背景光对图形视网膜电图空间调谐特性的影响

本工作应用背景光压抑闪光视网膜电图(FERG),考察对图形视网膜电图(PERG)的空间调谐特性的影响,并与无背景光时的结果进行比较。背景光使全屏幕闪光诱发的FERG基本压抑。在这种条件下,同屏幕的图形刺激所诱发的PERG的振幅显示一定的低空间频率衰减(LSFA)。FFT分析表明,当时间频率为3.91Hz时,PERG的二次谐波呈现十分明显的LS-FA,与无背景光时的结果吻合得很好;当时间频率为7.81Hz时,也表现出LSFA,而无背景光时则缺如。这些结果表明,无背景光时记录的PERG振幅在低空间频率区偏高确实是由于混杂有亮度特异性成分—FERG。但是,PERG振幅所显示的LSFA仍不如二次谐波那么明显,这可能是因为FERG中存在的非线性成分未为背景光所完全压抑,仍然混杂在低频区的PERG反应中。     

正常有色素家兔眼的视网膜功能研究

目的通过分析正常有色素家兔的mfERG的特征,了解其视网膜的功能特性。方法对9只兔18眼正常有色素家兔同时检测mfERG,并与正常成人的18只眼的检测结果对照分析。结果正常有色素家兔mfERG的P1波振幅密度三维地形图特点,可见黄斑区高峰,但较人黄斑区高峰为低,其他部分呈颞侧高鼻侧低的斜坡状,象限野mfERG颞侧半可见P1波振幅密度、P1波振幅、N1波振幅比鼻侧增高;也高于正常成人眼的颞侧值。五个环形野mfERG,P1波振幅密度,第1环最大,由1环到5环依次递减,P1和N1波振幅以4环和5环最大;与人相比环1的P1波振幅密度和振幅降低,环4和环5增高。结论兔眼颞侧视网膜功能高于鼻侧。兔眼视网膜存在类人黄斑区,但其黄斑中心视功能不如人眼,中心外视功能高于人眼。     

正常人和某些病变性眼的平均视网膜电图及振荡电位

本工作用全视野闪光刺激器结合微处理机平均技术对人视网膜电图(ERG)进行了记录和分析。正常人的 ERG 主要由 a 波、b 波和振荡电位组成。正常人的振荡电位的峰值时间及其间隔相当恒定,但在糖尿病性视网膜病和视神经萎缩病人,振荡电位减小或消失。本文对这种改变的可能原因进行了讨论。     

CERKL的过表达在人类视网膜变性疾病中的作用及其临床意义

目的:研究基因CERKL过度表达与色素性视网膜炎的关系及其临床治疗意义。方法:应用RT-PCR分析、比较基因序列CERKL的转录产物的结构差异。利用序列CERKL过表达的转化与细胞溶解,获得不同的CERKL蛋白,检测其体外磷酸化酶活性。结果:CERKL转录得到四种转录子,两个较长的CERKLa、CERKLb,两个较短的CERKLc、CERKLd。四种转录蛋白在体外酶活性实验中均没有产生酶解产物C1P,在300μM H2O2氧化条件下,转录子CERKL能有效抑制细胞凋亡,其24 h相对PARP清除率明显低于12 h相对PARP清除率(P0.01)。但在400μM H2O2氧化条件下其未能表现出抑制细胞凋亡的作用。结论:色素性视网膜炎有可能是基因CERKL转录时其终止子因无义突变导致缺陷,进而发生选择性转录,并由CERKL较长变异体(CERKLa、CERKLb)的缺失或CERKL较短变异体(CERKLc、CERKLd)毒性引起的。     

光化学反应诱导大鼠视网膜水肿模型

目的:通过静脉注射光敏剂Erythrosin B联合激光照射诱导大鼠视网膜水肿模型,并观察该模型中大鼠视网膜及血管形态改变.方法:大鼠尾静脉注射光敏剂Erythrosin B后,使用532nm Nd:YAG激光(1.95±0.05 mw)照射大鼠视网膜8分钟.分别于建模后第1、2、3、5、7及14天进行眼底照相、眼底荧光造影(FFA)及视网膜光学相干断层扫描(OCT)检查.并在OCT图像上测量大鼠视网膜厚度(RT).结果:眼底照相、FFA及OCT结果显示大鼠视网膜在激光照射后可立即出现血管损伤及视网膜厚度增加,但未发现视网膜血管栓塞.激光照射前RT为219±2 μm,激光照射后第1天RT即增加至283±6μm,并在第2天到达顶峰(302±7μm),之后开始下降,至第5天恢复到激光照射前水平(234±9 μm),到第14天时视网膜发生明显萎缩,RT较激光处理前减小(198±6μm).结论:这一通过光化学反应诱导的视网膜水肿模型具有可靠及可重复性,可被运用于视网膜水肿的病理学及动力学研究.     

