Biomedical subjects

Matthias Löhn

Publications and source records attributed to Matthias Löhn.

9 recordsLinked to original sources

PubMed2005

Biochemistry and pharmacology of novel anthranilic acid derivatives activating heme-oxidized soluble guanylyl cyclase.

The heme-enzyme soluble guanylyl cyclase (sGC) is an ubiquitous NO receptor, which mediates NO downstream signaling by the generation of cGMP. We studied the mechanism of action of the anthranilic acid derivatives 5-chloro-2-(5-chloro-thiophene-2-sulfonylamino-N-(4-(morpholine-4-sulfonyl)-phenyl)-benzamide sodium salt (HMR1766) (proposed international nonproprietary name, ataciguat sodium) and 2-(4-chloro-phenylsulfonylamino)-4,5-dimethoxy-N-(4-(thiomorpholine-4-sulfonyl)-phenyl)-benzamide (S3448) as a new class of sGC agonists. Both compounds activated different sGC preparations (purified from bovine lung, or crude from human corpus cavernosum) in a concentration-dependent and quickly reversible fashion (EC50 = 0.5-10 microM), with mixed-type activation kinetics. Activation of sGC by these compounds was additive to activation by NO donors, but instead of being inhibited, it was potentiated by the heme-iron oxidants 1H-[1,2,4]-oxdiazolo[3,4-a]quinoxalin-1-one (ODQ) and 4H-8-bromo-1,2,4-oxadiazolo(3,4-d) benz(b)(1,4)oxazin-1-one (NS2028), suggesting that the new compounds target the ferric heme sGC isoform. Protoporphyrin IX acted as a competitive activator, and zinc-protoporphyrin IX inhibited activation of heme-oxidized sGC by HMR1766 and S3448, whereas heme depletion of sGC by Tween 20 treatment reduced activation. Both compounds increased cGMP levels in cultured rat aortic smooth muscle cells; induced vasorelaxation of isolated endothelium-denuded rat aorta, porcine coronary arteries, and human corpus cavernosum (EC50 1 to 10 microM); and elicited phosphorylation of the cGMP kinase substrate vasodilator-stimulated phosphoprotein at Ser239. HMR1766 intravenous bolus injection decreased arterial blood pressure in anesthetized pigs. All of these pharmacological responses to the new compounds were enhanced by ODQ and NS2028. Our findings suggest that HMR1766 and S3448 preferentially activate the NO-insensitive heme-oxidized form of sGC, which exists to a variable extent in vascular tissues, and is a pharmacological target for these new vasodilator drugs.

Animals↗

PubMed2005

Long term Rho-kinase inhibition ameliorates endothelial dysfunction in LDL-Receptor deficient mice.

Chronic inhibition of Rho-kinase has been recently implicated in retardation of atherogenesis induced by high-fat diet in low-density lipoprotein receptor deficient (LDLR-/-) mice. However, it remains to be examined whether long-term Rho-kinase inhibition will reduce vascular dysfunction in this model. LDLR-/- mice on a high-fat diet were treated either with saline (LDLR-/-) or with the Rho-kinase inhibitor Fasudil (HA1077, 5-Isoquinolinesulfonyl homopiperazine, 100 mg/kg/day by gavage, LDLR-/- +Fasudil) for 10 weeks. Fasudil-treatment normalized endothelial function (measured by means of endothelium-dependent vasorelaxation) in LDLR-/- +Fasudil, to the level of controls (C57BL/6J). No tolerance toward Rho-kinase inhibition has been detected in Fasudil-treated animals. We conclude that long-term Rho-kinase inhibition normalizes endothelial function without development of tolerance.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed2005

Down-regulation of calpain 9 is linked to hypertensive heart and kidney disease.

Calpains are a family of 14 intracellular calcium-dependent proteases, which have been implicated in cardiovascular diseases. We aimed to analyze specifically the expressional regulation of the different calpain isoforms in hypertensive target organ damage. Using real-time PCR, we found calpain 6 and 9 down-regulated by more than 50% and the endogenous calpain inhibitor calpastatin up-regulated by 225%, respectively, in the hearts of Dahl salt-sensitive rats on a high salt (4% NaCl) compared to normal salt diet. On the protein level, calpain 9 but not calpastatin was regulated in the hypertensive target organs heart and kidney. Moreover, the myocardial expression of calpain 9 protein was inversely linked to left ventricular mass (r= -0.93, p<0.01), and renal expression of calpain 9 protein correlated inversely with albuminuria (r= -0.82, p<0.05). In the aorta, there was no regulation of calpain 9 on the protein level. We conclude that differential regulation of calpain 9 may play a role in hypertensive target organ damage.

