| Objective:This study explored the effects of CT23 expression on biological behavior of glioma cell lines(U251MG and U87MG).Methods:Screen two cell lines with an intense expression of CT23 both at the level of mRNA and protein.CT23 siRNA transfected into human glioma cell lines,using cationic liposome X-treme GENE siRNA transfection reagent.After transfection,the mRNA and protein levels expression of CT23 gene were detected by RT-PCR and Western blot,respectively.The cell growth curve was determined by CCK-8.The Flow Cytometry was used to detect the apoptosis and cell cycle of U251MG and U87MG.The expression of cyclin A,cyclin B and caspase 3 were tested by western blot.The cell migration and invasion capabilities were tested by wound healing assay,transwell migration assay and transwell matrigel invasion assay,respectively.The adhesion ability was detected by cell adhesion assay and the expression of MMP 2 and MMP 9 were tested by western blot.Results:CT23 was intensely expressed in glioma cell lines U251MG and U87MG in the levels of both mRNA and protein.Compared with the control group,the mRNA and protein level of CT23 were significantly decreased in the CT23 siRNA intervention group(P<0.01).The cell proliferation capability in CT23 siRNA intervention group was significantly inhibited compared with the control group(P<0.01).The CT23 siRNA intervention group has less S phase cells(%)and more G2/M phase cells(%)than the other groups(P<0.01).The expression of Cyclin A was reduced in the CT23 siRNA intervention group(P<0.01).The results of flow cytometry show that the CT23 siRNA intervention group has higher apoptosis rate(%)and more expression of caspase 3(P<0.01)compared with the other groups.Although in the wound healing assay,no significant differences was found in the first 12 hour,in the second 12 hour the migration rate(%)in the CT23 siRNA intervention group was much lower than the control group(P<0.01).In the transwell migration assay and transwell matrigel invasion assay,the cells passed the transwell membrane4 in the CT23 siRNA intervention group was less than the control groups(P<0.01).MMP2 and MMP-9 were doweexpressed in CT23 siRNA of two glioma cell lines(P>0.05).The results of adhesion assay show that the adhesion capablility in CT23 siRNA intervention group was significantly inhibited(P<0.01).Conclusion:Knock-down of CT23 by small interfering RNA(siRNA)attenuates malignant behaviors of glioma cells in vitro.These findings may shed light on the gene therapy about the CT23 expression in glioma. |
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