MAPK family is considered to play an important role in PKC-mediated signal pathways, therefore we examined the contributions of MAPKs to oridonin-induced autophagy and apoptosis. As shown in Fig. 4A, pretreatment of HeLa
Vanoxerine dihydrochloride with specific inhibitors of ERK (PD98059), JNK (SP600125), and p38 (SB203580) significantly increased the cell growth inhibitory ratio of oridonin. But neither PD98059 nor SB203580 influenced oridonin-induced autophagy, whereas SP600125 suppressed this autophagic ratio (Fig. 4B). The downregulations of Beclin 1 level and conversion from LC3-I to LC3-II further confirmed the negative regulatory effect of SP600125 on oridonin-induced autophagy (Fig. 4C). Also, SP600125 pretreatment resulted in an increase in the apoptotic ratio of oridonin-treated
cells (Fig. 4D). Subsequently, we investigated the effect of PKC on MAPKs by detection of their protein levels in oridonin-treated HeLa cells with or without staurosporine. As shown in Fig. 4E, staurosporine almost thoroughly reversed the oridonin-induced enhancements of JNK and p-JNK levels, whereas the decreases of ERK, p-ERK, p38 and p-p38 levels induced by oridonin did not change. In addition, the increased levels of JNK and p-JNK were further enhanced by PMA treatment compared with oridonin-treated alone (Fig. 4F). These results indicate that JNK but not ERK or p38 contributes to oridonin-induced autophagy, and this process is required for the regulation of PKC.
