视网膜光辐照实验模型研究进展

随着电子设备的普及与环境光污染现象的日趋严重, 视网膜光辐照引起的损伤及其对视网膜色素变性、年龄相关性黄斑变性等疾病的致病作用逐渐被关注。光-视网膜组织反应类型以光热反应及光化学反应为主, 其中可见光产生的光化学反应与视网膜疾病关系密切。光辐照参数包括光辐照波长、辐照能量及辐照时间等, 不同参数下光辐照对视网膜细胞的作用还可能受多种外界因素影响。较多视网膜光辐照细胞实验模型建立以研究光辐照, 尤其是蓝光对视网膜色素上皮细胞、光感受器细胞及神经细胞损伤的机制。光辐照会导致视网膜各细胞氧化应激、内质网应激及线粒体损伤激活自噬调控细胞凋亡。炎性小体激活及外泌体也参与调控光辐照对视网膜色素上皮细胞的损伤。也有研究探讨不同波长光源对细胞的潜在治疗作用。本文从光-视网膜组织反应类型、生物学研究中常用的光辐照参数, 以及视网膜色素上皮细胞光辐照模型、视网膜光感受器细胞光辐照模型、视网膜神经节细胞光辐照模型等方面就视网膜光辐照的实验模型, 探讨不同模型间细胞及光辐照参数的差异。     

氩绿激光致兔视网膜感光细胞凋亡的实验研究

目的:观察氩绿激光对兔视网膜损伤及修复的组织学改变及对兔视网膜感光细胞凋亡的影响。方法:将8只有色兔(16眼)中每只眼的上、下方视网膜随机分配为激光光凝区及空白对照区,光凝后24h、4wk通过光镜和电镜及TUNEL技术观察视网膜改变及感光细胞凋亡。结果:(1)光镜观察:光凝处视网膜损伤不明显但脉络膜小静脉充血,4wk后消退。(2)电镜观察:光凝处外节膜盘排列稀疏,神经节细胞轻微肿胀,外核层细胞异染色质增多,4wk后色素增殖,胶原增生。(3)TUNEL染色观察:光凝后24h,光凝处外颗粒层可见较多阳性细胞,内颗粒层偶见,4wk后接近正常。(4)感光细胞凋亡率:光凝后24h,感光细胞凋亡率较正常对照组增加,差异有统计学意义(P<0.01)。光凝后4wk较24h凋亡率显著降低,差异有统计学意义(P<0.01)。结论:轻度氩绿激光光凝视网膜损伤轻微。     

光损伤导致视网膜感光细胞凋亡的实验研究

目的探讨视网膜光损伤后感光细胞病理学改变的特征及其发生机制。方法以白色强光持续照射的方法制成大鼠视网膜光损伤模型并采用常规HE染色与TUNEL技术对光损伤后视网膜感光细胞的病理学改变进行动态观察研究。结果白色强光照射后视网膜感光细胞发生进行性的变性,TUNEL标记结果显示光损伤后视网膜外核层出现大量阳性着色细胞。结论持续高强度白光照射可选择性地导致视网膜感光细胞发生进行性的变性而凋亡是感光细胞退行性变性的重要发生机制。     

缺血再灌注损伤诱导大鼠视网膜细胞凋亡

目的 观察缺血再灌注大鼠视网膜损伤及细胞凋亡情况。 方法 采用升高大鼠眼压到109.725 mm Hg(1 mm Hg=0.133 kPa)持续1 h的方法制作视网膜缺血再灌注模型，采用常规眼球切片观察不同缺血和再灌注时间的视网膜损伤的组织病理改变；采用DNA琼脂糖凝胶电泳法检测视网膜神经元凋亡情况；采用DNA原位末端标记(terminal dUTP nick end labelling, TUNEL)法定位凋亡的视网膜细胞。 结果 缺血30 min 再灌注24、48 h的大鼠视网膜无明显的病理改变；缺血60 min再灌注24、48 h的大鼠视网膜神经节细胞层和内核层细胞明显变薄；缺血60 min再灌注12、24 h的大鼠视网膜有梯状条带。而正常对照组、缺血30 min再灌注24、48 h组及缺血60 min再灌注48 h组大鼠视网膜均无类似表现。TUNEL法显示视网膜内的细胞凋亡主要发生在节细胞和内核层光感受细胞。 结论 大鼠视网膜缺血再灌注主要是导致视网膜神经节细胞层和内核层细胞损伤，细胞凋亡可能是损伤的重要机制。 (中华眼底病杂志, 2002, 18: 296-298)     

