Collagen Production in Cardiac Fibroblasts During Inhibition of Aminopeptidase B

• Published in: Journal of the renin-angiotensin-aldosterone system 2005-09, Vol.6 (2), p.69-77
• Main Authors: Lijnen, Paul J, Petrov, Victor V, Turner, Marisa, Fagard, Robert H
• Format: Article
• Language: English
• Subjects: Alanine - metabolism; Aminopeptidases - antagonists & inhibitors; Animals; Arginine - metabolism; Cell Separation; Cells, Cultured; Collagen - biosynthesis; DNA - biosynthesis; DNA - genetics; Fibroblasts - drug effects; Fibroblasts - enzymology; Fibroblasts - metabolism; Gene Expression - drug effects; Guanidines - pharmacology; Heart - drug effects; Male; Matrix Metalloproteinases - metabolism; Myocardium - metabolism; Protease Inhibitors - pharmacology; Protein-Lysine 6-Oxidase - metabolism; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta - pharmacology; Transforming Growth Factor beta1
• ISSN: 1470-3203; 1752-8976
• DOI: 10.3317/jraas.2005.012

Objective. To determine whether the aminopeptidase B inhibitor, arphamenine A, could affect collagen production and expression in control and TGF-ß1-treated cardiac fibroblasts. Design and Methods. Cardiac fibroblasts from passage 2 from normal male adult rats were cultured to confluency and incubated with and without 600 pmol/l TGF-ß1 for 2 days in serum-free Dulbecco's modified Eagle's medium and then incubated with 100 µmol/l arphamenine A for 1 day in this medium added ascorbic acid, ß-aminopropionitrile and titriated proline. Soluble collagen was measured in the conditioned medium and non-soluble collagen in the cell layer. Aminopeptidase activity was estimated by spectrophotometric determination of the liberation of p-nitroaniline from alanine- or arginine-p-nitroanilide. Matrix metalloproteinase (MMP) and lysyl oxidase activity were assayed in the conditioned medium. A semi-quantitative reverse transcriptase- polymerase chain reaction was used to examine the expression of lysyl oxidase and collagen type I and III. Results. Arphamenine A dose-dependently inhibited basal and TGF-ß 1-stimulated aminopeptidase activity. Arphamenine A reduced soluble and non-soluble collagen production in control and TGF-ß1-treated cardiac fibroblasts, while it decreased collagen type I and III expression only in TGF-ß1-treated fibroblasts. Lysyl oxidase, MMP-1 and MMP-2 activity were inhibited by arphamenine A in the conditioned media of control and TGF-ß1treated cardiac fibroblasts. Conclusions. Our data show that the specific aminopeptidase B inhibitor, arphamenine A, reduces collagen production in cardiac fibroblasts and that this reduction is accompanied by a pronounced inhibition of lysyl oxidase.