Martin Gerl

Vasopeptidase inhibition prevents nephropathy in Zucker diabetic fatty rats

BACKGROUND Blocking the renin-angiotensin system is an established therapeutic principle in diabetic nephropathy. We investigated whether inhibition of both neutral endopeptidase and ACE (vasopeptidase inhibition) can prevent functional and morphological features of nephropathy in the Zucker diabetic fatty (ZDF) rat, an animal model of type II diabetes. METHODS Homozygous (fa/fa) ZDF rats (each n=15) aged 10 weeks were treated with placebo, ramipril (1 mg/kg/day in drinking water), or the vasopeptidase inhibitor AVE7688 (45 mg/kg/day in chow). Metabolic parameters and renal function (metabolic cages) were assessed at baseline (age 10 weeks), and at age 17, 27, and 37 weeks. Twenty heterozygous animals (fa/-) served as lean, nondiabetic controls. At age 37 weeks, the animals were sacrificed and the kidneys analyzed histopathologically. RESULTS Overt diabetes mellitus (blood glucose >20 mmol/l) was established at age 17 weeks in all homozygous ZDF rats. In the placebo group, urinary protein excretion increased progressively from 8+/-1 (baseline) to 342+/-56 mg/kg/day (week 37) whereas diabetes and proteinuria were absent in the lean control group. Ramipril tended to reduce albuminuria and morphological damage (p=ns) but AVE7688 virtually prevented albuminuria (33+/-12 mg/kg/day, p<0.05 vs. ZDF placebo) and drastically reduced the incidence and severity of glomerulosclerosis and tubulointerstitial damage. CONCLUSIONS In ZDF rats, development of diabetes mellitus is accompanied by functional and morphological kidney damage that resembles human diabetic nephropathy. Diabetic nephropathy can be prevented by chronic vasopeptidase inhibition.

The peroxisome proliferator-activated receptor-α (PPAR-α) agonist, AVE8134, attenuates the progression of heart failure and increases survival in rats

AbstractAim:To investigate the efficacy of the peroxisome proliferator-activated receptor-α (PPARα) agonist, AVE8134, in cellular and experimental models of cardiac dysfunction and heart failure.Methods:In Sprague Dawley rats with permanent ligation of the left coronary artery (post-MI), AVE8134 was compared to the PPARγ agonist rosiglitazone and in a second study to the ACE inhibitor ramipril. In DOCA-salt sensitive rats, efficacy of AVE8134 on cardiac hypertrophy and fibrosis was investigated. Finally, AVE8134 was administered to old spontaneously hypertensive rats (SHR) at a non-blood pressure lowering dose with survival as endpoint. In cellular models, we studied AVE8134 on hypertrophy in rat cardiomyocytes, nitric oxide signaling in human endothelial cells (HUVEC) and LDL-uptake in human MonoMac-6 cells.Results:In post-MI rats, AVE8134 dose-dependently improved cardiac output, myocardial contractility and relaxation and reduced lung and left ventricular weight and fibrosis. In contrast, rosiglitazone exacerbated cardiac dysfunction. Treatment at AVE8134 decreased plasma proBNP and arginine and increased plasma citrulline and urinary NOx/creatinine ratio. In DOCA rats, AVE8134 prevented development of high blood pressure, myocardial hypertrophy and cardiac fibrosis, and ameliorated endothelial dysfunction. Compound treatment increased cardiac protein expression and phosphorylation of eNOS. In old SHR, treatment with a low dose of AVE8134 improved cardiac and vascular function and increased life expectancy without lowering blood pressure. AVE8134 reduced phenylephrine-induced hypertrophy in adult rat cardiomyocytes. In HUVEC, Ser-1177-eNOS phosphorylation but not eNOS expression was increased. In monocytes, AVE8134 increased the expression of CD36 and the macrophage scavenger receptor 1, resulting in enhanced uptake of oxidized LDL.Conclusion:The PPARα agonist AVE8134 prevents post-MI myocardial hypertrophy, fibrosis and cardiac dysfunction. AVE8134 has beneficial effects against hypertension-induced organ damages, resulting in decreased mortality. The compound exerts its protective properties by a direct effect on cardiomyocyte hypertrophy, but also indirectly via monocyte signaling and increased endothelial NO production.

Determinants of intramyocellular lipid concentrations in rat hindleg muscle.

