Role of miR-145-5p Targeting ARK5 in Regulating the Proliferation and Apoptosis of Human Epithelial Ovarian Cancer Cells / 中国医学科学院学报

Objective To explore the effect of miR-145-5p on the proliferation and apoptosis of human ovarian cancer cells and the possible molecular mechanisms involved.Methods Real-time quantitative PCR was performed to detect the expression of miR-145-5p in ovarian epithelial cells and ovarian cancer cells.CCK-8 and flow cytometry were used to detect the effects of miR-145-5p overexpression on the proliferation and apoptosis of ovarian cancer cells.TargetScan was employed to predict the target genes of miR-145-5p.Western blotting,dual luciferase reporter assay and rescue experiment were employed to predict and verify the underlying molecular mechanism of miR-145-5p function.Results The expression of miR-145-5p in ovarian cancer cells was significantly lower than that in normal ovarian epithelial cells(

Gab2 effects the invasion and metastasis of breast carcinoma through PI3 K/Akt/ARK5/MMP signal pathway / 中国药理学通报

Aim To investigate the molecular mecha-nism of Gab2 in the invasion and metastasis of breast cancer and provide a theoretical basis for clinical pre-vention of breast cancer invasion and metastasis. Methods The Gab2 , MMP-2 and MMP-9 expressions in 80 cases of breast cancer were detected by immuno-histochemistry . Western blot was used to detect the ex-pression of Gab2 protein in MDA-MB-231 cells and MCF-7 cells. The siRNA plasmid was used to transfect MDA-MB-231 cells. Western blot was used to detect the proteins expression of Gab2 , MMP-2 and MMP-9 . Transwell in vitro experiment was used to detect the in-vasion ability of each group transfected MDA-MB-231 cells, Western blot was used to analyze phosphorylation of Akt and ARK5 induced by epithelial growth factor ( EGF ) in transfected cells ( SiGab2/MDA-MB-231 and Scr/MDA-MB-231 ) . Results The expression of Gab2 protein in invasive ductal carcinoma was signifi-cantly higher than in normal breast tissue ( P<0. 01 ) . The expression of Gab2 was dramatically correlated with lymph node metastasis, ER expression, tumor his-tological grade, MMP-2 and MMP-9 (P<0. 05). The expression of Gab2 protein in MDA-MB-231 cells was higher than in MCF-7 cells. The expression of Gab2, MMP-2 and MMP-9 decreased in SiGab2/MDA-MB-231 cells and the invasion ability of SiGab2/MDA-MB-231 cells was significantly decreased ( P<0. 05 ) , and after 5 minutes’ stimulating by EGF, the phosphoryla-tion of Akt and ARK5 was significantly reduced. Con-clusion Gab2 can promote the invasion and metasta-sis of breast cancer by effecting the expression of MMP-2 and MMP-9 through PI3 K/ Akt /ARK5 signal path-way.

Gab2-Akt-ARK5 signaling pathway is associated with the invasion of glioma cells / 中国肿瘤临床

Objective:This study aimed to investigate the effect and significance of a binding protein-2 (Gab2)-Akt-ARK5 signaling pathway on the invasion of glioma cells. Methods:Immunohistochemical methods were used to detect the expressions of Gab2 and ARK5 in 45 cases of glioma tissue. siRNA plasmid was used to transfect LN-229 cells, and western blot was performed to analyze the protein expressions of Gab2 and ARK5. In vitro Matrigel invasion assay was conducted to detect variations in the invasiveness of transfected cells. Western blot was also conducted to analyze the protein phosphorylation of Akt and ARK5 in the cells transfected with Gab2 plasmid. Results:Immunohistochemical assay revealed that the expressions of ARK5 and Gab2 in glioma cells were positively correlated, and both expressions were higher in high-grade glioma (WHO gradeⅢ,Ⅳ) than in low-grade glioma (WHO gradeⅠ,Ⅱ). LN-229 cells transfected with ARK5 plasmid, Gab2 plasmid, ARK5 and Gab2 plasmid, and control plasmid were named siARK5/LN-229, siGab2/LN-229, siARK5 and siGab2/LN-229, and SCR/LN-229, respectively. After transfection was performed, the protein expressions of ARK5 and Gab2 were respectively decreased in siARK5/LN-229 and siGab2/LN-229. The protein expressions of ARK5 and Gab2 in siARK5 and siGab2/LN-229 were also respectively decreased. After ARK5 or Gab2 was downregulated, the number of glioma cells, which invaded and penetrated Matrigel, was decreased (P<0.01). The number of glioma cells also decreased significantly after ARK5 and Gab2 were downregulated. The phosphorylation of Akt and ARK5 in siGab2/LN-229 cells was decreased after these cells were stimulated by insulin-like growth factor-1. Conclusion:The silencing of ARK5 or Gab2 impaired glioma cell invasiveness. The decreased protein expression of Gab2 inhibited the phosphorylation of Akt and ARK5. These results suggested that the Gab2-Akt-ARK5 signaling pathway could be relevantly involved in glioma cell invasion.

