Effects of long-term intravenous administration of adrenomedullin (AM) plus hANP therapy in acute decompensated heart failure: a pilot study.

BACKGROUND: It was reported previously that 30 min administration of adrenomedullin (AM) improves hemodynamics in chronic stable heart failure patients. The present study was designed to examine whether long-term AM + human atrial natriuretic peptide (hANP) administration can be used as a therapeutic drug in patients with acute decompensated heart failure (ADHF) in clinical setting. METHODS AND RESULTS: Seven acute heart failure patients (74 +/- 5 years) with dyspnea and pulmonary congestion were studied. AM (0.02 microg x kg(-1) x min(-1)) + hANP (0.05 microg x kg(-1) x min(-1)) was infused for 12 h and then hANP (0.05 microg x kg(-1) x min(-1)) was infused for 12 h. Hemodynamic, renal, hormonal and oxidative stress responses were evaluated. AM + hANP significantly reduced mean arterial pressure, pulmonary arterial pressure and systemic and pulmonary vascular resistance without changing heart rate, and increased cardiac output for most time-points compared with those at baseline. In addition, AM + hANP reduced aldosterone, brain natriuretic peptide and free-radical metabolites compared with those at baseline (all P<0.05). AM + hANP increased urine volume and U(Na)V compared with baseline data. CONCLUSIONS: In this small, pilot trial, AM + hANP therapy had beneficial hemodynamic and hormonal effects in ADHF. Intravenous infusion of AM with hANP could be used as a therapeutic drug in ADHF. These data are preliminary and require confirmation in a larger clinical study.

Stimulatory and Inhibitory regulation of lipolysis by the NPR-A/cGMP/PKG and NPR-C/G(i) pathways in rat cultured adipocytes.

OBJECTIVE: Recent studies have suggested the abundant expression of natriuretic peptide receptor in adipose tissue. This study was designed to investigate the levels of natriuretic receptor-A (NPR-A) and NPR-C gene expression during the process of preadipocyte differentiation and its role in adipogenesis and lipid metabolism. METHODS: We measured mRNA levels of NPR-A and NPR-C during the process of rat preadipocyte differentiation in vitro. We also measured the effects of ANP and C-ANP, a ligand for NPR-C, on preadipocyte differentiation. In addition, we assessed the effects of ANP and C-ANP on lipolysis and the cellular mechanism. RESULTS: The mRNA levels of NPR-A and NPR-C on day 3, 6, 10 are (-26%, +226%), (+6%, +568%), and (+207%, +3232%) respectively as compared with day 1. ANP (10(-)(7) M) and 8-bromo-cGMP (10(-)(4) M) significantly increased Oil Red positive area and cell number of matured-adipocytes. ANP and 8-bromo-cGMP also increased the mRNA levels of adipocyte-related genes such as PPARgamma, leptin, and adiponectin on day 3, whereas C-ANP did not change these parameters. ANP (10(-)(9)-10(-)(6) M) increased intracellular cGMP levels and promoted lipolysis in adipocytes and the effects were abolished by HS-142-1, and KT5823. Conversely C-ANP (10(-)(6) M) decreased intracellular cAMP levels and lipolysis and its effect was inhibited by PTX. CONCLUSION: Results suggest that ANP may promote adipocyte differentiation and lipolysis via the NPR-A/cGMP/PKG pathway. Direct action of ANP via NPR-C in adipogenesis may be either absent or barely present, but ANP may play a counter regulatory role in lipolysis via NPR-C/Gi pathway.

Fasudil, a Rho-kinase inhibitor, reverses L-NAME exacerbated severe nephrosclerosis in spontaneously hypertensive rats.

BACKGROUND: In this study, we tested the hypothesis that long-term Rho-kinase inhibition would reverse nitro-L-arginine methyl ester-exacerbated nephrosclerosis in spontaneously hypertensive rats and attempted to elucidate the mechanism involved. METHODS: Five groups (each n = 8) were studied: untreated spontaneously hypertensive rats; nitro-L-arginine methyl ester (50 mg/l in drinking water, for 3 weeks)-treated spontaneously hypertensive rats; nitro-L-arginine methyl ester with fasudil (10 mg/kg/day)-treated spontaneously hypertensive rats; nitro-L-arginine methyl ester for 3 weeks followed by fasudil for 3 weeks-treated spontaneously hypertensive rats (same doses), and nitro-L-arginine methyl ester for 3 weeks followed by untreated for 3 weeks. We examined renal function, blood pressure, histological features, oxidative stress markers, and mRNA expression in the renal cortex. RESULTS: Nitro-L-arginine methyl ester-treated spontaneously hypertensive rats had higher blood pressure, proteinuria, and serum creatinine and lower creatinine clearance, urinary NO3/NO2 ratio, and urinary cGMP excretion compared with control spontaneously hypertensive rats (all Ps < 0.05). Nitro-L-arginine methyl ester-treated spontaneously hypertensive rats also had increased free radical metabolites and abnormal morphological findings with increased nicotinamide adenine dinucleotide phosphate oxidase activity, phosphorylation of myosin phosphatase targeting subunit-1, and mRNA expression of RhoA, RhoB, RhoC, collagen I and III, transforming growth factor-beta, nicotinamide adenine dinucleotide phosphate subunit, endothelial nitric oxide synthase, plasminogen activator inhibitor, and intercellular adhesion molecule-1 in the renal cortex compared with control spontaneously hypertensive rats. Long-term co-treatment with fasudil slightly improved these indices, but most of them were not statistically significant. Late fasudil treatment significantly improved kidney function, morphological changes, and alterations of mRNA expression in the renal cortex, although late untreated controls did not show any improvement. CONCLUSION: These results suggest that Rho-kinase inhibition partly reverses hypertensive glomerulosclerosis. The renoprotective effect of the Rho-kinase inhibitor may have multiple mechanisms including inhibition of extracellular matrix production, oxidative stress, adhesion molecule production, and antifibrinolysis.

