胰高血糖素类肽-1缓释珠对大鼠视网膜神经节细胞的保护作用

目的探讨玻璃体内植入胰高血糖素类肽-1（GLP-1）缓释珠对大鼠视网膜神经节细胞的保护作用。设计实验研究。研究对象SPF级Sprague-Dawley（SD）雄性大鼠25只。方法将25只大鼠随机分为2组,实验组13只,对照组12只。实验组大鼠右眼玻璃体内植入4个GLP-1缓释珠,对照组右眼玻璃体内注入4μl复方氯化钠。GLP-1缓释珠直径600μm,内含3000个整合了GLP-1基因的人骨髓间充质干细胞,外被致密的藻酸盐外膜,以确保GLP-1产物可顺利释放而不引起免疫排斥。玻璃体内注射均在右眼视神经夹伤后立即进行。视神经夹伤后第23天用3％荧光金从双侧上丘做逆行标记,第28天取双眼球标本做视网膜铺片并在荧光显微镜下拍摄照片,采用人工双盲法进行视网膜神经节细胞计数。主要指标视网膜神经节细胞密度以及视网膜神经节细胞存活率。结果视网膜神经节细胞密度实验组与对照组分别为（2113±474）／mm2和（1734±424）／mm2,两组之间的差异有统计学意义（t=2．111,P=-0．046）。视网膜神经节细胞存活率实验组与对照组分别为（74±18）％和（57±16）％,两组之间的差异有统计学意义（t=-2．451,P=-0．022）。结论GLP-1缓释珠玻璃体内植入后对视神经夹伤大鼠视网膜神经节细胞具有保护作用,可以提高视网膜神经节细胞存活率。     

饱和氢气水对大鼠视神经夹伤模型视网膜神经节细胞保护作用的实验研究

 目的 探索饱和氢气水对大鼠视神经夹伤模型视网膜神经节细胞（RGC）的保护作用。设计 实验研究。研究对象 SPF级SD大鼠18只。方法 对18只大鼠采用随机数表法随机分为3组,每组6只。均选取右眼为实验眼,左眼为正常对照眼。使用40 g微型视神经夹在大鼠视神经球后2 mm处夹持60 s建立视神经夹伤模型。A组给予饱和氢气水腹腔注射,5 ml/kg,每日1次;B组和C组分别给予饱和氢气水和生理盐水滴眼,每次1滴,每日3次。用药第9天,麻醉下采用3%荧光金双上丘两点注射法逆行标记大鼠RGC,第14天深麻醉下取眼球并处死动物,行视网膜定向铺片,距离视乳头中心上下左右各2 mm 拍摄照片,盲法计数RGC。主要指标 RGC存活率。结果 A组、B组和C组RGC存活率分别为40.35%±13.04%、58.34%±14.00%和43.07%±7.80%（F=3.965, P=0.041）。其中B组与A组和C组之间均有显著性差异（P=0.020;P=0.042）;A组和C组之间无显著性差异（P=0.698）。结论 饱和氢气水滴眼2周对大鼠视神经夹伤模型视网膜神经节细胞可能具有一定的保护作用。（眼科,2014, 23： 107-110）     

葛根素对大鼠视网膜神经节细胞的作用

目的探讨葛根素对大鼠视神经夹伤模型视网膜神经节细胞(retinal ganglion cells,RGC)的保护作用.设计随机对照实验研究.对象40只SD大鼠.方法将40只大鼠随机分5组,每组8只,分别为生理盐水组,莫尼定组,葛根素高、中、低剂量组.右眼行视神经夹伤,左眼为正常对照.术后24天双侧上丘荧光金逆行标记.术后28天视网膜定向铺片、拍摄荧光照片及图像分析.RGC的存活率为右眼RGC密度除以左眼RGC密度,再乘以100%.主要指标RGC的存活率.结果40只SD大鼠RGC存活率生理盐水组为(53.99±6.69)%,葛根素低剂量组(58.67±6.55)%,中剂量组(63.85±5.75)%,高剂量组(69.66±4.79)%,莫尼定组(62.10±3.90)%.葛根素高、中剂量组及莫尼定组均与生理盐水组比较有显著性差异(P值均小于0.05).结论葛根素中剂量组、高剂量组及莫尼定组对视神经损伤后RGC的存活有明显的增强作用.     

