Medizin - Open Access LMU - Teil 09/22

Na+ channel currents of rat motor and sensory nerve fibres were studied with the patch-clamp technique on enzymatically demyelinated axons. Differences between motor and sensory fibres in multi-channel inactivation kinetics and the gating of late single-channel currents were investigated. In the axon-attached mode, inactivation of multi-channel Na+ currents in sensory axons was best fitted with a single time constant while for motor axons two time constants were needed. Late single-channel currents in sensory axons were characterized by short openings whereas motor axons exhibited additional long single-channel openings. In contrast, in excised, inside-out membrane patches, no differences between motor and sensory fibres were found: in both types of fibre inactivation of multi-channel Na+ currents proceeded with two time constants and late single-channel currents showed short and long openings. After application of the reducing agent glutathione to the cytoplasmic side of excised inside-out patches, inactivation of Na+ currents in both motor and sensory fibres proceeded with a single, fast exponential time constant and late currents appeared with short openings only. These data indicate that the axonal metabolism may contribute to the different inactivation kinetics of Na+ currents in motor and sensory nerve fibres.

Exposure of nervous tissue to hypoxia results in interstitial acidification. There is evidence for concomitant decrease in extracellular pH to the increase in tissue lactate. In the present study, we used double-barrelled pH-sensitive microelectrodes to investigate the link between lactate transport and acid-base homeostasis in isolated rat spinal roots. Addition of different organic anions to the bathing solution at constant bath pH caused transient alkaline shifts in extracellular pH; withdrawal of these compounds resulted in transient acid shifts in extracellular pH. With high anion concentrations (30 mM), the largest changes in extracellular pH were observed with propionate >l-lactate ≈ pyruvate >62; 2-hydroxy-2-methylpropionate. Changes in extracellular pH induced by 10 mMl- andd-lactate were of similar size. Lactate transport inhibitors α-cyano-4-hydroxycinnamic acid and 4,4′-dibenzamidostilbene-2,2′-disulphonic acid significantly reducedl-lactate-induced extracellular pH shifts without affecting propionate-induced changes in extracellular pH. Hypoxia produced an extracellular acidification that was strongly reduced in the presence of α-cyano-4-hydroxycinnamic acid and 4,4′-dibenzamidostilbene-2,2′-disulphonic acid. In contrast, amiloride and 4,4′-di-isothiocyanostilbene-2,2′-disulphonate were without effect on hypoxia-induced acid shifts. The results indicate the presence of a lactate-proton co-transporter in rat peripheral nerves. This transport system and not Na+/H+ or C1−/HCO−3 exchange seems to be the dominant mechanism responsible for interstitial acidification during nerve hypoxia.

Isolated ventral and dorsal rat spinal roots incubated in normal (2.5 mM) or high glucose (25 mM) concentrations or in high concentrations of other hexoses were exposed transiently to hypoxia (30 min) in a solution of low buffering power. Compound nerve action potentials, extracellular direct current potentials, and interstitial pH were continuously recorded before, during, and after hypoxia. Ventral roots incubated in 25 mM D-glucose showed resistance to hypoxia. Dorsal roots, on the other hand, revealed electrophysiological damage by hyperglycemic hypoxia as indicated by a lack of posthypoxic recovery. In both types of spinal roots, interstitial acidification was most pronounced during hyperglycemic hypoxia. The changes in the sensitivity to hypoxia induced by high concentrations of D-glucose were imitated by high concentrations of D-mannose. In contrast, D-galactose, L-glucose, D-fructose, and L-fucose did not have such effects. Resistance to hypoxia, hypoxia-generated interstitial acidification, and hypoxia-induced electrophysiological damage were absent after pharmacological inhibition of nerve glycolysis with iodoacetate. These observations indicate 1) that enhanced anaerobic glycolysis produces resistance to hypoxia in hyperglycemic peripheral nerves and 2) that acidification may impair the function of peripheral axons when anaerobic glycolysis proceeds in a tissue with reduced buffering power.

