Effects of down-regulation of sphingosine kinase-1 expression on cell proliferation and cell apoptosis of cervical carcinoma Hela cells and its molecular mechanism

Abstract

Objective To study the down-regulation of sphingosine kinase 1 ( SphK1 ) expression on cell proliferation and cell apoptosis of cervical carcinoma cell line Hela cells,and to explore its possible molecular mechanisms.Methods SphK1 siRNA and control siRNA were utilized to transfect to Hela cells,which was divided into three groups including untransfected group,siRNA control group and SphK1 siRNA group.Expressions of SphK1 mRNA and protein were detected by read-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting methods after transfection with SphK1 siRNA.Cell counting Kit-8 (CCK-8) and Flow cytometry methods was used to analyze the changes of cell proliferation and cell apoptosis.Further,expressions of cell proliferation and cell apoptosis related proteins were detected by Western blotting method.Results Relative levels of SphK1 mRNA and protein in SphK1 siRNA group were 0.153 ±0.037 and 0.123 ±0.037,respectively,which was significantly lower than those in untransfected group ( 1.000 ±0.000 and 0.688 ±0.049 ) and siRNA control group (0.932 ±0.039 and 0.673 ±0.057 ) (P ＜0.05 ).The results of cell proliferation demonstrated that compared with untransfected group and siRNA control group,the proliferation of Hela cells in SphK1 siRNA group was obviously inhibited ( P ＜0.05 ).The results of Flow Cytometry revealed that early apoptotic rate of Hela cells in SphK1 siRNA group ( 23.85 ± 0.72)％ were markedly higher than those in untransfected group ( 11.67 ± 1.02 ) ％ and s iRN A control group ( 11.85 ± 0.71 ) ％,and there was significant difference ( F =211.451,P＜0.05 ).Further,the results of Western blotting demonstrated that expressions of p21,caspase-3 and caspase-9 proteins in SphK1 siRNA group (0.381±0.025,0.406 ±0.041 and 0.374 ±0.035,respectively) were obviously higher than those in nntransfected group (0.127 ±0.039,0.064 ±0.023 and 0.115 ±0.039,respectively) and siRNA control group (0.125 ±0.034,0.060±0.018 and 0.126 ±0.038,respectively)( P ＜0.05 ).Conclusion SphK1 may be closely associated with the development and occurrence of cervical carcinoma,and its expression down-regulation mediated proliferation inhibition and cell apoptosis may be tightly related to elevation of p21,caspase-3 and caspase-9 expressions. Key words: Sphingosine kinase-1; Hela cells; Small interfering RNA; Cell proliferation; Cell apoptosis