The investigation of intramyocellular lipids (IMCLs) with proton MR spectroscopy (1H‐MRS) in humans has recently received increasing attention. IMCL levels correlate with insulin resistance and are affected by diet and exercise, making IMCL an interesting marker for metabolic investigations. In the present in vivo study, the feasibility of using 1H MRS for the detection of IMCL in rats is demonstrated, and the influence of various factors, such as age, gender, muscle type, and rat strain, on IMCL levels is systematically analyzed. In healthy Wistar and Sprague Dawley (SD) rats, the highest ratios of IMCL/tCr were found in young rats, and IMCL/tCr decreased with increasing age. In addition, IMCL concentration was clearly influenced by gender and muscle type. Insulin‐resistant, male, obese, Zucker diabetic fatty (ZDF) rats showed significantly higher IMCL levels than Wistar or SD rats. In conclusion, although IMCL levels are clearly influenced by insulin resistance, several other factors influence IMCL levels, such as age, gender, muscle type, and rat strain. Therefore, when using IMCL as a surrogate marker for insulin resistance, it is necessary to carefully compare results with age‐ and gender‐matched controls, and to use identical conditions. Magn Reson Med 50:242–248, 2003.

Inhibition of Na+–H+ exchange by cariporide reduces inflammation and heart failure in rabbits with myocardial infarction

The aim of this study was to assess the effects of the Na+–H+ exchange inhibitor cariporide on left ventricular (LV) morphology and function as well as inflammation in rabbits with heart failure. Rabbits with myocardial infarction (MI) and sham controls were randomized to receive either standard chow or chow supplemented with cariporide for 9 weeks. LV morphology was determined by echocardiography. LV systolic and diastolic function was assessed under load‐dependent and ‐independent conditions by analysis of LV pressure–volume loops using piezo‐electric crystals. Plasma concentrations of C‐reactive protein and aldosterone were measured. Rabbits with MI developed LV dilatation that was reduced by cariporide. Systolic and diastolic LV function was impaired in rabbits with MI when compared to sham, as indicated by a decreased dP/dtmax (MI: 3537±718 mmHg s−1, sham: 5839±247 mmHg s−1, P<0.05), the load‐independent preload recruitable stroke work (PRSW)(MI: 22±7 mmHg, sham: 81±23 mmHg, P<0.05) and a reduction in the time constant of relaxation tau (τ) (MI: 27±1 ms, sham: 17±1 ms, P<0.05), and significantly improved by cariporide (dP/dtmax: 4586±374 mmHg s−1, PRSW: 67±18 mmHg, τ: 20±2 ms; P<0.05 vs MI/control). Induction of MI was associated with an increase in aldosterone and CRP, indicating activation of the neurohormonal and the inflammatory system that were largely reduced by cariporide. Cariporide improves LV morphology and function post MI and suppresses inflammation and neurohormonal activation in congestive heart failure (CHF). Na+–H+ exchange inhibition may represent a new pharmaceutical approach for the treatment of CHF.

Vasopeptidase Inhibition Prevents Target Organ Damage and Improves Survival in Spontaneously Hypertensive Rats

Background. Vasopeptidase inhibition has been shown to be an effective antihypertensive principle but its long-term effects on hypertensive target organ damage are not known. We investigated the myocardial, vascular and renal effects of chronic vasopeptidase inhibition in arterial hypertension. Methods and Results. One hundred and thirty-nine male spontaneously hypertensive rats aged 15 months were treated chronically with either the pure angiotensin-converting enzyme (ACE) inhibitor, ramipril (1 mg/kg/d in drinking water, n=46), or the vasopeptidase inhibitor AVE7688 (30 mg/kg/d in chow, n=46), or placebo (n=47) and followed up until they died. After six months, both ramipril and AVE7688 had markedly reduced plasma ACE activity, normalised blood pressure (BP), reduced left ventricular mass and improved systolic function to similar extents. Acetylcholine mediated relaxation of aortic rings was improved by both ramipril and AVE7688. There was substantial albuminuria in the placebo group (albumin-to-creatinine ratio 107±54 µg/mg), which was significantly reduced by ramipril to 57±34 µg/mg, and practically abolished in the AVE7688 group (22±12 µg/mg, p<0.05 vs. placebo and ramipril).Tubulointerstitial damage (semi-quantitative score) was significantly reduced by AVE7688 and ramipril. significantly reduced byAVE7688 and ramipril. Overall mortality was markedly reduced in the ramipril and AVE7688 groups (13% and 16% at six months, respectively), both p<0.05 vs. placebo (71%). Conclusions. Vasopeptidase inhibition effectively controls BP and reduces myocardial, vascular and renal target organ damage, resulting in a markedly prolonged survival.At similar degrees of plasma ACE inhibition, AVE7688 compared to ramipril offers superior protection against hypertensive kidney damage.