光照对大鼠视网膜的影响及其机制研究

目的 探究光损伤对大鼠视网膜结构和功能的影响及其机制。方法 自制光照箱。取健康成年SD(Sprague-Dawley)雄性大鼠,随机分为正常组和实验组。正常对照组大鼠饲养在12 h明12 h暗的正常环境中;实验组大鼠首先暗适应24 h,后采用5 000 lx照度的可见光照射48 h,再暗适应24 h。麻醉大鼠,时间约30 min。首先,应用视网膜电生理(ERG)检测大鼠视网膜功能;其次,取出大鼠眼球后放入眼球固定液或4%多聚甲醛,制作石蜡切片行苏木精-伊红(HE)染色观察视网膜组织结构变化,冰冻切片行TUNEL染色,分别应用光镜和共聚焦显微镜观察。实时荧光定量反转录聚合酶链反应(qRT-PCR)及蛋白质印迹法(WB)检测光损伤后视网膜肿瘤坏死因子-α (TNF-α)含量。结果 HE染色显示光照后视网膜外核层较正常明显变薄,颞侧较鼻侧视网膜损伤较重;TUNEL染色显示光损伤后视网膜凋亡细胞数量明显增多;qRT-PCR及WB显示光损伤后视网膜组织表达TNF-α增多;ERG显示光损伤大鼠视网膜功能明显降低。以上结果差异均有统计学意义(P<0.05)。结论 光照后视网膜外核层损伤是导致视...     

视网膜光损伤的防治研究进展

光对视网膜的损伤能诱发活性氧自由基产生,使视网膜细胞处于氧化应激状态,从而造成细胞一系列损伤、凋亡、生物膜溶解和细胞坏死,导致感光细胞的凋亡和视网膜变性,引发眼病,甚至导致视力丧失。目前国内外对视网膜光损伤的防治有一些研究和报道。我们就从视网膜光损伤的机制和多方面的防治进行归纳研究,现综述如下。     

视网膜光损伤的药物防治

视网膜光化学损伤可致感光细胞凋亡和视网膜变性,严重导致视力丧失。近10余年来,人们对药物防治光损伤进行了大量研究,包括神经营养因子、抗氧化剂、自由基清除剂、钙通道阻滞剂及皮质激素等。本对各种药物的保护作用作一综述。     

地塞米松对光损伤大鼠视细胞凋亡的防治作用

目的 :研究地塞米松对光损伤及视细胞凋亡的防治作用 ,以探讨光损伤视细胞凋亡的发生机制。方法 :所有SD大鼠经循环光环境适应 7d ,实验前暗适应 36h。实验A组的大鼠只光照 ,实验B组的大鼠光照 6h后在暗箱中喂养 ,实验a组的大鼠在暗适应后腹腔注射地塞米松再光照 ,实验b组的大鼠光照 6h后在暗箱中喂养 ,且每天应用地塞米松。对经以上处理过的大鼠行灌流固定 ,摘除眼球。光镜标本在常规脱水、透明、石蜡包埋切片后 ,行HE、TUNEL法染色 ,光镜观察。应用CIAS 10 0 0图像分析系统定量检测外核层面积和视细胞凋亡指数 ,所得数据做统计学分析。结果 :在实验A组中 ,随着光照时间的增加 ,视网膜光损伤逐渐加重 ,视细胞凋亡逐渐增多 ,外核层面积逐渐减少。而在实验a组中 ,也出现如实验A组的规律性变化 ,但应用地塞米松后 ,视网膜光损伤程度减轻 ,发生视细胞凋亡的时间延迟 3h。两实验组定量检测结果经统计学分析表明 ,地塞米松对视网膜光损伤及视细胞凋亡有明显的预防作用。实验B组和实验b组的定量检测结果经统计学分析表明 ,地塞米松对视网膜光损伤及视细胞凋亡有明显的治疗作用。结论 :地塞米松在实验性大鼠视网膜光损伤及视细胞凋亡过程中发挥较好的防治作用     

