interventions to restore VSMC phenotype may ameliorate the propensity of disease progression. It is suggested that the transcriptome analysis may allow the discovery of potential signaling pathways involved in the dysregulation of transcription factors. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Single-cell Transcriptomic Analysis of Salivary Gland Endothelial Cells

O dÅ‚ugim Å¼yciu w zdrowiu z dr n. med. KarolinÄ… Karabin

Play Episode Listen Later Jun 24, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.06.22.545817v1?rss=1 Authors: Altrieth, A. L., Suarez, E., Nelson, D. A., Gabunia, S., Larsen, M. Abstract: Vascular endothelial cells have important functions in fibrosis via direct and indirect methods and in regeneration through secretion of tissue-specific, paracrine-acting angiocrine factors. In the salivary gland, endothelial cells are required for proper development, but their roles within adult glands are largely unknown. The goal of this work was to identify ligand-receptor interactions between endothelial cells and other cell types that are important during homeostasis, fibrosis, and regeneration. To model salivary gland fibrosis and regeneration, we utilized a reversible ductal ligation. To induce injury, a clip was applied to the primary ducts for 14 days, and to induce a regenerative response, the clip was subsequently removed for 5 days. To identify endothelial cell-produced factors, we used single-cell RNA-sequencing of stromal-enriched cells from adult salivary glands. Transcriptional profiles of homeostatic salivary gland endothelial cells were compared to endothelial cells of other organs. Salivary gland endothelial cells were found to express some unique genes and some that were similar to other fenestrated endothelial cells from the colon and small intestine. Comparison of the 14-day ligated, mock ligated, and 5-day deligated stromal-enriched transcripts and lineage tracing were used to identify evidence for a partial endoMT phenotype, which was observed in a small number of endothelial cell subsets with ligation. CellChat was used to predict changes in ligand-receptor interactions in response to ligation and deligation. CellChat predicted that endothelial cells are sources of protein tyrosine phosphatase receptor type m, tumor necrosis factor ligand superfamily member 13, and myelin protein zero signaling and targets for tumor necrosis factor signaling following ligation. CellChat also predicted that endothelial cells are sources of angiocrine factors, chemokine (C-X-C motif) ligand, and EPH signaling to promote regenerative responses following deligation. These studies will inform future endothelial cell-based regenerative therapies. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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#3 Celiakia - fakty i mity prof. dr hab. n. med. Bożena Cukrowska

Play Episode Listen Later Jun 21, 2023 103:10

Dieta bezglutenowa jest jedną z najpopularniejszych diet w ostatnich latach - stosują ją celebryci, piszą o niej kolorowe czasopisma i portale internetowe, a półki w księgarniach uginają się od poradników na temat diet bezglutenowych. Jednak ta moda ma swoje dobre i złe strony. Pozytywnym aspektem jest to, że celiakia – choroba której przyczyną jest gluten, przebiła się do społecznej świadomość. Celiakia to choroba glutenozależna, która do niedawna była postrzegana tylko jako choroba wieku dziecięcego, ale jak się okazuje rozpoznaje się ją także u dorosłym. W porównaniu z dziećmi u dorosłych mogą dominować objawy z poza przewodu pokarmowego, co może utrudniać rozpoznanie celiakii. Przypuszcza się, że wzrost wykrywalności przypadków celiakii jest częściowo wynikiem dostępu do coraz lepszych narzędzi diagnostycznych i częstszych badań przesiewowych u osób z grup ryzyka. Jednak mimo tak dużego postępu diagnostyka celiakii w niektórych przypadkach nie jest łatwa i może zająć nawet kilka lat! Gość: prof. dr hab. n. med. Bożena Cukrowska – pediatra i immunologiem. Pani Profesor kieruje Pracownią Immunologii w Instytucie "Pomnik - Centrum Zdrowia Dziecka" w Warszawie: https://www.czd.pl/strony/dzialalnosc-kliniczna/pracownie/pracownia-immunologii Posiada osiągnięcia związane z zastosowaniem probiotyków w leczeniu i profilaktyce alergii pokarmowej u dzieci oraz z wdrażaniem nowych metod diagnostycznych w gastroenterologii, hepatologii i onkologii dziecięcej. W odcinku znajdziesz odpowiedzi m. in. na następujące pytania: Kiedy odkryto celiakię? Jak często występuje choroba? Jakie mamy formy nietolerancji glutenu poza celiakią i czym się one różnią? Dlaczego gluten szkodzi w celiakii? Jakie są oczywiste i nieoczywiste objawy celiakii? Kto jest w grupie ryzyka zachorowani na celiakię? Czy celiakia może być przyczyną przewlekłego zmęczenia? Jak aktualnie diagnozujemy celiakię? Jak wybrać odpowiednie badanie genetyczne w kierunku celiakii? Czego nie robić przed diagnostyką celiakii? Jaki jest związek zonuliny z celiakia? Lista publikacji o których wspominamy w podcaście: Majsiak E. i wsp. Clinical Manifestation and Diagnostic Process of Celiac Disease in Poland—Comparison of Pediatric and Adult Patients in Retrospective Study. Nutrients 2022 Jan 23;14(3):491. Majsiak E. i wsp. The impact of symptoms on quality of life before and after diagnosis of coeliac disease: the results from a Polish population survey and comparison with the results fromthe United Kingdom. BMC Gastroenterol. 2021 Mar 4;21(1):99. Sowińska A. i wsp. Transcriptional and Ultrastructural Analyses Suggest Novel Insights into Epithelial Barrier Impairment in Celiac Disease. Cells 2020, 9(2), 516. Tye-Din J.A. i wsp. Celiac Disease: A Review of Current Concepts in Pathogenesis, Prevention, and Novel Therapies. Front Pediatr. 2018 Nov 21;6:350. Husby S. i wsp. European Society Paediatric Gastroenterology, Hepatology and Nutrition guidelines for diagnosing coeliac disease 2020. J Pediatr Gastroenterol Nutr 2020;70(1):141-56. Cukrowska B. Celiakia – zasady diagnostyki według nowych wytycznych ESPGHAN 2020. Pediatria Po Dyplomie kwiecień 2020. König J. i wsp. Randomized clinical trial: Effective gluten degradation by Aspergillus niger-derived enzyme in a complex meal setting.Sci Rep. 2017; 7: 13100.

Spatial Transcriptional Mapping Reveals Site-Specific Pathways Underlying Human Atherosclerotic Plaque Rupture

JACC Podcast

Play Episode Listen Later Jun 12, 2023 9:52

Commentary by Dr. Valentin Fuster

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ARID1B controls transcriptional programs of axon projection in the human corpus callosum

Play Episode Listen Later May 4, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.05.04.539362v1?rss=1 Authors: Martins-Costa, C., Pham, V. A., Wiegers, A., Sidhaye, J., Doleschall, B., Novatchkova, M., Lendl, T., Piber, M., Peer, A., Moeseneder, P., Stuempflen, M., Chow, S. Y. A., Seidl, R., Prayer, D., Hoeftberger, R., Kasprian, G., Ikeuchi, Y., Corsini, N., Knoblich, J. A. Abstract: Mutations in ARID1B, a member of the mSWI/SNF complex, cause severe neurodevelopmental phenotypes with elusive mechanisms in humans. The most common structural abnormality in the brain of ARID1B patients is agenesis of the corpus callosum (ACC). This condition is characterized by a partial or complete absence of the corpus callosum (CC), an interhemispheric white matter tract that connects distant cortical regions. Using human neural organoids, we identify a vulnerability of callosal projection neurons (CPNs) to ARID1B haploinsufficiency, resulting in abnormal maturation trajectories and dysregulation of transcriptional programs of CC development. Through a novel in vitro model of the CC tract, we demonstrate that ARID1B mutations reduce the proportion of CPNs capable of forming long-range projections, leading to structural underconnectivity phenotypes. Our study uncovers new functions of the mSWI/SNF during human corticogenesis, identifying cell-autonomous defects in axonogenesis as a cause of ACC in ARID1B patients. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Transcriptional dissection of symptomatic profiles across the brain of men and women with depression

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Play Episode Listen Later Apr 22, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.21.537733v1?rss=1 Authors: mansouri, s., Pessoni, A. M., Rivera, A. M., Tamminga, C. A., Parise, E., Turecki, G., Nestler, E. J., Chen, T.-H., Labonte, B. Abstract: Major depressive disorder (MDD) is one of the most important causes of disability worldwide. While recent work provides insights into the molecular alterations in the brain of patients with MDD, whether these molecular signatures can be associated with the expression of specific symptom domains in males and females remains unclear. Here, we identified sex-specific gene modules associated with the expression of MDD, combining differential gene expression and co-expression network analyses in six cortical and subcortical brain regions. Our results show varying levels of network homology between males and females across brain regions, although the association between these structures and the expression of MDD remains highly sex-specific. We refined these associations to several symptom domains and identified transcriptional signatures associated with distinct functional pathways, including GABAergic and glutamatergic neurotransmission, metabolic processes, and intracellular signal transduction, across brain regions associated with distinct symptomatic profiles in a sex-specific fashion. In most cases, these associations were specific to males or to females with MDD, although a subset of gene modules associated with common symptomatic features in both sexes was also identified. Together, our findings suggest that the expression of distinct MDD symptom domains is associated with sex-specific transcriptional structures across brain regions. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Epigenetic Landscapes During Cancer (Luciano Di Croce)

Play Episode Listen Later Apr 20, 2023 48:02

In this episode of the Epigenetics Podcast, we caught up with Luciano Di Croce from the Center of Genomic Regulation in Barcelona to talk about his work on epigenetic landscapes in cancer. The Di Croce Lab focuses on the Polycomb Complex and its influence on diseases like cancer. Luciano Di Croce started out his research career investigating the oncogenic transcription factor PML-RAR. They could show that in leukemic cells knockdown of SUZ12, a key component of Polycomb repressive complex 2 (PRC2), reverts not only histone modification but also induces DNA de-methylation of PML-RAR target genes. More recently the team focused on two other Polycomb related proteins Zrf1 and PHF19 and were able to characterize some of their functions in gene targeting in different disease and developmental contexts. References Di Croce, L., Raker, V. A., Corsaro, M., Fazi, F., Fanelli, M., Faretta, M., Fuks, F., Lo Coco, F., Kouzarides, T., Nervi, C., Minucci, S., & Pelicci, P. G. (2002). Methyltransferase recruitment and DNA hypermethylation of target promoters by an oncogenic transcription factor. Science (New York, N.Y.), 295(5557), 1079–1082. https://doi.org/10.1126/science.1065173 Richly, H., Rocha-Viegas, L., Ribeiro, J. D., Demajo, S., Gundem, G., Lopez-Bigas, N., Nakagawa, T., Rospert, S., Ito, T., & Di Croce, L. (2010). Transcriptional activation of polycomb-repressed genes by ZRF1. Nature, 468(7327), 1124–1128. https://doi.org/10.1038/nature09574 Jain, P., Ballare, C., Blanco, E., Vizan, P., & Di Croce, L. (2020). PHF19 mediated regulation of proliferation and invasiveness in prostate cancer cells. eLife, 9, e51373. https://doi.org/10.7554/eLife.51373 Related Episodes Oncohistones as Drivers of Pediatric Brain Tumors (Nada Jabado) Transcription and Polycomb in Inheritance and Disease (Danny Reinberg) Targeting COMPASS to Cure Childhood Leukemia (Ali Shilatifard) Contact Epigenetics Podcast on Twitter Epigenetics Podcast on Instagram Epigenetics Podcast on Mastodon Active Motif on Twitter Active Motif on LinkedIn Email: podcast@activemotif.com