A radioimmunoassay for the N-terminal propeptide of rat procollagen type III. Application to the study of the uptake of the N-terminal propeptide of procollagen type III in isolated perfused rat liver.

Antibodies against a synthetic peptide representing the 14 C-terminal amino acids of the N-terminal propeptide of rat and bovine procollagen type III were raised in rabbits and used to develop a radioimmunoassay. N-Terminal propeptide of procollagen type III, purified from calf skin, served as standard and tracer material. The IC50 of the standard inhibition curve was 2.1 micrograms/l, the lower limit of detection about 0.4 microgram/l, interassay variation was 8.5% and the intraassay variation 6.6% in typical experiments. Three peaks of antigenicity were detected in rat serum after gel chromatography. One peak coeluted with purified N-terminal propeptide of procollagen type III, one peak contained material approximately twice this size, and one peak eluted close to the void volume of the column. The antigen concentration in rat serum decreased in an age dependent manner. Rat, bovine, sheep and minipig serum antigen was sufficiently crossreactive to allow the application of the assay to these species, whereas human, goat and guinea pig samples were not. The degradation product Col 1, causing non-parallel inhibition in commercially available assays for human samples was not recognized since it does not contain the epitope represented by the synthetic peptide. The assay was used to study the half-life of bovine and endogenous N-terminal propeptide of procollagen type III in isolated perfused rat liver. [125I]-labeled antigen was cleared rapidly from the perfusate (t1/2 less than 5 min). The bovine antigen was removed from the perfusate with a half-life of 15 +/- 4 min. Endogenous propeptide was perfused for 120 min with little change in concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of combined inhibition of the Na+–H+ exchanger and angiotensin‐converting enzyme in rats with congestive heart failure after myocardial infarction

We investigated the single vs the combined long‐term inhibition of Na+–H+ exchanger‐1 (NHE‐1) and ACE in rats with congestive heart failure induced by myocardial infarction (MI). Rats with MI were randomized to receive either placebo, cariporide (3000 p.p.m. via chow), ramipril (1 mg kg−1 day−1 via drinking water) or their combination for 18 weeks starting on day 3 after surgery. Cardiac morphology and function was assessed by echocardiography and by means of a 2.0 F conductance catheter to determine left ventricular (LV) pressure volume relationships. MI for 18 weeks resulted in an increase in LV end‐diastolic diameter (LVDed) in the placebo‐treated group when compared to sham (placebo: 1.1±0.04 cm; sham: 0.86±0.01; P<0.05). Combined inhibition of NHE‐1 and ACE, but not the monotherapies, significantly reduced LVDed (1.02±0.02 cm). Preload recruitable stroke work (PRSW), dp/dtmax (parameter of systolic function) and end‐diastolic pressure volume relationship (EDPVR, diastolic function) were significantly impaired in placebo‐treated MI group (PRSW: 39±7 mmHg; dp/dtmax: 5185±363 mmHg s−1; EDPVR: 0.042±0.001 mmHg μl−1; all P<0.05). Cariporide treatment significantly improved PRSW (64±7 mmHg), dp/dtmax (8077±525 mmHg s−1) and EDPVR (0.026±0.014 mmHg μl−1), and reduced cardiac hypertrophy in rats with MI. Combined inhibition of NHE‐1 and ACE had even a more pronounced effect on PRSW (72±5 mmHg) and EDPVR (0.026±0.014 mmHg μl−1), as well as cardiac hypertrophy that, however, did not reach statistical significance compared to cariporide treatment alone. The NHE‐1 inhibitor cariporide significantly improved LV remodeling and function in rats with congestive heart failure induced by MI. The effect of cariporide was comparable or tended to be stronger (e.g. systolic function) compared to ramipril. Combined treatment with cariporide and ramipril tended to be more effective on LV remodeling in rats with heart failure than the single treatments. Thus, inhibition of the NHE‐1 may be a promising novel therapeutic approach for the treatment of congestive heart failure.