杞贞胶囊对视网膜神经节细胞保护作用及其作用机制的实验研究

目的 探索杞贞胶囊对大鼠视神经夹伤模型视网膜神经节细胞的保护作用及其作用机制。设计 实验研究。研究对象 SPF级SD大鼠72只。方法 72只SD大鼠随机分为2组：用药组36只;对照组36只。两组大鼠右眼行视神经夹伤,于球后2 mm处用40 g微型视神经夹夹伤视神经60 s。左眼作为正常对照。夹伤后2小时及此后每日予以灌胃给药一次。用药组给予20%杞贞溶液2.5 ml/kg,对照组给予生理盐水2.5 ml/kg。给药第28天取眼球标本,用药组和对照组各取24只行HE染色﹑Tunel试剂盒染色﹑Caspase-3免疫组化染色;剩余每组12只分离视网膜提取mRNA,测定Bax和Bcl-2基因的表达量。主要指标 视网膜厚度﹑Bax和Bcl-2基因表达量。结果 用药组视网膜厚度平均为（109.0±4.4）μm;对照组视网膜厚度为（101.8±7.6）μm（F=29.497,P=0.028）。两组间Bax基因表达差异具有统计学意义（t=1.089,P=0.028）;Bcl-2基因表达差异未见统计学意义（t=0.553,P=0.692）。结论 杞贞胶囊对大鼠视神经夹伤后的视网膜神经节细胞具有保护作用,可能通过下调Bax基因表达和抑制Caspase蛋白活性从而减少视网膜神经节细胞凋亡。     

金丝桃素对视神经损伤大鼠视网膜节细胞的保护作用

目的观察金丝桃素对视神经损伤大鼠视网膜节细胞的保护作用。方法24只SD大鼠随机分为正常对照组、单纯夹伤组、生理盐水对照组、金丝桃素治疗组4组,每组6只（12眼）。对所有大鼠行双上丘注射2％荧光金逆行标记节细胞,7d后,对单纯夹伤组、生理盐水对照组、金丝桃素治疗组进行球后视神经钳夹．同时在生理盐水对照组、金丝桃素治疗组玻璃体内分别注入生理盐水和金丝桃素5ul,14d后进行视网膜节细胞的计数。采用SPSS13．0统计软件对所得数据进行t检验。结果视神经夹伤后14d,存活的视网膜节细胞显著减少。单纯夹伤组节细胞存活率为50％,生理盐水对照组节细胞存活率为52％,金丝桃素治疗组节细胞存活率为68％。金丝桃素治疗组相比单纯夹伤组和生理盐水对照组,存活的节细胞明显要多（P〈0．05）。结论玻璃体内注射金丝桃素能减少大鼠视神经损伤后视网膜神经节细胞的死亡率．对视网膜节细胞有保护作用。     

蛇毒神经生长因子对大鼠视神经夹伤保护的电镜观察

目的研究蛇毒神经生长因子在视神经损伤后对视网膜神经节细胞的保护作用。方法将Wistar大鼠40只随机分为实验对照组和实验治疗组。制作实验性视神经夹伤模型,用头部宽1mm的微型血管夹夹伤大鼠右眼视神经后,实验治疗组向伤眼玻璃体腔内注入蛇毒神经生长因子100BU(0.025mL)。实验对照组向伤眼玻璃体腔内注入0.025mL平衡盐液。于损伤后第3d、7d、14d、30d、60d取材,用透射电镜观察不同时间段各组视网膜形态学变化。结果电镜下大鼠视网膜改变:实验治疗组和对照组电镜下均可见坏死和凋亡。伤后14d,实验治疗组视网膜微管数目比实验对照组较多,排列比较整齐。结论在视神经损伤早期,蛇毒神经生长因子能减轻视神经夹伤后微管的损坏,提高视网膜神经节细胞的存活数量,对视网膜神经节细胞有明显的保护作用。     

