Figure 7. Effect of KRGP treatment on Klotho-induced FoxO3a binding to the MnSOD promoter in Tac-treated HK-2 cells. HK-2 cells were seeded in culture plates at 90% confluence. On the next day, the cells were treated with Tac (50 μg/mL) in the absence or presence of 10 μg/mL KRGP. After 12 h, the cells were harvested to detect MnSOD transcription activity after nuclear FoxO3a translocation using a ChIP assay and quantitative real-time PCR (q-PCR) for MnSOD expression. (A) The FoxO3a protein/MnSOD promoter complex was analyzed by q-PCR with primer pairs for the MnSOD-promoter region containing a FoxO3a-binding element. (B) MnSOD mRNA was detected by q-PCR in whole-cell lysates. (C and D) MnSOD mRNA and protein expression in mitochondrial fraction from each group. Relative MnSOD protein expression was presented after normalization to CoX-IV expression Relative MnSOD expression was presented after normalization to cyclophilin A expression. Data are presented as mean ± SE and are representative of at least three independent experiments. #P < 0.05.