一种视网膜锥体细胞退行性变大鼠基础生理值的测定

目的对视网膜锥体细胞退行性变（retinal cone degeneration,RCD）大鼠的主要脏器及血液、生化指标进行检测。方法随机选择BN（Brown Norway）-RCDF2代大鼠,视网膜电图鉴定,取静脉血检测生化指标,并对重要脏器进行准确称量。结果此大鼠脏器质量在雌雄之间除体重、大脑和肾差异有显著性外（P〈0.05）,其余均无差异;脏器系数脑干和小脑雌性显著大于雄性（P〈0.01）;生化指标碱性磷酸酶、葡萄糖、白蛋白、尿素氮、总蛋白雌性高于雄性,总胆固醇、天冬氨酸转氨酶、丙氨酸转氨酶、甘油三酯雌性低于雄性,其中雌性碱性磷酸酶活性显著高于雄性;雌性离子钙浓度显著高于雄性。结论BN-RCD大鼠基础生理值和生化参数均在正常范围内,其中一些参数在性别之间有差异,但与正常SD大鼠相比,天冬氨酸转氨酶、丙氨酸转氨酶、肌酐、钾离子差异显著,葡萄糖、甘油三酯、离子钙有差异。RCD大鼠为遗传性视网膜疾病的研究提供了一种新的突变模式动物。     

视紫红质及RPE65蛋白在光致视网膜变性疾病中的作用机制

视紫红质是感光细胞中的一种视色素,在光线的接收和视觉电位的产生方面具有重要的生理作用,由视紫红质介导的过度光信号传导是光性视网膜变性的主要原因。近年的研究表明,视网膜色素上皮细胞中的RPE65蛋白作为影响视紫红质再生的关键因素,与视网膜光损伤的易感性密切相关。就视紫红质和RPE65蛋白在光致视网膜变性中的作用机理作一探讨。     

糖尿病性视网膜病变早期mfERG的研究

目的:探讨糖尿病患者糖尿病性视网膜病变早期多焦视网膜电图(multifocal electroretinogram,mfERG)的特点。方法:应用德国ROLAND公司RETIscan系统对24例40眼糖尿病患者,同时对27例40眼正常对照眼记录mfERG。按机器的默认值一阶反应进行记录。整个记录过程中可以分为12个时段,每个时段记录为时间47秒。结果:mERG测得的糖尿病患者组1～5环所在位置的P1波的振幅密度随离心度的增加而降低,P1波和N1波幅值降低和潜伏时延迟。各象限的P1、N1波振幅密度、波幅值降低,潜伏时延迟。结论:糖尿病性视网膜病变受累的区域多发生在黄斑部,功能上的改变早于形态学的变化;病变的早期功能改变最先出现于视网膜的后极偏颞上方;糖尿病患者在未出现眼底改变时各个象限视网膜的外层已经严重受累。     

正常猕猴与人的视网膜电图比较

目的比较正常猕猴与人视网膜电图异同,为进一步利用猕猴建立动物模型研究视网膜疾病打下基础。方法健康成年猕猴7只（14只眼）与8例（16只眼）正常人进行视网膜电图检测,对两者Rod-ERG中的b波,Max-ERG的a、b波,Cone-ERG的b波峰时值及波幅和OPs的O2值,Flicker-ERG的P值进行统计学检验。结果猕猴与人的视网膜电图波形结果较为相似,各指标与人的结果相比,潜伏期短,幅值低,但Cone-ERG和Flicker-ERG两者幅值差异不具有统计学意义。结论视网膜电图检测从功能上证明猕猴较其他常用实验动物更接近人,尤其表现在视锥细胞及黄斑区功能,可用作人类视网膜疾病尤其是黄斑区病变的良好动物模型。     

