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. 2018 Sep 10;10(9):320. doi: 10.3390/cancers10090320

Figure 5.

Figure 5

Melatonin induced expression of lncRNA RAD51-AS1, causing RAD51 downregulation. (A,B) Huh7 and HepG2 cells were treated with 1 mM melatonin for 48 h, and expression of RAD51-AS1 and RAD51 was examined by quantitative real-time RT-PCR. p < 0.001 (***) (C) Human serum melatonin levels and hepatic RAD51-AS1 levels were measured by ELISA and real-time RT-PCR, respectively (n = 33). RAD51-AS1 expression was positively correlated with serum melatonin levels. (D) RAD51-AS1 and RAD51 expression in human liver tissues was analyzed by real-time RT-PCR (n = 33). RAD51-AS1 expression was negatively correlated with RAD51 expression. (E) Real-time RT-PCR results showed that silencing RAD51-AS1 expression had no effect on RAD51 mRNA levels. (F) Real-time RT-PCR results showed that overexpression of RAD51-AS1 had no effect on RAD51 mRNA levels. (G) Huh7 cells were transfected with 50 nM synthesized random control siRNA (si-CTR) or RAD51-AS1-specific siRNA (siRAD51-AS1) and then cotreated with 1 mM melatonin. After 48 h of treatment, cells were harvested and subjected to western blotting to detect RAD51 levels; β-actin served as an internal control (left panel). Quantitative results are shown in the right panel. p < 0.001 (***). (H) Huh7 cells were transfected with a RAD51-AS1-expressing plasmid or the empty vector; after 24 h and 48 h, the cells were harvested and subjected to western blotting to examine RAD51 protein levels (left panel). Quantification of the western blotting results are shown in the right panel. p < 0.05 (*), p < 0.01 (**), p < 0.001 (***). All experiments were performed in triplicate.