Fig. 7.

RBM3 activates cell proliferation through SCD-circRNA 2. EdU experiments were conducted to verify whether SCD circRNA-2 is associated with the cell cycle. SCD-circRNA 2 groups revealed high levels of DNA replication, while low replication levels were observed in siRNA groups relative to those in control (a-b). Data are presented as means ± SD; n = 3. Student's t-test was used. * P < .05. Scale bars, 100 μm. (c) The effect of RBM3 shRNA on the proliferation of HepG2 and Huh7 cell lines is reversed by SCD circRNA-2 overexpression. Scale bars, 100 μm. Statistics for quantitative data are shown in (d). Data are presented as means ± SD; n = 3. Student's t-test was used. To further demonstrated that SCD-circRNA2 was the downstream of RBM3, we established subcutaneously implanted tumor models with HepG2 cells infected with overexpression negative control lentivirus (HepG2 Over-NC), HepG2 cells infected with RBM3 overexpression lentivirus and shRNA negative control lentivirus (HepG2 Over-1 / shRNA NC), and RBM3 overexpressed HepG2 cells infected with SCD-circRNA 2 shRNA lentivirus (HepG2 Over-1 / SCD-circRNA 2 shRNA) in nude mice. Tumor images are presented in (e). Volume (f) and weights (g) of subcutaneous tumors derived from HepG2 Over-NC, HepG2 Over-1 / shRNA NC and HepG2 Over-1 / SCD-circRNA 2 shRNA cells at four weeks after neoplasm seeding. For (f-g), data were presented as means ± SEM. Mann-Whitney U test was used. All P values are defined as: *P < .05, **P < .01 and ***P < .001. (h) RBM3-overexpressing HepG2 and Huh7 cells were transfected with SCD-circRNA 2 siRNA. Forty-eight hours after transfection, cells were harvested. ERK, p38 and JNK phosphorylation was assessed by western blotting analysis with the indicated antibodies. (i) The overexpression of SCD-circRNA 2 was performed as described in Fig. 6a. The relative expression of phosphorylated ERK significantly increased after SCD-circRNA 2 overexpressing in HepG2 (up) and Huh7 (down) cells.