多能干细胞分化来源视网膜色素上皮细胞移植治疗视网膜变性研究进展

视网膜色素上皮（RPE）对视觉功能的维持起着至关重要的作用。视网膜变性是全球不可治愈性致盲疾病的重要原因,它由视网膜色素上皮功能失常所引起。因此,视网膜色素上皮移植是视网膜变性患者恢复视力的一种最有前景的手段之一。随着干细胞技术的快速发展,从多能干细胞（PSC）到有功能的视网膜色素上皮细胞的体外分化诱导技术已经成熟,其中包括胚胎干细胞（ESCs）和诱导多能干细胞（iPSCs）等。此外,从患者特异性iPSCs分化而来的RPE更能用于阐明发病机理并有针对性地个体治疗。更值得一提的是,经诱导得到RPE的移植不论在动物模型中,还是在临床试验里都已经得到了可喜的治疗效果。本文回顾PSC来源RPE干预治疗视网膜变性的最新研究进展。     

SD大鼠与Wistar大鼠脑胶质瘤动物模型的建立及比较

目的比较利用SD大鼠、Wistar大鼠建立脑胶质瘤动物模型的不同,为研究脑胶质瘤的发病机制及治疗方法提供操作平台。方法利用立体定向仪建立SD大鼠、Wistar大鼠大脑皮层接种C6细胞（2.5×105个细胞/只）,建立脑胶质瘤动物模型,利用组织病理学、免疫组织化学以及核磁共振成像等技术,比较两种动物模型在成瘤率、肿瘤生长状况、死亡率以及动物一般情况等方面的异同。结果SD大鼠组、Wistar大鼠组的成瘤率均为100%,两组均未见转移;但SD大鼠组肿瘤成瘤时间较长,且部分肿瘤有自愈倾向,而Wistar大鼠组则未出现类似情况。结论Wistar大鼠大脑皮层脑胶质瘤动物模型的肿瘤性状更接近于人的脑胶质瘤,因此更适合探索和研究脑胶质瘤的发病机制和治疗方法;而SD大鼠的肿瘤由于性状类似转移瘤,且有自愈倾向,不适合作为上述相关研究的动物模型。     

玻璃体腔注射对裸小鼠视网膜组织形态学的影响

目的观察单纯的玻璃体腔注射对裸小鼠视网膜组织形态学的影响,为建立简单的制作视神经损伤动物模型奠定实验基础。方法在全身麻醉配合眼部局部麻醉情况下,利用微量注射器往裸小鼠玻璃体腔内迅速注入10μL生理盐水,然后在不同的时间点取注射眼进行固定、切片和HE染色,观察视网膜特别是视神经节细胞的变化。结果正常对照组视网膜层次清晰,各层排列整齐而致密,视网膜神经节细胞呈单层排列,大小不一,染色质分布均匀。实验组于注射后第1天、第3天和第5天视网膜神经节细胞减少的情况不明显,十层结构仍相对清晰。但于第7天,视网膜神经节细胞出现细胞明显缺失的现象,第14天为最严重,第30天和第60天与第14天相比无明显差别。结论玻璃体腔注射过量的生理盐水能够损伤视网膜组织,造成神经节细胞减少,有可能成为一种简单的制作视神经损伤动物模型的方法。     

Correlations between Photodegradation of Bisretinoid Constituents of Retina and Dicarbonyl Adduct Deposition

Non-enzymatic collagen cross-linking and carbonyl adduct deposition are features of Bruch''s membrane aging in the eye, and disturbances in extracellular matrix turnover are considered to contribute to Bruch''s membrane thickening. Because bisretinoid constituents of the lipofuscin of retinal pigment epithelial (RPE) cells are known to photodegrade to mixtures of aldehyde-bearing fragments and small dicarbonyls (glyoxal (GO) and methylglyoxal (MG)), we investigated RPE lipofuscin as a source of the reactive species that covalently modify protein side chains. Abca4^−/− and Rdh8^−/−/Abca4^−/− mice that are models of accelerated bisretinoid formation were studied and pre-exposure of mice to 430 nm light enriched for dicarbonyl release by bisretinoid photodegradation. MG protein adducts were elevated in posterior eyecups of mutant mice, whereas carbonylation of an RPE-specific protein was observed in Abca4^−/− but not in wild-type mice under the same conditions. Immunolabeling of cryostat-sectioned eyes harvested from Abca4^−/− mice revealed that carbonyl adduct deposition in Bruch''s membrane was accentuated. Cell-based assays corroborated these findings in mice. Moreover, the receptor for advanced glycation end products that recognizes MG and GO adducts and glyoxylase 1 that metabolizes MG and GO were up-regulated in Abca4^−/− mice. Additionally, in acellular assays, peptides were cross-linked in the presence of A2E (adduct of two vitamin A aldehyde and ethanolamine) photodegradation products, and in a zymography assay, reaction of collagen IV with products of A2E photodegradation resulted in reduced cleavage by the matrix metalloproteinases MMP2 and MMP9. In conclusion, these mechanistic studies demonstrate a link between the photodegradation of RPE bisretinoid fluorophores and aging changes in underlying Bruch''s membrane that can confer risk of age-related macular degeneration.     

