The Effect Of MiR-224 On Autophagy Related Protein P62 Expression In HeLa Cell

Objective:According to recent study,ectopic expression of micro RNA could lead to cervical cancer.Some studies showed that compared with normal cervical tissue,the level of miR-224 expression was obviously higher in cervical cancers.The present experiment aimed to detect the expression level of cervical caner cells and normal epithedlial cells.The result of the detection will validate the results of previous experimental.And the experiment will study the effect of autophagy by interfering the expression of miR-224 in He La cells.Methods:(1)qRT-PCR technology was used to detect the expression level of miR-224 in cervical cancer cell line(He La)and normal epithelial cell line(293T).(2)Western blot technology was used to detect the expression level of p62 protein in He La and 293 T cells.(3)In He La cells,miR-224 over expression and low expression were achieved by transfection with chemical synthetic miR-224 mimic and miR-224 inhibitor oligo nucleotides with siRNA-Mate transfection reagents.The He La cells were divided into 4groups:miR-224 mimic group(Group M),miR-224 mimic negative control group(Group NC),miR-224 inhibitor group(Group IN),miR-224 inhibitor negative control group(Group INC).Real-time PCR technology was used to detect the expression level of miR-224.Western blot technology was used to detect the expression level of p62 protein in each group.Results:(1)The relative expression of miR-224 in 293 T and He La were 0.151±0.014,6.732±1.33,respectively.Compared with normal epithelial cell,the expression of miR-224 is significantly up-regulated in cervical cancer cell(P<0.05).(2)The level(0.608±0.021)of p62 in Hela was less than the level(0.888 ±0.033)of p62 in 293T(P<0.05).(3)In transfection of miR-224 mimic group,compared with negative control,the expression of miR-224 showed about 25.13 fold up-regulation(P<0.05),the expression of p62 protein showed a 74% decrease(P<0.05).In contrast,the transfection of miR-224 inhibitor made the expression of miR-224 showed a 24% decrease(P < 0.05),the expression of p62 protein showed a 1.71 fold up-regulation(P<0.05).Conclusion:(1)MiR-224 is significantly up-regulated in He La cells.It is suggested that it may play a role in promoting cancer in cervical cancer.(2)The level of autophagy activity is up-regulated in He La cells.In He La,the over expression of miR-224 can decreased the expression of p62 protein;to reduce the expression of miR-224 can increased expression of p62 protein.This suggests that miR-224 might regulate the level of autophagy activity in He La cells.It is suggested that miR-224 may affect the biological behavior of cervical cancer cells by affecting the the autophagy activity of cells.