ABSTRACT
Background The primary pathologic mechanism of posterior capsular opacification (PCO) is proliferation,migration and epithelial-mesenchymal transition (EMT) of residuary lens epithelial cells (LECs) after cataract extract surgery.Researches showed that MG132,a proteasome inhibitor,can attenuate the proliferation of bovine LECs,but its effect on human LECs remains unclear.Objective This study was to investigate the inhibitory effect of MG132 on proliferation,migration and differentiation of human LECs in vitro.Methods Human lens capsule were collected during the surgery.Human LECs were primarily cultured by explant method and passaged.The second or third generation of cells were incubated to 96-well plates at the density of 5×105/ml (200 μl/well) for 24 hours.Fibroblast growth factor-2 (FGF-2,10 mg/L),MG132 (10 μmol/L) or MG132+FGF-2 was added into the culture medium for 24 hours separately,and regular cultured cells served as the control group.The proliferation value (absorbance,A490) of the cells was assayed by MTT colorimetric method.A bare area was made by a sterile cotton swab in the cell layer,and migrated cell number in the blank zone was counted to evaluate the migration ability of the cells after 24 hours.Transforming growth factor-32(TGF-β2),MG132 or MG132+TGF-β2 was added into the culture medium for 24 hours separately,and the expression of fibronectin (FN) in the cells was detected using immunochemistry.Results The proliferation values (A490) of the cells were 0.582±0.020,0.723±0.010,0.434± 0.011 and 0.465±0.008 in the control group,FGF-2 group,MG132 group and MG132 + FGF-2 group,respectively,showing a significant difference among the groups (F =110.482,P＜0.01).The A value was significantly higher in the FGF-2 group and lower in the MG132 group and MG132+FGF-2 group than that of the control group (all at P＜ 0.05).The migrated cell number was 8.67 ± 1.08,11.58 ± 1.59,2.67 ± 0.09 and 2.75 ± 0.09 in the control group,FGF-2 group,MG132 group and MG132+FGF-2 group,respectively,with a significant difference among the groups (F=34.301,P＜0.01),and more cells in the blank zone were seen in the FGF-2 group and less cells were in the MG132 group and MG132+FGF-2 group in comparison with the control group (all at P＜0.05).Compared with the control group,the proliferative rate and migrating rate of the cells declined by 25.4％ and 75.0％ in the MG132 group as well as 20.1％ and 68.3％ in the MG132+FGF-2 group,but in the FGF-2 group,they increased by 24.2％ and 33.6％.The expressing levels (A value) of FN in the LECs were 1.242±0.023,2.329±0.113,1.043 ±0.021 and 1.163±0.018 in the control group,TGF-β2 group,MG132 group and MG132 +TGF-β2 group,respectively,with a significant difference among the groups (F =113.752,P＜0.01),a considerably increased expressing value was seen in the TGF-β2 group and decreased value was in the MG132 group and MG132+TGF-β2 group when compared with the control group (all at P＜0.05).Conclusions MG132 can effectively inhibit the proliferation,migration and differentiation of human LECs in vitro.