Effect of tetrandrine eye drops on expression of apoptotic protein and transforming growth factor-β2 in human Tenon capsule fibroblast in vitro

Abstract

Background Our previous study determined that tetrandrine (Tet) has an inhibitory effect on the proliferation of human Tenon capsule fibroblasts ( TCFs ) in vitro,but its mechanism is poorly understood.Objective This study was to investigate the effect mechanism of Tet on human TCFs. Methods Human TCFs were isolated and cultured from scleral tissue of donor using explant technique.The cells were identified by vimentin antibody staining and morphology.The third generation of cells were seeded in the culture plate at the density of 1 × 105 cells/ml.Twenty-four hours after inoculation,the Tet of 1 × 10-5 mol/L was added in the well of culture plate,and the cells cultured only in 1640 medium served as the control group.The apoptosis of the cells was assessed by TUNEL,and the expressions of bax,bel-2,transforming growth factor-β2 (TGF-β2 ) in TCFs were detected using immunochemistry. Results The cultured cells showed the features of the fibroblasts in shape with the positive response for vimentin.A number of TUNEL positive cells were seen in Tet group and no TUNEL positive response was found in control group.The expression levels (A value) of bax,bcl-2 and TGF-~ protein in TCFs were 0.577 ± 0.009,0.430±0.012 and 0.341 ±0.017 in Tet group,and those in control group were 0.320±0.015,0.819±0.021 and 0.624±0.014 respectively,showing statistically significant differences between two groups( t =33.277,-35.356,-28.093,P＜0.01 ). Conclusions Tet suppresses the proliferation of human TCFs through up-regulating the expression of bax and down-regulating the expressions of bcl-2 and TGF-β2 in vitro. Key words: Tetrandrine; Tenon capsule fibroblast; Apoptosis-related protein/bax, bcl-2; Tansforming growth factor β2 protein