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Learning induces unique transcriptional landscapes in the auditory cortex

Play Episode Listen Later Apr 15, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.15.536914v1?rss=1 Authors: Graham, G.-E., Chimenti, M. S., Knudtson, K., Grenard, D. N., Co, L., Sumner, M., Tchou, T., Bieszczad, K. M. Abstract: Learning can induce neurophysiological plasticity in the auditory cortex at multiple timescales. Lasting changes to auditory cortical function that persist over days, weeks, or even a lifetime, require learning-induced gene expression. Indeed, de novo transcription is the molecular determinant for whether transient experiences transform into long-term memories with a lasting impact on behavior. However, auditory cortical genes that support auditory learning, memory, and acquired sound-specific behavior are largely unknown. This report is the first to identify genome-wide changes in learning-induced gene expression within the auditory cortex thought to underlie the formation of auditory memory. Bioinformatic analyses on gene enrichment profiles from RNA sequencing identified biological pathways that include cholinergic synapses and neuroactive receptor interactions. The findings characterize key candidate effectors underlying changes in cortical function that support the formation of long-term auditory memory in the adult brain. The molecules and mechanisms identified are potential therapeutic targets to facilitate long-term and sound-specific changes to auditory function in adulthood and are now prime for future gene-targeted investigations. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Early-life stress and ovarian hormones alter transcriptional regulation in the nucleus accumbens resulting in sex-specific responses to cocaine

stress llc hormones regulation copy cocaine responses conversely suzuki cham early life resulting ovarian nucleus biorxiv transcriptional

Play Episode Listen Later Apr 15, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.14.536984v1?rss=1 Authors: Rocks, D., Jaric, I., Bellia, F., Cham, H., Greally, J. M., Suzuki, M., Kundakovic, M. Abstract: Early-life stress and ovarian hormones contribute to increased female vulnerability to cocaine addiction. Here we reveal molecular substrates in the key reward area, the nucleus accumbens, through which these female-specific factors affect immediate and conditioning responses to cocaine in mice. We find shared involvement of X chromosome and estrogen signaling gene regulation in enhanced conditioning responses seen after early-life stress and during the low-estrogenic state in females. During the low-estrogenic state, females respond to acute cocaine exposure by increasing the accessibility of neuronal chromatin enriched for the binding sites of {Delta}FosB, a transcription factor implicated in chronic cocaine response and addiction. Conversely, high-estrogenic females respond to cocaine by preferential closing of neuronal chromatin, providing a mechanism for limiting cocaine-driven chromatin and synaptic plasticity. We find that physiological estrogen withdrawal, exposure to early-life stress, and absence of the second X chromosome all nullify the protective effect of high-estrogenic state on cocaine conditioning in females. Our findings offer a molecular framework to understand sex-specific neuronal mechanisms underlying cocaine use disorder. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Transcriptional Regulation of the P-bodies Associated Psoriasis Candidate Gene CCHCR1

Play Episode Listen Later Apr 8, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.08.535951v1?rss=1 Authors: Ling, Y. H., Chen, Y. Y., Leung, K. N., Chan, K. M., Liu, W. K. Abstract: CCHCR1 (coiled-coil alpha-helical rod protein 1) is a candidate gene of psoriasis and was recently identified as a novel component in P-bodies, the site for regulating mRNA turnover, and a protein in centrosomes. Little is known about its transcriptional regulation or its functions in cell cycle progression. In this study, CCHCR1 was found in a close (287-bp) head-to-head orientation with its neighboring gene, TCF19. The transcription of the CCHCR1/TCF19 gene pair was controlled by a shared bidirectional promoter and was induced at the G1/S transition of the cell cycle. The 287-bp intergenic sequence was sufficient for the G1/S expression of both genes, but the expression of CCHCR1 was further promoted by the presence of exon 1 of TCF19 and CCHCR1. The expression of the CCHCR1/TCF19 gene pair was dependent on the E2F1 transcription factor. E2F1 binding sites were predicted in the CCHCR1/TCF19 bidirectional promoter by in silico analysis. Overexpression of E2F1 induced the expression of CCHCR1 and TCF19. In addition, E2F1 knockdown inhibited both CCHCR1 and TCF19 expression. Knockdown of CCHCR1 or TCF19 reduced cell count but only depletion of CCHCR1 significantly induced p21 expression, implying that CCHCR1 and TCF19 may both regulate cell growth but in divergent pathways. Taken together, we revealed a bidirectional regulation of the CCHCR1/TCF19 gene pair in the G1/S transition and provide a new perspective to understand the role of CCHCR1 as a candidate gene of psoriasis. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Strain-specific changes in nucleus accumbens transcriptome and motivation for food reward in mice exposed to maternal separation

Play Episode Listen Later Apr 6, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.06.535855v1?rss=1 Authors: Henry, M., Fneich, S., Mathou, A., Xia, L., Foury, A., Benoit, S., Jouin, M., Junien, C., Capuron, L., Jouneau, L., Moisan, M.-P., Delpierre, C., Gabory, A., Darnaudery, M. Abstract: Adversity in childhood exerts enduring effects on brain and increases the vulnerability to psychiatric diseases. It also leads to a higher risk for eating disorders and obesity. We hypothesised that neonatal stress in mice affects motivation to obtain palatable food in adulthood and changes gene expression in reward system. Male and female pups from C57Bl/6J and C3H/HeN mice strains were subjected to a daily maternal separation (MS) protocol from PND2 to PND14. In adulthood, their motivation for palatable food reward was assessed in operant cages. Compared to control mice, male and female C3H/Hen mice exposed to MS significantly did more lever presses to obtain palatable food especially when the effort required to obtain the reward is high. Transcriptional analysis reveals 375 genes differentially expressed in the nucleus accumbens of male MS C3H/HeN mice compared to the control group, some of these being associated with the regulation of the reward system (e.g. Gnas, Pnoc). Interestingly, C57Bl/6J mice exposed to MS did not show any alteration in their motivation to obtain a palatable reward nor significant changes in gene expression in the nucleus accumbens. In conclusion, neonatal stress produces lasting changes in motivation for palatable food in C3H/HeN offspring but has no impact in C57Bl/6J offspring. These behavioural alterations are accompanied by drastic changes in gene expression specifically within the nucleus accumbens, a key structure in the regulation of motivational processes. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Transsynaptic labeling and transcriptional control of zebrafish neural circuits

Play Episode Listen Later Apr 3, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.03.535421v1?rss=1 Authors: Coomer, C., Naumova, D., Talay, M., Zolyomi, B., Snell, N., Sorkac, A., Chanchu, J.-M., Cheng, J., Roman, I., Li, J., Robson, D., Halpern, M. E., Barnea, G. Abstract: Deciphering the connectome, the ensemble of synaptic connections that underlie brain function is a central goal of neuroscience research. The trans-Tango genetic approach, initially developed for anterograde transsynaptic tracing in Drosophila, can be used to map connections between presynaptic and postsynaptic partners and to drive gene expression in target neurons. Here, we describe the successful adaptation of trans-Tango to visualize neural connections in a living vertebrate nervous system, that of the zebrafish. Connections were validated between synaptic partners in the larval retina and brain. Results were corroborated by functional experiments in which optogenetic activation of retinal ganglion cells elicited responses in neurons of the optic tectum, as measured by trans-Tango-dependent expression of a genetically encoded calcium indicator. Transsynaptic signaling through trans-Tango reveals predicted as well as previously undescribed synaptic connections, providing a valuable in vivo tool to monitor and interrogate neural circuits over time. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Efficient genetic editing of human intestinal organoids using ribonucleoprotein-based CRISPR

Play Episode Listen Later Mar 31, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.31.535108v1?rss=1 Authors: Skoufou-Papoutsaki, N., Adler, S., D'Santos, P., Mannion, L., Mehmed, S., Kemp, R., Smith, A., Perrone, F., Nayak, K., Russell, A., Zilbauer, M., Winton, D. J. Abstract: Organoids are currently one of the most widely used ex vivo models in epithelial biology. Combined with genetic editing strategies, organoids offer a promise of rapid and efficient investigation of gene function in many models of human disease. However, to date, the editing efficiency of organoids with the use of non-viral electroporation methods has been only up to 30%, with implications for the subsequent need for selection including including turnaround time and exhaustion or adaptation of the organoid population. Here, we describe an efficient method of intestinal organoid editing using a Ribonucleoprotein CRISPR-based approach. Editing efficiencies of up to 98% in target genes were robustly achieved across different anatomical gut locations and developmental timepoints from multiple patient samples with no off-target editing. The method allowed us to study the effect of the loss of the tumour suppressor gene, PTEN, in normal human intestinal cells. Analysis of PTEN deficient organoids defined phenotypes that likely relate to its tumour suppressive function in vivo, such as a proliferative advantage and increased organoid budding. Transcriptional profiling revealed differential expression of genes in pathways commonly known to be associated with PTEN loss including mTORC1 activation. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Coordinated stimulation of axon regenerative and neurodegenerative transcriptional programs by Atf4 following optic nerve injury

Play Episode Listen Later Mar 31, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.29.534798v1?rss=1 Authors: Somasundaram, P., Farley, M. M., Rudy, M. A., Stefanoff, D. G., Shah, M., Goli, P., Heo, J., Wang, S., Tran, N. M., Watkins, T. A. Abstract: Previously we showed that neurodegeneration initiated by axonal insults depends in part on the stress-responsive kinase Perk (Larhammar et al., 2017). Here we show that Perk acts primarily through Activating Transcription Factor-4 (Atf4) to stimulate not only pro-apoptotic but also pro-regenerative responses following optic nerve injury. Using conditional knockout mice, we find an extensive Perk/Atf4-dependent transcriptional response that includes canonical Atf4 target genes and modest contributions by C/ebp homologous protein (Chop). Overlap with c-Jun- dependent transcription suggests interplay with a parallel stress pathway that couples regenerative and apoptotic responses. Accordingly, neuronal knockout of Atf4 recapitulates the neuroprotection afforded by Perk deficiency, and Perk or Atf4 knockout impairs optic axon regeneration enabled by disrupting the tumor suppressor Pten. These findings contrast with the transcriptional and functional consequences reported for CRISPR targeting of Atf4 or Chop and reveal an integral role for Perk/Atf4 in coordinating neurodegenerative and regenerative responses to CNS axon injury. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Mapping RUNX2 transcriptional dynamics during multi-lineage differentiation of human mesenchymal stem cells

