缺血-再灌注大鼠视网膜TPA变化与视网膜水肿的关系

目的　探讨缺血-再灌注对大鼠视网膜分泌组织型纤溶酶原激活物(TPA)的影响及其与视网膜水肿的关系。方法　采用提高眼压法造成视网膜缺血后,恢复眼压形成血流再灌注。实验分正常对照组、缺血 1h再灌注 1h组、缺血 1h再灌注 2h组、缺血 2h再灌注 1h组和缺血 2h再灌注 2h组。每组各取 10例测试视网膜组织TPA的活性和含水量。结果　缺血-再灌注后,大鼠视网膜组织TPA的活性和含水量随缺血和再灌注时间的延长而升高(P<0 01)。 结论　缺血-再灌注可引起视网膜组织TPA的活性升高和视网膜水肿,是视网膜组织结构和功能损伤的因素之一。     

缺血预处理对大鼠视网膜缺血再灌注损伤保护作用

目的：探讨缺血预处理是否对视网膜缺血再灌注损伤有保护作用及其机理,方法：利用前房灌注生理盐水形成高眼压的视网膜缺血再灌注损伤的动物模型,视网膜缺血时间为1h分别于缺血前30min、24h或72h对大鼠一只眼5min短暂缺血即预处理,24h或72h后行视网膜电图（ERG）、电镜、光镜、丙二醛（MDA）及热休克蛋白70（HSP70）检测,或者一侧眼行5min假处理,24h后行1h缺血,24h或72h再行上述检测,所有对侧眼不作处理作对照,结果：与假处理相相比,缺血前24、72h进行预处理后的大鼠视网膜光镜、电镜表现损害明显减轻,ERGb波明显恢复（P＜0．01）,MDA含量降低（P＜0．01）,缺血前30min预处理的视网膜表现严重的损害,ERGb波几安全消失,结论：缺血预处理对视网膜缺血再灌注损伤有保护作用,且有一定时限性。     

缺血再灌注大鼠视网膜细胞线粒体钙含量与组织型纤溶酶原激活物活性的相关性

目的观察缺血再灌注大鼠视网膜细胞线粒体钙含量与组织型纤溶酶原激活物（tissue-type plasminogena etivator，TPA）活性的变化，探讨两者之间的相关性。方法采用提高眼压法造成视网膜缺血后，恢复眼压形成血流再灌注。实验分正常对照组和缺血1h再灌注1h组、缺血1h再灌注2h组、缺血2h再灌注1h组、缺血2h再灌注2h组四个实验组，每组各取10例测定视网膜细胞线粒体钙浓度与TPA活性的变化，并进行相关性分析。结果缺血再灌注后，大鼠视网膜细胞线粒体钙含量与TPA活性随缺血和再灌注时间的延长而升高（P〈0．01），两者之间呈显著相关性（r=0．524，P〈0．01）。结论缺血再灌注可引起视网膜细胞线粒体钙含量的升高，从而导致视网膜组织TPA活性的增高，引起视网膜组织结构和功能的损伤。     

视网膜静脉阻塞患者血浆t－pA和PAI活性检测及临床意义

采用发色底物法检测６１例视网膜静脉阻塞（ＲＶＯ）患者血浆组织纤溶酶原激活剂（ｔ－ＰＡ）及纤溶酶原激活剂抑制物（ＰＡＩ）活性。正常对照组ｔ－ｐＡ活性２．０７±０．４０ＩＵ／ｍｌ，ＰＡＩ活性７．３３±０．６７ＡＵ／ｍｌ。ＲＶＯ患者ｔ－ＰＡ活性显著降低（１．６９±０．５６ＩＵ／ｍｌ，ｐ＜０．０１），ＰＡＩ活性显著增高（８．８０±１．６０ＡＵ／ｍｌ，Ｐ＜０．０１）。其中缺血型较非缺血型ｔ－ｐＡ活性更低（分别为１．３５±０．４３ＩＵ／ｍｌ，１．９２±０．５３ＩＵ／ｍｌ，Ｐ＜０．０１），ＰＡＩ活性更高（分别为９．３５±１．３７ＩＵ／ｍｌ，８．４２±１．２９ＩＵ／ｍｌ，Ｐ＜０．０１）．并且，这些改变随病情严重程度的加重而愈显著。     

