Inhibitory effect of tetrandrine on the apoptosis of cultured human Tenon's capsule fibroblasts

Background: Excessive conjunctival scar formation is a main cause of filtering surgical failure in glaucoma. It has been reported that the failure rate of filtering surgery is a tough problem in the research of glaucoma. Research showed that tetrandrine (Tet) suppress the proliferation of cultured human fibroblasts of Tenons capsule (TCFs) in vitro, but its possible mechanism is still unclear up to now. Objective: The aim of this study is to investigate the inhibitory effects of Tet on the apoptosis of fibroblasts of Tenons capsule and mechanism. Methods: The Tenon's capsule was obtained from donor eyes of Zhongshan Eye Center Eye Bank for further use. Human Tenon's capsule fibroblasts were cultured by explant culture method in mixed medium in vitro and the third to seventh generations of cells were collected for the experiment. The subcultured cells were identified by morphology observation and Vimentin staining. The apoptosis of cultured fibroblasts in Tet-treated group and control group was studied by using TUNEL (TdT-mediated dUTP nick end labling, TUNEL), flow cytometry (FCM) and transmission electron microscope. The cell number in different cellular cycle was calculated in Ted-treated group and control group. Results: The cultured cells reached confluence in two weeks after cultured and presented the spindle or triangle shape with the radial-like or vortexin-like arrangement. The cells showed the positive staining for Vimentin. Apoptosis changes of cultured cell and apoptosis bodies were seen under the transmission electron microscope. Apoptotic cell nuclei were observed in Tet group by TUNEL. FCM result showed that the cells at G0/G1 phase decreased by 22.2%, and the cells at S and G2/M phases increased by 20.53% and 1.6% in Ted-treated group. Significant differences in the numbers of cells in different cellular cycles and cell numbers of apoptosis were found between Tet-treated group and control group (P=0.000). Conclusion: Tet can inhibit the proliferation of human TCFs by inducing apoptosis in vitro.