Fri, 1 Jan 1993 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5943/1/5943.pdf Henriksen, Jens H.; Gerbes, Alexander L.; Bendtsen, Flemming ddc:610,

Previous studies suggested the expression of mRNA, coding for CNP, exclusively in the central nervous system. In the present study, using the polymerase chain reaction (PCR) technique instead of the less sensitive Northern blot hybridization, CNP-specific sequences have also been detected in rat atria and ventricles of the heart as well as in organs of the immune system (thymus, spleen and lymph nodes). Parallel PCR-assays documented ANP-mRNA in these tissues. To verify specificity of the PCR-products, Southern blots have been hybridized with a third internal oligonucleotide and amplification products have been sequenced. The relative level of CNP-mRNA in these tissues was estimated to be in the range of 1-9% of total brain CNP transcripts. The results suggest that the peptide may have a peripheral as well as a central site of action. In light of its pronounced effect on cell proliferation, particular interest should focus on a possible role of CNP in the immune system.

Fri, 1 Jan 1993 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5971/1/Gerbes_ALexander_5971.pdf Gerbes, Alexander L.; Bendtsen, Flemming; Henriksen, Jens H.

Torasemide is a new loop diuretic with a longer half-life and longer action than furosemide in healthy subjects. In order to evaluate the pharmacodynamic effects, single oral doses of furosemide (80 mg) and torasemide (20 mg), which were equipotent in healthy subjects, were given to 14 patients with cirrhosis and ascites. Before the study patients underwent an equilibration period of 4 days without diuretics. The drugs were alternated following a randomized double-blind cross-over design after a wash-out period of at least 2 days. Urine was collected at defined intervals for 24 h after drug administration and blood samples were taken before, 6 h and 24 h after medication. Torasemide induced greater cumulative 24 h diuresis (2863 ± 343 vs. 2111 ± 184 ml, p < 0.01) than furosemide. Torasemide did not differ from furosemide for cumulative 0–6 h sodium excretion (96 ± 17 vs. 92 ± 23 mmol sodium) but caused a more pronounced cumulative 6–24 h natriuresis (38 ± 11 vs. 17 ± 4 mmol, p < 0.05). Five patients exhibited a weak response to furosemide (0–36 mmol sodium/24 h, median 24 mmol; 690–1460 ml urinary volume/24 h, median 1325 ml). These patients showed significantly higher natriuresis and diuresis following torasemide (26–136 mmol sodium/24 h, median 78 mmol, p < 0.05; 1670–3610 ml urinary volume/24 h, median 2200 ml, p < 0.05). Twenty-four hours after administration of both drugs there were no significant changes in hemodynamic, renal or hormonal parameters. No adverse effects were noted with either treatment. These findings suggest that torasemide might be more advantageous than furosemide in the treatment of ascites due to cirrhosis.

Fri, 1 Jan 1993 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5984/1/Gerbes_ALexander_5984.pdf Nemer, M.; Gerbes, Alexander L.

Fri, 1 Jan 1993 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5985/1/Juengst_Dieter_5985.pdf Jüngst, Dieter; Wernze, Heinrich; Pilz, A.; Gerbes, Alexander L.

Inhaltsverzeichnis fehlt.

Fri, 1 Jan 1993 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5995/1/5995.pdf Gerbes, Alexander L.; Bendtsen, Flemming; Henriksen, Jens H. ddc:610, Medizin

Fri, 1 Jan 1993 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6065/1/6065.pdf Gerbes, Alexander L.; Paumgartner, Gustav ddc:610, Medizin

Sat, 5 Dec 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7630/1/Neupert_Walter_7630.pdf Horwich, Arthur; Hartl, Franz-Ulrich; Neupert, Walter; Martin, Jörg; Clark, Deborah J.; West, Ann H.

The import of proteins into mitochondria is an intricate process comprised of multiple steps. The first step involves the sorting of cytosolically synthesized precursor proteins to the mitochondrial surface. There precursor proteins are recognized by specific receptors which deliver them to the general import site present in the outer membrane. The second stage of import involves a series of complex intraorganelle sorting events which results in the delivery of the proteins to one of the four possible submitochondrial destinations, namely the outer and inner membranes, the matrix and intermembrane space. Here in this review, we discuss the current knowledge on these intramitochondrial sorting events. We especially focus on targetting of proteins to the intermembrane space. Sorting to the intermembrane space represents a particularly interesting situation, as at least three separate targetting pathways to this subcompartment are known to exist.