缺血再灌注损伤对大鼠视网膜功能的影响

目的 探讨缺血再灌注损伤对大鼠视网膜功能的影响。 方法 70只健康 Wistar 大鼠，预实验随机抽取 20 只分为正常对照组和单纯灌注组，记录视网膜电图（electroret inography，ERG）并测定b波峰值，经统计学处理无差异后，其余50只随机分为10组，用升高眼压1 h 的方法建立右眼视网膜缺血模型，分别于缺血后1 h及再灌注3、6、12 、24 h、3、5、7、14、21 d记录双眼暗视闪光 ERG并测定b波峰值。 结果 正常对照组动物左右眼ERG b波峰值无差异；单纯灌注眼与正常对照眼ERG b波峰值无差异；单纯缺血组实验眼ERG各波消失，再灌注实验眼组ERG b波部分恢复，但随再灌注时间的延长b波峰值呈进行性下降. 结论 视网膜缺血再灌注损伤可导致大鼠视网膜功能持续渐进性的影响。（中华眼底病杂志，2003，19：201-268）     

大鼠视网膜缺血再灌注后IL-1β的免疫组化研究

目的：观察大鼠视网膜缺血再灌注(retinal isehemia reperfusion，RIR)后，白细胞介素1β(IL-1β)多肽在视网膜表达变化。方法：采用前房灌注生理盐水，形成130mmHg(17．3kPa)高眼压，诱导大鼠视网膜缺血60min，解除高眼压，建立RIR模型。缺血60min，再灌注12h、48h作视网膜冰冻切片，IL-1β免疫组化观察。结果：正常对照组未见IL-1β表达，RIR后12h、48h，视网膜神经节细层可见IL-1β表达。结论：结果提示：IL-1β多肽在蛋白质水平参与RIR损伤发生。     

黄芪对大鼠视网膜缺血再灌注损伤的影响

目的:观察黄芪对大鼠视网膜缺血再灌注损伤的影响及其作用机制。方法:将90只大鼠随机分为3组:对照组、缺血组、保护组,采用前房灌注液体形成14.63kPa高眼压60min的方法建立模型,连续7d每天1次黄芪注射液6g/kg,ip,于手术前30min加注1次,对照组给予同等剂量的生理盐水。两组缺血60min后,分别再灌注0,2,6,12,24,48,72h,用比色法进行视网膜SOD、MDA、NO的测定,用光镜测量包埋切片的平均视网膜内层厚度。结果:黄芪能显著对抗大鼠视网膜缺血再灌注后MDA、NO水平的升高和SOD水平的降低,同时能改善平均视网膜内层厚度的变化。结论:黄芪可减轻大鼠视网膜缺血再灌注后的损伤,其机制可能与黄芪清除自由基,拮抗NO的毒性作用有关。     

瞬间性视网膜缺血重新灌注对大鼠视网膜细胞凋亡的影响

我们以升高眼压的方法造成视网膜瞬间缺血重新灌注的动物模型 ,对瞬间性视网膜缺血重新灌注所引起的视网膜细胞凋亡及其相应的一些改变进行研究。1　材料和方法1.1　动物模型及标本制作2 5 0～ 30 0ｇ雌性ＳＤ大鼠 2 0只 ,均以 1只眼用于实验观察。盐酸氯胺酮 35ｍｇ/ｋｇ及盐酸甲苯噻嗪 5ｍｇ/ｋｇ联合肌肉注射麻醉 ,前房刺入联有灌注液的 2 5号针头 ,调整灌注液的高度 ,升高眼压至 6 5ｍｍＨｇ(1ｍｍＨｇ =0 .133ｋＰａ)造成瞬间性视网膜缺血1ｍｉｎ或 5ｍｉｎ ,然后恢复眼压使血流重新灌注。手术前后常规用氯霉素眼液点眼。视网膜缺血重…     