Effect of PTTG1 in the invasion of glioma cells / 中国癌症杂志

Background and purpose:Numerous researches indicated that the expression of pituitary tumor transforming gene1 (PTTG1) was correlated with the severity of glioma tumors. However the specific mechanism of PTTG1 is not clear in glioma. In this study, we explored the role and significance of PTTG1 in the invasion of glioma cells. Methods:Western blot was used to detect the expression of PTTG1 protein in various glioma cell lines. siRNA plasmid was used to transfect U87 cells. Western blot was used to analyze the expression of PTTG1 protein in transfected U87 cells. Matrigel invasion assay was used to detect the invasive ability in the cells being transfected in vitro. Western blot was used to analyze epithelial growth factor (EGF) induced protein phosphorylation of ARK5 and Akt in the cells being transfected PTTG1 plasmid (siPTTG1/U87) and scrambled siRNA (Scr/U87). Results:The expression of PTTG1 protein was higher in all glioma cell lines. After transfection, the invasion of siPTTG1/U87 was obviously decreased after 5 min with EGF stimulation than the Scr/U87, the phosphorylation of ARK5 and Akt was significantly enhanced. However, whether or not the existence of EGF, the phosphorylation of ARK5 and Akt had no differences in siPTTG1/U87. Conclusion:In glioma cells, PTTG1 protein is high expressed and maybe have an important function in glioma cells invasion through Akt-ARK5 signaling pathway.

Expression of ARK5 in hepatocellular carcinoma tissue and its effect on growth of SMMC-7721 cells / 吉林大学学报(医学版)

Objective To detect the expression of ARK5 in hepatocellular carcinoma (HCC)tissue and hepatoma SMMC-7721 cells,and to investigate its effect on the growth of hepatoma cells.Methods The expression levels of ARK5 mRNA and protein were determined by RT-PCR and Western blotting in 30 cases of HCC tissue, paracarcinoma tissue,SMMC-7721 cells,and hepatic cells LO2.The SiRNA of ARK5 and negative control (NC) siRNA were constructed and transfected into the SMMC-7721 cells,and used as experimental group and negative control group;at the same time blank control group was set up. The proliferation activity and apoptotic rate of transfected cells were detected by MTT assay and flow cytometry (FCM).Results The PCR and Western blotting results showed that the expression levels of ARK5 mRNA and protein in HCC tissue and SMMC-7721 cells were significantly higher than those in paracarcinoma tissue and LO2 cells (P<0.05 ). The MTT assay results demonstrated that the inhibitory rates of growth of transfected cells in experimental group at 24,48 and 72 h were (19.39±5.42)%, (23.19±0.53)%,and (20.74±1.23)%;there were significant differences compared with blank control group and negative control group (P<0.01).The FCM results indicated that the apoptotic rate of the transfected cells in experimental group was (15.017±0.945)%,there were significant differences compared with blank control group (8.770%± 0.656 )% and negative control group (8.763%± 1.201%) (P<0.05 ). Conclusion The ARK5 expression level is significantly increased in HCC tissue and hepatoma SMMC-7721 cells;the inhibition of ARK5 expression could suppress the growth of hepatoma cells and induce apoptosis. So ARK5 maybe act as a cancer-promoting gene and induce hepatocellular carcinogenesis.