Experimental Research of Apoptosis in Pterygium Fibroblasts Induced by 5F and Effect on the Expression of Caspase-3

OBJECTIVE To observe the induced apoptosis effect of 5F on HPF cultured in vitro and the expression of caspase-3.METHODS Cell were treated with different concentration of 5F(0,8,32 and 128 mg·L~(-1))with different time.The effect of 5F on cell proliferation was detected by thiazole blue chromatomentry.Cell apoptosis was observed by Hoechst33258 fluorescent staining;flow cell cycle and apoptosis were investigated by flow cytometry.The transcription level of caspase-3 were determined by RT-PCR;The expression of caspase-3 protein was detected by Western blot;The change of caspase-3′s activity during the process of cell apoptosis was valued by chemistry chromatometry.RESULTS Compared with the control group,the cell growth in experimental group was inhibited significantly(P0.01),and the cell proliferation inhibition rate showed a dose-and time-dependent manner.Hoechst33258 fluorescent staining displayed that cells in experimental group partly showed typical apoptosis.Under the effect of 5F with the concentration of 32.18 mg·L~(-1),the peak of apoptosis was reached at 12 h and was(12.48±1.29)%;RT-PCR showed that the expression of caspase-3 mRNA in cells was increased.Western blot showed that the expression of caspase-3 protein was increased.The caspase-3 activity was increased dose-and time-dependerthy.The activity was 3.52 times as control group when the concentration of 5F was 128 mg·L~(-1).CONCLUSION 5F can inhibit the proliferation of PHF,induce cell apoptosis,increases the activity of caspase-3,and show a dose-and time-dependent manner.The mechanism of 5F inducing cell apoptosis may relate with the increasing of caspase-3 activity,facilitating of transcription of caspase-3 and protein translation,increasing of casepase-3 in order to accelerate the cell apoptosis.