小干扰RNA保护大鼠视网膜神经节细胞的实验研究

 目的 通过大鼠视神经夹伤模型,研究小干扰RNA（siRNA）对视网膜神经节细胞（RGC）的保护作用。设计 实验研究。 研究对象 SPF级SD大鼠54只。方法 54只SD大鼠随机分为A、B、C三组,每组18只。均选取右眼为实验眼,左眼为正常对照。在球后2 mm处用40 g压力微型视神经夹夹持视神经60 s,做视神经夹伤模型。建立模型后当天,A、B、C三组分别给予玻璃体注射10 μg、20 μg siRNA和生理盐水。视神经夹伤后10天,每组取6只大鼠用荧光金做逆行标记,14天时取标记后的大鼠双眼眼球标本做视网膜铺片并拍摄照片,RGC计数。计算RGC存活率（右眼RGC数/左眼RGC数×100%）。每组其余12只大鼠进一步用蛋白印迹法检测视网膜组织中caspase-3蛋白的表达水平。主要指标 RGC存活率,caspase-3蛋白的表达水平。结果 A、B、C组RGC存活率分别为53.63%±7.35%、57.86%±6.00%、45.00%±4.37%（F=7.11,P=0.029）,其中A组与C组（P=0.025）,B组和C组（P=0.002）之间均有显著性差异;A 组和B组之间无显著性差异（P=0.24）。A、B、C三组视网膜组织中Caspase-3蛋白与内参灰度比值分别为0.20±0.02、0.19±0.02、0.24±0.03（F=9.73,P=0.02）。其中A组与C组（P=0.005）,B组和C组（P=0.001）之间均有显著性差异;A 组和B组之间无显著性差异（P=0.418）。结论 小干扰RNA能有效保护大鼠视神经夹伤模型的RGC,提高RGC的存活率。     

玻璃体内注射酵母多糖对视神经夹伤大鼠RGCs的保护作用

目的 观察眼内注射酵母多糖对视神经夹伤大鼠视网膜神经节细胞(RGCs)的保护作用.方法 SD大鼠45只,随机分为3组,以荧光金逆行标记RGCs 1周后,行左眼视神经钳夹伤,右眼为假手术对照.3 d后分别于各对应组大鼠左眼玻璃体腔注射PBS或酵母多糖5μL,所有大鼠存活21 d处死,每组各取5只大鼠行视网膜冰冻切片,余做视网膜铺片,计算RGCs标识率(左眼RGCs数μ右眼RGCs数)× 100%,并行统计学分析.结果 酵母多糖组、PBS组和单纯钳夹组中RGCs标识率分别为:(19.22±2.51)%、(1.86 ±3.09)%和(1.78±2.61)%,酵母多糖组中存活RGCs较单纯钳夹组和PBS组差异有统计学意义(P=0.000),单纯钳夹组和PBS组差异无统计学意义(P=0.925).结论 酵母多糖玻璃体腔注射后,能诱导眼内炎症反应,从而对视神经夹伤大鼠RGCs的存活具有一定的保护作用.     

杞贞胶囊对大鼠视网膜节细胞的保护作用

目的通过大鼠视神经夹伤模型,探讨杞贞胶囊对视网膜神经节细胞的保护作用。方法50只大鼠随机分为5组,每组10只,右眼制作视神经夹伤模型。阴性对照组、阳性对照组及低、中、高剂量组分别用生理盐水、4.69%益脉康及低(16.4g生药/100ml)、中(32.8g生药/100ml)、高(65.6g生药/100ml)剂量的杞贞胶囊溶液灌胃给药,每日1次,持续4周。动物安死术前5d逆行标记节细胞,用彩色颗粒分析软件计数节细胞。结果阴性对照组、阳性对照组和低、中、高剂量组节细胞平均存活率分别为(40.88±12.50)%、(59.34±9.19)%、(52.66±12.52)%、(59.39±7.45)%和(63.00±8.95)%。阴性对照组与低剂量组之间差异具有显著性(P=0.014),阴性对照组与阳性对照组、中、高剂量组之间差异有非常显著性(P<0.01)。结论杞贞胶囊能有效地保护视网膜神经节细胞,随着给药剂量的增加,作用逐渐增强。益脉康也具有视网膜神经节细胞保护作用。     

