RECEPTORS FOR THE ANAPHYLATOXIN C5A (CD88) ON HUMAN MESANGIAL CELLS

In these studies, we determined whether there are receptors for the an aphylatoxin C5a (C5aR, CD88) on human mesangial cells (HMC). To prepar e Abs to C5aR, we first synthesized an immunogenic peptide spanning re sidues 8-32 of the molecule, and this peptide was used to immunize rab bits. Anti-C5aR antiserum, but not preimmune serum, stained fixed and unfixed HMC in culture. By Western blotting anti-C5aR, Abs identified a 49.6-kDa protein in HMC. By reverse-transcription PCR, a cDNA produc t of 558 bp was amplified corresponding to the expected size of C5aR c DNA. A cDNA of the same size was amplified simultaneously from human P BL, Restriction mapping of the products amplified from HMC and from PB L gave restriction fragments of the same size, incubation of HMC with increasing doses of C5a caused a progressive increase in the levels of the transcription factors activator protein-1 (AP-1) and cAMP respons e element binding protein (CREB), but C5a had no effect on the level o f nuclear factor-kappa B (NF-kappa B), The effects of C5a on AP-1 were concentration and time dependent and peaked after 60 min. In contrast , the C5a metabolite C5adesArg had no significant effect on AP-1 level s. Preincubation of HMC with rabbit anti-C5aR antiserum inhibited part ially the effect of C5a on AP-1. However, anti-C5aR Abs alone had no a ppreciable effects on AP-1, C5a caused a significant up-regulation of mRNA for the early response genes c-jun and c-fos on NMC. These result s provide evidence for the presence of C5aR in adult BMC in culture an d indicate that, after binding to C5aR, the anaphylatoxin C5a causes s ignificant up-regulation of certain transcription factors and early re sponse genes.