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)     

视网膜缺血再灌注损伤和细胞因子的研究进展

视网膜缺血再灌注（retinal ischemia reperfusion,RIR)损伤是由多种因素，如自由基，炎性细胞因子，白细胞等介导的复杂病理生理过程，RIR损伤和细胞因子之间存在密切联系，一方面，RIR损伤可以诱导多种炎性细胞因子，如白细胞介素1和6的表达，另一方面，多种外源性神经营养因子，如碱性成纤维细胞生长因子，重组人肝细胞生长因子，脑源性神经生长因子等，具有保护RIR损伤的作用，研究RIR损伤和细胞因子关系具有重要意义，因此我们报告了国外有关RIR损伤和细胞因子的最新研究进展。     

Ischemia Induces Significant Changes in Purine Nucleoside Concentration in the Retina-Choroid in Rats

Adenosine, produced from the decomposition of adenosine triphosphate, is believed to provide protective effects during ischemia. On the other hand, adenosine metabolites may serve as precursors for oxygen free radical formation. The time course of formation of adenosine and its purine metabolites was studied during retinal ischemia in rats. Concentrations of adenosine and its purine nucleoside metabolites inosine, hypoxanthine, and xanthine in the retina-choroid of ketamine/xylazine-anesthetized rats were measured during retinal ischemia using high performance liquid chromatography. Quantitative measurements were made possible in the small tissue mass through the use of internal standards. Ischemia was induced by ligation of the central retinal artery. In each rat, one eye was ischemic while the other served as a non-ischemic control. Eyes were frozen in situ at 1, 5, 10, 20, 30, 60, and 120 min of ischemia. The retina-choroid was then removed from the frozen eyes and analysed. Significant increases in the concentrations of adenosine, inosine, and hypoxanthine in ischemic compared to control retina-choroid were detectable within 1 to 5 min of the onset of ischemia, and within 10 min for xanthine. Increase in adenosine concentration in ischemic relative to control retina-choroid plateaued at 30 min of ischemia, while inosine and hypoxanthine concentrations increased continuously. The increase in xanthine concentration was exponential throughout the measurement period. This study documented the time-related changes in purine nucleoside concentration during ischemia. Prolonged ischemia results in ongoing production of xanthine, which by serving as a precursor for oxygen free radical formation, could be a pathogenic factor in prolonged retinal ischemia.     

Triamcinolone Acetonide Prevents Enhancement of Hypoxia-induced Neuronal and Inducible Nitric Oxide Synthases in the Retinas of Rats with Oxygen-induced Retinopathy

Purpose

We investigated whether oxygen-induced retinopathy (OIR) results in changes in the protein expression of neuronal and inducible nitric oxide synthases (nNOS and iNOS, respectively) in rat model of OIR. In addition, we evaluated whether treatment of rats with triamcinolone acetonide (TA) prevents this response.

Methods

To promote OIR, Sprague-Dawley rats were exposed to hyperoxia from postnatal day 2 (P2) to P14. They were then returned to normoxia after P15. TA was injected into the right vitreous of P15 rats, while saline was injected into the left vitreous. At P18 the expression of nNOS and iNOS was determined using Western blotting and immunostaining techniques in retinas obtained from control rats.

Results

In P18 OIR rats, the abundance of nNOS and iNOS protein was significantly increased compared with controls. These increases were not observed in the retinas of rats treated with TA. The change in expression of nNOS and iNOS were specific to parvalbumin and glial fibrillary acidic protein-positive cells. Treatment with TA prevented the increased expression of nNOS and iNOS in all samples.

Conclusions

Hypoxia upregulates expression of nNOS and iNOS in OIR rat retinas, which is can be prevented by treatment with TA.     