RPE细胞发生上皮间充质转化机制及PVR的治疗研究进展

增殖性玻璃体视网膜病变(proliferative vitreoretinopathy,PVR)是眼外伤、糖尿病性视网膜病变、血管性视网膜病变和炎症性视网膜病变等眼部疾病的严重并发症,也是孔源性视网膜脱离手术失败的最重要原因,对视功能的危害较大。大量研究已证明PVR发生的主要危险因素是视网膜损伤后血视网膜屏障受损,视网膜色素上皮(retinal pigment epithelial,RPE)细胞受到玻璃体腔内细胞因子的刺激,RPE细胞发生上皮间充质转化(epithelial-mesenchymal transition,EMT),转分化为成纤维样细胞,细胞的形态发生了变化,细胞间的紧密连接消失,细胞极性丧失,增殖、迁移、侵袭能力增强。在视网膜前表面或视网膜下形成具有收缩性的纤维增殖膜,形成的纤维增殖膜会使视网膜形成皱褶,牵拉视网膜导致视网膜脱离,最终会导致患者视力下降甚至失明。国内外对预防和治疗PVR进行了大量的研究,本文对RPE细胞发生上皮间充质转化相关信号通路及PVR的治疗进行简要综述。     

兔视网膜光凝术后组织形态学改变的动态观察

目的:研究532nm激光TsoⅢ级光斑光凝色素家兔视网膜后光凝斑组织形态改变的病理特征。方法:(1)18只色素家兔按光凝后观察时间1,3,7,14,21d及28d分为6组,每组3只。每只(双眼)用532nm激光TsoⅢ级光斑光凝色素家兔视网膜后极部30点(532nm,450mW,100μm,0.05s)。(2)光凝后不同观察时间点行荧光素眼底血管造影(fluorescence fundus angiography,FFA)观察光凝斑FFA特点;取光凝区眼球壁制作标本,进行光镜和透射电镜观察并定量分析视网膜光凝斑大小、视网膜细胞存活率的动态变化。结果:(1)荧光血管造影:光凝后3d,部分光凝斑为高荧光,荧光渗漏随时间延长逐步减轻,至光凝后14d,荧光渗漏基本消失。(2)组织病理学检查:光凝斑区域视网膜各层组织细胞结构破坏;光凝斑周围光感受器细胞的凋亡或坏死和神经节细胞的损伤;继后色素上皮细胞、M櫣ller细胞和成纤维细胞增生修复破坏区。(3)形态学定量指标:光凝后第1d视网膜光凝斑直径最大为116.4±9.6μm,比参照光斑(75μm)增大了55.2%,随后有下降的趋势,21d后为82.8±5.4μm趋于稳定。视网膜细胞存活率光凝后第1d最小为(29.5±4.2)%,随后有上升的趋势,21d为(48.2±4.4)%趋于稳定。结论:视网膜光凝TsoⅢ级光斑会导致光凝斑周围视网膜感觉神经细胞的非选择性、扩展性损伤。     

Failure of ascorbate to protect against broadband blue light-induced retinal damage in rat

