Nd:YAG激光致视网膜感光细胞损伤及相关基因的表达

Injury Induced by Low Dose Nd:YAG Laser and Expression of Related Genes in Photoreceptor Cells of Rabbit Retina

目的:观察兔眼视网膜激光损伤后不同时间点感光细胞的改变以及相关基因的表达变化。方法:北京青紫蓝灰兔随机分为正常对照组和激光损伤组,在调Q倍频Nd:YAG激光损伤后不同时间采用超微结构观察、凝胶电泳、TUNEL染色、原位杂交(ISH)以及Northernblot等方法检测了感光细胞损伤特点并分析相关基因的表达改变。结果:电镜、核酸电泳和TUNEL结果表明视网膜感光细胞照射后出现明显的凋亡特征,在伤后不同时间,其分布和强度都有改变。而杂交结果表明c-fos和bax在照后表达增强,尤其以照后第1天为最,而bcl-2和p53在损伤后无明显改变。结论:大量感光细胞的凋亡是临床上低剂量倍频Nd:YAG激光致视网膜损伤、视功能下降的主要机制。c-fos和bax基因的表达可能介导了视网膜细胞凋亡的过程。

Objective:While widely used in clinical medicine,laser can also cause accidental eye injury.This paper is to study the pathologic mechanism of retinal injury induced by low dose laser irradiation and to observe the variation of related genes expression associated with this process.Methods:The rabbits were divided randomly into injury group and normal group.The retinal injury model was introduced with the double frequency Nd:YAG laser(λ=532nm) for 100 plus per eye.The animals were killed at different time intervals after laser exposure and the retinas were excised for further investigation.The examination of histopathology,TdTmediated dUTP nick and labeling(TUNEL),electrophoresis were applied to detect apoptosis of photoreceptors.In situ hybridization(ISH) and Northern blot were performed to determine the related genes.Results:Morphological and biochemical analysis show that during 6h,1d and 3d after laser exposure,rabbits retinas demonstrated a spectrum of apoptotic changes.The TUNELpositive photorecepor nuclei were present in all the retina layers ,mainly in the out nuclear layer(ONL).Neutral gel electrophoresis showed DNA damage in the retinas following laser injury and exhibited a timerelated increase in the pattern of 180 bp DNA fragments indicative of endonuclease activity.In situ hybridization histochemistry(ISH) revealed cfos,bax genes expression were upregulated and that was further confirmed by Northern blot,while bcl2 and p53 genes showed no alteration.Conclusions:These results suggest that apoptosis of photoreceptor cells is an important mechanism in retinal laser injury when even there is no macroscopic damage.and the two genes cfos,bax seem to be involved in the signaling pathways of low dose laserinduced photoreceptor cells apoptosis and might be the potential target genes for therapy while p53 and bcl2 have little relevancy to this process.