Neutral endopeptidase expression in mesangial cells

In the kidney, neutral endopeptidase (NEP) is implicated in the metabolism of several peptides involved in blood pressure and sodium homeostasis control, such as the atrial natriuretic peptide, bradykinin and angiotensin I. Due to its physiological importance in the modulation of pressor responses, the presence of NEP in mouse mesangial cells has been investigated, since these cells control glomerular function and are able to synthesise components of the renin-angiotensin system. A NEP-like activity (NEP-like) that cleaves the fluorogenic substrates Abz-BKQ-EDDnp and AbZ-DRRL-EDDnp was purified from mesangial cell lysate by ion-exchange, followed by gel filtration chromatography. The enzyme was able to hydrolyse bradykinin at the G(4)-F-5 peptide bond and was inhibited by thiorphan. A pH study established that enzyme activity was maximal at pH 7.5 and the determined K-m was 4.86 muM using AbZ-DRRL-EDDnp as substrate. NEP-like was recognised by monoclonal anti-NEP and had a molecular mass of 95 kDa. The purified enzyme was sequenced and showed similarity with human, rat, mouse and rabbit NEPs. We isolated, for the first time, NEP-like from mesangial cells. This enzyme could have an important role in the renal physiology by its action upon different peptides that are able to alter renal haemodynamics.Univ Fed Sao Paulo, Dept Med, Disciplina Nefrol, Div Nephrol, BR-04023900 Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Biophys, Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Med, Disciplina Nefrol, Div Nephrol, BR-04023900 Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Biophys, Sao Paulo, BrazilWeb of Scienc