· Background: Excessive generation of free radicals due to light absorption is proposed as the most likely mechanism for photochemical retinal damage. The observed reduction of green light-induced retinal injury after ascorbate treatment is believed to be an antioxidative effect. The aim of the present study was to evaluate the possible protection of ascorbate against blue light-induced photoreceptor damage. · Methods: Cyclic light-reared albino rats were injected intraperitoneally with either ascorbate (1 mg/g body weight) or, as placebo, physiological saline 24 h before and just prior to exposure to blue light. After 20–22 h of dark adaptation, two groups of the rats were exposed in pairs to the blue light (400–480 nm) for 6 h at an average irradiance of 0.7 W/m² in the cage. Six days after light exposure, all rats were killed and retinal samples were analyzed. · Results: Diffuse blue light irradiation resulted in an uneven distribution of damage in the retina. As judged from the pathological changes in the retina irradiated, no microscopic difference was observed between the two groups. The preserved thickness of the outer nuclear layer was on average 61.3% in the ascorbate-treated and 66.4% in the placebo-treated group. The photoreceptor loss was not significantly different between the two groups. · Conclusion: The ascorbate did not protect the retina from blue-light induced damage. This favors the assumption that the mechanisms for blue light-induced retinal damage might differ from that for green light. Received: 13 October 1998 Revised version received: 22 January 1999 Accepted: 18 February 1999     

N-乙酰半胱胺酸对大鼠挫伤性视网膜病变的保护作用

目的:研究凋亡相关基因Bcl-2/Bax在大鼠挫伤性视网膜病变中的表达,以及N-乙酰半胱氨酸(N-acetyl-cysteine,NAC)对其表达的影响。方法:自由落体法制作视网膜挫伤模型。将SD大鼠随机分为正常组6只、模型组24只及治疗组24只。每组按挫伤后不同时间段分为1,3,7,14d,每个观察时段6只大鼠。HE染色光镜观察视网膜组织学变化,应用SABC免疫组织化学法检测大鼠视网膜挫伤后视网膜组织Bcl-2/Bax蛋白表达的变化。结果:挫伤性视网膜病变的损害主要集中在神经纤维层。挫伤后视网膜组织水肿,细胞紊乱,胞浆空泡样变,视网膜组织变薄,细胞丢失。NAC治疗组视网膜水肿程度有所改善,细胞紊乱,胞浆空泡样变,细胞丢失有所恢复。正常组及NAC治疗组视网膜组织Bax均未见表达,Bcl-2低表达。视网膜挫伤后1d时Bax表达开始增多,3d强阳性表达,7d表达有所减少,14d时表达进一步减少。Bcl-2低表达,未见明显变化。NAC治疗组各观察指标变化趋势基本与视网膜挫伤组相似,但表达明显减弱,两组相比较,于挫伤后1,3d和7d时差异有统计学意义(P<0.05)。Bcl-2在各时段的表达均较模型组有所增强,两组相比较,于挫伤后1,3d和7d时差异有统计学意义(P<0.01)。结论:在大鼠挫伤性视网膜病变中,NAC能够改善视网膜组织病理学损害并通过调节Bcl-2/Bax蛋白表达而抑制细胞凋亡,对视网膜挫伤具有保护作用。     

In-vivo-Imaging retinaler Zellapoptose nach akuter Lichtexposition

Purpose

Outer nuclear apoptosis following acute light exposure has previously only been shown histologically. This study investigated whether in vivo detection with DARC (detection of apoptosing retinal cells) technology could identify cells undergoing apoptosis.

Methods

Acute blue light damage (λ=405 nm; 3.2 mW/cm²) was applied to eyes of dark Agouti rats over 2 h. In vivo retinal imaging using confocal scanning laser ophthalmoscopy was performed before and directly after light exposure as well as after 24 h of dark adaptation. Development of retinal cell apoptosis was then assessed using intravitreal fluorescent-labeled annexin-5 with DARC technology in vivo.

Results

Directly after light exposure, no pathological retinal changes were observed by in vivo imaging. However, retinal flattening and the development of apoptosis within the irradiated retina occurred 1 day later and following dark adaptation. Confocal live scanning through the exposed retina revealed hyperfluorescent apoptotic cells at the level of the outer retina. Histological analysis confirmed the occurrence of photoreceptor cell death and the development of cellular damage at the outer retina.

