Isoform‐specific modulation of CYP450 omega hydroxylases by cyclosporine in glomerular mesangial cells

Abstract

Cyclosprine A (CSA), an immunosuppressive agent used in the treatment of many proteinuric conditions, protects the glomerular filtration barrier. We have shown that 20‐hydroxyeicosatetraenoic acid (20‐HETE), an eicosanoid generated by the CYP4a and 4f omega hydroxylase activities, maintains and protects the glomerular filtration barrier. We hypothesize that CSA induces expression of CYP4a and/or 4f in mesangial cells to protect the filtration barrier and mitigate proteinuria. We incubated murine mesangial cells in CSA (10 nM, 100 nM, 1 μM, 10 μM) or vehicle for 1, 8 or 24 hrs. We then isolated total RNA and performed real time RT‐PCR using specific primers to measure expression of CYP4f13, CYP4a12a and CYP4a12b. CYP4f13 expression showed the greatest response to CSA treatment. At 8 hr CSA 1 μM and 10 μM increased CYP4f13 by 91% and 74%, respectively. At 24 hr CSA 100 nM, 1 μM and 10 μM increased CYP4f13 by 36%, 268% and 200%, respectively. At 24 hr, CSA 10nM and 100 nM increased expression of CYP4a12a by 75% and 88%, respectively, while CSA 10 μM appeared to suppress expression to 48% of control at 24 hr. CSA 10 μM increased expression of CYP4a12b by 85% at 8 hr; this concentration appeared to suppress expression at 24 hr. Thus, mesangial cells express CYP4a and 4f, and CSA modulates the expression of these CYP isoforms in this cell type in a time and dose‐dependent manner.Supported by NIH/NIDDK R01 DK064969.