Cyclosporine A가 PEGF 및 IL - 1a 유발 메산지움 세포내 Cytoslic 및 Group - II Secretory Phospholipase A2 활동도에 미치는 영향

Abstract

The utility of Cyclosporine A(CsA) in various therapeutic applications is chiefly limited by consi- derable nephrotoxicity which seems to be related to its interference with the synthesis of vasoactive prostanoids. Mesangial cells have high levels of cy- tosolic PLAz(cPLAz), the regulatory enzyme in arachidonic acid(A.A.) release and prostaglandin(PG) synthesis, and group-II secretory PLAz(sPLAz), me- diating inflammatory reaction. In order to investigate the molecular mechanism of the effect of CsA on PG synthesis, we evaluated the effect of CsA on the activity of PLAz stimulated by platelet-derived growth factor(PDGF) and interleukin-la(IL-1), which are known to play important roles in glomerular processes. The activity of PLAz was measured in cytosolic fraction and culture supematant, and as- sayed by the release of free A.A. from exogenously added [C]arachidonylphosphatidylcholine(cPLAz) and ethanolamine(sPLAz). Compused to control PDGF(10ng/ ml, 10min.) significantly stimulated cPLAz activity(M ±S.D, 22.5±8.4 pmol of AA/min/mg of protein vs 50.1±11.7, p<0.05), but did not increase sPLAz ac- tivity. IL-1(10ng/ml) treatment also significantly en- hanced cPLAz activity(22.5±8.4 vs 45.4±10.6, p<O.06). IL-1 treatment for 24 hours significantly increased the release of sPLAz into the culture supernatant (47.5±7.7 fmol of A.Ajhour/mg of supernatant pro- tein vs 200.8±24.3, p<0.05). The administration of CsA(100ng/ml) markedly inhibited the activity of cPLAz following stimulation of PDGF or IL-1(PDGF; 50.1±11.7 vs 20.3±12.8, IL-la: 45.5±10.6 vs 25.2± 13.3, p<0.05), but failed to prevent the increase of sPLAz activity(IL-la; 200.8±24.3 vs IL-la+CsA; 180.0± 19.6, p>0.05). There was no significant change of lactic dehydrogenase concentration in culture su- pernatant after exposure to cydosporine for 24 hours(control; 61.4±9.8 IU/L vs IL-la+CsA; 70.4±12.1, p>0.05). On gel filtration chromatography, PDGF-and IL-la-stimulated cPLAz activity migra- ted as a single peak, suggesting tbe molecular mass of approximately 60 kD, and the sPLAz activity was detected as a single peak with an estimated mole- cular mass of 15 kD. The administration of CsA did not change the position of peaks of both PLAz activity. These data suggest that while CsA does not influence the activity of sPLAz, the inhibitory effect of CsA on cPLAz activity result from the modification of enzyme without change of molecular weight. We speculate that cPLAz in mesangial cells should be a molecular target for CsA providing a possible mechanism for interference of the drug with the balance of vasoactive prostanoids, but further studies on cyclooxygenase activity and PG measure- ment are needed.