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Biomedical subjects

C Gros

Publications and source records attributed to C Gros.

At least 37 records · Page 2Linked to original sources
PubMed1996

Effect on prolonged inhibition of neutral endopeptidase on cardiac hypertrophy in rats with myocardial infarction.

Myocardial infarction was induced by rats by ligation of the left coronary artery. Treatment with TM1, a prodrug of SQ 28,603, an inhibitor of neutral endopeptidase (NEP, EC 3.4.24.11), was started 18-20 hours after ligation and was continued for 4 weeks (100 mg/kg, orally, twice daily). Morphological and biochemical parameters were assessed at the endo of therapy. The treatment resulted in a significant reduction of heart hypertrophy, which was restricted to the parts of myocardium hemodynamically upstream of the infarcted left ventricle. The weights of the right ventricle and atria were reduced by 15-20%, whereas the treatment had no effect on the left ventricle and septum weights. Treatment led to an almost complete inhibition of plasma NEP activity and to a slight decrease (-14%, p < 0.05) in plasma ACE activity. Plasma ANF level increased 3.8-fold after ligation, and treatment resulted in a slight ( + 29%) and nonsignificant additional increase in the ANF level. The amount of hydroxyproline in the right ventricle was enhanced by + 207% in control ligated rats and by +140% (NS) in treated rats. These data indicated that prolonged NEP inhibition exerts a favorable effect in heart failure by reducing the development of right ventricular and atrial hypertrophy. These effects may result from an improvement in hemodynamic conditions, leading to a reduction in cardiac preload.

Alanine↗
PubMed1996

A novel [125I]iodinated carboxypeptidase A substrate detects a metallopeptidase activity distinct from carboxypeptidase A in brain.

We have designed two radioactive substrates, hippuryl-L-[3H]phenylalanine and 3-(p-hydroxy, m-[125I]phenyl)propionic acid ([125I]Bolton reagent) derivative of L-arginyl-L-phenylalanine, i.e. [125I]BRF, for a highly sensitive assay of carboxypeptidase A (CPA) activity. After cleavage of the C-terminal phenylalanine residue by CPA, the radioactive product of the reaction was conveniently separated by polystyrene bead chromatography. Using [125I]BRF, typical CPA activity inhibited by 1 microM 2-benzyl-3-mercaptopropanoic acid was detected in extracts from rat pancreas or intestine. In brain and some other tissues, however, [125I]BRF-hydrolyzing activity was only inhibited by this compound in 1000-fold higher concentration, suggesting the participation of a metallopeptidase distinct from CPA.

Animals↗
PubMed1995

Purification, cDNA cloning, functional expression, and characterization of a 26-kDa endogenous mammalian carboxypeptidase inhibitor.

The recent demonstration of the occurrence in rat brain and other nonpancreatic tissues of carboxypeptidase A (CPA) gene transcripts without associated catalytic activity could be ascribed to the presence of a soluble endogenous protein inhibitor. This tissue carboxypeptidase inhibitor (TCI), detected by the inhibition of added bovine pancreatic CPA, was purified from rat brain. Peptides were obtained by partial proteolysis of purified TCI, a protein of approximately 30 kDa, and starting from their sequences, a full-length cDNA encoding a 223-amino acid protein containing three potential phosphorylation sites was cloned from a cDNA library. Its identity with TCI was shown by expression in Escherichia coli of a recombinant protein recognized by antibodies raised against native TCI and display characteristic CPA-inhibiting activity. TCI appears as a hardly reversible, non-competitive, and potent inhibitor of CPA1 and CPA2 (Ki approximately 3 nM) and mast-cell CPA (Ki = 16 nM) and inactive on various other proteases. This pattern of selectivity might be attributable to a limited homology of a 11-amino acid sequence with sequences within the activation segments of CPA and CPB known to interact with residues within their active sites. The widespread expression of TCI in a number of tissues (e.g., brain, lung, or digestive tract) and its apparently cytosolic localization point to a rather general functional role, e.g., in the control of cytosolic protein degradation.

Amino Acid Sequence↗
PubMed1995

Carboxypeptidase A isoforms produced by distinct genes or alternative splicing in brain and other extrapancreatic tissues.