The phenotype of a fully differentiated, mature Schwann cell is appar-ently largely determined by Schwann cell-axon interactions. In vitro, elevation of intra-cellular cAMP levels in Schwann cells induces a phenotype which resembles that of a mature, i.e., axon-related, Schwann cell. Therefore, an important role for cAMP as a second messenger of axon-Schwann cell interactions in vivo is assumed. However, the effects of cAMP on Schwann cells are not restricted to induction of features of a mature phenotype. For example, elevation of intracellular cAMP levels results of also in a markedly increased synthesis of nerve growth factor (NGF) mRNA, which is barely detectable in intact sciatic nerves of adult animals. Furthermore, since cAMP induces myelin gene expression in cultured Schwann cells, additional regulatory mechanisms have to be postulated for the induction and maintenance of a mature non-myelinating Schwann cell phenotype. Here we show that a soluble protein growth factor can partially induce a non-myelinating mature Schwann cell phenotype in vitro. Treatment with transforming growth factor 1 (TGF-1) results in a marked and rapid downregulation of the low affinty NGF receptor (NGFR) on cultured Schwann cells without induction of PO gene expression. In contrast, in agreement with previous studies, an increase in PO mRNA levels and a reduction in NGFR mRNA after cAMP elevation is much slower when compared with the effect of TGF-1, suggesting the involvement of different intracellular mechanisms. Consistent with this hypothesis, we did not observe an induction of mRNA coding for TGR- isoforms after cAMP elevation in cultured Schwann cells which constitutively synthesize TGF-1 mRNA

Mitochondrial protein import involves the recognition of preproteins by receptors and their subsequent translocation across the outer membrane. In Neurospora crassa, the two import receptors, MOM19 and MOM72, were found in a complex with the general insertion protein, GIP (formed by MOM7, MOM8, MOM30 and MOM38) and MOM22. We isolated a complex out of S. cerevisiae mitochondria consisting of MOM38/ISP42, the receptor MOM72, and five new yeast proteins, the putative equivalents of N. crassa MOM7, MOM8, MOM19, MOM22 and MOM30. A receptor complex isolated out of yeast cells transformed with N. crassa MOM19 contained the N. crassa master receptor in addition to the yeast proteins. This demonstrates that the yeast complex is functional, and provides strong evidence that we also have identified the yeast MOM19.

Mon, 1 Jun 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7625/1/Neupert_Walter_7625.pdf Neupert, Walter; Martin, Jörg; Hartl, Franz-Ulrich ddc:610, Medizin

Subchondral bone density (by means of CT osteoabsorptiometry), and cartilage thickness (directly measured on photocopies of frozen sections), were examined in 30 human patellae, with an age range from 47 to 90 y. A surface demonstration of the distribution was prepared, and representative pictures produced by summation with a computer. Subchondral bone-density maxima were found in the proximal part of the lateral facet, and the density pattern interpreted as the expression of the long-term distribution of stress in the joint. It is reasonable to assume that cartilage thickness, of which two-thirds of the maximum values occupy a lateral position, is also dependent on the local stress. The 2 distributions show correlation coefficients > 0.5 in approximately a third of the cases we examined. Displacement of the higher values of cartilage thickness relative to the subchondral density maxima is attributed to incongruence in the medial part of the joint.

The plasmid-encoded outer membrane protein YadA of enteropathogenic yersiniae is associated with pathogenicity. Recently, collagen binding of YadA-positive yersiniae was reported without detailed characterization (L. Emödy, J. Heesemann, H. Wolf-Watz, M. Skurnik, G. Kapperud, P. O'Toole, and T. Wadström, J. Bacteriol. 171:6674-6679, 1989). To elucidate the nature of collagen binding to YadA, we used a recombinant Yersinia strain expressing the cloned YadA gene. Direct binding of YadA-positive yersiniae to collagens was demonstrated in affinity blot experiments on nitrocellulose filters. A spectrum of collagen types in a wide concentration range were tested for their ability to block binding of 125I-labeled collagen type II to YadA-positive yersiniae. The results indicate a specific binding site(s) for YadA in collagen types I, II, III, IV, V, and XI. In contrast, collagen type VI did not bind to YadA. To characterize the binding site(s) more precisely, isolated collagen chains and cyanogen bromide fragments were investigated. These studies revealed that binding of YadA to collagen type I is confined to the alpha 1(I) chain, whereas the binding site within collagen type XI is localized in the alpha 3(XI) chain. alpha 2(I), alpha 1(XI), and alpha 2(XI) did not bind to YadA. Most interestingly, in the alpha 1(II) chain the specific binding site for YadA resides in the cyanogen bromide fragment CB10. The latter might indicate a binding site that does not depend on conformation. Based on these findings, further fragmentation and the synthesis of peptides may allow definition of the peptide sequence(s) relevant for YadA binding.