Play Episode Listen Later Mar 30, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.30.534618v1?rss=1 Authors: Govindaraj, K., Kannan, S., Karperien, M., Post, J. N. Abstract: The multi-lineage differentiation capacity of human mesenchymal stem cells (hMSCs) enables its potential for tissue engineering and regenerative medicine. Master transcription factors play a key role during development, differentiation, homeostasis and disease pathology. RUNX2 is the master transcription factor for bone development, and it regulates several important signaling pathways during chondrogenic and osteogenic differentiation of hMSCs. However, modulation of RUNX2 activity during hMSC differentiation into various lineages is not yet fully described. We differentiated hMSCs into chondro-, osteo-, and adipogenic lineages and studied RUNX2 protein dynamics using Transcription Factor - Fluorescence Recovery After Photobleaching (TF-FRAP) at different time points. The TF-FRAP method can capture the dynamic changes of RUNX2 protein mobility at the single cell level resolution, and cluster analysis shows how RUNX2 dynamics change at subpopulation level in proliferating and differentiating hMSCs. Our data show that although whole hMSC population is exposed to differentiation stimuli, some subpopulations in hMSCs do not respond to environmental cues. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Transcriptional Activation of Regenerative Hematopoiesis via Vascular Niche Sensing

Play Episode Listen Later Mar 29, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.27.534417v1?rss=1 Authors: Itkin, T., Houghton, S., Schreiner, R., Lin, Y., Badwe, C., Voisin, V., Murison, A., Seyedhassantehrani, N., Kaufmann, K. B., Garcia-Prat, L., Booth, G. T., Geng, F., Liu, Y., Gomez-Salinero, J. M., Shieh, J.-H., Redmond, D., Xiang, J. Z., Josefowicz, S. Z., Trapnell, C., Spencer, J. A., Zangi, L., Hadland, B., Dick, J. E., Xie, S. Z., Rafii, S. Abstract: Transition between activation and quiescence programs in hematopoietic stem and progenitor cells (HSC/HSPCs) is perceived to be governed intrinsically and by microenvironmental co-adaptation. However, HSC programs dictating both transition and adaptability, remain poorly defined. Single cell multiome analysis divulging differential transcriptional activity between distinct HSPC states, indicated for the exclusive absence of Fli-1 motif from quiescent HSCs. We reveal that Fli-1 activity is essential for HSCs during regenerative hematopoiesis. Fli-1 directs activation programs while manipulating cellular sensory and output machineries, enabling HSPCs co-adoptability with a stimulated vascular niche. During regenerative conditions, Fli-1 presets and enables propagation of niche-derived Notch1 signaling. Constitutively induced Notch1 signaling is sufficient to recuperate functional HSC impairments in the absence of Fli-1. Applying FLI-1 modified-mRNA transduction into lethargic adult human mobilized HSPCs, enables their vigorous niche-mediated expansion along with superior engraftment capacities. Thus, decryption of stem cell activation programs offers valuable insights for immune regenerative medicine. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Microbial short-chain fatty acids regulate drug seeking and transcriptional control in a model of cocaine seeking

Play Episode Listen Later Mar 24, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.22.533834v1?rss=1 Authors: Meckel, K. R., Simpson, S., Godino, A., Peck, E. G., George, O., Calipari, E. S., Hofford, R. S., Kiraly, D. Abstract: Cocaine use disorder represents a public health crisis with no FDA-approved medications for its treatment. A growing body of research has detailed the important connections between the brain and the resident population of bacteria in the gut, the gut microbiome in psychiatric disease models. Acute depletion of gut bacteria results in enhanced reward in a mouse cocaine place preference model, and repletion of bacterially-derived short-chain fatty acid (SCFA) metabolites reverses this effect. However, the role of the gut microbiome and its metabolites in modulating cocaine-seeking behavior after prolonged abstinence is unknown. Given that relapse prevention is the most clinically challenging issue in treating substance use disorders, studies examining the effects of microbiome manipulations in relapse-relevant models are critical. Here, Sprague-Dawley rats received either untreated water or antibiotics to deplete the gut microbiome and its metabolites. Rats were trained to self-administer cocaine and subjected to either within-session threshold testing to evaluate motivation for cocaine or 21 days of abstinence followed by a cue-induced cocaine-seeking task to model relapse behavior. Microbiome depletion did not affect cocaine acquisition on an FR1 schedule. However, microbiome-depleted subjects exhibited significantly enhanced motivation for low dose cocaine on a within-session threshold task. Similarly, microbiome depletion increased cue-induced cocaine-seeking following prolonged abstinence. In the absence of a normal microbiome, repletion of bacterially-derived SCFA metabolites reversed the behavioral and transcriptional changes associated with microbiome depletion. These findings suggest that gut bacteria, via their metabolites, are key regulators of drug-seeking behaviors, positioning the microbiome as a potential translational research target. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Notch1 forms nuclear transcriptional condensates that drive target gene expression.

Play Episode Listen Later Mar 18, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.17.533124v1?rss=1 Authors: Foran, G., Hallam, R. D., Megaly, M., Turgambayeva, A., Li, Y., Necakov, A. Abstract: The Notch receptor is a titratable, context-specific counter of intercellular interactions that translates productive interactions with ligands on neighbouring cells into corresponding changes in gene expression via the nuclear localization of the Notch intracellular Domain (NICD). Using an Optogenetic Notch1 construct in combination with a live imaging transcriptional reporter and super-resolution imaging, we show that the N1ICD activates gene expression through spontaneous self-assembly into transcriptional condensates whose phase separation is driven by C-terminal Intrinsically Disordered Regions (IDR) of the N1ICD. N1ICD condensates recruit and encapsulate a broad set of core transcriptional proteins, thereby facilitating gene expression and promoting super enhancer-looping. We produced a model of Notch1 activity, whereby discrete changes in nuclear NICD abundance is translated into precise changes in target gene expression through the assembly of phase separated N1ICD molecular crucibles that catalyze gene expression in a concentration-dependent manner by enriching essential transcriptional machineries at target genomic loci. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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The FSGS disease gene product and nuclear pore protein NUP205 regulates nuclear localization and activity of the transcriptional regulators YAP and TAZ in podocytes

Play Episode Listen Later Mar 7, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.07.531564v1?rss=1 Authors: Ester, L., Cabrita, I., Ventzke, M., Christodoulou, M., Fabretti, F., Benzing, T., Habbig, S., Schermer, B. Abstract: Background: Mutations in genes encoding nuclear pore proteins (NUPs) cause steroid-resistant nephrotic syndrome (SRNS) and focal and segmental glomerulosclerosis (FSGS). The mechanisms of how NUP deficiency may cause podocyte dysfunction and failure of the kidney filtration barrier are elusive. The tightly controlled activity of the transcriptional effectors of the evolutionarily conserved Hippo pathway YAP and TAZ is essential for podocyte homeostasis. Here we analyze the role of NUPs in controlling YAP/TAZ nuclear import and activity in podocytes. Methods: We used quantitative label-free mass spectrometry to characterize the YAP/TAZ interactomes in podocytes, particularly identifying NUP interactions. Moreover, we specifically studied NUP205 in controlling YAP/TAZ nuclear import and YAP/TAZ-dependent target gene expression. Results: Here we identify the disease-causing nuclear pore proteins NUP107, NUP133, NUP205, and XPO5 as components of YAP and TAZ protein complexes in podocytes. We demonstrate that NUP205 is essential for YAP/TAZ nuclear import. The nuclear interaction of YAP/TAZ with TEAD1 and their transcriptional activity were dependent on NUP205 expression. Furthermore, we identify a feedback regulatory mechanism that controls YAP activity depending on TAZ-mediated NUP205 expression. Conclusion: This study links the disease protein NUP205 with the activity of the transcriptional regulators and Hippo effectors YAP and TAZ and suggests a pathogenic role of YAP/TAZ-deregulation in podocytes in patients with NUP205 mutations. Moreover, this study suggests an important role of YAP/TAZ signaling in human FSGS. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Widespread post-transcriptional regulation of co-transmission

Play Episode Listen Later Mar 2, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.01.530653v1?rss=1 Authors: Chen, N., Zhang, Y., Rivera-Rodriguez, E., Yu, A., Hobin, M., Rosbash, M., Griffith, L. Abstract: While neurotransmitter identity was once considered singular and immutable for mature neurons, it is now appreciated that one neuron can release multiple neuroactive substances (co-transmission) whose identities can even change over time. To explore the mechanisms that tune the suite of transmitters a neuron releases, we developed transcriptional and translational reporters for cholinergic, glutamatergic, and GABAergic signaling in Drosophila. We show that many glutamatergic and GABAergic cells also transcribe cholinergic genes, but fail to accumulate cholinergic effector proteins. Suppression of cholinergic signaling involves posttranscriptional regulation of cholinergic transcripts by the microRNA miR-190; chronic loss of miR-190 function allows expression of cholinergic machinery, reducing and fragmenting sleep. Using a translation-trap strategy we show that neurons in these populations have episodes of transient translation of cholinergic proteins, demonstrating that suppression of co-transmission is actively modulated. Posttranscriptional restriction of fast transmitter co-transmission provides a mechanism allowing reversible tuning of neuronal output. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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Esm-1 mediates transcriptional polarization associated with diabetic kidney disease

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Play Episode Listen Later Mar 2, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.01.530562v1?rss=1 Authors: Gaudet, A., Zheng, X., Kambham, N., Bhalla, V. Abstract: Background: Esm-1, endothelial cell-specific molecule-1, is a susceptibility gene for diabetic kidney disease (DKD) and is a cytokine- and glucose-regulated, secreted proteoglycan, that is notably expressed in kidney and attenuates inflammation and albuminuria. Esm1 has restricted expression at the vascular tip during development but little is known about its expression pattern in mature tissues, and its precise effects in diabetes. Methods: We utilized publicly available single-cell RNA sequencing data to explore the characteristics of Esm1 expression in 27,786 renal endothelial cells obtained from four adult human and three mouse databases. We validated our findings using bulk transcriptome data from an additional 20 healthy subjects and 41 patients with DKD and using RNAscope. Using correlation matrices, we relate Esm1 expression to the glomerular transcriptome and evaluated these matrices with systemic over-expression of Esm-1. Results: In both mice and humans, Esm1 is expressed in a subset of all renal endothelial cell types and represents a minority of glomerular endothelial cells. In patients, Esm1(+) cells exhibit a highly conserved enrichment for blood vessel development genes. With diabetes, these cells are fewer in number and profoundly shift expression to reflect chemotaxis pathways. Analysis of these gene sets highlight candidate genes such as Igfbp5 for cross talk between cell types. We also find that diabetes induces correlations in the expression of large clusters of genes, within cell type-enriched transcripts. Esm1 significantly correlates with a majority genes within these clusters, delineating a glomerular transcriptional polarization reflected by the magnitude of Esm1 deficiency. In diabetic mice, these gene clusters link Esm1 expression to albuminuria, and over-expression of Esm-1 reverses the expression pattern in many of these genes. Conclusions: A comprehensive analysis of single cell and bulk transcriptomes demonstrates that diabetes correlates with lower Esm1 expression and with changes in the functional characterization of Esm1(+) cells. Esm1 is both a marker for glomerular transcriptional polarization, and a mediator that re-orients the transcriptional program in DKD. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Distinct disease mutations in DNMT3A result in a spectrum of behavioral, epigenetic, and transcriptional deficits