The presence of carboxypeptidase A (EC 3.4.17.1; CPA) gene transcripts and corresponding catalytic activity was investigated in brain and other extradigestive rat tissues in which presence of the pancreatic enzyme had not been reported so far. Transcripts of two known CPA genes, CPA1 and CPA2, were identified in extremely low abundance in brain and several other extrapancreatic tissues using Northern blot and reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Whereas the CPA1 gene transcripts in brain, heart, stomach, or colon had a size similar to that in pancreas (1.35 kilobases), the CPA2 gene transcripts in brain, testis, or lung were of a smaller size (1.1 kilobases). Northern blot analysis using various probes, RT-PCR, and 5'-rapid amplification of cDNA 5'-end (5' RACE analysis) all indicated that this smaller size of the brain transcript was attributable to production by alternative splicing of the pro-mRNA. This process corresponds to deletion of the first four exons, leading to a mRNA encoding a protein in which the signal peptide and activation peptide of prepro-CPA2 are absent but the active site remains. The prediction that the shorter CPA2 isoform, designated CPA2(S), should correspond to a cytoplasmic metallopeptidase that does not require tryptic activation was verified by characterization of the recombinant protein and comparing it with the native CPA-like activity in brain. Both recombinant CPA2(S) generated in Escherichia coli and a soluble protein from brain displayed similar sizes on Western blots (32 kDa to be compared to 34 kDa for pancreatic CPA2). Recombinant CPA2(S) and a soluble CPA-like activity from brain displayed similar sensitivity to a series of inhibitors, contrasting with that of the pancreatic enzyme. It is concluded that alternative splicing produces a truncated CPA2 with distinct subcellular localization and modified catalytic activity. In spite of the presence of the CPA1 mRNA, no corresponding CPA activity could be detected in brain extracts, even after tryptic activation. This apparent discrepancy seems attributable to the presence of an endogenous peptide inhibitor which remains to be identified.

Alternative Splicing↗
PubMed1995

Large scale purification of rubella virus and the isolation of native viral core protein.

A number of structural analyses of viruses are dependent on the availability of purified virus and of pure viral components in milligram amounts. In order to allow such analyses of the Rubella togavirus we have identified a virus-cell-system which produces large amounts of Rubella virus in tissue culture and we have developed a rapid and efficient procedure of Rubella virus purification which involves adsorption and elution of virus to fixed erythrocytes. Furthermore, we describe a procedure which allows the extraction of native core protein from viral cores and its chromatographic purification.

Animals↗
PubMed1994

Effects of alatriopril, a mixed inhibitor of atriopeptidase and angiotensin I-converting enzyme, on cardiac hypertrophy and hormonal responses in rats with myocardial infarction. Comparison with captopril.

The aim of the study was to compare, in a rat model of congestive heart failure, the effect of captopril, a selective angiotensin-converting enzyme (ACE; EC 3.4.15.1) inhibitor, to that of alatriopril, a mixed inhibitor of ACE and atriopeptidase (EC 3.4.24.11), an enzyme implicated in the degradation of atrial natriuretic factor (ANF). Myocardial infarction was induced by ligation of the left coronary artery. Groups of rats received orally twice daily captopril (10 mg/kg), alatriopril (100 mg/kg) or vehicle. Treatments were started 18 to 20 h after ligation and continued for 4 weeks. Hypertrophic and hormonal changes reflecting congestive heart failure were assessed in rats with large infarcts by measuring the relative weight of cardiac tissues as well as by assaying ANF in heart and plasma and by measuring renin activity in plasma. Both treatments significantly reduced cardiac hypertrophy, but alatriopril showed a greater efficacy than captopril--the increase in relative heart weight reaching 38% with captopril and only 22% with alatriopril (P < .05). The hypertrophy of right ventricle was reduced by 47% with alatriopril and by 35% with captopril (N.S.), whereas the corresponding reductions for atria were 47% vs. 21% (P < .05). Both treatments prevented the ligation-induced increase of ANF level in the right ventricle. In contrast, plasma ANF level was significantly reduced after captopril but not after alatriopril treatment, a difference that probably reflects the protection of endogenous ANF in circulation resulting from atriopeptidase inhibition. Plasma renin was increased by 36-fold after captopril but only by 1.6-fold after alatriopril, a difference that presumably reflects the inhibition of renal renin secretion by endogenous ANF after alatriopril. These data suggest that enhancement of ANF levels in circulation via atriopeptidase inhibition magnifies the capacity of ACE inhibitors to prevent cardiac hypertrophy, and they show the potential therapeutic value of mixed ACE-atriopeptidase inhibitors in congestive heart failure.

Alanine↗
PubMed1993

Inhibition of neutral endopeptidase stimulates renal sodium excretion in patients with chronic renal failure.