香菇多糖对视神经损伤视网膜节细胞保护作用的实验研究

目的 评估香菇多糖(LNT)对视神经损伤后的保护作用.方法 成年纯种新西兰大耳白兔按视神经夹伤后存活时间不同(1,3,7,14 d)随机分为4组,每只兔右眼为治疗眼(实验组),于伤后玻璃体腔内注射LNT;左眼为对照眼(对照组),伤后玻璃体腔内注射生理盐水.观察LNT实验组与对照组视网膜形态学改变,用TUNEL染色法对视网膜神经节细胞(RGC)做凋亡细胞的定位和计数,评估香菇多糖对视网膜神经节细胞的保护作用.结果 正常组视网膜内界膜平滑完整,三层细胞分界清楚,节细胞呈单层排列,内、外丛状层厚度、染色均匀.对照组损伤后视网膜厚度较正常组变薄,细胞排列疏松紊乱,节细胞数目减少,以伤后7 d最为严重.各时间点实验组与对照绀比较节细胞存活数增加,网膜厚度大于对照组,且细胞排列相对整齐.结论 兔视神经损伤后玻璃体腔内注射LNT能有效保护视网膜神经节细胞.     

Neuroprotective effect of sulfhydryl reduction in a rat optic nerve crush model

PURPOSE: The signaling of retinal ganglion cell (RGC) death after axotomy is partly dependent on the generation of reactive oxygen species. Shifting the RGC redox state toward reduction is protective in a dissociated mixed retinal culture model of axotomy. The hypothesis for the current study was that tris(2-carboxyethyl)phosphine (TCEP), a sulfhydryl reductant, would protect RGCs in a rat optic nerve crush model of axotomy. METHODS: RGCs of postnatal day 4 to 5 Long-Evans rats were retrogradely labeled with the fluorescent tracer DiI. At approximately 8 weeks of age, the left optic nerve of each rat was crushed with forceps and, immediately after, 4 muL of TCEP (or vehicle alone) was injected into the vitreous at the pars plana to a final concentration of 6 or 60 microM. The right eye served as the control. Eight or 14 days after the crush, the animals were killed, retinal wholemounts prepared, and DiI-labeled RGCs counted. Bandeiraea simplicifolia lectin (BSL-1) was used to identify microglia. RESULTS: The mean number of surviving RGCs at 8 days in eyes treated with 60 microM TCEP was significantly greater than in the vehicle group (1250 +/- 156 vs. 669 +/- 109 cells/mm(2); P = 0.0082). Similar results were recorded at 14 days. Labeling was not a result of microglia phagocytosing dying RGCs. No toxic effect on RGC survival was observed with TCEP injection alone. CONCLUSIONS: The sulfhydryl-reducing agent TCEP is neuroprotective of RGCs in an optic nerve crush model. Sulfhydryl oxidative modification may be a final common pathway for the signaling of RGC death by reactive oxygen species after axotomy.     

大鼠视神经压榨伤模型的建立

目的建立大鼠标定性视神经损伤模型.方法健康SD大鼠28只,7只为正常对照组,只进行双上丘注射3%快蓝逆行标记视网膜神经节细胞(retinal ganglion cells,RGCs),另21只为标定性视神经损伤组,依损伤后存活时间的不同再分为A组(4d组)、B组(14d组)及C组(21d组),每组7只.21只大鼠以夹持力为40 g的特制视神经夹,在大鼠眼球后2 mm处夹持视神经4s,制成大鼠标定性视神经压榨伤模型,于处死前3d采用双上丘直接注射3%快蓝(fast blue)法标记双眼RGCs,将全视网膜铺片置于荧光显微镜下,在距视乳头1 mm处的颞上、颞下、鼻下、鼻上4处作荧光摄影(400 ×),并输入计算机经图像分析仪计数RGCs,按RGCs标识率进行统计学比较.RGCs标识率=损伤眼(右眼)RGCs数/未损伤眼(左眼)RGCs数×100%.结果正常大鼠的RGCs标识率右眼RGCs数/左眼RGCs数为99.79%±13.05%,左眼RGCs数/右眼RGCs数为101.86%±13.91%,无论是用左眼的RGCs数比右眼的RGCs数,或用右眼的RGCs数比左眼的RGCs数,其结果无显著性差异(P＞0.5).视神经损伤组的RGCs标识率A组(4d组)RGCs标识率为77.79%±7.11%;B组(14d组)RGCs标识率为63.76%±3.79%;C组(21d组)RGCs标识率为54.66%±4.75%.以上显示,损伤各组的RGCs标识率明显低于正常对照组(P＜0.05),且随着时间的推移,损伤A、B、C组的RGCs标识率渐进性降低.结论用特制的夹持力为40 g的视神经夹,夹持正常大鼠视神经4s,可造成部分性RGCs丧失,随大鼠存活时间的推移,RGCs呈渐进性丧失.眼科学报2001;1799～102.     