Play Episode Listen Later Feb 28, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.27.530041v1?rss=1 Authors: Beard, D. C., Zhang, X., Wu, D. Y., Martin, J. R., Hamagami, N., Swift, R. G., McCullough, K. B., Ge, X., Bell-Hensley, A., Zheng, H., Lawrence, A. B., Hill, C. A., Papouin, T., McAlinden, A., Garbow, J. R., Dougherty, J. D., Maloney, S. E., Gabel, H. W. Abstract: Phenotypic heterogeneity is a common feature of monogenic neurodevelopmental disorders that can arise from differential severity of missense variants underlying disease, but how distinct alleles impact molecular mechanisms to drive variable disease presentation is not well understood. Here, we investigate missense mutations in the DNA methyltransferase DNMT3A associated with variable overgrowth, intellectual disability, and autism, to uncover molecular correlates of phenotypic heterogeneity in neurodevelopmental disease. We generate a DNMT3A P900L/+ mouse model mimicking a disease mutation with mild-to-moderate severity and compare phenotypic and epigenomic effects with a severe R878H mutation. We show that the P900L mutation leads to disease-relevant overgrowth, obesity, and social deficits shared across DNMT3A disorder models, while the R878H mutation causes more extensive epigenomic disruption leading to differential dysregulation of enhancers elements. We identify distinct gene sets disrupted in each mutant which may contribute to mild or severe disease, and detect shared transcriptomic disruption that likely drives common phenotypes across affected individuals. Finally, we demonstrate that core gene dysregulation detected in DNMT3A mutant mice overlaps effects in other developmental disorder models, highlighting the importance of DNMT3A-deposited methylation in neurodevelopment. Together, these findings define central drivers of DNMT3A disorders and illustrate how variable disruption of transcriptional mechanisms can drive the spectrum of phenotypes in neurodevelopmental disease. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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The Kinase Module of the Mediator regulates cell cycle re-entry and transcriptional response following DNA damage

Play Episode Listen Later Feb 26, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.26.530133v1?rss=1 Authors: Memisoglu, G., Bohn, S., Krogan, N., Haber, J. E., Ruthenburg, A. J. Abstract: When faced with a DNA double strand break, cells activate an elaborate signaling cascade called the DNA damage response to protect genomic integrity. To identify novel factors that modulate the DNA damage response to DNA double strand breaks, we performed an epistatic miniarray profile analysis of Mec1 and Rad53, two essential kinases that coordinate the DNA damage response in budding yeast. Through this analysis, we discovered a genetic interaction between the kinase module (CKM) of the Mediator of transcription and Rad53. We find that all four subunits of the CKM, as well as CKM's kinase activity are critical for cell cycle re-entry following a DNA break, whereas the core Mediator subunits are dispensable. Notably, CKM mutants do not impair DNA repair by homologous recombination or confer sensitivity to DNA damaging reagents, suggesting that CKM specifically impinges on DNA damage signaling. In support of this, we find that Rad53 and CKM physically interact in response to DNA damage. Following the induction of a DNA break, CKM is a critical regulator of global transcription inhibition. In addition to this global effect, we illustrate that CKM functions locally at DNA breaks together with the core Mediator. In the absence of catalytically active CKM, the CKM-Mediator complexes at DNA breaks are replaced by RNAPII. Taken together, our results reveal a previously uncharacterized role for CKM in the DNA damage response. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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A Single Cell Time Course of Senescence Uncovers Discrete Cell Trajectories and Transcriptional Heterogeneity

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Play Episode Listen Later Feb 18, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.17.529001v1?rss=1 Authors: Ciotlos, S., Wimer, L., Campisi, J., Melov, S. Abstract: Senescent cells (SnCs) are typically studied as endpoints of a complex transformational process, owing to their frequent maladaptive effects on surrounding tissue and cells. SnCs accumulate with age, and while they ultimately comprise a small percentage of cells in tissues, they have important roles in age associated pathologies. Several obstacles remain in understanding the heterogeneous nature of senescence, and formulating potent beneficial intervention strategies. One approach targets and kills senescent cells (senolysis), and is often driven by a low resolution understanding of SnC identity, which risks both incomplete clearance and off-target effects. Cellular senescence is not a singular binary response, but a range of response trajectories that vary by multiple parameters including inducer and initial cell state. In order to elucidate the developmental trajectories of SnCs, we performed single-cell RNA sequencing on IMR90 lung fibroblasts senescencing across a 12 day time period. Our analysis reveals substantial heterogeneity in gene expression within timepoints and across the full time-course. We uncovered unique markers and differentially regulated pathways in cell populations within each timepoint. Supervised trajectory inference of the time-course data uncovered the root-origin and fates of distinct SnC lineages over 3 stages of senescence induction. Altogether our data provide a novel approach to stud SnC development, identifying cell states of interest, and differentiating between SnCs and quiescent cells. This will aid in identifying key targets for therapeutic intervention in senescence. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Transcriptional heterogeneity in diabetic foot wounds

Play Episode Listen Later Feb 18, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.16.528839v1?rss=1 Authors: Sandoval-Schaefer, T., Phan, Q., Dash, B., Prassinos, A., Duan, K., Gazes, M. I., Vyce, S. D., Hsai, H. C., Driskell, R., Horsley, V. Abstract: Wound repair requires the coordination of multiple cell types including immune cells and tissue resident cells to coordinate healing and return of tissue function. Diabetic foot ulceration is a type of chronic wound that impacts over 4 million patients in the US and over 7 million worldwide (Edmonds et al., 2021). Yet, the cellular and molecular mechanisms that go awry in these wounds are not fully understood. Here, by profiling chronic foot ulcers from non-diabetic (NDFUs) and diabetic (DFUs) patients using single-cell RNA sequencing, we find that DFUs display transcription changes that implicate reduced keratinocyte differentiation, altered fibroblast function and lineages, and defects in macrophage metabolism, inflammation, and ECM production compared to NDFUs. Furthermore, analysis of cellular interactions reveals major alterations in several signaling pathways that are altered in DFUs. These data provide a view of the mechanisms by which diabetes alters healing of foot ulcers and may provide therapeutic avenues for DFU treatments. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

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The Zinc-BED transcription factor Bedwarfed promotes proportional dendritic growth and branching through transcriptional and translational regulation in Drosophila

Play Episode Listen Later Feb 15, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.15.528686v1?rss=1 Authors: Bhattacharjee, S., Iyer, E. P. R., Iyer, S. C., Nanda, S., Rubaharan, M., Ascoli, G. A., Cox, D. N. Abstract: Dendrites are the primary points of sensory or synaptic inputs to a neuron and play an essential role in synaptic integration and neural function. Despite the functional importance of dendrites, relatively less is known about the underlying mechanisms regulating cell-type specific dendritic patterning. Herein, we have dissected functional roles of a previously uncharacterized gene, CG3995, in cell-type specific dendritic development in Drosophila melanogaster. CG3995, which we have named bedwarfed (bdwf), encodes a zinc-finger BED-type protein which is required for proportional growth and branching of dendritic arbors, exhibits nucleocytoplasmic expression, and functions in both transcriptional and translational cellular pathways. At the transcriptional level, we demonstrate a reciprocal regulatory relationship between Bdwf and the homeodomain transcription factor (TF) Cut. We show that Cut positively regulates Bdwf expression and that Bdwf acts as a downstream effector of Cut-mediated dendritic development, whereas overexpression of Bdwf negatively regulates Cut expression in multidendritic sensory neurons. Proteomic analyses revealed that Bdwf interacts with ribosomal proteins and disruption of these proteins produced phenotypically similar dendritic hypotrophy defects as observed in bdwf mutant neurons. We further demonstrate that Bdwf and its ribosomal protein interactors are required for normal microtubule and F-actin cytoskeletal architecture. Finally, our findings reveal that Bdwf is required to promote protein translation and ribosome trafficking along the dendritic arbor. Taken together, these results provide new insights into the complex, combinatorial and multi-functional roles of transcription factors (TFs) in directing diversification of cell-type specific dendritic development. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

growth llc factor bed regulation copy cox promotes zinc branching iyer nanda translational drosophila tfs proportional biorxiv dendritic transcriptional proteomic

Transcriptional networks predating cognition-associated pyramidal lineages are restructured by erythropoietin

Play Episode Listen Later Feb 5, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.04.527116v1?rss=1 Authors: Singh, M., Zhao, Y., Daguano Gastaldi, V., Wojcik, S. M., Curto, Y., Kawaguchi, R., Merino, R. M., Fernandez Garcia-Agudo, L., Taschenberger, H., Brose, N., Geschwind, D., Nave, K.-A., Ehrenreich, H. Abstract: Recombinant human erythropoietin (rhEPO) has potent procognitive effects,hematopoiesis-independent, but underlying mechanisms and physiological role of brain-expressed EPO have remained obscure. Here, we provide encyclopedic transcriptional hippocampal profiling of mice treated with rhEPO. Based on ~108,000 single nuclei, we unmask multiple pyramidal lineages with their comprehensive molecular signatures. By temporal profiling and gene regulatory analysis, we build a developmental trajectory of CA1 pyramidal neurons derived from multiple predecessor lineages and elucidate gene regulatory networks underlying their fate determination. With EPO as tool, we discover novel populations of newly differentiating pyramidal neurons, overpopulating to ~200% upon rhEPO with upregulation of genes crucial for neurodifferentiation, dendrite growth, synaptogenesis, memory formation, and cognition. Using a Cre-based approach to visually distinguish pre-existing from newly formed pyramidal neurons for patch-clamp recordings, we learn that rhEPO treatment differentially affects excitatory and inhibitory inputs. Our findings provide mechanistic insight into how EPO modulates neuronal functions and networks Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

llc networks cognition cre zhao merino nave curto epo brose wojcik restructured lineages ehrenreich biorxiv transcriptional pyramidal geschwind predating ca1 erythropoietin

FDX1 regulates cellular protein lipoylation through direct binding to LIAS.