内毒素在Lewis鼠诱导的角膜改变——角膜平片的免疫组织化学研究

目的:探讨内毒素全身注射所致的角膜改变。方法:使用抗单核细胞、巨噬细胞和MHC-Ⅱ类抗原阳性细胞的单克隆抗体,用标准的ABC方法于内毒素注射前、后制备的角膜平片上进行免疫组织化学染色。结果:发现在正常角膜中央区实质内有散在分布的巨噬细胞,越近角膜缘分布越密集,MHC-Ⅱ类抗原阳性细胞则仅存在于角膜缘;内毒素注射后,中央区角膜单核巨噬细胞增多,这些细胞于角膜实质内发生了一系列形态学改变,MHC-Ⅱ类抗原阳性细胞仅见于注射后早期的中央区角膜内皮面。结论:内毒素诱导的角膜中巨噬细胞的增多可能是机体应激状态下的重要防御机制,角膜实质中MHC-Ⅱ类抗原的缺如则对维持角膜局部免疫微环境的稳定性有重要意义。眼科学报1996;12:70—74。     

Changes in the choroidal circulation of rabbit following RPE removal

Purpose To examine the effects of surgical removal of the retinal pigment epithelium (RPE) on the choroidal circulation of the rabbit.Methods The retina and choroid were examined by biomicroscopy, scanning laser ophthalmoscopy, fluorescein and indocyanine green angiography and histology at various times after the surgical removal of the RPE by gentle aspiration following a prior local vitrectomy and bleb detachment. Comparison was made between small and large areas of RPE removal, or only slight pressure to the retina without removal of RPE.Results Removal of the RPE layer causes transient leakage of vascular fluid into the subretinal space for at least a week after surgery and loss of perfusion in the underlying choroidal vessels and choriocapillaris at the débridement site. This reduction in local choroidal blood flow can occur within 15 min after RPE débridement and can be transient or permanent. Histology indicates that permanent changes are due to fibroblastic infiltration that compresses the choroidal vessels. Permanent changes tend to occur after removal of relatively large areas of RPE. The removal of small areas of RPE or slight pressure on the retina causes a transient loss of local choroidal perfusion, and fibroblastic infiltration into the choroid does not occur. Conclusion Removal of the RPE causes changes throughout the underlying choroid. It reduces the circulation in the large choroidal vessels as well as the choriocapillaris. If large areas of RPE are removed, this choroidal non-perfusion can be permanent due to fibroblastic infiltration. Small areas of RPE removal or slight pressure on the retina lead to only transient reduction of the choroidal flow. The rapidity with which the choroidal blood flow can be reduced implies a reflex mechanism that responds to sudden RPE pressure and/or trauma. These changes are best observed with ICG angiography.     

视神经损伤后脑衰反应调节蛋白-2表达及其磷酸化修饰的变化及意义

背景 脑衰反应调节蛋白-2(CRMP-2)具有促进中枢神经轴突生长的作用,但中枢神经损伤后在细胞周期素依赖蛋白激酶5(CDK5)诱导下CRMP-2会发生过度磷酸化修饰,从而导致生长锥塌陷,阻碍神经系统的修复.视神经作为一种特殊的中枢神经组织,其损伤后是否发生CRMP-2表达的变化和磷酸化修饰鲜见研究报道. 目的 探讨视神经钳夹伤小鼠模型视神经组织中CRMP-2表达及其磷酸化修饰水平的动态变化及意义.方法 选取8～9周龄健康BALB/c小鼠48只,雌雄不限.采用随机数字表法将小鼠随机分为假手术组和损伤后3、7、14 d组,每组12只.各损伤组小鼠右眼术中暴露视神经,用小号动脉夹于球后2 mm处夹持视神经10s,假手术组小鼠手术操作同各损伤组,但不钳夹视神经.分别于术后3、7、14 d获取小鼠视神经组织,采用实时荧光定量PCR法检测各组小鼠视神经组织中CRMP-2 mRNA的相对表达量;采用Western blot法检测各组小鼠视神经组织中CRMP-2蛋白、磷酸化CRMP-2(p-CRMP-2)及CDK5的表达变化,检测结果进行组间比较.结果 假手术组及损伤后3、7、14d组小鼠视神经组织中CRMP-2 mRNA及蛋白相对表达量的总体比较差异均无统计学意义(CRMP-2 mRNA:F=2.971,P=0.097;CRMP4蛋白:F=1.202,P=0.370).假手术组及损伤后3、7、14 d组小鼠视神经组织中p-CRMP-2蛋白的相对表达量分别为0.001±0.000、0.064±0.003、0.136±0.005和0.346±0.012,CDK5蛋白的相对表达量分别为0.440±0.009、0.723±0.011、0.874±0.015和0.952±0.019,总体比较差异均有统计学意义(p-CRMP-2:F=445.600,P＜0.001;CDK5:F=186.600,P＜0.001),其中损伤后3、7、14 d组小鼠视神经组织中p-CRMP-2和CDK5蛋白的相对表达量均明显高于假手术组,差异均有统计学意义(均P＜0.01).结论 小鼠视神经钳夹伤后视神经组织中CRMP-2的表达无明显变化,但视神经组织中CDK5和p-CRMP-2蛋白表达均明显上调,且随着损伤后时间的延长上调更为明显.     

