[Effects of SMP-30 overexpression on apoptosis of human lens epithelial cells induced by ultraviolet B irradiation].

Objective: This study was to observe the effect of SMP-30 on ultraviolet B (UVB)-induced apoptosis of human lens epithelial cells(HLE-B3) in vitro. Methods: Experimental study. The SMP-30 cDNA was amplified by PCR and inserted into the pRFP-N1 expressing vector which had been double digested by XhoI/HindIII. HLE-B3 cells were cultured and divided into three groups: normal group, pRFP-N1 vector plasmid group and pRFP-N1-SMP-30 plasmid group (SMP-30). Then cells were exposed to UVB and the survival rate of cells was detected by MTT assay. The effects of SMP-30 on UVB-induced HLE-B3 apoptosis were measured by the Cell Death Detection ELISA kit. Meanwhile, the influence of SMP-30 on UVB-induced apoptosis-relative protein expression in HLE-B3 cells was tested by Western blots. Moreover, 2', 7'-Dichlorofluorescin diacetate staining was performed to monitor the protective effects of SMP-30 on UVB-induced HLE-B3 reactive oxygen species(ROS). One-way analysis of variance combined with Dunnett's statistical method were performed to analyze the data. Results: The full length of PSF cDNA fragment was correctly inserted into the pRFP-N1 vector, which was confirmed by DNA sequencing. The SMP-30 fragment was inserted to the plasmid pRFP-N1 correctly, which was also confirmed by DNA sequencing. The PRFP-N1-SMP-30 plasmid was transfected into HLE-B3 successfully. SMP-30 expression was up-regulated in the transfection group, compared with the control group. Data showed that the survival rate of HLE-B3 after the pRFP-N1-SMP-30 plasmid transfection was 0.90±0.14, while the apoptosis rate was 0.43±0.06 and the ROS production was 0.52±0.02, showing significant difference in comparison with the vector plasmid group and the normal group(t=5.830, 9.934, 12.19, P<0.05). In the meantime, SMP-30 overexpression down-regulated the levels of Bax and cleav-caspase-3, but up-regulated the Bcl-2 and Pro-caspase-3 expression levels under UVB irradiation. Conclusion: SMP-30 overexpression plays a protective role in UVB-induced apoptosis via regulating the expression of apoptosis-related proteins and inhibiting the production of ROS in HLE-B3 cells. (Chin J Ophthalmol, 2017, 53: 835-841).