During storage of CPD-A1 preserved whole blood factors of the complement cascade become activated, as evidenced by a rapid increase in the concentrations of C3a-desArg and C4a-desArg. After 10 to 14 days of whole blood storage, the elevations of C3a and C4a levels were highly significant. This increase was paralleled by an increase in the concentration of the lysosomal proteinase elastase from polymorphonuclear (PMN) granulocytes. By contrast, the concentration of the C3 activator complex C4b2b remained unchanged even after 3 weeks of storage. The supplementation of the anticoagulant CPD-A1 with the polyvalent-proteinase-inhibitor aprotinin and the specific elastase- inhibitor eglin C failed to inhibit complement activation, whereas leukocyte depletion could partially abolish the increase of the concentration of C4a, but had no effect on C3a concentrations. These observations support the notion that cleavage of C4 during storage of whole blood is partially leukocyte dependent, whereas the activation of C3 is possibly caused by the activation of the alternate pathway of the complement system by contact of plasma with plastic surfaces.

The relation of (multiple) organ failure (OF) to the release of inflammatory mediators and the incidence of infection and sepsis was studied prospectively in 100 patients with multiple trauma (injury severity score=37). Sixteen patients died of OF, 47 patients survived OF, and 37 patients had no OF. Fifteen (24%) of the patients with OF showed no signs of infection. In patients with early onset of OF (n=45), infection followed with a lag of 2 or more days. In 16 (44%) of these patients, infection led to a deterioration in organ function. With late onset of OF (n=18), infection preceded OF in nine patients. Polymorphonuclear leukocyte—elastase, neopterin, C-reactive protein, lactate, antithrombin III, and phospholipase A discriminated significantly among the three outcome groups. Of all factors, only polymorphonuclear leukocyte—elastase showed a difference between patients with and without infection or sepsis, respectively. These data indicate that infection might not play a crucial role in the pathogenesis of posttraumatic OF in a substantial portion of patients with trauma. Early OF, especially, seems to be mainly influenced by the direct sequelae of tissue damage and shock (eg, the release of inflammatory mediators). Since infection and sepsis did not lead to an augmented release of mediators in patients with trauma, the role of both entities remains unclear.

Cytochrome b2 reaches the intermembrane space of mitochondria by transport into the matrix followed by export across the inner membrane. While in the matrix, the protein interacts with hsp60, which arrests its folding prior to export. The bacterial-type export sequence in pre-cytochrome b2 functions by inhibiting the ATP-dependent release of the protein from hsp60. Release for export apparently requires, in addition to ATP, the interaction of the signal sequence with a component of the export machinery in the inner membrane. Export can occur before import is complete provided that a critical length of the polypeptide chain has been translocated into the matrix. Thus, hsp60 combines two activities: catalysis of folding of proteins destined for the matrix, and maintaining proteins in an unfolded state to facilitate their channeling between the machineries for import and export across the inner membrane. Antifolding signals such as the hydrophobic export sequence in cytochrome b2 may act as switches between these two activities.

Fri, 20 Mar 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7628/1/Neupert_Walter_7628.pdf Neupert, Walter; Klei, Ida J. van der; Rassow, Joachim; Pfanner, Nikolaus ddc:610, Medizin

Cytochrome c heme lyase (CCHL) catalyses the covalent attachment of the heme group to apocytochrome c during its import into mitochondria. The enzyme is membrane-associated and is located within the intermembrane space. The precursor of CCHL synthesized in vitro was efficiently translocated into isolated mitochondria from Neurospora crassa. The imported CCHL, like the native protein, was correctly localized to the intermembrane space, where it was membrane-bound. As with the majority of mitochondrial precursor proteins, CCHL uses the MOM19-GIP receptor complex in the outer membrane for import. In contrast to proteins taking the general import route, CCHL was imported independently of both ATP-hydrolysis and an electrochemical potential as external energy sources. CCHL which lacks a cleavable signal sequence apparently does not traverse the inner membrane to reach the intermembrane space; rather, it translocates through the outer membrane only. Thus, CCHL represents an example of a novel, 'non-conservative' import pathway into the intermembrane space, thereby also showing that the import apparatus in the outer membrane acts separately from the import machinery in the inner membrane.