两种实验白内障晶状体及血清多元素含量的动态变化

研究亚硒酸钠或半乳糖诱发大鼠白内障形成中晶状体和血清的Fe、Al、Zn、Cu、Mg、Ca 和P 含量的动态变化。结果发现,用两种因子诱发白内障后,Ca/P 比值明显升高;在亚硒酸钠诱发的白内障晶状体中Zn 和Ca 含量增加,Zn、Cu、Mg、Ca 和P 元素与半乳糖诱发的白内障发展有密切关系。结果证明,这些元素可能是白内障的继发因子。     

Dysbiosis in the Gut Microbiome in Streptozotocin-Induced Diabetes Rats and Follow-Up During Retinal Changes

PurposeTo analyze the gut bacterial microbiome of streptozotocin-induced diabetic rats and rats with retinal changes.MethodsInduction of diabetes was confirmed by an increase in blood sugar (>150 mg/dL), and the progression of diabetes with retinal changes was assessed by histology and immunohistochemistry of retinal sections. Microbiomes were generated using fecal DNA, and the V3–V4 amplicons were sequenced and analyzed by QIIME and R.ResultsDysbiosis in the gut microbiome of diabetic rats and diabetic rats with retinal changes was observed at the phylum and genus levels compared with the control rats. Heat-map analysis based on the differentially abundant genera indicated that the microbiomes of controls and diabetic rats separated into two distinct clusters. The majority of the microbiomes in diabetic rats with retinal changes also formed a distinct cluster from the control rats. β-diversity analysis separated the microbiome of control rats from the microbiome of diabetic rats and diabetic rats with retinal changes, but the microbiomes of diabetic rats and diabetic rats with retinal changes showed an overlap. Functional analysis indicated that the enhanced inflammation in diabetic rats showing retinal changes could be ascribed to a decrease in anti-inflammatory bacteria and an increase in pathogenic and proinflammatory bacteria.ConclusionsThis study showed that the gut bacterial microbiome in diabetic rats with retinal changes was different compared with control rats. The results could help develop novel therapeutics for diabetics and diabetic individuals with retinal changes.     

Changes in macular pigment optical density and serum concentrations of its constituent carotenoids following supplemental lutein and zeaxanthin: the LUNA study

