Figure 4

MicroRNA-34a induction during cisplatin nephrotoxicity is p53 dependent. (A) Effects of pifithrin-a on cisplatin-induced miR-34a expression in BUMPT cells. BUMPT cells were treated with 40 µmol/L cisplatin for 12 h in the presence or absence of 20 µmol/L pifithrin-α. Total RNA was isolated for real-time PCR analysis of miR-34a using specific primers. (B) Effects of pifithrin-α on cisplatin-induced p53 activation. BUMPT cells were treated with 40 µmol/L cisplatin for 0–20 h in the presence or absence of 20 µmol/L pifithrin-α. Whole cell lysates were collected for immunoblot analysis of phospho-p53 (ser-15) and β-actin. Upper panel, immunoblots; lower panel, densitometry analysis. For densitometry, the phosph-p53 signal of each condition was normalized with its β-actin signal and then compared with that of the control (0 cisplatin), which was arbitrarily set as one. (C) miR-34a induction during cisplatin treatment of wild-type and p53-deficient mice. Male wild-type and p53-deficient mice (age 8–10 wks) were injected with 30 mg/kg body weight cisplatin. Kidneys were collected at d 3 of cisplatin injection to isolate total RNA for real-time PCR analysis of miR-34a. (A,C) The miR-34a signals were normalized with that of control samples, which were arbitrarily set as one. Data are expressed as mean ± SD, n = 3; *, significantly different from the cisplatin and wild-type groups.