Mitochondria contain a complex machinery for the import of nuclear-encoded proteins. Receptor proteins exposed on the outer membrane surface are required for the specific binding of precursor proteins to mitochondria, either by binding of cytosolic signal recognition factors or by direct recognition of the precursor polypeptides. Subsequently, the precursors are inserted into the outer membrane at the general insertion site GIP (general insertion protein. Here we report the analysis of receptors and GIP by crosslinking of translocation intermediates and by coimmunoprecipitation. Surface-accumulated precursors were cross-linked to the receptors MOM19 and MOM72, suggesting a direct interaction of preproteins with surface receptors. We identified three novel mitochondrial outer membrane proteins, MOM7, MOMS, and MOM30 that, together with the previously identified MOM38, seem to form the GIP site and are present in the mitochondrial receptor complex.

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9804/2/9804.pdf Fink, Edwin; Hoffmann, H.; Spannagl, Michael; Siebeck, Matthias ddc:610, Medizin

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9810/1/9810.pdf Fritz, Hans; Bernett, P.; Jochum, Marianne; Riel, K. A. ddc:610

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9815/1/9815.pdf Tanagho, Emil A.; Lue, Tom F.; Stief, Christian Georg; Bénard, Francois; Aboseif, Sherif R.; Takahashi, Y.

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9816/1/9816.pdf Jonas, Udo; Stief, Christian Georg; Allhoff, E. P.; Seabert, J.; Djamilian, Mohamad H.; Krah, H. ddc:610, Medizin

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9833/1/9833.pdf Schweiberer, Leonhard; Fritz, Hans; Schramm, Wolfgang; Spannagl, Michael; Machleidt, W.; Duswald, Karl-Heimo; Jochum, Marianne; Waydhas, Christian; Nast-Kolb, Dieter

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9834/1/9834.pdf Fritz, Hans; Redl, H.; Waydhas, Christian; Machleidt, W.; Joka, T.; Inthorn, D.; Jochum, Marianne

The effect of different aprotinin applications on hemmtatic changes and blood product requirements in orthotopic liver transplantation was investigated in a prospective, open, and randomized study. From November 1989 to June 1990, 13 patients received aprotinin as a bolus of 0.5 Mill, kallikrein inac-tivator units (KIU) on three occasions in the course of an OLT, whereas 10 other patients were treated with continuous aprotinin infusion of 0.1-0.4 Mill. KIU/hr. Before and after reperfusion of the graft liver, signs of hyperfibrinolysis, measured by thrombelastography, were significantly lower in the infusion group. Tissue-type plasminogen activator (t-PA) activity increased during the anhepatic phase but to a significantly lesser extent in the infusion group. Blood product requirements during OLT were tendentiously higher in the bolus group but not significantly so. However, the use of packed red blood cells was significantly lower in the postoperative period, whereas there was no significant difference in fresh frozen plasma requirements between the two groups. All 23 patients have survived, and only one woman of each group required retransplantation due to severe host-versus-graft reactions. Furthermore, we investigated the perfusate of the graft liver in both groups and detected signs of a decreased t-PA release in the infusion group. Our results demonstrate an advantage of aprotinin given as continuous infusion over bolus application in OLT.

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9837/1/9837.pdf Schmitt, M.; Jänicke, F.; Jochum, Marianne; Assfalg-Machleidt, I.; Machleidt, W. ddc:610,

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9842/1/9842.pdf Fritz, Hans; Jochum, Marianne; Riel, K. A.

Kein Inhaltsverzeichnis.

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9961/1/9961.pdf Schuh, Angela ddc:610, Medizin

Die Gesundheitsvorsorge zielt sowohl in die somatisch-körperliche als auch in die seelisch-psychische Richtung ab. Bezüglich der somatischen Ebene stehen Rhythmisierung, Schlafförderung, körperliche Aktivität und Entspannung sowie der Begriff der Abhärtung im Vordergrund der Ordnungstherapie. Die Gesundheitserziehung und -bildung am Kurort und durch den Hausarzt rundet die Gesundheitsvorsorge ab.