Animals↗

PubMed2004

Inhibition of Rho-kinase stimulates nitric oxide-independent vasorelaxation.

Vasoconstrictor factors, like urotensin, angiotensin and catecholamines, activate Rho-dependent serine-threonine kinase (Rho-kinase) and inhibition of this pathway represents a novel therapy for cardiovascular diseases with hypertensive syndrome. The disbalance of relaxing endothelial nitric oxide (NO)-producing and vasoconstrictive pathways can be especially important in diseases where hypertension is accompanied by endothelial dysfunction that compromises NO generation. However, a recent study reported that the efficacy of the Rho-kinase inhibitor (R)-(+)-trans-N-(4-Pyridyl)-4-(1-aminoethyl)cyclohexanecarboxamide (Y27632) is dramatically attenuated upon removal of endothelium or inhibition of endothelial NO synthase (eNOS). This raises the question whether Rho-kinase inhibition could be an effective treatment in case of hypertension associated with endothelial dysfunction. The purpose of the present study was to determine whether the vasorelaxing effect of Rho-kinase inhibition is mediated through eNOS-dependent mechanisms. We show here that in the models of genetically reduced endothelial NO production (eNOS-/- mice and spontaneous hypertensive rats (SHR)) or in models of pharmacologically reduced endogenous NO production (N(omega)-nitro-L-arginine methyl ester (LNAME) treatment), Rho-kinase inhibition induced a strong vasodilation and reduction of blood pressure indicating independence of Rho-kinase pathway from eNOS. An additional important finding of our study is that Rho-kinase inhibitors induce a strong vasorelaxation and blood pressure reduction upon intravenous injection not only in hypertensive but in normotensive animals, as well. Inhibition of Rho-kinase represents a promising possibility to treat hypertension that is accompanied by endothelial dysfunction.

Animals↗

PubMed2004

Chronic vasopeptidase inhibition normalizes diabetic endothelial dysfunction.

Type 2 diabetes mellitus is a major cause of vascular morbidity but animal models for this disease have not been adequately characterized. We demonstrate that endothelial dysfunction is present in the Zucker diabetic fatty (ZDF) rat. Vasopeptidase inhibition with AVE7688 (7-[[(2S)-2-(acetylthio)-1-oxo-3-methylpropyl]amino]-1,2,3,4,6,7,8,12b-octahydro-6-oxo-(4S,7S,12bR)-pyrido[2,1-a][2]benzazepine-4-carboxic acid), 45 mg/kg/day in chow for 6 weeks, normalized acetylcholine mediated relaxation of mesenteric artery rings. Thus, chronic vasopeptidase inhibition may prevent vascular complications related to type 2 diabetes mellitus.

Animals↗

PubMed2002

Cell cycle retardation in monocytoid cells induced by aminopeptidase N (CD13).

Aminopeptidase N (APN, CD13) is highly expressed on human monocytes. Activation of leucocyte subpopulations, such as T-cells, can induce CD13 expression. However, little is known about the physiological role of CD13 expression on human leucocytes. It was suggested that CD13, similar to other peptidases, could be involved in control of cell growth. The hypothesis that CD13 influences proliferation of monocytoid cells by retarding the velocity of the cell cycle was tested. It was shown that CD13 expression was modulated within the cell cycle. Cells entering the S-phase of cell cycle decreased their CD13 surface expression. Occupation of the active center of CD13 with artificial ligands such as monoclonal antibodies (mab) or the low molecular weight inhibitor actinonin induced a prolongation of cell cycle and decreased the rate of cell growth. Additionally, after ligation of CD13 by mab the complex of CD13 and monoclonal antibody was actively internalized into the cell. We suggest that CD13 could have important functions for the proliferation of human monocytoid cells. Here we show for the first time that occupation of the CD13 active center by antibodies or inhibitors influences the cell cycle and thereby diminishes cell growth rate. Occupation of the active center by antibodies or inhibitors might prevent cleavage or binding and internalization of still unknown natural substrate(s) and could evoke a deceleration of the cell cycle and reduced cell growth rate.

Antibodies, Monoclonal↗

PubMed2002

Periadventitial fat releases a vascular relaxing factor.

