Suppression by cyclosporin A of interleukin 1β-induced expression of group II phospholipase A₂ in rat renal mesangial cells

Abstract

1. We investigated whether cyclosporin A, a potent immunosuppressive drug, affects group II phospholipase A₂ (PLA₂; EC 3.1.1.4) induction in rat renal mesangial cells.

2. Previously we showed that the expression of group II PLA₂ in rat renal mesangial cells is triggered by exposure of the cells to inflammatory cytokines such as interleukin 1β (IL-1β) or tumour necrosis factor α and agents that elevate cellular levels of cyclic AMP. Treatment of mesangial cells with IL-1β for 24 h induced PLA₂ activity secreted into cell culture supernatants by about 16 fold. Incubation of mesangial cells with cyclosporin A inhibited IL-1β-induced PLA₂ section in a dose-dependent fashion, with an IC₅₀ value of 4.3 μM. Cyclosporin A did not directly inhibit enzymatic activity of PLA₂.

3. Immunoprecipitation of radioactively labelled PLA₂ protein from mesangial cell supernatants revealed that the inhibition of PLA₂ activity is due to a suppression of PLA₂ protein levels. This effect was preceded by a reduction of PLA₂ mRNA steady state levels, as demonstrated by Northern blot analyses of total cellular RNA isolated from stimulated mesangial cells.

4. In order to evaluate whether cyclosporin A would affect the transcriptional activity of the PLA₂ gene, we performed nuclear run on transcription experiments and provided evidence that the transcription rate of the PLA₂ gene is reduced by cyclosporin A.

5. Previously we found that the nuclear transcription factor κB (NFκB) is an essential component of the IL-1β-dependent upregulation of PLA₂ gene transcription. By electrophoretic mobility shift analysis, we demonstrated that cyclosporin A diminishes the formation of NFκB DNA-binding complexes, thus suggesting that this transcription factor is a target for cyclosporin A-mediated repression of PLA₂ gene transcription.

6. The data presented in this study strongly suggest that the cellular mechanism involved in the IL1β - dependent transcriptional upregulation of the PLA₂ gene in mesangial cells is a target for the action of cyclosporin A.

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