Discussion

This study confirms acute light-induced outer nuclear apoptosis using in vivo DARC technology. This may open new and promising ways to assess programmed cell death of the photoreceptor cells, which – until now – was possible only with postmortem analysis.     

硫辛酸烟酸二联体对蓝光致大鼠视网膜损伤的防治作用

目的:探讨硫辛酸烟酸二联体(N2L)对蓝光致SD大鼠视网膜损伤的防治作用及最佳药物剂量,探寻其可能存在的保护机制。方法:选取150-200 g的SPF级雄性SD大鼠36只,随机分为正常对照组、蓝光损伤组、N2L低剂量组(1.0 mg/kg)、N2L中剂量组(2.5 mg/kg)、N2L高剂量组(5.0 mg/kg)及生理盐水组,每组各6只。正常对照组12 h明暗循环饲养,其余组每日接受9 h日常光照,3 h波长455 nm、强度3000±50 lx蓝光照射及12 h黑夜来建立损伤模型,持续14 d。同时每日腹腔注射1 mL对应剂量的药物。14 d后,所有组常规12 h明暗循环再饲养5 d,采用视网膜电图检查。过量麻醉法处死大鼠制备标本,HE染色,在光学显微镜下观察外核层厚度;CheKine^TM超氧化物歧化酶(SOD)活性检测试剂盒检测SOD活性;Western Blot检测大鼠视网膜Caspase-3、醌氧化还原酶1(NQO1)、谷胱甘肽巯基转移酶(GST)、Bcl-2和Bax蛋白表达量。结果:蓝光损伤组暗视ERG 3.0、10.0 (cd·s)/m^2<...     

枸杞子提取物对光诱导视网膜损伤的保护作用

目的　探索枸杞子提取物对人视网膜色素上皮（ARPE-19）细胞及C57BL/6J小鼠视网膜光诱导损伤的保护作用。方法　ARPE-19细胞分为正常细胞对照组,光诱导细胞损伤组,细胞低、中、高剂量组（0.1 g·L^-1、0.5 g·L^-1、1.0 g·L^-1枸杞子提取物+光诱导细胞损伤）,测定各组细胞活力以及细胞内活性氧（ROS）含量的变化。40只C57BL/6J小鼠随机分为正常动物对照组,光诱导动物损伤组,动物低、中、高剂量组（280 mg·kg^-1、370 mg·kg^-1、460 mg·kg^-1枸杞子提取物+光诱导动物损伤组）,每组8只。各剂量枸杞子提取物干预组小鼠在6周龄开始给予枸杞子提取物灌胃,8周后再给予10 000 lux光照射24 h;光照结束后ERG评估各组小鼠视网膜功能,OCT检测视网膜外核层（ONL）厚度,FFA检测视网膜血管渗漏情况,HE染色后对视网膜ONL细胞进行计数,同时检测血清中丙二醛（MDA）含量和超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）活性。结果　光诱导细胞损伤组ARPE-19细胞活力下降为正常细胞对照组的61.88%,细胞内ROS含量为正常细胞对照组的1.52倍;细胞低、中、高剂量组ARPE-19细胞活力较光诱导细胞损伤组均明显上升,细胞内ROS含量均明显下降,并均呈剂量依赖性（均为P＜0.05）。与动物对照组相比,光诱导动物损伤组小鼠ERG暗适应a波、b波振幅和明适应b波振幅均明显下降,动物低、中、高剂量组各波的振幅均得到不同程度改善。光诱导动物损伤组小鼠视网膜出现萎缩灶、血管渗漏和血管末端成珠样结构,视网膜ONL厚度变薄,为（52.18±4.23) μm,ONL每列细胞数明显减少,为（17.63±1.30）个;动物低、中、高剂量组小鼠视网膜病理改变及视网膜ONL厚度与ONL每列细胞数均得到不同程度改善,尤其动物高剂量组改善最为明显,ONL厚度为（59.72±2.76）μm, ONL每列细胞数为（20.00±1.51）个,与光诱导动物损伤组相比差异均有统计学意义（P=0.007、0.004）。相对于光诱导动物损伤组,动物低、中、高剂量组小鼠血清SOD、GSH-Px活性均明显提高,MDA含量均明显下降,且均呈剂量依赖性（均为P＜0.05）。结论　枸杞子提取物可以一定程度上保护视网膜免受光损伤。     