Play Episode Listen Later Feb 4, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.02.03.526472v1?rss=1 Authors: Dreishpoon, M., Bick, N., Petrova, B., Warui, D. M., Cameron, A., Booker, S. J., Kanarek, N., Golub, T., Tsvetkov, P. Abstract: Ferredoxins are a family of iron-sulfur (Fe-S) cluster proteins that serve as essential electron donors in numerous cellular processes that are conserved through evolution. The promiscuous nature of ferredoxins as electron donors enables them to participate in many metabolic processes including steroid, heme, vitamin D and Fe-S cluster biosynthesis in different organisms. However, the unique natural function(s) of each of the two human ferredoxins (FDX1 and FDX2) are still poorly characterized. We recently reported that FDX1 is both a crucial regulator of copper ionophore induced cell death and serves as an upstream regulator of cellular protein lipoylation, a mitochondrial lipid-based post translational modification naturally occurring on four mitochondrial enzymes that are crucial for TCA cycle function. Here we show that FDX1 regulates protein lipoylation by directly binding to the lipoyl synthase (LIAS) enzyme and not through indirect regulation of cellular Fe-S cluster biosynthesis. Metabolite profiling revealed that the predominant cellular metabolic outcome of FDX1 loss-of-function is manifested through the regulation of the four lipoylation-dependent enzymes ultimately resulting in loss of cellular respiration and sensitivity to mild glucose starvation. Transcriptional profiling of cells growing in either normal or low glucose conditions established that FDX1 loss-of-function results in the induction of both compensatory metabolism related genes and the integrated stress response, consistent with our findings that FDX1 loss-of-functions is conditionally lethal. Together, our findings establish that FDX1 directly engages with LIAS, promoting cellular protein lipoylation, a process essential in maintaining cell viability under low glucose conditions. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

llc copy protein booker binding cellular lias tca fes golub regulates petrova bick biorxiv transcriptional metabolite

Redefining hematopoietic progenitors cells and reaffirming lineage commitment by transcriptional atlas

llc commitment redefining copy cells rna lineage reaffirming biorxiv hematopoietic transcriptional progenitors

Play Episode Listen Later Jan 28, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.01.27.524347v1?rss=1 Authors: Yue, Y. Abstract: Tracing the differentiation of the hematopoietic progenitor cells in human blood, particularly with the scarcity in adults have been challenging owing to the technical limitations and sensitivity. Often, the use of classic and traditional surface markers lacks the sensitivity and specificity to clearly define the hematopoietic progenitors. With the evolution of single cell sequencing approaches, including single cell RNA sequencing (scRNA seq) technologies, this have enable the opportunities to study the heterogeneity and hierarchy of HPCs. Here, scRNA-seq was used to capture, identify and define the characterization of HPCs in adults peripheral blood. In addition, HPCs lineage were reaffirmed by unique makers and key factor genes. Several key factors, including NFE2, LYL1, MYB, and RUNX1 of myeloid and RUNX2, HOXA9, and BCL11A of lymphoid progenitors, were identified as major switches that controlled hematopoietic lineage formation. The reaffirmed lineage commitment also revealed hematopoiesis was a continuum procedure controlled by the activation and suppression of transcription factors, with identified boundary. With these information, an elaborate and comprehensive transcriptional atlas of all types of hematopoietic progenitors was then constructed. Importantly, this newly created atlas can enable accurate definition, hierarchy, and biological insights of hematopoietic progenitors for facilitating new research development in cell engineering, regenerative medicine, and disease treatment. Moreover, this open a wide variety of possibilities for artificial precise control progenitor cells differentiation and targeted treatment of immune disease, providing hope for untreatable diseases. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Wingless signaling promotes lipid mobilization through signal-induced transcriptional repression

Play Episode Listen Later Jan 26, 2023

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.01.25.525602v1?rss=1 Authors: Liu, M., Hemba-Waduge, R.-U.-S., Li, X., Huang, X., Liu, T.-H., Han, X., Wang, Y., Ji, J.-y. Abstract: Conserved Wnt/Wingless signaling plays pivotal roles in regulating normal development and energy metabolism in metazoans, and aberrant activation of Wnt signaling drives the pathogenesis of many diseases including cancer. However, the role of Wnt signaling in regulating cellular lipid homeostasis, particularly lipid mobilization, remains poorly understood. Here we show that canonical Wg signaling inhibits lipid accumulation in Drosophila larval adipocytes by stimulating lipid catabolism while simultaneously inhibiting lipogenesis. Using a combination of RNA-sequencing and CUT&RUN assays, we identified a battery of Wg target genes encoding key factors required for lipogenesis (such as FASN1 and AcCoS), lipolysis (such as lipid droplet-associated proteins Lsd-1 and Lsd-2), and fatty acid {beta}-oxidation in the mitochondria and peroxisome (e.g., CPT1 and CRAT), most of which are directly repressed by active Wg signaling. Furthermore, lipid accumulation defects caused by active Wg signaling are rescued by either ectopically expressing Lsd-1 and Lsd-2 or depleting the transcriptional repressor Aef1, whose binding motif was identified in 52% of Wg signaling-repressed genes. These findings suggest that active Wg signaling reduces intracellular lipid accumulation by inhibiting lipogenesis and fatty acid {beta}-oxidation and by promoting lipolysis and lipid mobilization, and Wg signaling-induced transcriptional repression play a prominent role in these converging mechanisms. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

llc run copy li lsd wang rna huang induced promotes liu ji repression wg mobilization signaling lipids drosophila wnt biorxiv wingless transcriptional crat

