FIGURE 4.

Hippo signaling participates in MR-mediated protection against H₂O₂-induced NP cell senescence. rNP cells were pretreated with the indicated concentration of MR (200 μM) for 2 h, followed by 200 μM H₂O₂ for another 2 h. (A) Western blot analysis results. (B,C) SA-β-gal staining analysis. rNP cells were pretreated with 1 μM Yap/Taz inhibitor-1 (YTI) and 200 μM MR for 2 h and then treated with 200 μM H₂O₂ for 2 h. (D) qPCR assay was applied to detect the expression of p53 and p21 mRNA levels in rNP cells. (E) Western blot results of p53 protein expression in response to 1 μM YTI treatment. (F) p53 luciferase reporter gene activity results. rNP cells were transiently transfected with a pp53-TA-luc-CP reporter plasmid and cultured with or without H₂O₂ (200 μM) or MR (200 μM) or 1 μM YTI for 48 h. Total cell lysates were subjected to luciferase assay. (G) Western blot results of p53 and p21 protein expressions in response to 200 μM MR with siRNA-mediated knockdown of Taz or Yap. rNP cells were treated with or without H₂O₂ and MR (200 μM) after 24 h of siRNA transfection. Data are representative of 3 independent repeat experiments. Values are expressed as mean ± SD. *p < 0.05, **p < 0.01 compared with control group. #p < 0.05, ##p < 0.01 compared with H₂O₂-treated cells. &p < 0.05, &&p < 0.01 compared with MR- and H₂O₂-treated cells.