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. 2023 Feb;384(2):265–276. doi: 10.1124/jpet.122.001429

Fig. 6.

Fig. 6.

miR-9-3p and miR-9-5p inhibitors increase TOP2β/180 protein and enhance XK469-induced DNA damage in K/VP.5 cells. (A) Representative immunoassay (from five experiments performed on separate days) using cellular lysates from K/VP.5 cells transfected (48 hours) with control, miR-9-3p, or miR-9-5p inhibitors (50 nM). Blots were probed with antibodies specific for TOP2β or for GAPDH. (B) Expression of TOP2β/180 protein levels in K/VP.5 cells transfected (48 hours) with either control, miR-9-3p, or miR-9-5p inhibitors (50 nM). Results shown are the mean ± S.D. from five experiments performed on separate days, comparing control inhibitor versus miR-9-3p inhibitor (P = 0.043) and miR-9-5p mimic (P = 0.014), respectively. Statistical analysis was performed using a two-tailed paired Student’s t test, as documented in Materials and Methods. All data points represent biologic replicates from the separate experiments performed on separate days. (C) TOP2β/180 mRNA levels in K/VP.5 cells transfected (48 hours) with either control, miR-9-3p, or miR-9-5p inhibitors (50 nM). Results shown are the mean ± S.D. from four experiments performed on separate days, comparing control mimic versus miR-9-3p mimic (P = 0.427) and miR-9-5p mimic (P = 0.605), respectively. Statistical analysis was performed using a two-tailed paired Student’s t test, as documented in Materials and Methods. Data points represent biologic replicates from the separate experiments performed on separate days. (D) K/VP.5 cells transfected (48 hours) with either control, miR-9-3p, or miR-9-5p inhibitors (50 nM) were incubated with XK469 (250 µM) or DMSO for 1 hour, followed by performance of alkaline Comet assays. Results shown are the mean ± S.D. from five experiments performed on separate days, comparing effects of XK469 in miR-inhibitor controls versus miR-9-3p inhibitor (P = 0.016) and mir-9-5p inhibitor (P = 0.011), respectively. *P < 0.05. Statistical analysis was performed using a two-tailed paired Student’s t test, as documented in Materials and Methods. All data points represent biologic replicates from the separate experiments performed on separate days.