Regulation of endothelin production and secretion in cultured collecting duct cells by endogenous transforming growth factor-β

Abstract

Confluent cultures of two renal collecting duct cell lines (M-1 and mIMCD-K2 cells derived from cortical and inner medullary collecting ducts, respectively) express endothelin1 (ET1), transforming growth factor-β (TGFβ; both TGFβ1 and TGFβ2), and both types of the TGFβ receptor. Experiments were performed to test whether endogenous TGFβ may be a paracrine modulator of ET1 expression in these cells. Treatment of M-1 and mIMCD-K2 cells with TGFβ2 antisense oligodeoxynucleotides (ODN) significantly reduced ET1 messenger RNA (mRNA) and ET secretion (as well as TGFβ2 mRNA) in a concentration-dependent manner, whereas control ODN were without significant effects. To produce ET inhibition, antisense ODN had to be present in the basolateral medium, whereas its sole presence in the apical medium was without effect. In addition, a pan-specific TGFβ antibody caused a significant reduction of ET1 mRNA expression and ET1 secretion. M-1 cells were found to express high levels of the mRNA for plasminogen activator of both tissue and urokinase types. Addition of the nonspecific serine protease inhibitor aprotinin (50 μg/ml) to the medium for 24 h significantly reduced the secretion of ET1. These results suggest that secretion of endogenous TGFβ, at least in part activated by the plasminogen/plasmin system, participates in the regulation of ET1 synthesis and secretion by collecting duct cell lines.