635nm激光致新西兰兔晶状体氧化损伤效应初步研究

Preliminary study on effects of oxidative damage to crystalline lens in New Zealand rabbits induced by 635 nm laser beam

目的探索波长为635 nm的激光对新西兰兔眼晶状体的氧化损伤效应。方法 1在体激光暴露实验。健康雌性新西兰兔5只,随机取2只兔为空白对照组(4只眼),另3只兔的右眼为暴露组(3只眼),左眼为自身对照组(3只眼);暴露组兔眼以照射功率为20 m W的635 nm激光连续照射2 h/d,每周6 d,共60 d,2个对照组兔眼均不进行激光暴露;实验结束后分离各组晶状体进行透明度检查,采用酶标仪测定晶状体超氧化物歧化酶(SOD)活力和丙二醛(MDA)、水溶性蛋白(WSF)、脲溶性蛋白(USF)的水平,计算WSF占总蛋白百分比(%)。2离体激光暴露实验。健康雌性新西兰兔4只,分离晶状体置于无菌M199培养基孵育后,以左眼为自身对照组(4只眼),右眼为暴露组(4只眼);暴露组晶状体以照射功率为20 m W的635 nm激光照射8 h/d,连续照射5 d;自身对照组晶状体不进行激光暴露。采用酶标仪测定晶状体SOD活力和MDA水平。结果在体激光暴露实验中,暴露组1只晶状体后囊部位出现轻微混浊,Geraldin分级为1级,其余各组晶状体均未发现浑浊现象,Geraldin分级为0级;暴露组、自身对照组和空白对照组3组晶状体的SOD活力和MDA水平分别比较,差异均无统计学意义[SOD:(1.803±0.542)vs(1.982±0.513)vs(1.247±0.235)mmol/(min·gprot),P>0.05;MDA:(0.078±0.017)vs(0.080±0.005)vs(0.077±0.006)μmol/gprot,P>0.05];3组晶状体WSF、USF的水平和WSF占总蛋白百分比分别比较,差异均无统计学意义[WSF:(113.31±14.70)vs(118.75±12.91)vs(114.05±14.41)g/L,P>0.05;USF:(3.23±0.22)vs(3.04±0.48)vs(2.52±0.50)g/L,P>0.05;WSF占总蛋白百分比:(97.33±0.30)%vs(97.39±0.25)%vs(97.79±0.70)%,P>0.05]。离体激光暴露实验中,暴露组晶状体SOD活力低于自身对照组[(0.144±0.004)vs(0.172±0.004)mmol/(min·gprot),P<0.05];2组晶状体MDA水平比较,差异无统计学意义[(0.031±0.002)vs(0.030±0.003)μmol/gprot,P>0.05]。结论 635 nm激光暴露可能会导致晶状体氧化损伤反应,具体机制有待进一步的研究。

Objective To explore the effects of oxidative damage in crystalline lens of New Zealand rabbits induced by 635 nm laser beam. Methods Female New Zealand rabbits were used in the present study, i） In vivo study. Five rabbits were used, 2 of them were randomly chosen as the blank control group （4 eyes） , on the other 3 rabbits, the right eyes （3 eyes） were regarded as the exposed group and left （3 eyes ） as the self-control group. The eyes in exposed group was directly exposed to 635 nm laser beam with irradiation power 20 mW, 2 hours a day for 60 continuous days. The eyes in control groups were treated like the exposed ones but without exposure to the laser beam. Crystalline lens was separated and their transparencies were detected. The levels of superoxide dismutase （SOD）, malondialdehyde （MDA）, water solublefraction （WSF） and urea soluble fraction （USF） of crystalline lens were detected by microplate spectrophotometer, and the percentage level of WSF in total protein were calculated, ii） In vitro study. The crystalline lens excised from 4 rabbits were cultured in medium 199, the left crystalline lens regarded as the self-control group and the right ones as the exposed group. The crystalline lens in exposed group were exposed to 635 nm laser beam with irradiation power 20 mW, 8 hours a day for 5 continuous days while the control group was not exposed to laser beam. The levels of SOD and MDA were detected by microplate spectrophotometer. Results i） In vivo study. One crystalline lens in the exposed group displayed slightly opacification in posterior capsule, the Geraldin grade 1 was recorded, whereas crystalline lens in other groups still kept clear, with the Geraldin grade 0. No statistical significant differences of crystalline lens SOD activity and MDA level among the exposed, self-control and blank control groups were found respectively [ SOD ： （ 1. 803± 0. 542 ） vs （ 1. 982 ± 0. 513 ） vs （ 1. 247± 0. 235 ） retool/（ rain · gprot） ,P 〉 0. 05 ; MDA ： （0. 078± 0. 017 ） vs （0. 080 ±0. 005 ） vs （0. 077± 0. 006 ） μmol/gprot, P 〉 0. 05 ] , while the same results were obtained in the levels of WSF, USF and the percentage levels of WSF in total protein respectively [ WSF： （ 113.31 ± 14.70） vs （ 118.75 ±12.91 ） vs （ 114.05 ±14.41 ） g/L, P 〉 0. 05 ; USF： （3.23±0. 22） vs （ 3.04±0. 48 ） vs （ 2. 52±0. 50） g/L, P 〉 0. 05 ; the percentage levels of WSF ： （ 97. 33±0. 30 ） % vs （97. 39±0. 25）% vs （97. 79±0. 70）%, P 〉0. 051 . ii） In vitro study. Crystalline lens SOD level in exposure group was significantly lower than that in the self-control group [ （ 0. 144±0. 004 ） vs （ 0. 172± 0. 004 ） retool/（ rain·gprot ）, P 〈 0. 05 ], but there was no statistical difference in the MDA level between the two groups [ （ 0. 031± 0. 002 ） vs （ 0. 030±0. 003） μmol/gprot, P 〉 0. 051].Conclusion It is possible that the oxidative damage of lens could be induced by the 635nm laser beam, and the concrete mechanism is needed to be studied.