Circulation November 15, 2022 Issue

Circulation on the Run

Play Episode Listen Later Nov 14, 2022 23:43

This week, please join authors Qiang Zhang and Matthew Burrage as well as Senior Associate Editor Victoria Delgado as they discuss the article "Artificial Intelligence for Contrast-free MRI: Scar Assessment in Myocardial Infarction Using Deep Learning-Based Virtual Native Enhancement." 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 cohosts. I'm Dr. Carolyn Lam, associate editor from the National Heart Center and Duke National University of Singapore. Dr. Peder Myhre: And I'm Dr. Peder Myhre from University of Akershus University Hospital in Norway. Dr. Carolyn Lam: Peder, today's feature discussion is on AI for contrast-free MRI. Isn't that so cool, using AI to perhaps understand what we could see only with contrast, but now in a contrast-free manner. Now I know that sound a bit confusing, but I hope very, very enticing, because everyone's going to have to wait for a little while before we get to that interesting feature discussion. And for now, let's talk about some of the papers we have in today's issue, shall we? Dr. Peder Myhre: Yes, Carolyn, I can't wait for the feature discussion, but we're going to start with some of the other papers in this week's issue, and we're going to start in the world of preclinical science with a paper looking at human cardiac reprogramming, because Carolyn, direct cardiac reprogramming of fibroblasts into cardiomyocytes has emerged as one of the promising strategies to remuscularize the injured myocardium. Yet it is still insufficient to generate functional induced cardiomyocytes from human fibroblasts using conventional reprogramming cocktails and underlying molecular mechanisms are not really well understood. Transcriptional factors often act in concert and form tightly controlled networks featuring with common targets among different transcriptional factors. Therefore, missing one component during heart development could lead to heart function defects and congenital heart disease. And in this study by corresponding author Yang Zhou from the University of Alabama at Birmingham, the authors perform transcriptomic comparison between human induced cardiomyocytes and functional cardiomyocytes to assess additional factors that govern transcriptional activation of gene programs associated with sarcomere contractility. Dr. Carolyn Lam: Wow. Really nicely explained. Thanks, Peder. So what did they find? Dr. Peder Myhre: So Carolyn, through these computational analysis of transcriptomic data, the authors identified TBX20 as the most under expressed transcription factor in human induced cardiomyocytes compared to endogenous cardiomyocytes. They also demonstrated that TBX20 enhances human cardiac reprogramming and improves contractility and mitochondrial function in the reprogrammed cardiomyocytes. Dr. Carolyn Lam: Nice. Could you summarize the clinical implications, please? Dr. Peder Myhre: Yes. So the clinical implications are that enhancing the efficiency and quality of direct cardiac reprogramming for human fibroblast is a critical step in the clinical translation of this technology, and better understanding of this synergistic regulation of key cardiac transcription factors during reprogramming will provide new insights into the genetic basis in normal and diseased hearts. Well, Carolyn, please tell me about your next paper. Dr. Carolyn Lam: Thanks, and we're moving now to kidney disease. Now end stage renal disease is associated with a high risk of cardiovascular events, but what about mild to moderate kidney dysfunction? Is it causally related to coronary heart disease and stroke? Well, today's authors give us a clue, and it's from corresponding author Dr. Di Angelantonio from University of Cambridge and colleagues who took a very unique combined approach to answer this question. They first conducted observational analyses using individual level data from four huge population based data sources, namely the emerging risk factors collaboration, Epic CVD, Jillion Veteran Program and UK Biobank. Can you imagine this comprised almost 650,000 participants with no history of cardiovascular disease or diabetes at baseline, yielding almost 43,000 and 15,700 incident coronary heart disease and stroke events respectively during a 6.8 million person years of follow up. So huge observational study, which they then followed with a Mendelian randomization analyses using a genetic risk score of 218 variants for GFR and involving participants in Epic CVD Million Veterans Program and the UK Biobank. Dr. Peder Myhre: Wow, Carolyn, this is a topic that I think many of us have really been wondering and thinking about. The mild to moderate kidney dysfunction, what does it really mean? And what a beautiful study to answer this. So what did they find? Dr. Carolyn Lam: First, there was a U-shaped association of creatinine-based GFR with coronary heart disease and stroke with higher risk in participants with GFR values below 60 or more than 105 mills per minute per 1.73 meters squared. Mendelian randomization analyses for coronary heart disease showed an association among participants with GFR below 60, but not for those with GFR above 105. Results were not materially different after adjustment for traditional cardiovascular risk factors and the Mendelian randomization results for stroke were nonsignificant but broadly similar to those for coronary heart disease. So in summary, in people without manifest cardiovascular disease or diabetes, mild to moderate kidney dysfunction is causally related to the risk of coronary heart disease, highlighting the potential value of preventive approaches that preserve and modulate kidney function. Dr. Peder Myhre: Thank you, Carolyn, for such a great summary and an important result from that study. I'm going to now take us back to the world of preclinical science and talk about diabetic cardiomyopathy and exercise. And we both know that patients with diabetes are vulnerable to development of myocardial dysfunction, and that exercise, our favorite thing, for maintaining cardiovascular health, especially in patients with diabetes. And despite a wealth of evidence supporting that cardiometabolic benefits of exercise, the precise exercise responsive signals that confer the beneficial effects of exercise in cardiomyocytes to remain poorly defined. And previous studies have identified fibroblast growth factor 21, FGF21, a peptide hormone with pleiotropic benefits on cardiometabolic hemostasis as an exercise responsive factor. And in this study from Aimin Xu from the University of Hong Kong, the authors investigated a six-week exercise intervention program in FGF21 knockout mice and wild-type litter mates that all had diabetic cardiomyopathy induced by high fat diet and injection of streptozotocin. Dr. Carolyn Lam: Nice. So what did they find? Dr. Peder Myhre: Yeah, the authors found that exercise lowers circulating FGF21 levels, therefore remodeling the heart as an FGF21 sensitive target organ. And the protective effects of exercise against diabetic cardiomyopathy are therefore compromised in mice with deficiency of FGF21. They also identified Sirtuin-3 as an obligor downstream effector on FGF21, preserving mitochondrial integrity and cardiac function. Finally, the authors demonstrated that FGF21 induces Sirtuin-3 expression through AMPK-FOXO3 signaling access. Dr. Carolyn Lam: So could you put that together for us better? So what are the clinical implications? Dr. Peder Myhre: So the clinical implications from this paper is that circulating FGF21 is a potential biomarker for assessment of exercise efficacy in improving cardiac functions. And exercise is a potent FGF21 sensitizer in cardiomyocyte and has the potential to enhance the therapeutic benefits of FGF21 analogs in diabetic cardiomyopathy, and selective activation of FGF21 signal in cardiomyocytes may serve as exercise mimetics and represent a promising targeted intervention for precise management of diabetic cardiomyopathy. Dr. Carolyn Lam: Oh my goodness. That is fascinating. Thank you, Peder. Well let's wrap up with what else there is in today's issue. There's an On My Mind paper by Dr. Weir entitled, “The Emperor's New Clothes: Aren't We Just Treating Grades of Heart Failure with Reduced Ejection Fraction.” Dr. Peder Myhre: And there is a Research Letter by Dr. James Martin from Baylor College of Medicine entitled “Gene Therapy Knockdown of Hippo Signaling Resolves Arrhythmic Events in Pigs after Myocardial Infarction.” Dr. Carolyn Lam: Very nice. Thanks, Peder. So wow, let's go onto a featured discussion on AI for contrast-free MRI and a virtual native enhancement here coming right up. Dr. Peder Myhre: Awesome. Dr. Carolyn Lam: Now we all know that myocardial scar is currently assessed non-invasively using cardiac MRI with late gadolinium enhancement as what we would call the imaging gold standard. Wouldn't it be amazing to have a contrast-free approach, which could provide the same information with many advantages such as a faster or cheaper scan, and without contrast associated problems? Well guess what? We're about to discuss that today in a feature publication in today's issue, and I am so pleased to have the co first authors with us today. They are Dr. Qiang Zhang and Dr. Matthew Burridge, both from University of Oxford, and to discuss it as well, our senior associate editor, Dr. Victoria Delgado from Barcelona. So welcome, everyone. Qiang Zhang, could I start with you and ask you, I understand you're a machine learning expert, which means you're probably smarter than all of us here. Could you maybe explain in simple terms what made you and Dr. Burridge do the study? Dr. Qiang Zhang: First? Thank you so much, Carolyn and Victoria, for the invitation. As you have mentioned, late gadolinium enhancement, or LGE, has been the imaging gold standard in clinical practice for myocardial catheterization including scar assessment for patients with myocardial infarction. However, LGE requires the injection for gadolinium contrast, and this is cautioned in some patient groups and increases the scan time and cost. On the other hand, pre-contrast CMR such as Sydney T1-T2 mapping, a gadolinium-free alternative for myocardial catheterization. But their clinical use has been hindered by confounding factors and a lack of clear interpretation. So with our cross deceptor team at Oxford, we developed an artificial intelligence, virtual native enhancement technique VNE. It can produce a sort of a virtual LGE image but without the need for gadolinium contrast. And we have previously tested it in patients with hypertrophic cardiomyopathy as published in this journal last year. And in this new study together with Matt here, we tested in patients with history of chronic or prior myocardial infarction. Dr. Carolyn Lam: Oh wow. Cool. So audience, you heard it. Instead of LGE, we now have VNE, virtual native enhancement. That's super cool. Thank you. Matt, could I bring you in here? So tell us a little bit more about the population you studied and what you both found. Dr. Matthew Burrage: Yeah, absolutely. And thank you so much for the invitation as well. So as Chang has said, this was a single sensor study that we performed at the University of Oxford and specifically targeting assessing myocardial scar in patients with a history of chronic or prior MI. So we had two sources for our population data. Well, first we used our real world clinical service data from our institution. So we screened 11 years worth of patient data for presence of MI. So patients were included. There was a evidence of a previous MI based on an ischemic pattern of LGE, but we specifically excluded patients who had an acute presentation, or if there were features of acute MI on the CMR scan such as presence of myocardial edema or microvascular obstruction. The reason for this is we wanted to keep this as a clean population to avoid the potential confounding effects of myocardial edema or MVO on native T1 values. And so we also excluded other myocardial pathologies such as underlying cardiomyopathies and infiltrative diseases. A second population dataset came from the OX Army study, which is a single center prospective study of patients presenting with acute MI. And for these patients we used their six month follow up scan to again avoid the confounding effects of edema and pathology. So overall we had a total of 912 patients who have contributed over 4,000 image data sets. The patient characteristics, 81% were male, they had a mean age of 64 years and there were cardiovascular risk factors such as diabetes melitis, hypertension, hypercholesterolemia in 20 to 40% of patients, while just over half had a history of previous revascularization. We also separately applied the VNE technology to a pig model of myocardial infarction, which was thanks to our collaborator, Rohan Domakuma in the US. And so those were scans performed eight to nine weeks after an induced MI in the LAD territory in a series of pigs. And so this gave us the ability to provide a direct comparison between LGE, VNE, and histopathology in this model. Dr. Carolyn Lam: Wow. And results? Dr. Matthew Burrage: So what we found and the key results were firstly that VNE provided significantly better image quality than LGE, and this was on blinded analysis by five independent operators from our test data sets. Secondly, the VNE correlated strongly with LGE in terms of quantifying infarct size and the degree of transmurality, so the extent of the MIs in our test data set. We had pretty good overall accuracy of 84% for VNE in detecting scar compared to LGE with no false positive VNE cases. And finally there was also excellent visuospatial agreement with the histopathology in the pig model of myocardial infarction. So really this, we think, is a technology that provides clinicians with images in a format that firstly they're familiar with, which looks like LGE, provides essentially the same information as LGE, but it can be achieved without the need for any gadolinium contrast agents and can be acquired in a fraction of the time. So it takes less than one second to generate the VNE image. So as we've said before, we feel there's a lot of potential here for this technology to potentially eliminate the need for gadolinium contrast in a significant proportion of CMR scans, reduced scan times and costs, increased clinical throughput and hopefully improve the accessibility of CMR for patients in the near future. Dr. Carolyn Lam: Oh wow. That is tremendous. So first of all, congratulations to both of you. Before I ask Victoria for some thoughts, could I also just check with Qiang Zhang, because all AI algorithms need to be externally validated or surely there's some catch to it, or so-called limitations, or something else you may study. Could you maybe round up by saying is there anything that clinicians should not be applying it to or be aware of some limitations or? Dr. Qiang Zhang: Thank you, Carolyn. So a limitation of this study is that the dataset that is used for developing the models, the majority of them are patients around six month after the acute infarction. So where the myocardial infarction is still evolving, which may include residual edema and microvascular obstruction, and that is difficult to assess using the current VNE model. And also we found it challenging to assess small sub endocardial infarction and actually to address those limitations, we are working on improving the VNE models, training it on even larger data sets and training it on LGE to detect small sub endocardial function. And we will further develop it to detect, for example, acute edema and a microvascular obstruction, and in the meantime develop quality control driven AI models to inform the clinical users of and unreliable results. Dr. Carolyn Lam: Wow, thank you. So Victoria, now I'm dying to hear your thoughts. How do you think this fits in the landscape of all AI imaging now? Dr. Victoria Delgado: I think that it's an excellent development and I congratulate the others for the article and the proof of concept that we can move away from the late enhancement and the use of gadolinium enhancement. I think that this is a major step forward because as Matt said, they are going to decrease very much the time of scanning and the post processing because is automatically done as far as I understand. So even if you can interpret yourself the amount of so-called virtual enhancement, the system gives you a value for that extension of the virtual in non-gadolinium enhancement. So that reduces very much the variability that can be in each observer if that is done automatically. But my question to them is also if that can be influenced by the type of scanner that you use, for example on echocardiography, that's much more my field of interest, it depends very much sometimes how the images are processed of which are the vendors that we have used to acquire the images. Is this a limitation for your software? Can you foresee there some variability or is completely independent? Dr. Qiang Zhang: Thank you, Victoria. So we are aware of actually the difference of the data produced by different scan of vendors and the advantage of AI-driven methods is that it is data driven. So we plan to incorporate dataset from other vendors so that the trend that VNE models can work with like multiple scanner vendors. This actually will be done alongside the ongoing standardization program of T1 mapping in our group, which is the underpinned technology for VNE. And this is led by Professor Stephan Pitchnik and Vanessa Farrera. And we actually hope the VNE technology as AI driven methods could contribute to a solution to the CMO standardization between the scanner vendor. Dr. Victoria Delgado: And another question, if I may follow in this CMR, it has been proposed as a very valuable imaging technique to assess infarct size and to see the efficacy of some therapies to reduce the myocardial infarction size. How do you think that this new methods will impact in future trials and the way we have been interpreting the previous trials, like for example, the one that you use for the validation? Dr. Matthew Burrage: Yeah, thanks Victoria. It's a really, really excellent question. I think there's a lot of potential for the new VNE technology to also become a clinical endpoint in some of these trials in terms of reduction in infarct size, because the information that we get is more or less the same as we get from the LGE. So there's lots of potential that we can, again, use this as a biomarker in trials for looking at reduction in infarct size and reperfusion therapies. But it has the benefit that it can be done quicker and without gadolinium contrast. Dr. Victoria Delgado: This is amazing guideline and really I would have a lot of questions for them as well. And knowing the literature, for example, in the Scenic center in Madrid that they have been scanning the evolution of myocardial infarction from 0.02 weeks to see how this would translate with your technique. That will be amazing to understand how this can be done. Dr. Carolyn Lam: Oh wow, there you go. New research idea right there. Well how about if we end with a very quick question for each of the first authors. So maybe Matt, you could start, I mean is this ready for primetime and clinical use? And if it's not, what needs to be done to get there? In other words, where are you headed as the next step? Dr. Matthew Burrage: So again, thank you, Carolyn, that's a really excellent question and I think the next step before this becomes ready for primetime clinical use is validating this technology really across the spectrum of other myocardial pathologies. So the next work that we are developing this on is in patients with acute myocardial infarction, and then extending this to sort of acute inflammatory conditions like myocarditis, other non-ischemic cardiomyopathies, things like amyloidosis as well. So this will be the next step into rollout and we are looking to track things like VNE burden and how that relates to clinical outcomes, similar to the previous LGE papers have done across different myocardial pathologies, but then ultimately aiming towards clinical rollout within the next few years. Dr. Qiang Zhang: Yeah, I think pretty much what Matt has said, we're going to develop the deep learning methods and test it further on pretty much the whole spectrum of commonly encountered diseases, and then more complex pathologies such as acute pathologies like edema, microvascular obstruction, and then we test on large population study like UK Biobank and other prospective clinical trials. And of course the most importantly is to roll out for real world clinical use. And as Matt said, we are aiming to do this within the next two to five years. Dr. Carolyn Lam: Wow, this is amazing. Both Victoria and I said thank you, congratulations on this landmark piece of work. Thank you for publishing it in circulation. Audience, thank you for joining us today from Greg, Peder, myself. You've been listening to Circulation on the Run, and don't forget to tune in again next week. Dr. Greg Hundley: 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, please visit ahajournals.org.