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/9963/1/9963.pdf Schuh, Angela ddc:610, Medizin

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/10116/1/10116.pdf Senn, E.; Schuh, Angela ddc:610, Medizin

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/10293/1/10293.pdf Schönharting, M.; Dammer, U.; Arbogast, Helmut; Stiegler, H.; Strohmenger, R.; Meier-Ewert, H.; Dendorfer, A.; Nees, S.

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/10294/1/10294.pdf Schildberg, F. W.; Heim, G.; Arbogast, Helmut; Hiller, E.; Stiegler, H.

We report a 25-year-old woman presenting with sarcoidosis and bilateral sacroiliitis. Her sarcoidosis related symptoms (malaise, cough and dyspnoea) improved dramatically under treatment with steroids but severe back pain persisted. Only seven similar cases have been described over the last 40 years and the question of a possible association between the two diseases has been raised. However, prevalence data from the literature and the apparent lack of genetic links are better arguments for coincidence than for association.

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/10322/1/10322.pdf Jonas, Udo; Stief, Christian Georg; Thon, Walter F.; Djamilian, Mohamad H.; Freymann, H. ddc:610

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/10325/1/10325.pdf Jonas, Udo; Stief, Christian Georg; Thon, Walter F.; Djamilian, Mohamad H.; Truß, M. ddc:610, Medizin

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/10336/1/10336.pdf Jonas, Udo; Stief, Christian Georg; Thon, Walter F. ddc:610, Medizin 0

Wed, 1 Jan 1992 12:00:00 +0100 https://epub.ub.uni-muenchen.de/10338/1/10338.pdf Merckx, L.; Stief, Christian Georg

Loperamide, an opiate agonist of high specificity for p-receptors, was recently reported to suppress ACTH and cortisol levels in normal subjects, but not in patients with proven ACTH-dependent Cushing’s disease. However, there is little information on the site of action of loperamide in the hypothalamo-pituitary-adrenal axis of man. We investigated the effect of loperamide on pituitary hormone secretion in uiuo and in vitro. In seven normal subjects, basal ACTH plasma levels were significantly suppressed 3 h after loperamide administration (16 mg, orally) from 5 + 1 to 2 f 0 pmol/L (P < 0.0001). After the combined pituitary stimulation test (100 pg human CRH, 100 rg GnRH, 100 pg GH-releasing hormone, and 200 pg TRH), the ACTH peak (maximum increase at 30 min) was significantly blunted by loperamide from 9 + 1 to 4 of: 1 pmol/L (P < 0.001) and the area under the curve of ACTH from O-120 min was reduced from 35 + 5 to 23 + 4 pmol/L.2 h (P < 0.05). In the insulin-hypoglycemia test (0.15 IU/kg BW), neither the ACTH peak nor the area under the curve of ACTH was affected by loperamide. In six patients with Cushing’s disease and one patient with secondary adrenal insufficency due to hypothalamic failure, neither basal ACTH and cortisol levels nor CRH-stimulated levels were influenced by loperamide. In four cultured human corticotropic adenomas, loperamide was not able to reduce basal and CRH-induced ACTH secretion. In summary, loperamide is able to reduce basal and CRHinduced ACTH and cortisol levels in normal subjects, but not in patients with Cushing’s disease or secondary adrenal failure of hypothalamic origin. Loperamide has no significant effect on insulin-hypoglycemia- induced ACTH and cortisol levels and, therefore, no effect on stress-induced elevation of cortisol levels. Loperamide might act at a suprapituitary site in man in viuo, but, nevertheless, a pituitary site cannot be excluded.

