Background Plasma membrane calcium ATPase 3 (PMCA3) participates in the regulation of Ca2+ level in lens epithelial cells (LECs),which may be associated with the pathogenesis of cataract.It has been proved that ultraviolet B (UVB) is one of causing-factors of cataract.However,the effect of UVB on the expression of PMCA3 in LECs is unclear.Objective This study was to investigate the effects of UVB irradiation on the expression of PMCA3 in human LECs B-3.Methods HLE B-3 cells were cultured and passaged.The cells were exposed to 0,5,10 and 20 mJ · s/cm2 UVB for 0,20,40,80 s,respectively and further cultured for 24,48 and 72 hours.MTT assay was used to detect the cell proliferation rate.JC-1 staining was used to detect the changes of mitochondrial membrane potential (△ψm) in the cells.The intracellular reactive oxygen species (ROS) level was detected by DCFH-DA staining,and cell apoptosis was evaluated using annexin V-FITC/PI staining.In addition,the intracellular calcium ion (Ca2+) concentration in the cells was assayed with Fluo-3/AM staining.The expression levels of PMCA3 mRNA and PMCA protein in the HLE B-3 cells were detected by real-time PCR and Western blot,respectively.Results The survival rates of the cells were significantly reduced with the increase of irradiated intensity of UVB and the lapse of time (Fgroup =72.411,P =0.000 ; Ftime =36.588,P =0.000),and the survival rates of the cells in the 10 mJ · s/cm2 and 20 mJ · s/cm2 UVB for 24 hours were (75.3 ± 2.2) ％ and (48.7 ±4.5) ％,respectively,which were significantly lower than those in the 0 mJ · s/cm2 UVB group ([100.0±0.0] ％) (P=0.001,0.000).The survival rates of the cells in the 5,10,20 mJ · s/cm2 UVB for 48 hours were (84.9± 1.2) ％,(69.3±17.4)％ and (32.8±4.5)％,showing significant declines in comparison with the 0 mJ · s/cm2 UVB group ([100.0±0.0] ％) (P =0.047,0.000,0.000).In 72 hours following 5,10,20 mJ · s/cm2 UVB irradiation,the survival rates of the cells were (55.1 ± 3.0) ％,(42.1 ± 1.9) ％ and (26.1 ±4.7) ％,respectively,with significant differences in comparison with the 0 mJ · s/cm2 UVB group ([100.0 ± 0.0] ％) (P =0.000,0.000,0.000).JC-1staining exhibited the intracellular red fluorescence in the normal cells group.However,in the 5 mJ · s/cm2 UVB group,weak green fluorescence was seen,and the green fluorescence was enhanced in the 10 mJ · s/cm2 and 20 mJ · s/cm2 UVB groups.After irradiated by 5,10 and 20 mJ · s/cm2 UVB,the ROS levels in the cells increased from 0.4％ to 35.8％,51.9％ and 76.7％,respectively.The apoptosis and necrosis rate of the cells was 2.0％ in the 0 mJ · s/cm2 UVB and 4.2％,7.6％,15.1％ in the 5,10,20 mJ · s/cm2 UVB groups,respectively.The Ca2+ level raised by (1.2±0.1) and (1.3±0.1) folds in the 10 and 20 mJ · s/cm2 UVB groups more than that in the 0 mJ · s/cm2 group (P =0.039,0.004).The expression levels of PMCA3 mRNA in the cells were significantly reduced (P=0.001,0.004,0.000),and the expression levels of the PMCA protein were declined in the 5,10 and 20 mJ · s/cm2 UVB groups compared with the 0 mJ · s/cm2 UVB group (P=0.000,0.000,0.001).Conclusions UVB irradiation causes cataract probably through downregulating the expression of PMCA3 in human LECs and inducing apoptosis of LECs in a dose-and time-dependent manner.

AIM: To investigate the effect of electroacupuncture on the expression of matrix metalloproteinase-3(MMP-3), tissue inhibitor of metalloproteinase-3(TIMP-3)and collagen type III alpha 1(Col3α1)in the retina of guinea pigs with lens-induced myopia.METHODS:A total of 80 guinea pigs were randomly divided into the normal control(NC)group, negative lens-induced myopia(LIM)group, electro-acupuncture(EA)group, and sham group, with 20 guinea pigs in each group. The NC group did not receive any intervention, the LIM, EA and the sham groups were all wearing -6.0 D lens in the right eye and no lens in the left eye. The EA group was given electroacupuncture stimulation at Hegu Point and temple, and guinea pigs in the sham group were given intervention at false points. The refraction was measured by optometry, the axial length was measured by A-ultrasound, and the changes in retinal tissue structure were observed by HE staining. Quantitative polymerase chain reaction(Q-PCR)and western blot(WB)were used to detect the expression of MMP-3, TIMP-3, and Col3α1 at mRNA and protein levels before modeling and at 2 and 4 wk after modeling. RESULTS:The axial length of the negative LIM group was significantly increased compared with the NC group at 2 and 4 wk after modeling(all P<0.05), and the diopter was significantly decreased(all P<0.05); the axial length of the eye in the EA group was decreased compared with the negative LIM group(all P<0.05), and the diopter was increased(all P<0.05). HE staining showed that the boundaries of retinal tissue in the NC group were obvious and arranged regularly. The retinal thickness, the thickness of the inner and outer nuclear layers, and the number of cells were reduced and irregularly arranged in the negative LIM group. The overall structure of the retina in the group EA was relatively perfect, the arrangement was more regular, and the morphological structure of each layer of the tissue did not appear obvious abnormalities. Q-PCR and WB detection results showed that the mRNA and protein relative expression levels of MMP-3, TIMP-3, and Col3α1 in the negative LIM group were significantly higher than those in the NC group(all P<0.05); the expression levels of the EA group was significantly decreased after intervention compared with the negative LIM group(all P<0.05).CONCLUSION: Electroacupuncture can delay axial growth in negative LIM guinea pigs, and downregulate negative LIM induced expression of MMP-3, TIMP-3, and Col3α1 in the retina.