三种不同辐照度的460～480nm波长光对SD大鼠视网膜组织结构的影响

Influence of 460 -480 nm wavelength light at three different irradiance on retina tissue of SD rats

目的探讨健康成年sD大鼠视网膜对波长为460～480nm的蓝光照射的耐受性。方法实验研究。选取健康6周龄sD雄性大鼠76只，分为正常对照组（4只）和实验组（18组，每组4只）。选取460～480nm波长照射光，以辐照度0．6、1．5、10．0W／m2分别照射实验组动物3h和12h，于光照后4、24h和3d处死动物并取视网膜。通过HE染色、原位末端转移酶标记法染色、免疫荧光染色观察SD大鼠视网膜全层组织结构改变，并检测各层细胞凋亡情况及Mtiller细胞相关蛋白表达水平的变化。对不同取材时间各光照组视网膜神经节细胞数目进行单因素方差分析，各组问神经节细胞数日比较采用LSD-t检验分析。结果当光辐照度为0．6W／m2，光照时间为3h或12h时，大鼠视网膜各层组织结构均未见损伤。当光辐照度为10．0W／m2，光照时间为3h或12h时，大鼠视网膜各层均有损伤，以光感受器细胞及视网膜色素上皮细胞为主，同时内核细胞层Mtiller细胞亦有损伤，损伤主要表现为细胞凋亡，凋亡于光照后4h开始，24h达到最强。当光辐照度为1．5W／m2，光照3h时，大鼠视网膜各层组织结构未见损伤；光照12h时，大鼠视网膜各层可观察到轻微损伤改变，程度较光辐照度为10．0W／m2组明显减轻。无论光照时间为3h或12h，光照结束后3d取材时，各组神经节细胞数目组问差异均有统计学意义[0．0、0．6、1．5、10．0W／m2组光照时间为3h时视网膜神经节细胞数日分别为（41．42±0．17）、（40．58±0．50）、（40．92±0．32）、（22．83±0．79）个；F=1305．86，P=0．000；0．0、0．6、1．5、10．0W／m2组光照时间为12h时视网膜神经节细胞数目分别为（41．42±0．17）、（40．83±0．69）、（41．08±0．17）、（22．75±0．83）个；F=1095．78，P=0．0001。辐照度为10．0W／m2组的神经节细胞较正常组明显减少，差异有统计学意义（光照时间为3h时：t=52．32，P=0．000；光照时间为12h时：t=47．58，P=0．000）。结论波长为460～480nm蓝光，辐照度为0．6W／m2时，不引起SD大鼠视网膜组织结构损伤的安全照射时间为12h。辐照度升高至1．5W／m2时，安全照射时间缩短至3h。

Objective To investigate the survival of SD rat retinal cells after irradiated with 460 - 480 nnl wavelength blue light at three different irradiance. Methods Experimental study. Seventy-six SD rats six weeks old were divided into control group （4 animals ） and experimental groups （ 18 different subgroups） with 4 animals for each group. Healthy SD rats were exposed to the 460 -480 nm blue light for 3 or 12 hours on three different irradiance of 0. 6, 1.5 and 10. 0 W/m2 respectively. After recovery in darkness for 4 hours, 24 hours or 3 days, retinal tissue was collected after the eyes were enucleated. Hematoxylin-eosin staining was performed to evaluate the general anatomical changes of the rat retinal layers. TdT-mediated dUTP nick-end labeling was performed to detect apoptotic cells and immunofluorescence staining was conducted to illustrate the expression and location of related proteins in Mtiller cells. The number of ganglion cells was analyzed by one-way ANOVA. Results No injmy was observed in SD rat retina at the irradiance of 0. 6 W/m2. Retinal cells were damaged with irradiauce at 10. 0 W/m2 , which mainly affected photoreceptors and retinal pigment epithelial cells, while Mtiller ceils of retinal inner nuclear layer were also involved. Apoptosis began after 4 hours of exposure and peaked at 24 hours. Moderate damage of retina was observed at the irradiance of 1.5 W/m2 after 12 hours rather than 3 hours exposure. The number of ganglion cells was significantly different among the different groups 3 days after exposure. When the exposure time was 3 hours, the number of retinal ganglion cells under different irradiance at 0. 0, 0. 6, 1.5 and 10. 0 W/m2 was respectively 41.42 ±0. 17, 40. 58 ±0. 50, 40. 92 ＋0. 32 and 22. 83 ± 0. 79 （F = 1305.86,P = 0. 000）. When the exposure time was 12 hours, the number of retinal ganglion cells under different irradiance at 0. 0, 0. 6, 1.5 and 10. 0 W/m2 was respectively 41.42 ±0. 17, 40. 83 ±0. 69, 41.08 ±0. 17 and 22. 75 ± 0. 83 （ F = 1095.78, P = 0. 000）. The number of retinal ganglion cells showed significant decrease 3 days after exposure at the ilTadiance 10. 0 W/hi2 （ exposure time 3 h： t = 52. 32,P =0. 000;exposure time 12 h：t =47. 58,P =0. 000）. Conclusions The safe exposure time to SD rat retina layers is 12 hours for the blue light with wavelength 460 - 480 nm at the in＇adiance 0. 6 W/m2. And the safe exposure time descends to 3 hours at the irradiance 1.5 W/m2.