Role of adrenomedullin system in lipid metabolism and its signaling mechanism in cultured adipocytes.

We investigated the levels of adrenomedullin (AM) system during the process of preadipocyte differentiation and its role in lipid metabolism and cellular signaling mechanism in differentiated adipocytes. We cultured rat preadipocytes and measured the following during the process of differentiation: two molecular forms of AM in the culture medium using a specific immunoradiometric assay and gene expression of AM and its receptor component using RT-PCR analysis. In differentiated adipocytes, we measured the effects of AM on the intracellular cAMP level, lipolysis, glucose incorporation, and the protein levels. Two molecular forms of AM were secreted into the medium, and the AM-mature/AM-total ratio was increased after 6 days of differentiation. Cultured rat preadipocytes highly expressed the genes of AM and its receptor components at day 1, and they increased at day 10. Administration of AM to preadipocytes increased the number of Oil Red O-positive adipocytes and spectrophotometric absorbance of Oil Red O. AM dose dependently increased cAMP level and lipolysis, and its effect was blocked by CGRP(8-37). Isoproterenol increased lipolysis, and AM had additive effects on isoproterenol-induced lipolysis. KT5720 and U0126 significantly inhibited the AM-induced lipolysis, whereas KT5720, but not U0126, significantly inhibited the isoproterenol-induced lipolysis. AM increased glucose incorporation and its effect was blocked by wortmannin. Western blot analysis revealed that AM increased phospho PKA, ERK, and Akt. These results indicate that AM and its receptor component are highly expressed in cultured adipocytes and may play a role in lipid metabolism via a different signaling pathway.

Inhibition of Rho-kinase attenuates nephrosclerosis and improves survival in salt-loaded spontaneously hypertensive stroke-prone rats.

OBJECTIVES: We examined whether the Rho/Rho-kinase pathway is involved in the pathogenesis of nephrosclerosis in severely hypertensive rats and assessed the effects of long-term treatment with a Rho-kinase inhibitor, fasudil, on kidney function, histological findings, gene expressions, and survival. We also attempted to elucidate the mechanisms involved. METHODS: We studied the following four groups: control Wistar-Kyoto rats (WKY), untreated salt-loaded spontaneously hypertensive stroke-prone rats (SHR-SP), low-dose fasudil (15 mg/kg per day)-treated SHR-SP, and high-dose fasudil (30 mg/kg per day)-treated SHR-SP. After 8 weeks' treatment, the effects of fasudil were examined. RESULTS: Untreated SHR-SP were characterized by increased blood pressure without circadian variation, decreased kidney function, abnormal renal morphological findings, and increased messenger RNA expression levels of transforming growth factor beta, collagen I, collagen III, p40phox, p47phox, plasminogen activator inhibitor 1, and intracellular adhesion molecule 1 in the renal cortex, compared with WKY. Long-term high-dose fasudil treatment significantly improved renal function (serum creatinine -32%, creatine clearance +39%), proteinuria (-92%) and histological findings (glomerular injury score -57%, arteriolar injury score -55%, fibrous area -40%, ED-1-positive cells -43%) without changing blood pressure or circadian variation, compared with untreated SHR-SP. In addition, fasudil significantly improved increased mRNA expression levels in the renal cortex. Furthermore, high-dose fasudil significantly prolonged survival time compared with untreated SHR-SP (P < 0.01). Low-dose fasudil treatment improved these variables slightly, but did not affect most significantly. CONCLUSION: The Rho/Rho-kinase pathway participates in the pathogenesis of nephrosclerosis in SHR-SP independently of blood pressure-lowering activity, partly by upregulation of the gene expressions of extracellular matrix, oxidative stress, adhesion molecules, and antifibrinolysis.