远红外线及近红外线的光生物学效应在眼病的应用研究

低能量的远红/近红外光(far-red/near-infrared,FR/NIR)通过减少细胞死亡、增强线粒体功能等光生物学效应(photobiomodulation,PBM)来促进组织伤口愈合.因其能够穿透组织到达视网膜,近年应用于眼病治疗,特别是眼底病,如甲醇中毒性视网膜病变、视网膜光损伤、遗传性视网膜病变、年龄相关性黄斑病变、糖尿病视网膜病变、早产儿视网膜病变等.PBM不会对视网膜及其他组织造成损伤,从而为眼病带来一种新型、非侵入性、安全有效的治疗方法.     

Minocycline protects photoreceptors from light and oxidative stress in primary bovine retinal cell culture

PURPOSE: To determine whether minocycline, a compound known to protect the retina against light-induced damage in rodent models, and its structurally related analogues would protect photoreceptor cells in primary bovine retinal cell culture against light and oxidative stress. METHODS: Minocycline and its analogues were tested in primary retinal cell culture to see whether they would inhibit light or oxidative stress-induced cell death. Primary cell cultures composed of photoreceptors, bipolar cells, and glial cells were prepared from bovine retinas. The extent of cell death induced by light or oxidative stress was assessed by using Sytox Green (Invitrogen-Molecular Probes, Eugene, OR) a nucleic acid dye uptake assay. Differential protection of photoreceptor cells from stress were examined using immunocytochemistry. RESULTS: Minocycline and methacycline were cytoprotective against light- or oxidative stress-induced damage of bovine primary photoreceptors in culture with an EC(50) < 10 microM. In contrast, structurally related analogues such as demeclocycline, meclocycline, and doxycycline were phototoxic at >3 to >10 microM. Though demeclocycline was found to be phototoxic, it was cytoprotective (EC(50) = 5 microM) against oxidative stress in the absence of exposure to light. CONCLUSIONS: The protective action of minocycline against light-induced damage in the cell-based assays agrees with earlier reports in animal models and suggests that the in vitro assay using bovine primary retinal cell culture is a suitable model for evaluating compounds for retinal protection. Cellular protection or toxicity produced by structurally related compounds show that minor structural modifications can alter the function of minocycline and lead to potent retinal protective compounds.     

Mechanisms of intravitreal toxicity of indocyanine green dye: implications for chromovitrectomy

PURPOSE: Indocyanine green (ICG) dye was shown to improve the visualization of preretinal tissues during chromovitrectomy. However, controversy arose regarding the safety of intravitreal ICG application, because worse functional outcomes and a higher incidence of retinal pigment epithelium (RPE) changes and visual field defects were reported. The mechanisms of ICG-related toxicity and their relevance for chromovitrectomy are reviewed. METHODS: A literature search was performed from 1998 through 2005 for relevant information related to the mechanisms of intravitreal ICG toxicity. Animal and clinical data on intravitreal ICG-related toxicity were collected to clarify the mechanisms of the risk of intravitreal ICG injection. RESULTS: Over 80 controversial in vitro, ex vivo, and in vivo animal investigations as well as clinical reports on intravitreal ICG staining were found in the literature. The main postulated mechanisms of intravitreal ICG-related toxicity were as follows: biochemical direct injury to the ganglion cells/neuroretinal cells, RPE cells, and superficial retinal vessels; apoptosis and gene expression alterations to either RPE cells or neuroretinal cells; osmolarity effect of ICG solution on the vitreoretinal interface; light-induced injury; and mechanical cleavage effect to the internal limiting membrane/inner retina. Whereas the exact mechanism of intravitreal ICG-related damage remains yet to be determined, most animal experiments proposed that ICG dye has a dose-dependent toxic effect on retinal tissue. This hypothesis was supported by clinical data, because better functional outcomes were obtained when low dye concentrations and short incubation times were reported. CONCLUSIONS: Much evidence supports that ICG dye has a dose-dependent toxic effect on the retina. Therefore, the following recommendations to minimize toxic effects on the retina are proposed: dye injection in concentrations as low as possible; avoidance of repeated ICG injections onto bare retina; dye injection far from the macular hole to prevent direct dye contact with the RPE; short incubation time of ICG in the vitreous cavity to diminish the concentration in contact with the retinal tissue; and the light pipe kept far from the retina throughout the whole surgical procedure.     