Using gene-specific synthetic oligonucleotides the expression and regulation of kallikrein-like genes in the human prostatic cancer cell line LNCaP were studied. Prostate-specific antigen (PSA) and human glandular kallikrein (hGK-1) together constitute a subfamily of serine proteases exclusively produced in the human prostate. RNA analysis revealed that both genes are expressed in LNCaP cells with PSA basal levels being 2-fold higher than hGK-1 levels. Both mRNAs are induced over a period of 24 h in the presence of 3.3 nM of the synthetic androgen mibolerone. Stimulation of PSA RNA is about 5- fold,whereas hGK-1 stimulation is less pronounced. Nuclear run-on analysis revealed that androgen induction of kallikrein-like genes in LNCaP cells is a rapid event (c3 h) occurring at the level of transcription initiation. Treatment of cells with cycloheximide demonstrates that, while PSA/hGK-1 basal transcription strictly depends on continuous protein synthesis, transcriptional induction by androgen does not. This suggests the direct involvement of the androgen receptor in the induction process independent of additional labile protein factors necessary for kallikrein basal transcription. A binding motif is present in the PSA and hGK-1 promoters, closely resembling the consensus sequence for steroidresponsive elements. The androgen antagonist cyproterone acetate was also able to stimulate transcription of kallikrein-like genes in LNCaP cells. In contrast, androgen-dependent transcriptional suppression of the protooncogene c-myc was strongly counteracted by cyproterone acetate. Thus, antiandrogens act differentially on androgen-regulated prostate-specific (PSA, hGK-1) and growthrelated (c-myc) gene expression in LNCaP cells.

Cultured cardiac myocytes from adult Sprague-Dawley rats express both insulin-like growth factor-I (IGF-I) receptors and insulin-like growth factor-II/mannose 6-phosphate (IGF-II/Man6P) receptors and respond to IGF-I with a dose-dependent accumulation of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and inositol 1,4-bisphosphate [Ins(1,4)P2]. Specific binding of [125I]IGF-I to isolated membranes from cultured cardiac myocytes amounted to 1-1.2%. Binding of [125I]IGF-I was inhibited by unlabeled IGF-I at nanomolar concentrations and insulin at much higher concentrations. These data suggest that IGF-I binds to its own receptor on rat cardiac myocytes. Competitive binding studies using isolated membranes from cardiac myocytes and [125I]IGF-II showed 2-4% specific binding. Binding of [125I]IGF-II was inhibited by IGF-II and much less potently by IGF-I and insulin. Immunoglobulin G (IgG) 3637 (an IgG directed against the IGF-II/Man6P receptor) partially inhibited binding of [125I]IGF-II whereas nonimmune IgG did not. Affinity cross-linking studies with [125I]IGF-II and cardiac myocyte membranes and subsequent analysis of the ligand-receptor complex using SDS-PAGE and autoradiography showed a radiolabeled band of approximately 250 kilodalton (kDa). The formation of the [125I]IGF-II-receptor complex was inhibited by incubation with IGF-II and IgG 3637 but not by insulin or nonimmune IgG. Western blotting of protein extracts from cultured cardiac myocytes was performed using IgG 3637 and an immunoperoxidase technique for the visualization of the IGF-II/Man6P receptor protein. A specific band at 220 kDa under nonreducing conditions was detected on the blots, providing further evidence for the expression of the IGF-II/Man6P receptor by cardiac myocytes. The effect of IGFs on the accumulation of inositol phosphates was measured by HPLC analysis of perchloric acid extracts from myo-[3H]inositol-labeled cultured cardiac myocytes. IGF-I (50 ng/ml) stimulated the accumulation both of Ins(1,4,5)P3 and Ins(1,4)P2 after 30 sec by 43% and 63%. IGF-II (up to 500 ng/ml) had no significant effect on inositol phosphate accumulation under the same conditions. However, in the presence of millimolar concentrations of Man6P, IGF-II (500 ng/ml) also increased Ins(1,4,5)P3 accumulation by 59%. We conclude that cardiac myocytes from adult rats express IGF receptors and respond to IGFs with the accumulation of Ins(1,4,5)P3 and Ins(1,4)P2. This effect seems to be mediated by an IGF-I receptor-specific pathway.

Wed, 1 Jan 1992 12:00:00 +0100 https://www.thieme-connect.de/ejournals/abstract/10.1055/s-2006-936401 https://epub.ub.uni-muenchen.de/17947/1/Schmidt_17947.pdf Schmidt, Josef M.

Wed, 1 Jan 1992 12:00:00 +0100 https://www.thieme-connect.de/ejournals/abstract/10.1055/s-2006-936409 https://epub.ub.uni-muenchen.de/17948/1/Schmidt_17948.pdf Schmidt, Josef M.