Effect of cyclosporin A on endothelin synthesis by cultured human renal cortical epithelial cells.

Abstract

We report here for the first time that human renal proximal tubular cells secrete endothelin, clear evidence of de-novo endothelin synthesis by these cells and the effect of cyclosporin A (CsA) on endothelin synthesis both in short-term (24 h) and medium-term (5-day) culture. Human renal cortical epithelial cells were cultured and shown to possess proximal tubular characteristics. These cells produced endothelin in culture in a time-dependent manner, as measured by radioimmunoassay (291.6 +/- 51.4 pg/well/24 h). Furthermore, endothelin production by these cells was significantly decreased by up to 80% by cycloheximide (1051.8 +/- 54.9 pg/mg cell protein/24 h versus 253.2 +/- 12.6 pg/mg cell protein/24 h), showing that these cells actively synthesize endothelin. In short-term culture (24 h), CsA significantly inhibited endothelin synthesis at a medium concentration of 10,000 micrograms/l. No change in endothelin synthesis was seen at lower CsA concentrations. In contrast, over a 5-day period, a non-significant increase in endothelin synthesis was observed at CsA concentrations of 2000 micrograms/l (152.5 +/- 20.4%); however, cell growth was significantly decreased at this concentration (71.33 +/- 6.39%). Using a newly developed two-site immunoradiometric assay specific for endothelin-1 (ET-1), we demonstrate that ET-1 is the major endothelin isoform produced by human renal proximal tubular cells.