October 2022 Discover Circ Res

Discover CircRes

Play Episode Listen Later Oct 20, 2022 29:58

This month on Episode 41 of Discover CircRes, host Cynthia St. Hilaire highlights four original research articles featured in the September 30 and October 14 issues of Circulation Research. This episode also features an interview with Dr Kory Lavine and Dr Chieh-Yu Lin from Washington University St. Louis, to discuss their study, Transcriptional and Immune Landscape of Cardiac Sarcoidosis. Article highlights: Tian, et al. EV-Mediated Heart Brain Communication in CHF Wleklinski, et al. Impaired Dynamic SR Ca Buffering Causes AD-CPVT2 Masson, et al. Orai1 Inhibition as a Treatment for PAH Li, et al. F. Prausnitzii Ameliorates Chronic Kidney Disease Cindy St. Hilaire: Hi, and welcome to Discover Circ Res, the podcast of the American Heart Association's journal, Circulation Research. I'm your host, Dr Cynthia St. Hilaire from the Vascular Medicine Institute at the University of Pittsburgh, and today I'm going to highlight articles from our September 30th and October 14th issues of Circulation Research. I'm also going to have a chat with Dr Kory Lavine and Dr Chieh-Yu Lin from Washington University St. Louis, and we're going to discuss their study Transcriptional and Immune Landscape of Cardiac Sarcoidosis. But before I get to the interview, I'm going to highlight a few articles. Cindy St. Hilaire: The first article I'm going to share is Extracellular Vesicles Regulate Sympathoexcitation by Nrf2 in Heart Failure. The first author of this study is Changhai Tian, and the corresponding author is Irving Zucker, and they are at University of Nebraska. After a myocardial infarction, increased oxidative stress in the heart can contribute to adverse cardiac remodeling, and ultimately, heart failure. Nrf2 is a master activator of antioxidant genes, suggesting a protective role, but studies in rats have shown its expression to be suppressed after MI, likely due to upregulation of Nrf2-targeting microRNAs. These microRNAs can also be packaged into vesicles and released from stressed heart cells. Now, this group has shown that rats and humans with chronic heart failure have an abundance of these microRNA-containing EVs in their blood. In the rats with chronic heart failure, these extracellular vesicles were found to be taken up by neurons of the rostral ventrolateral medulla, RVLM, wherein the microRNA suppressed Nrf2 expression. The RVLM is a brain region that controls the sympathetic nervous system, and in the presence of EVs, it is ramped up by sympathetic excitation. Because such elevated sympathetic activity can induce the fight or flight response, including increased heart rate and blood pressure, this would likely worsen heart failure progression. The team, however, found that inhibiting microRNAs in the extracellular vesicles prevented Nrf2 suppression in the RVLM and sympathetic activation, suggesting the pathway could be targeted therapeutically. Cindy St. Hilaire: The next article I want to highlight is titled, Impaired Dynamic Sarcoplasmic Reticulum Calcium Buffering in Autosomal Dominant CPVT2. The first author of this study is Matthew Wleklinski, and the corresponding author is Bjӧrn Knollmann, and they are at Vanderbilt University. Exercise or emotional stress can prompt the release of catecholamine hormones, which induce a fast heart rate, increased blood pressure, and other features of the fight or flight response. For people with catecholaminergic polymorphic ventricular tachycardia, or CPVT, physical activity or stress can cause potentially lethal arrhythmias. Mutations of calsequestrin-2, or CASQ2, which is a sarcoplasmic reticulum calcium-binding protein, is a major cause of CPVT, and can be recessive or dominant in nature. For many recessive mutations, disease occurs due to loss of CASQ2 protein. This group investigated a dominant lysine to arginine mutation in this protein, and found by contrast, protein levels remain normal. In mice carrying the mutation, not only was the level of CASQ2 comparable to that in control animals, but so, too, was the protein's subcellular localization. The mutation instead interfered with CASQ2's calcium binding or buffering capability within the sarcoplasmic reticulum. The result was that upon catecholamine injection or exercise, the unbound calcium released prematurely from the sarcoplasmic reticulum, triggering spontaneous cell contractions. In uncovering this novel molecular etiology of CPVT, the work provides a basis for studying the consequences of other dominant CASQ2 mutations. Cindy St. Hilaire: The next article I want to highlight is from our October 14th issue of Circulation Research, and the title of the article is ORAI1 Inhibitors as Potential Treatments for Pulmonary Arterial Hypertension. The first author is Bastien Masson, and the corresponding author is Fabrice Antigny, and they're from Inserm in France. In pulmonary arterial hypertension, the arteries of the lungs become progressively obstructed, making it harder for the heart to pump blood through them, ultimately leading to right ventricular hypertrophy and heart failure. A contributing factor in the molecular pathology of pulmonary arterial hypertension is abnormal calcium handling within the pulmonary artery smooth muscle cells. Indeed, excess calcium signaling causes these cells to proliferate, migrate, and become resistant to apoptotic death, thus leading to narrowing of the vessel. This group now identified the calcium channel ORAI1 as a major culprit behind this excess signaling. Samples of lung tissue from pulmonary arterial hypertension patients and a pulmonary arterial hypertension rat model had significantly upregulated expression of this channel compared with controls. And in patient pulmonary arterial smooth muscle cells, the high ORAI1 levels resulted in heightened calcium influx, heightened proliferation, heightened migration and reduced apoptosis. Inhibition of ORAI1 reversed these effects. Furthermore, in pulmonary hypertension model rats, ORAI1 inhibition reduced right ventricle systolic pressure and attenuated right ventricle hypertrophy when compared with untreated controls. This study indicates that ORAI1 inhibitors could be a new potential target for treating this incurable condition. Cindy St. Hilaire: The last article I want to share is titled Faecalibacterium Prausnitzii Attenuates CKD via Butyrate-Renal GPR43 Axis. The first author of this study is Hong-Bao Li, and the corresponding author is Tao Yang, and they are from the University of Toledo. Progressive renal inflammation and fibrosis accompanied by hypertension are hallmarks of chronic kidney disease, which is an incurable condition affecting a significant chunk of the world's population. Studies indicate that chronic kidney disease is linked to gut dysbiosis. Specifically, depletion of lactobacillus bifidobacterium and faecalibacterium, prompting investigations into the use of probiotics. While supplements including lactobacillus and bifidobacterium have shown little effectiveness in chronic kidney disease, supplementations with F. prausnitzii have not been investigated. Now, this group has shown in a mouse model of chronic kidney disease that oral administration of F. prausnitzii has beneficial effects on renal function, reducing renal fibrosis and inflammation. This bacterial supplementation also produced the short chain fatty acid butyrate, which was found to be at unusually low levels in the blood samples from the CKD model mice and from chronic kidney disease patients. Oral supplementation with this bacterium boosted butyrate levels in the mice, and in fact, oral administration of butyrate itself mimicked the effects of the bacteria. These findings suggest that supplementation with F. prausnitzii or, indeed, butyrate could be worth investigating as a treatment for chronic kidney disease. Cindy St. Hilaire: Today I have with me Dr Kory Lavine and Dr Chieh-Yu Lin from Washington University St. Louis, and we're going to talk about their paper, Transcriptional and Immune Landscape of Cardiac Sarcoidosis. This is in our September 30th issue of Circulation Research. Welcome, and thank you for taking the time to speak with me today. Chieh-Yu Lin: Thank you for inviting us. It's a great honor to be here today. Kory Lavine: Thank you. Cindy St. Hilaire: Really great paper, ton of data, and hopefully, we can pick some of it apart. But before we get into it, I actually want to just talk about sarcoidosis generally. I know it's a systemic inflammatory disease that has this kind of aggregation of immune cells as its culprit, and it can happen in a bunch of different organs. It's mostly in the lung, but it's also, like you're studying, in the heart. Can you just give us a little bit of background? What is sarcoidosis, and how common is cardiac sarcoidosis? Chieh-Yu Lin: Well, this is actually a great question, and I'll try to answer it. You actually capture one of the most important kind of features for sarcoidosis. It happens in all kind of organ system, mostly commonly in lung, in lymph nodes, but also in heart, spleen, even in brain, or even orbit, like eyes. It's really a truly multisystemic disease that has been characterized by this aggregate of macrophages, or myeloid cells, with scattered multinucleated giant cells, as the name implies, have multiple nuclear big, chunky, cells that form an aggregate. That's kind of like a pathognomonic feature for sarcoidosis, whether it's happening in lung, in the heart. When any organ system, a lot of studies has been done, but as of now, a very clear pathogenesis or mechanism has been, I would say, still pretty elusive, or still remain quite unclear, despite all the great effort has been made in this field. The other thing is that a lot of the studies actually focusing on pulmonary sarcoidosis for good reasons. Actually, that's one of the most common manifestations. For cardiac sarcoidosis, although it's only effect in probably, I would say depends on the data, 20% to 30% of the outpatient that with sarcoidosis, with or without lung involvement. It's actually carry a very significant clinical implications as of matter that the presentation of cardiac sarcoidosis can be devastating and sometimes actually fatal. Some of the study actually show that cardiac sarcoidosis actually higher, up to 80%, just because the first presentation's actually, unfortunately, sudden cardiac death. That's why Kory and I, we teamed up. I'm a cardiothoracic pathologist, so in my clinical practice I see specimens and samples from human body, from patient suffer from sarcoidosis, both in lung, lymph node, and heart. Kory is an outstanding heart failure, heart transplant cardiologist, see the other end, which is the patient care. This disease, specifically in heart, its presentation and its pathogens in heart, really attracts our attention. Cindy St. Hilaire: Do we know any or some of the potential causes? Why it would start, maybe in a different patient population, but also in the heart versus the lung? Do we know anything about that process? Kory Lavine: We know nothing about it. Sarcoid has no known etiology. There's been thoughts in the past that it may be driven by infection, the typical pathogens or autoimmune ideologies, but really, there's little data out there to support those possibilities. Right now, the field's wide open. The other challenge is we don't really have a good way to treat this disease, so a lot of the therapies available are things like steroids, which can have some effect on the disease but carry a lot of risk of complications. The other agents that we sometimes use to lower the doses of steroids, things like methotrexate and azathioprine, are only modestly effective. These are really the motivation for Chieh-Yu and myself to pursue this. We don't really know what causes the disease, and we don't really have very good treatments. We really wanted to take the first step, that's to study the real disease, and understand what are the pathologic cell types that are present within the granuloma, which is these aggregation of immune cells that Chieh-Yu was speaking about. Cindy St. Hilaire: What is actually happening at the beginning of this disease? These granulomas form, and then what is the pathological progression in the heart? What goes on there? Chieh-Yu Lin: This is actually another great question that I will say there's not much that has been discovered because, especially in human tissue, every time we have a sample, it's actually a kind of time point. We cannot do a longitudinal study. But in general speaking, very little is known about how it's initiated because it will need to accumulate to a certain disease burden for this to have a clinical symptom sign and be manifested, and then being clinically studied. We do know that in both heart and lung after treatment of progressions, it's usually in, a general speaking, going through a phase from a more proliferative means that it's creating more granulomas, more inflammatory cell aggregate, to a more fibrotic phase. Means that sometimes you actually see the granuloma start to disappear or dissipate, and then showing this kind of dense collagen and fibrosis. That has been commonly documented in both lung and heart sarcoidosis. The other things is that very difficult to study this disease that we do not have a great animal model, so we cannot use animal model to try to approximate or really study the disease pathogenesis. There are several animal models they try to use microbacteria or infectious agents, and these infectious agents can create morphologically similar granuloma, per se, but just like in human body. For instance, patients suffer from TB in their lung, biopsy will show this. But clinically, these are two very distinct disease entities, even though they look alike. Even in the heart, one of the conditions that we study in our paper is giant cell myocarditis, as the name implying having multinucleated giant cells granuloma. It looks really alike under microscopy for pathologists like me, but their clinical course in response to treatment is drastically different. This type of barriers and in the current limitations of our study tool makes, as Kory just said, this is really a wide open. We just know so little despite all the effort. Cindy St. Hilaire: Yeah. I'm guessing based on this granuloma information, to start with, the obvious question you went after is going after the