Fig. 8. Induction of p8 is required for ET-1-stimulated renal mesangial cell hypertrophy. (A) dsRNA-dependent RNAi of p8 ablates expres sion of p8. Renal mesangial cells were mock transfected with oligofectAMINE or transfected with oligofectAMINE plus p8 siRNA at the indicated concentrations before serum starvation (0.5%) for 24 h. As a control, a separate Petri dish was untreated but starved (0.5%) or left proliferating in normal serum (10%). RNA was extracted and subjected to northern analysis with a p8 probe as described above. (B) p8 RNAi blocks both basal and ET-1-dependent induction of p8 expression. Renal mesangial cells were pre-treated with p8 siRNA (10 µg/6 cm dish), serum starved for 48 h and then treated with ET-1 for the indicated times (0, 24 and 48 h). RNA was extracted and subjected to northern analysis with p8 or gapdh probes. (C) p8 RNAi completely inhibits ET-1-induced renal mesangial cell hypertrophy. p8 siRNA was introduced into the cells and, after 48 h of serum starvation, mesangial cells were subjected to the hypertrophy protocol. ET-1-stimulated [³H]leucine and [³H]thymidine incorporation were monitored as described in Figure 1.