Experiment research in the regulation of Maspin gene promoter methylation targeted by short hairpin RNAs on HN13 cells migration and invasion ability

Objective To observe the effect of Maspin gene promoter methylation targeted by short hairpin RNAs on HN13 cells migration and invasion ability. Methods Both specific (sequence 1,sequence 2,sequence 3,sequence 4) and nonspecific (sequence 0) shRNA sequences were designed in view of the Maspin gene promoter region methylation site,which were soon used to conduct recombinant adenovirus and transfected to HN13 cells,using no-load adenovirus as the controls.The transfection rate,cell growth curve and the degree of methylation were observed;At the same time,the mRNA and protein expression level were detected by RT-PCR and WB assay,the HN13 cells migration and invasion ability were detected by Transwell chamber experiment. Results The transfection efficiency of Maspin-shRNA recombinant adenovirus was above 95%.After the transfection,the cell growth rate of sequence 1,2,4 increased,which was obviously different to the controls,and their methylation levels were higher than the controls too (P<0.05).After 7 days'culture,the Maspin mRNA and protein expression level of sequence 0 were both higher than the controls,and the Maspin mRNA and protein expression level of sequence 1,2,4 were both lower than the controls,all the difference had statistical significance (P<0.05).The number of cell migration and invasion of sequence 0 were both lower than the controls,and the number of cell migration and invasion of sequence 1,2,4 were both higher than the controls,all the difference had statistical significance (P<0.05). Conclusion Maspin gene promoter methylation could promote the migration and invasion ability of HN13 cells.