Virtually all blood vessels are surrounded by adventitial fat. Adipocytes produce a host of vasoactive substances that may influence vascular contraction. We tested whether or not perivascular adipose tissue modulates contraction of aortic ring preparations. We studied aortic rings surrounded by periadventitial adipose tissue from adult Sprague-Dawley rats. At a maximum concentration of 300 nM angiotensin II, 6.5 microM serotonin, and 5 microM phenylephrine, the contractile response of intact rings was 95%, 80%, and 30% lower than that of vessels without periadventitial fat. The anticontractile effect of periadventitial fat was reduced by inhibition of ATP-dependent K+ channels with glibenclamide (3 microM) and by the tyrosine kinase inhibitor genistein (10 microM). Blocking NOS, cyclo-oxygenase, cytochrome P450, or adenosine receptors did not restore the vascular response in intact vessels. The anticontractile effect of perivascular fat was present in Zucker fa/fa rats, suggesting that leptin receptors were not responsible. Transferring the bath solution from intact vessels, isolated periadventitial tissue, and cultured rat adipocytes to precontracted vessels lacking periadventitial fat resulted in a rapid relaxation. We suggest that perivascular adventitial adipose tissue releases a transferable adventitium-derived relaxing factor that acts by tyrosine kinase-dependent activation of K+ channels in vascular smooth muscle cells.

Adipocytes↗

PubMed2002

Cilnidipine is a novel slow-acting blocker of vascular L-type calcium channels that does not target protein kinase C.

Cilnidipine is a novel dihydropyridine (DHP) antagonist. However, its pharmacological effects on vascular DHP-sensitive L-type channels and protein kinase C (PKC)-mediated arterial contraction is incompletely understood. To address this issue, we studied the effects of cilnidipine on multi-subunit, C-class L-type Ca2+ channels in rat aortic A7r5 cells, as well as on Ca2+ channel (L-type) alpha1C-b and (T-type) alpha1G subunits in the Xenopus oocyte expression system. Cilnidipine dose- and time-dependently inhibited Ba2+ currents in A7r5 cells, with half-maximal inhibitions (IC50) at 10 nmol/l after 10 min. Unlike classical pharmacological Ca2+ channel blockers, cilnidipine's block of Ca2+ currents did not reach steady-state levels within 10 min, indicating steady-state half-maximal inhibition of native, multi-subunit L-type channels at < 10 nmol/l. In contrast, smooth muscle alpha1Cb currents were blocked by cilnidipine at much higher doses (steady-state IC50, 20 micromol/l) whereas alpha1G currents were not inhibited by cilnidipine (30 micromol/l). Cilnidipine dose-dependently inhibited depolarization- and Ca2+-induced contractions of rat aortic rings, with an IC50 of 10 nmol/l at 10 min. However, the onset of the effects was very slow, with approximately 71% inhibition by 3 nmol/l cilnidipine after 90 min exposure to cilnidipine. In contrast, cilnidipine did not inhibit phorbol 12-myristate-13-acetate (100 nmol/l)-mediated contractions. We conclude that cilnidipine represents an extremely slow-acting DHP that targets multi-subunit L-type channels, but not PKC in arterial smooth muscle. Because cilnidipine is less potent in cells expressing the pore-forming alpha1C-b subunit, the data further suggest that this unique slow-acting mechanism of cilnidipine is mediated by a complex interaction of cilnidipine with alpha1C-b and accessory channel subunits.

Animals↗

PubMed2002

Regulation of arterial tone by smooth muscle myosin type II.

The initiation of contractile force in arterial smooth muscle (SM) is believed to be regulated by the intracellular Ca2+ concentration and SM myosin type II phosphorylation. We tested the hypothesis that SM myosin type II operates as a molecular motor protein in electromechanical, but not in protein kinase C (PKC)-induced, contraction of small resistance-sized cerebral arteries. We utilized a SM type II myosin heavy chain (MHC) knockout mouse model and measured arterial wall Ca2+ concentration ([Ca2+](i)) and the diameter of pressurized cerebral arteries (30-100 microm) by means of digital fluorescence video imaging. Intravasal pressure elevation caused a graded [Ca2+](i) increase and constricted cerebral arteries of neonatal wild-type mice by 20-30%. In contrast, intravasal pressure elevation caused a graded increase of [Ca2+](i) without constriction in (-/-) MHC-deficient arteries. KCl (60 mM) induced a further [Ca2+](i) increase but failed to induce vasoconstriction of (-/-) MHC-deficient cerebral arteries. Activation of PKC by phorbol ester (phorbol 12-myristate 13-acetate, 100 nM) induced a strong, sustained constriction of (-/-) MHC-deficient cerebral arteries without changing [Ca2+](i). These results demonstrate a major role for SM type II myosin in the development of myogenic tone and Ca2+ -dependent constriction of resistance-sized cerebral arteries. In contrast, the sustained contractile response did not depend on myosin and intracellular Ca2+ but instead depended on PKC. We suggest that SM myosin type II operates as a molecular motor protein in the development of myogenic tone but not in pharmacomechanical coupling by PKC in cerebral arteries. Thus PKC-dependent phosphorylation of cytoskeletal proteins may be responsible for sustained contraction in vascular SM.

Animals↗