immune cell populations that possibly contribute to sarcoidosis. To do this, because you have the human tissue, you went for single cell transcriptional profiling, which is a great use of the technology. But what biological sources did you use, and how did you go about choosing patient? Because the great thing about single cell is you can do just that, you can look at however many thousands of cells in one patient. But how do you make sure or check that that is broadly seen versus just a co-founding observation in that patient? Kory Lavine: We use explanted hearts and heart tissue from patients that underwent either heart transplantation or implementation of LVADs. It's a pretty big hunk of myocardium, and we're lucky to work with outstanding pathologists both at WashU, JU, as well as our collaborators at Duke. Between the two institutions, we're able to pull together a collection of tissues where we knew there were granulomas within that piece of tissue we analyzed. You bring up an important challenge. You need to make sure the disease and cause of the disease is present in the tissue that you're analyzing, otherwise you'll not come up with the data that really is informative. Chieh-Yu Lin: Kory beautifully answered the question, but I just wanted to add one little thing, and that's also why we use various different modalities. Some of them is more inside you, like the NanoString Technologies' spatial transcriptomic. You can visualize and confirm that we are studying the phenomenon that has been described for sarcoidosis, and then using multichannel immunofluorescence to validate our sequencing data, to complement such limitations of certain technology. Cindy St. Hilaire: Especially, I feel like with this diseased tissue that it's such a large tissue, there's so much information, it's really hard to dig in and figure out where the signal is. This was a wonderful paper for kind of highlighting, integrating all these new technologies with also just classical staining. Makes for great pictures as well. How does this cellular landscape of cardiac sarcoidosis compare to a normal heart? What'd you find? Chieh-Yu Lin: This is a great question. Compared to normal heart, we have been talking about this accumulation of macrophages with scattered multinucleated giant cells. For the similar landscape, first and foremost, you do not see those type of accumulations in brain microscopy or by myeloid markers in the heart. Although, indeed, in even normal heart tissue we have rest and macrophages. It just doesn't form such morphological alterations. But then we dive deep into it, and then we found that from a different cell type perspective, we realized that the granuloma is composed by several different type of inflammatory cells, with most of the T cells and NKT cells kind of adding periphery. The myeloid cells, including the multinucleated giant cells also, are kind of in the center of the granuloma of the sarcoidosis. Then, we further dive in and realize that there are at least six different subtype of myeloid cells that is contributing to the formation of this very eye-catching distinctive granular malformations, and to just never feel first off and foremost, of course, is those multinucleated giant cells that is really distinct, even on the line microscopy] routine change stand. And then we have a typical monocyte that's more like a precursor being recently recruited to the heart, and we finally sent the other four different type of myeloid cell that carry different markers, and then improving the resident macrophages. Especially for me as a pathologist, I'm using my eye and looking at stand every day, is actually these six type of cells, myeloid cells, actually form a very beautiful special kind of distribution with the connections or special arrangement with all different type, kind of like multinucleated giant cell in the middle, flanked by HLA-DR positive epithelioid macrophages, kind of scatter, and then with dendritic cells and a typical monocyte at the peripheral, and then resident macrophage kind of like in the mix of the seas of granuloma information. All these are distinct from normal heart tissues that does carry a certain amount of macrophages, but just don't form this orchestrated architectural distinct structure that's composed of this very complicated landscape. Cindy St. Hilaire: Those images, I think it was figure six, it's just gorgeous to look at, the model you made. One of the questions I was thinking is there must be a significance between these cells that are on the periphery and those that are in the center of this granuloma. Do you have an idea or can we speculate as to are some more cause and some more consequence of the granuloma? Were you able to capture any more information about maybe the initiating steps of these from your study? Kory Lavine: That's a great question, and a question the field has had for a long time. Now, we know there's different populations of cells. The single cell data allows us to understand what are the transcriptional differences and distinctions between them to gain some insights. One thing that we do know from the field is that disease activity correlates with mTOR activity within these granulomas. We took advantage of phospho-S6 kinase staining as a downstream marker of mTOR activity, and Ki-67 is a marker of self proliferation. Which of these populations within the granuloma might be most active with respect to mTOR and respect to proliferation? If you ask most people in the field, they would jump up and say, "It's the giant cell in the middle." We found that that's not actually the case at all. It's the macrophages that surround the giant cell, the ones that are HLA-DR positive, the epithelioid macrophages, and the ones that are SYLT-3 positive that are scattered around them. That's really interesting and could make a lot of sense, and leads to hypothesis that perhaps activation mTOR signaling within certain parts of the granuloma might be sufficient to set up the rest of the architecture. That's something that we can explore in animal models, and are doing so to try to create a cause and effect relationship. Cindy St. Hilaire: Yeah, and I was actually thinking about this, too, in relation to kind of the resident macrophages versus infiltrating macrophages or even just infiltrating immune cells. Do you know the original source of the cells that make up the granuloma? Is it mostly resident immune, or are they recruited in? Kory Lavine: We can make predictions from the single cell data where you can use trajectory analysis to make strong predictions about what the origin of different populations might be. What those analyses predicted is that the giant cells and the cells that surround the giant cells, the HLA-DR positive and SYLT-3 positive macrophages, come from monocytes. That's the prediction, and, of course, resident macrophages do not. However, that prediction has to be tested, and that's the beauty and importance of developing animal models. The wonderful thing today is we now have genetic tools to do that. We can ask that question. Cindy St. Hilaire: I don't know. Maybe you don't want to spoil the lead of the next paper, but what kind of mouse model are you thinking about trying? Kory Lavine: Yeah. First of all, let me talk about the tools that are available, because they're published in Circulation Research, of course. We have a nice tool to specifically mark, track and delete in tissue resident macrophages using a CX3CR1 ERT pre-mouse, and taking advantage of the concept that tissue macrophages don't turn over from monocytes and turn over from themselves. We can give tamoxifen to label all monocytes macrophages in Dcs with that CRE, and then wait a period of time where only the resident macrophages remain labeled. We can use that trick to modulate mTOR signaling as a first step, and ask whether mTOR signaling is required in that population. We've now developed a new genetic tool to do the same thing in just recruited macrophages. Cindy St. Hilaire: What was the most challenging aspect of this study? There's a lot of moving parts. I'm sure probably the data analysis alone is challenging, but what would you say is the most challenging? Kory Lavine: I think you alluded to this early on, but the most challenging thing is collecting the right tissues to analyze, and that's not a small feat or a small effort here. All the technologies are a lot of fun, and everything works so well today compared to many years ago when we trained, so it's an exciting time to do science. The most challenging and time-consuming component was assembling a group of tissues that we could do single-cell sequencing on between our group and our colleagues at Duke, and then creating validation cohorts that we did across several different institutions, including our own as well as Stanford. That team effort in building that team is the most important, challenging, and honestly, enjoyable part of this. Chieh-Yu Lin: I cannot agree more what Kory just said. I think that that's the challenging and the fun part, and that we're very fortunate to really have a great team to tackle this questions in multiple from multiple institute. I just want to add one more thing that, particularly for me as a cardiopathologist, one of the hardest things is I've known how to look or diagnose sarcoidosis for years, but seeing the data emerging that is so complicated and then beyond my reliable eyes in understanding, it's kind of mentally very challenging but very fun to really open and broaden the vision. It's not just how it looks like just giant cells in macrophages. Cindy St. Hilaire: What do you think about in terms of diagnostics or even potential therapies? How do you think this data that you have now can be leveraged towards those objectives, whether it's screening for new cell types that are really key to this granuloma formation versus therapeutically targeting them? Kory Lavine: This study opens new doors, and right now, diagnosis of sarcoids islimited by trying to biopsy, which, in the heart, is limited by sample bias. You certainly can biopsy the wrong area because you don't know whether a granuloma is in the area or not. We do do some cardiac and other imaging studies like FDG-PET scans, which are helpful but are not perfect, and each of them has their individual limitations. One of the beauties of our study is it identifies new markers of macrophage populations that live within the granuloma, many of which are unique to this disease. That suggests that there's maybe an opportunity to develop imaging tracers that can identify those populations more specifically than our current PET imaging studies do, which rely simply on glucose uptake. It also opens up the possibility that we may able to take blood samples and identify some of these cell types within the blood, and have more simple testing for our patients. I think in terms of therapy, you alluded to it earlier, these concepts about mTOR signaling, that could be a new therapeutic avenue that needs to be rigorously explored in preclinical models. We're lucky already to have very good mTOR inhibitors available in clinical practice today. Cindy St. Hilaire: Obviously, opening new doors is amazing because it's more information, but often a good study leads to even more questions to be asked. What question, or maybe what questions, are you guys going to go after next? Chieh-Yu Lin: Well, that list is very long, and then that's actually the exciting thing about doing this research. There's no bad questions, in some sense. All the way from diagnosis, management, monitoring, therapeutic, how we predict where the patient can respond, that's the whole clinical side. Even the basic science side, we still haven't really answered the question, although our data suggests where that multinucleated giant cells coming from. It's very eye catching. How do they form, even though our data suggests it's from the recruited macrophages. But that's still a long way from the recruited macrophage, monocyte to that gigantic bag of nuclei in the very fluffy cytoplasm. And then, how the granuloma, as we discussed earlier in this discussion, really initially from a relatively normal background myocardium to form this disease process. There are just so many questions that we can ask. There are, of course, several fronts that we would like to focus on. Kory already nicely listed some of them. First and foremost is actually to establish animal model to enable us to do more details in mechanistic studies, because human tissue, as good as it is, it's kind of like a snapshot, just one time point, and it really limits our ability to test our hypothesis. Animal model, certainly, is one of the major directions that we are going forward, but also the other side, like more clinical science also to develop novel noninvasive methodologies to diagnose and to hopefully monitor this patient population in a better way. Cindy St. Hilaire: Well, it's beautiful work. I was actually reading this paper this weekend at a brunch place just next door to my house, and the guy sitting next to me happened to see over my shoulder the title and said that his father had passed away from it. This is hopefully going to help lots of people in the future, and really help to make the models that we need to ask, "What's happening in this disease?" Thank you so much for taking the time to speak with me, and congratulations on what seems to be a landmark study in understanding what's going on in this disease. Chieh-Yu Lin: Thank you so much. It's a pleasure. Cindy St. Hilaire: That's it for our highlights from the September 30th and October 14th issues of Circulation Research. Thank you so much for listening. Please check out the Circ Res Facebook page, and follow us on Twitter and Instagram with the handle @CircRes, and hashtag Discover Circ Res. Thank you so much to our guests, Dr Kory Lavine and Dr Chieh-Yu Lin from Washington University St. Louis. This podcast is produced by Ashara Retniyaka, edited by Melissa Stoner, and supported by the editorial team of Circulation Research. Some of the copy texts for highlighted articles was provided by Ruth Williams. I'm your host, Dr Cynthia St. Hilaire, and this is Discover Circ Res, your on-the-go source for the most 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 of the American Heart Association. For more information, please visit ahajournals.org.

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