The effect of C5aR knockdown on the phosphorylation of p38 MAPK, ERK1/2 and JNK as well as the expression of IL-6 and TNF-α in C5a-induced GMC.

siC5aR was transfected into the GMC to silence C5aR gene followed by C5a stimulation at the dose of 50 ng/ml for different time points, and then above-mentioned molecules were examined by Western blot, RT-PCR and ELISA respectively. (A-D) Western blot was performed to detect the expression of C5aR and the phosphorylation levels of p38 MAPK (A and B), ERK1/2 (A and C) and JNK (A and D) in the GMC after C5a stimulation for 7.5 min (C and D) or 15 min (B). (E and F) RT-PCR was done to determine the mRNA levels of IL-6 and TNF-α in the GMC after C5a stimulation for 4 h. (G) ELISA was used to detect the release of IL-6 and TNF-α from the GMC after C5a stimulation for 8 h. Results were represented as means ± SD (n = 3 in each group). Representative photographs were shown. ** P<0.01 versus C5a group and siCTR + C5a group.