The Protective Role of Mecobalamin Following Optic Nerve Crush in Adult Rats

IntroductionProgressivelossofganglioncellsandtheiraxonsisafeatureofoculardiseasessuchasopticnervedamage,glaucoma,ischemia,andinfla鄄mmation.Thelossofganglioncellsaffectsbothopticnervefibersandtheirsheaths.Thisresultsinalossofneuralnutritionandalossofbloodandoxygensupplythatcanleadtotheaccum鄄ulationofextracellularglutamateresultinginexcitoxitywhichinfluencestheconductionofvisualelectricsignals.Thesetwofactorsresultinapoptosisofretinalganglioncells[1].Furthermore,degenerationcontinuestoprogres…     

The effect of ginkgo biloba on the rat retinal ganglion cell survival in the optic nerve crush model

Purpose: To investigate the effect of ginkgo biloba on the retinal ganglion cell survival in a rat optic nerve crush model. Methods: Twenty‐four Sprague–Dawley rats were divided randomly into a study group of 12 animals receiving intraperitoneal injections of ginkgo biloba and a control group of 12 animals receiving intraperitoneal saline injections. All injections were performed 1 hr before the optic nerve crush and daily afterwards. For each animal, the right optic nerve was crushed closely behind the globe for 60 seconds using a microclip with 40 g power. The left optic nerve was kept intact. At 23 days after the optic nerve crush, the retinal ganglion cells were labelled retrogradely by injecting 3% fluorogold into both sides of the superior colliculus of the brain. At 4 weeks after the optic nerve crush, the animals were killed. Photographs taken from retinal flat mounts were assessed for the number and density of the retinal ganglion cells. Results: The survival rate, defined as the ratio of the retinal ganglion cell density in the right eye with the optic nerve crush divided by the retinal ganglion cell density in left eye without an optic nerve trauma, was significantly (p = 0.035) higher in the study group with ginkgo biloba than in the control group (60.0 ± 6.0% versus 53.5 ± 8.0%). Conclusion: The results suggest that intraperitoneal injections of a ginkgo biloba extract given prior to and daily after an experimental and standardized optic nerve crush in rats were associated with a higher survival rate of retinal ganglion cells.     

A new calcium channel antagonist, lomerizine, alleviates secondary retinal ganglion cell death after optic nerve injury in the rat

PURPOSE: We investigated whether lomerizine, a new diphenylmethylpiperazine calcium channel blocker, exerted a neuroprotective effect on axonal or retinal damage induced by optic nerve injury in the rat. METHODS: A partial crush lesion was inflicted unilaterally on the optic nerve, 2 mm behind the globe, in adult Wistar albino rats. Animals were treated with the vehicle, 10 or 30 mg/kg lomerizine. Each solution was given orally twice daily for 4 weeks. One week before euthanization, Fluoro-Gold (FG) was injected into both superior colliculi to retrogradely label surviving retinal ganglion cells (RGCs). Approximately 1 month after the optic nerve injury, the retinal damage was assessed morphologically, and the optic nerve axons surrounding the initial lesion were examined histologically. RESULTS: The mean RGC density in the control group decreased to 65.9 +/- 1.32% of the contralateral eye, whereas the systemic application of 10 or 30 mg/kg of lomerizine significantly enhanced the RGC survival to 88.1 +/- 0.38% and 89.8 +/- 0.28%, respectively. Histological examination of damaged axons revealed no significant enhancement of the density or total number of axons of the retinal ganglion cells in the lomerizine-treated group. The crush force we employed caused no significant morphological differences in the retinal layers between the sham-operated animals and the animals from the experimental groups. CONCLUSIONS: Our findings suggest that lomerizine alleviates secondary degeneration of RGCs induced by an optic nerve crush injury in the rat, presumably by improving the impaired axoplasmic flow.