Role of receptor tyrosine kinase Mer in regulating proliferation of rat Schwann cell line RSC96 cultured in a high-glucose environment
Objective To observe the regulatory role of receptor tyrosine kinase Mer(Mertk)in the proliferation of the rat Schwann cell line RSC96 under high glucose condition.Methods RSC96 cells were divided into groups one,two,three,four and five and were cultured in medium containing 25(normal glucose concentration),50,75,100 and 125 mmol/L glucose,respectively,and Mertk was detected in cells of each group by Western blotting at 48 h of incuba-tion,and the highest concentration of Mertk protein relative expression was selected to be the high-glucose concentration for the subsequent study.RSC96 cells were starved for 4 h,and were placed in the glucose medium containing 100 mmol/L glucose,and then were cultured for 0,24,36,48,and 60 h.Western blotting was used to detect the Mertk of cells in each group,and the time with the highest relative expression of Mertk protein was screened as the time of culture for the subsequent study.RSC96 cells were taken and divided into the control group,high glucose group,high glucose knock-down group and knockdown group:cells in the high glucose group were starved for 4 h and cultured in medium containing 100 mmol/L glucose;cells in the high glucose knockdown group were starved for 4 h,and were placed in medium contain-ing 100 mmol/L glucose,and then,5 µL of siRNA solution with knockdown of Mertk expression was added;cells in the knockdown group were starved for 4 h,and 5 µL of siRNA with knockdown of Mertk expression was added;cells in the control group were cultured in the normal medium.The cells in each group were taken at 48 h of culture,and the prolifera-tion of each group was measured by Edu method,and Mertk,phosphorylated nuclear transcription factor-κB(P65)and tu-mor necrosis factor-α(TNF-α)in each group were detected by Western blotting.Results Compared with the group one,the relative expression levels of Mertk protein were higher in cells of groups four and five(all P<0.05);the relative expres-sion of Mertk was higher in RSC96 cells at 48 and 60 h of culture in comparison with that at 0 of culture(all P<0.05).Compared with the control group,the proliferation activity of cells in the high glucose group was lower(P<0.05),and the proliferation activity of cells in the knockdown group was higher(P<0.05);compared with the control group,the relative expression levels of P65 and TNF-α of cells in the high glucose knockdown group were higher(both P<0.05),the relative expression of Mertk of cells in the knockdown group was lower,the relative expression levels of P65 and TNF-α of cells in the knockdown group were higher(all P<0.05),and the relative expression levels of Mertk,P65,and TNF-α of cells in the high glucose group were higher(all P<0.05).Conclusions RSC96 cells cultured in high glucose environment with knockdown of Mertk gene expression has high proliferative capacity and high cellular P65 and TNF-α expression.Mertk may promote the proliferation of RSC96 cells cultured in high glucose environment by promoting cellular P65 and TNF-α expression.
万方数据
维普
