Authors: Guo Chenjun, Guo Yong, Wang Song, Zhang Jie, Yan Hong
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To observe the damage of human lens epithelial cells (LECs) induced by ultraviolet (UV)B radiation and the expression changes of glutaredoxin 2 (Grx2) in the cells, and to investigate the protective effects of Grx2 on human LECs against UVB-induced apoptosis.
Human LECs (HLE-B3) were cultured and exposed to different energy of UVB (0, 10, 30, 50 mJ/cm2) with the wavelength of 297 nm.The morphology of the cells was examined under the optical microscope 2, 4, 8, 12 and 16 hours after irradiation of UVB.The survival rate of the cells was evaluated with cell counting kit 8 (CCK8). The apoptosis rate of the cells was detected by TUNEL assay.Real-time quantitative PCR and Western blot were employed to detect the expressions of Grx2 mRNA and Grx2 protein in the cells, respectively.The cells were transfected with pcDNA3.1-Grx2 plasmid by lipofectamine 2000 as the Grx2 transfected group, and pcDNA3.1 plasmid was transfected into cells as the empty plasmid group.The cells were irradiated by 50 mJ/cm2 for 4 hours, and the apoptosis rate of human LECs was detected by TUNEL assay.
Cultured cells grew well with the green fluorescence for Grx2 expression in the non-UVB exposure group, and shrinkage and death of the cells were found after UVB irradiation.The survival rate of the cells was gradually reduced after irradiation of 10, 30, 50 mJ/cm2 UVB as the increase of UVB energy and lapse of time.The expression level of Grx2 mRNA were 2.53±0.48 and 3.53±0.14 in the 10 mJ/cm2 UVB group and 30 mJ/cm2 UVB group 4 hours after irradiation, which were significantly higher than 1.01±0.08 and 1.00±0.09 in the non-UVB exposure group (all at P<0.05). The expression level of Grx2 mRNA in the cells was 15.30±3.01 at 1 hour after irradiation in the 50 mJ/cm2 UVB group, which was significantly higher than 1.00±0.07 in the non-UVB exposure group (P<0.05). The expression levels of Grx2 protein showed the same tendency to Grx2 mRNA.The apoptosis rate of the cells in the Grx2 transfected group was (15.34±1.71)%, and that in the empty plasmid group was (22.11±2.46)% at 4 hours after 50 mJ/cm2 UVB irradiation, with a significant difference between them (t=3.189, P<0.05).
UVB irradiation induced damage of human LECs in dose- and time-dependent manner.However, the expression of Grx2 in human LECs is up-regulated transiently after exposure of UVB, which has a protective effect on the cells against the UVB-induced apoptosis.