The Expression Of MicroRNA-140 In Osteosarcoma And The Effect Of MicroRNA-140 On Osteosarcoma Cells Biological Behaviors In Vitro

Objective: To detect the expression of miR-140 in osteosarcoma tissues and osteosarcoma cell line;Investigate the biological effects of miR-140 on the proliferation,apoptosis,invasiveness of osteosarcoma cells(U2OS)in virto;Elucidate underlying mechanisms that miR-140 played in osteosarcoma cells.Methods: 40 cases of osteosarcoma tissue specimens diagnosed by pathological examination were collected from the first affiliated hospital of nanchang university,the second affiliated hospital of nanchang university and jiangxi province tumor hospital.Adopt the method of real time fluorescence quantitative RT-PCR detection the expression levels of miR-140 of 40 cases of osteosarcoma tissue,edge of tumor tissue and osteosarcoma cell line(U2OS).By using Lipofectamine2000 according to the manufacturer's instruction,The miRNA-140 mimics and si RNA-HDAC4 were transfected in U2 OS cells respectively,Experiments were randomly allocated into 3groups: the transfection group,NC group and blank group,The expression levels of miR-140 and HDAC4 m RNA in U2 OS cells were detected by the Realtime-PCR.The protein change of HDAC4 and downstream genes HIF-1 of U2 cells after transfection was detected by Western blot,The proliferation of U2 OS cells after transfection was detected by CCK-8 assay,the apoptosis of U2 OS cells after transfection was assessed by FCM,the invasion ability of U2 cells were assessed by Transwell chamber assay.Luciferase reporter assays were performed to further confirm that HDAC4 is a direct target for miRNA-140.Result: Under-expression of miRNA-140 was observed in 29 of 40 osteosarcoma tissues in comparison with the edge of tumor tissue.The expression levels of miR-140 in U2 OS cells with transfection miR-140 mimics is significantly increased(p<0.05),but the expression levels of HDAC4 m RNA in U2 OS cells with transfection miR-140 mimics is significantly decreased(p<0.05),.U2 OS cells with transfection miR-140 mimics proliferation significantly decreased(p<0.05),,cell early apoptosis rate increased(p<0.05),,invasive ability decreased obviously(p<0.05),.Dual-luciferase reporter results show HDAC4 is a direct target gene ofmicro RNA-140.After U2 OS cells were transfected with si-HDAC4,the cell proliferation and invasion was inhibited(p<0.05),and cell apoptosis was induced(p<0.05),the protein expression levels of HDAC4 and downstream genes HIF-1.(p<0.05),Conclusion: MiR-140 is low expressed in osteosarcoma tissue.HDAC4 is a target genes of miR-140,The biological behavior of osteosarcoma cells after suppressing HDAC4 is consistent with over-expression of miR-140.miR-140 in osteosarcoma possibility play a role of tumor suppressor genes that may partially attribute to miR-140 characters of targeting and inhibiting the meaning of the HDAC4 expression.