Changes in proliferation,migration,and invasion of human cervical cancer cell line HeLa with overexpression of miR-216a/CAPN6 gene and their targeted relationship
Objective To observe the changes in proliferation,migration,and invasion of human cervical cancer cell line HeLa with overexpression of microRNA-216a(miR-216a)/Calpain 6(CAPN6)gene and their targeting relation-ships,and to explore the effects of miR-216a on the proliferation,migration,and invasion of HeLa cells.Methods HeLa cells in the logarithmic growth phase were divided into five groups:groups Ⅰ,Ⅱ,Ⅲ,Ⅳ and Ⅴ,with 6 well plates in each group.Cells in the group Ⅰ were transfected with miR-216a mimic,cells in the group Ⅱ were transfected with pcDNA3.1-CAPN6 plasmid,cells in the group Ⅲ were sequentially transfected with miR-216a mimic and pcDNA3.1-CAPN6,cells in the group Ⅳ were transfected with pcDNA3.1,and cells in the group Ⅴ were transfected with miR-216a mimic NC.At 48 h of cultivation,the proliferation,migration,and invasion of cells in each group were observed by CCK-8 method,Scratch healing test and Transwell test.At 24 h of cultivation,miR-216a in each group was detected by qRT-PCR,and CAPN6 and signal transducer and activator of transcription 3(STAT3)proteins in each group were detected by Western blotting.In addi-tion,HeLa cells in the logarithmic growth phase were divided into four groups:cells in the group A were sequentially trans-fected with miR-216a mimic and pGL3-CAPN6-WT plasmids,cells in the group B were sequentially transfected with miR-216a mimic and pGL3-CAPN6-MUT plasmids,cells in the group C were sequentially transfected with miR-216a mimic NC and pGL3-CAPN6-WT,and cells in the group D were sequentially transfected with miR-216a mimic NC and pGL3-CAPN6-MUT plasmids.At 36 h of cultivation,cells from each group were collected and the relative luciferase activity was calculat-ed by the dual-luciferase reporter gene assay kit.Results Compared with the group V,group I had lower cell prolifera-tion activity and percentage of scratch front migration distance,and fewer invasive cells at 48 h of cultivation(all P<0.01).Compared with the group IV,group II had higher cell proliferation activity and percentage of scratch front migra-tion distance,and more invasive cells(all P<0.01).Compared with the group I,group III had higher cell proliferation ac-tivity and percentage of scratch front migration distance,and more invasive cells(all P<0.01).Compared with the group II,group III had lower cell proliferation activity and percentage of scratch front migration distance,and fewer invasive cells(all P<0.01).Compared with group V,the relative expression level of miR-216a of cells in the group I was much higher(P<0.01).Compared with the group IV,the relative expression levels of CAPN6 protein and p-STAT3/STAT3 in the group II were much higher at 24 h of cultivation,while the relative expression levels of CAPN6 protein and p-STAT3/STAT3 were lower in the group III.Compared with the group V,the relative expression levels of CAPN6 protein and p-STAT3/STAT3 in the group I were lower.Compared with group I,the relative expression levels of CAPN6 protein and p-STAT3/STAT3 in the group III were higher.Compared with the group B,group A had lower cell luciferase activity(P<0.05).Conclusions Over-expression of miR-216a may inhibit the proliferation,invasion,and migration of Hela cells.There is a targeted relationship between miR-216a and CAPN6 gene in HeLa cells.MiR-216a may inhibit the prolifera-tion,migration,and invasion of HeLa by regulating the expression of CAPN6 gene.
microRNAmicro RNA-216acalpain 6cell proliferationcell invasioncell migrationcervical cancer
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