1. The acute effects of a single oral dose of sinorphan (100 mg), an inhibitor of neutral endopeptidase, on the plasma atrial natriuretic factor level and the fractional excretion of sodium were examined in 12 patients with severe chronic renal failure who were not on maintenance haemodialysis and who ingested a normal sodium diet. The drug was administered against placebo by a double-blind cross-over protocol. 2. Basal plasma atrial natriuretic factor level and fractional excretion of sodium were high (23.2 +/- 3.7 pmol/l and 2.64 +/- 0.38%, respectively). Sinorphan inhibited plasma neutral endopeptidase activity by 68-75% 30 min after ingestion. This effect persisted for at least 4 h. There were simultaneously increases in plasma atrial natriuretic factor and cyclic GMP levels to 1.9 and 1.4 times the basal values, respectively. Fractional excretion of sodium increased during the second and third hour periods after ingestion of the drug with a peak of 1.9 times the basal value in the second period. Changes in fractional excretion of sodium were significantly correlated with those in plasma atrial natriuretic factor and cyclic GMP levels. Plasma aldosterone level, creatinine clearance and mean blood pressure were unchanged, whereas plasma renin activity increased slightly. An increase in urinary cyclic GMP excretion was observed in parallel with the increase in plasma cyclic GMP level.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Binding of [125I]atrial natriuretic factor to mouse lung membranes in vivo: characterization and effects of peptidase inhibitors.

After i.v. injection of 125I-labeled rat atrial natriuretic factor ([125I] ANF; 99-126) in tracer dose to mice, a saturable binding to lung membranes was evidenced using a filtration assay. Analysis of the membrane-bound radioactivity by high-pressure liquid chromatography indicated that it corresponded to the intact hormone in sinorphan-treated mice. [125I]rANF binding was inhibited completely by i.v. administration of rANF with an ED50 of 1.0 +/- 0.1 nmol/kg, a value obtained in sinorphan-treated mice. SC 416,542, an ANF analog with a four amino acid deletion in its ring, representing a selective ligand of ANF clearance receptors, was as potent as rANF in inhibiting the in vivo binding. By contrast, ANF fragments produced by enkephalinase (EC 3.4-24.11, membrane metalloendopeptidase) were less potent or even inactive in competing with [125I]rANF. It is concluded that [125I]rANF binding to lung membranes in vivo occurs to clearance receptors. [125I]rANF binding was enhanced by more than 2-fold in mice receiving enkephalinase inhibitors such as sinorphan and, although to a lesser extent, aminopeptidase inhibitors; on the other hand inhibitors of a variety of other peptidases were ineffective. These data confirm by a novel approach that enkephalinase plays a key role in the inactivation of circulating ANF. Hence, the in vivo binding test can be used to assess the activity of clearance receptor ligands and peptidase inhibitors, two classes of drugs affecting ANF metabolism, with potential clinical utility in cardiovascular and salt-retaining diseases.

Animals↗
PubMed1991

Mixed inhibitors of angiotensin-converting enzyme (EC 3.4.15.1) and enkephalinase (EC 3.4.24.11): rational design, properties, and potential cardiovascular applications of glycopril and alatriopril.

Angiotensin-converting enzyme (ACE) and enkephalinase, two cell surface metallopeptidases, are responsible for angiotensin II formation and atrial natriuretic factor (ANF) degradation, respectively, and thereby play a critical role in the metabolism of hormonal peptides exerting essentially opposite actions in cardiovascular regulations. To affect simultaneously both hormonal systems by a single molecular structure, we have designed glycoprilat and alatrioprilat [(S)-N-[3-(3,4-methylene-dioxyphenyl)-2-(mercaptomethyl)-1-oxoprop yl] glycine and -alanine, respectively]. In vitro the two compounds inhibit both ACE and enkephalinase activities with similar, nanomolar potencies, and in vivo, glycopril and alatriopril, the corresponding diester prodrugs, occupy the two enzyme molecules in lung at similar low dosages (0.2-0.5 mg/kg of body weight, per os). The high potency of these compounds is attributable to interaction of the methylenedioxy group with the S1 subsite of ACE and of the aromatic ring with the S1' subsite of enkephalinase. In rodents, low doses of these mixed inhibitors exert typical actions of ACE inhibitors--i.e., prevention of angiotensin I-induced hypertension--as well as of enkephalinase inhibitors--i.e., protection from 125I-ANF degradation or enhancement of diuresis and natriuresis following acute extracellular volume expansion. In view of the known counterbalanced physiological actions of the two hormonal peptides, whose metabolism is controlled by ACE and enkephalinase, mixed inhibitors of the two peptidases show promise for the treatment of various cardiovascular and salt-retention disorders.

Alanine↗