Fig. 4. Effect of p38 and JNK inhibitors on Fas-mediated activation of caspases. (A) Fas-mediated activation of caspase-8 (lane 2) was increased to a greater extent in cells treated with SP600125 in the absence (lane 4) or presence of PD169316 (lane 5) than that seen with the latter alone (lane 5). (B) Caspase-9 activation was potentiated by SP600125 (lane 4). PD169316 inhibited caspase-9 activation in Fas-activated cells both in the absence (lane 2) and in presence of SP600126 (lane 5). (C) Caspase-3 activity was higher in Fas-activated cells in presence of SP600125 (lane 4) than PD169316 (lane 3). PD169316 blunted the potentiating effect of SP600125 (lane 5). (D) Immunoblot analysis demonstrating the effects of PD169316 and SP600125 on the formation of cleaved fragments of caspase-8 (blot 1), caspase-9 (blot 3) and caspase-3 (blot 5). Increased presence of cleaved caspase-9 and caspase-3 was detected in cells treated with SP600125 (lane 4), but not PD169316 alone (lane 3) or in the presence of the former (lane 5). The differential effects of these inhibitors occurring distal to caspase-8 were confirmed by the differences in cyt c release (blot 4) and PARP cleavage (blot 6) and the lack of a difference in the extent of cleavage of Bid into tBid (blot 2). (Data are representative of three experiments).