Macular pigment (MP), consisting of lutein (L) and zeaxanthin (Z), is believed to protect the retina from photo-oxidative damage. The current study investigates, in terms of MP optical density (MPOD) and serum concentrations of its constituent carotenoids, response to supplemental L and Z, and co-antioxidants. An intervention (I) group, consisting of 108 subjects (mean [+/-SD] age: 71.5 [+/-7.1] years), of which 92.6% exhibited features of age-related macular degeneration (AMD), received a daily supplement consisting of 12 mg L and 1 mg Z, both provided as ester 120 mg vitamin C, 17.6 mg vitamin E, 10 mg zinc, 40 microg selenium (Ocuvite Luteintrade mark) for a period of 6 months. MPOD was measured, by 2-wavelength autofluorescence (AF), on five occasions during the period of supplementation, and once again 3 months following discontinuation of the supplement. A control (C) group of 28 subjects (mean [+/-SD] age: 71.0 [+/-8.1] years), who received no dietary supplementation or modification, was examined at baseline and once again after a mean of 29.4 (+/-9.3) weeks. At baseline, mean (+/-SD) MPOD (at 0.5 degrees) was 0.504 (+/-0.197) and 0.525 (+/-0.189) in the I and C groups, respectively. There was a statistically significant increase in MPOD (at 0.5 degrees) for the I group (0.1 [+/-0.009]; p<0.0008), whereas no significant increase was seen in the C group (0.03 [+/-0.02]; p>0.05), over the period of supplementation. In order to classify supplemented subjects into quartiles, in terms of MPOD response, we calculated the difference between MPOD (at 0.5 degrees) at visit 6 and at baseline (visit 1). Quartile 1 (the "non-responder" quartile) displayed no increase in MPOD (at 0.5 degrees), in spite of rises seen in serum concentrations of L and Z. The three "responder" quartiles reached similar final plateaus of MPOD (at 0.5 degrees), reflected in final mean (+/-SEM) values of 0.59 (+/-0.04) optical density unit (ODU), 0.64 (+/-0.03) ODU and 0.64 (+/-0.03) ODU for quartiles 2, 3 and 4, respectively. Subjects with low baseline MPOD were more likely to exhibit a dramatic rise in MPOD, or to exhibit no rise in MPOD, in response to supplements than subjects with medium to high baseline MPOD values. Supplementation with 12 mg L and 1 mg Z, combined with co-antioxidants, resulted in an increase of MPOD at 0.5 degrees eccentricity in a majority of subjects, including those afflicted with AMD. However, there remains a substantial proportion of subjects for whom, in spite of rises in serum concentrations of L and Z in these subjects, MPOD augmentation in response to supplemental L, Z and co-antioxidants could not be detected over the study period, thus indicating that intestinal malabsorption of these carotenoids is not responsible for the lack of a macular response to such supplements. Further, our results suggest that saturable mechanisms play a role in the retinal capture and/or stabilisation of the macular carotenoids.     

TgAPPswePS1转基因鼠视网膜的功能损伤和Aβ斑块沉积

目的：初步研究TgAPPswePS1 转基因鼠眼内阿尔兹海默病(AD)相关的病理变化和视网膜功能改变,探讨淀粉样蛋白β（Aβ）对视网膜结构损伤的作用机制。方法：实验研究。选择月龄为24个月的老龄TgAPPswePS1 转基因小鼠为实验组,同月龄同种系野生型小鼠作为对照组,对2 组小鼠检测视网膜电图（ERG）,再取眼球样本,制备切片。观察视网膜各层结构,用免疫组织化学法检测视网膜切片上的淀粉样前体蛋白(APP)和Aβ表达水平,并观察分布部位及分布形态特点,再测量视网膜厚度。各测量值比较采用Student's t检验进行统计学分析。结果：实验组的Rod的a波和b波波幅与对照组相比均存在下降,其中b波差异存在统计学意义（t=5.23,P=0.002）;实验组Max反应的a波和b波波幅与对照组相比均存在下降,差异均存在统计学意义（t=15.69、15.76,P＜0.001）。对照组和实验组视网膜切片各个层次均完整,未见明显缺失,实验组的厚度较对照组薄,但差异无统计学意义（t=1.85,P=0.087）。在对照组视网膜中,APP存在微弱的背景免疫阳性反应,在实验组视网膜神经节细胞及内颗粒层细胞内呈强阳性反应。Aβ在对照组视网膜中无免疫阳性反应,在实验组视网膜中存在强阳性表达,呈弥散性和斑块状沉积2种形态,斑块主要沉积在神经节细胞层、内丛状层、内核层以及光感受器层。结论：在老龄TgAPPswePS1 转基因小鼠视网膜上存在典型AD病理特征改变,视网膜厚度变薄,同时存在功能改变,提示视网膜上的病理改变可能是AD疾病视功能障碍的基础。