The Preliminary Study On UVR-induced Rat Corneal Epithelial Cell Apoptosis

Cornea is extremely sensitive to UV radiation (UVB). Suffering from UVR for a long time will lead to damage and even apoptosis of corneal cells, and then results in kinds of pathological changes in cornea. By means of operation on standard weight female Wistar rat, and cultured with corneal epithelium, we obtained primary rat corneal epithelial cells (RCE cells). By staining with Wright/Giemsa, neutral red, HO33258and DAPI to observe cell morphologies, DAPI-staining and MTT method to draw the growth curve of cells, Western Blot to analyse expression changes of caspase-3, flow cytometry to detect cell apoptosis. These results shew that, normal RCE cells were polygonal after being adherent, the pseudopodiums were long and wide, a few nucleoli could be found in stained cells. The effects of UVB to RCE cells were time and intensity dependent. When the radiation time was more than90min (108mJ/cm2), the pseudopodiums drew back, cytoplasm shrank, while the time exceeded150min (180mJ/cm2), patches of cells floated up and entered into death. In apoptotic cells, the agglutinated chromatin fractured into several big fragments, the nuclear membranes appeared gaps and disappeared gradually. The growth curve of normal cells was "S"-like. The cells entered into logarithmic phase after being cultured for about40h, and reached a plateau after about70h. The number of irradiated cells reached minimum after being recultured for about30-40h. The expression of caspase-3was induced to increase by UVB radiation, the120min was the highest, while the60min and180min were low, and the Oh of recultured after irradiation was supreme, the48h was minimum. The number of early apoptotic cells increased along with radiation time, while later apoptotic cells and necrotic cells were few.All the results suggest that, UVB radiation could lead to RCE cells apoptosis. When the intensity is20μW/cm2, the effects of UVB radiation to RCE cells is time dependent. The time range of UVB radiation that induces RCE cells apoptosis is90-150min (108-180mJ/cm2). Additionally, the apoptotic response of RCE cells to UVB radiation is associated with the proliferation ability and cell density.