The mouse ERG before and after light damage is independent of p53

Death of retinal photoreceptors by apoptosis is observed under many physiological and pathological conditions such as histogenesis, retinal dystrophies and light-induced photoreceptor degeneration. To date, little is known about regulatory mechanisms for apoptosis in the retina. The tumor suppressor gene p53 is a regulator of apoptosis in a number of systems, however, p53-independent apoptosis has also been described. We have therefore investigated whether the lack of p53 influences the dark-adapted ERG in C57BL/6 p53^−/− mice compared to p53^+/+ control littermates under physiological (regular light-dark cycle) conditions. We also recorded ERGs at 12 to 14 h in darkness following diffuse bright light exposure to 8′000 or 15'000 lux for 2 h. ERG analysis over a range of 6 logarithmic units of light intensity revealed normal and virtually identical a-, b-, c-waves and oscillatory potentials in dark-adapted p53^+/+ and p53^−/− mice. After exposure to diffuse white fluorescent light strong decreases of all ERG components were found to be very similar in both genotypes. These data support the notion that the p53 protein is neither essential for normal retinal function nor for processes involved in light-induced depression of the ERG in mice. This revised version was published online in July 2006 with corrections to the Cover Date.     

Decreased Retinal Neuronal Cell Death in Caspase-1 Knockout Mice

Purpose

To determine whether apoptosis of retinal neurons induced by excessive light exposure and ischemia–reperfusion injury is altered in caspase-1 knockout mice.

Methods

Eight- to 10-week-old caspase-1 knockout mice (Casp1^–/–) and wild-type (WT) mice (C57BL/6) were exposed to diffuse, cool, white fluorescent light of 25 000 lux for 2?h. Other mice were subjected to retinal ischemia by increasing the intraocular pressure to 110?mmHg for 45?min. Electroretinograms (ERGs) were recorded before and after the light exposure. TdT-dUTP terminal nick-end labeling (TUNEL) was performed to identify the apoptotic cells after the insults. The inner retinal thickness was measured to evaluate the retinal injury after the ischemia–reperfusion. Expression of caspase-1 protein was studied by immunohistochemical analysis and Western blotting. Caspase-1-like protease activity was determined by a colorimetric tetrapeptide substrate.

Results

The morphology of the retina and the amplitudes of the a and b waves of the ERGs of Casp1^–/– mice did not differ from those of WT mice. After the light exposure, TUNEL-positive cells were observed in the outer nuclear layer of the WT mice retina. The number of TUNEL-positive photoreceptor nuclei after the light exposure, and the number of nuclei in the inner nuclear layer after the ischemia–reperfusion injury, were significantly less in Casp1^–/– mice than in WT mice. There were more caspase-1-positive photoreceptor cells in WT mice after the light injury. The inner retinal layer of Casp1^–/– mice was significantly thicker in Casp1^–/– mice than in WT mice 2 weeks after the ischemic insult.

Conclusions

Retinal neuronal apoptosis was less prominent in Casp1^–/– mice after excessive light exposure and ischemia–reperfusion injury. These data indicate that caspase-1 plays a role in retinal neuronal apoptosis.?Jpn J Ophthalmol 2006;50:417–425 © Japanese Ophthalmological Society 2006