Antiproliferative effect of curcumin on human retinal pigment epithelial cell / 中华实验眼科杂志

ABSTRACT

Background Curcumin derives from the rhizome of curcuma longa.It has proven to have an antiproliferative effect in previous studies on vast majority of endothelial and epithelial cells,however,the study of its inhibiting effect on the proliferation of retinal pigment epithelial (RPE) cells and underlying mechanism is rare.Objective Aim of this study was to investigate the potential inhibitory effect of curcumin on the proliferation of cuhured human RPE cells in vitro and its possible mechanism.Methods Human RPE cells harvested by trpsinEDTA were suspended in DMEM/F12 medium with serial dilutions of curcumin (5,10,15,20 mg/L),and the human RPE cells cultured by DMEM/F12 without curcumin were used as control.The proliferation value of human RPE cells (A value) was measured by water-soluble tetrazole-1 (WST-1) assay,the optimized dose of antiproliferation of curcumin was determined and applied for further experimental process.Apoptosis and cell cycle of human RPE cells were detected by flow cytometric analysis at 48 hours and 72 hours after curcumin treatment.The ultrastructure profile of the cells were examined by transmission electron microscopy (TEM).Western blot analysis was performed to measure the relative expressing level of the pro-apoptotic factors p53,p21 WAF1/CIP1 and proliferating cell nuclear antigen (PCNA) in the cells,respectively.Factorial design of two factor analysis of variance of SPSS 17.0 software was used to compare the difference of A values of the cells among the various groups and time points,and independent-sample t test was used to compare the differences of apoptosis rate and cell ratio in different cycles between curcumin group and control group.Results WST-1 assay showed that the A value was gradually reduced with the increase of curcumin dose (F tion =96.55,P =0.00),and gradually increased with the lapse of time (Ftime =4634.28,P =0.00).The early apoptotic rate of the cells was (13.37±1.26) ％ in the curcumin group 48 hours after treated by 15 mg/L curcumin,and that of the control group was (7.03 ±0.37) ％,with a significant difference between them (t =8.33,P=0.00).In 72 hours after treated by 15 mg/L curcumin,the early and middle-late apoptotic rates of the cells were (15.97±0.16) ％ and (0.26±0.03) ％,which were significantly higher than those of the control group (7.29±0.37) ％ and (0.14±0.02) ％ (t=37.80,P=0.00;t=7.44,P=0.00).The cell ratio of G0/G1 phase in the curcumin group was (57.17±1.17)％ 48 hours after treated by 15 mg/L curcumin,and that in the control group was (67.73± 1.10)％,showing a significant difference (t =11.40,P =0.00).M itochondrial swelling and vacuolar degeneration were seen in the cells after treated by 15 mg/L curcumin.The relative expression levels of p53 and p21WAF1/CIP1 protein in the cells were higher in the curcumin group than those of the control group at 24,48 and 72 hours (all at P＜0.05),but the expression levels of PCNA protein were lower in the curcumin group than those of the control group in various time points (all at P ＜ 0.05).Conclusions Curcumin can effectively inhibit the proliferation of human pigment epithelial cells in a dose-and time-dependent manner.P53 pathway may participate in anti-proliferating process.