HFJ和Wistar大鼠脂肪肝及胰岛素抵抗动物模型的比较

目的建立近交系HFJ大鼠和封闭群Wistar大鼠脂肪肝胰岛素抵抗动物模型及比较其生物学特性。方法雄性HFJ大鼠和Wistar大鼠,分别随机分为模型组和正常组,模型组喂养高脂饮食,正常组喂养普通饮食,两组脂肪分别占摄入能量的44.2%和19.2%,共饲养12周。每周称体质量,测定血糖、甘油三酯、胆固醇、ALT、AST、HDLC、LDLC和血胰岛素水平。实验期末处死动物摘取肝脏并称质量,计算肝指数;鼠肝脏用10%甲醛固定,石蜡包埋切片,HE染色,光镜下评估肝脂肪变性和炎症活动情况。结果镜下可见,HFJ和Wistar大鼠模型组肝细胞均呈现弥漫性脂肪变性,小叶内可见炎症细胞浸润,HFJ比Wistar脂肪变性较重,对照组肝脏均未见异常。两种动物的模型组ALT、AST、肝指数、HOMA-IR指数均显著高于其正常组,HFJ和Wistar种系间各指标的差异无显著性;HFJ大鼠模型组体重和正常组体重具有显著性差异(P<0.01),而Wistar大鼠模型组体重与正常组体重间无显著差异(P>0.05);HFJ大鼠TG和TC含量均显著高于Wistar大鼠。结论通过高脂饮食喂养成功建立了HFJ大鼠和Wistar大鼠脂肪肝与胰岛素抵抗疾病动物模型,与Wistar大鼠比较,HFJ大鼠具有自发性高血脂特征,造模更易成功,可为胰岛素抵抗和脂肪肝的发病机制研究、防治高血脂药物筛选提供一种新的实验动物。     

Influence of Toxoplasma gondii Acute Infection on Cholinesterase Activities of Wistar Rats

Several studies have shown the mechanisms and importance of immune responses against Toxoplasma gondii infection and the notable role of cholinesterases in inflammatory reactions. However, the association between those factors has not yet been investigated. Therefore, the aim of this study was to evaluate the acetylcholinesterase (AChE) activity in blood and lymphocytes and the activity of butyrylcholinesterase (BChE) in serum of rats experimentally infected with T. gondii during the acute phase of infection. For that, an in vivo study was performed with evaluations of AChE and BChE activities on days 5 and 10 post-infection (PI). The activity of AChE in blood was increased on day 5 PI, while in lymphocytes its activity was enhanced on days 5 and 10 PI (P<0.05). No significant difference was observed between groups regarding to the activity of BChE in serum. A positive (P<0.01) correlation was observed between AChE activity and number of lymphocytes. The role of AChE as an inflammatory marker is well known in different pathologies; thus, our results lead to the hypothesis that AChE has an important role in modulation of early immune responses against T. gondii infection.     

几种椎间盘退变动物模型的建立和应用

椎间盘退变是一种年龄相关的退行性疾病,是引起下腰痛的主要因素,严重影响病人的生活质量,并显著增加家庭的经济负担。目前,缺少椎间盘退变的有效干预和治疗手段,部分原因是其发病机制尚未阐明。椎间盘退变动物模型的构建对于阐明该疾病的病理机制至关重要。椎间盘退变是一个复杂的过程,受机械应力、结构损伤、生物化学与基因表达等多种因素的影响。本文总结了应用异常机械应力、结构损伤、生物化学或化学诱导和基因敲除等方式构建的椎间盘退变动物模型。生物力学是维持椎间盘稳态的重要因素,异常的机械应力会导致椎间盘退变。同时,椎间盘退变常伴随结构性损伤,椎间盘结构破坏也会导致椎间盘发生退变。此外,生物化学或化学诱导和关键基因敲除也会导致椎间盘退变。本文按照造成异常机械应力的因素将机械应力模型分为加压模型和失稳模型;按照椎间盘结构将结构损伤模型分为髓核与纤维环损伤模型和软骨终板损伤模型。总结了生物化学或化学诱导模型以及新型的基因敲除模型。讨论了不同类型椎间盘退变动物模型的可能应用和局限性。     

Trans-Corneal Subretinal Injection in Mice and Its Effect on the Function and Morphology of the Retina

Purpose

To introduce a practical method of subretinal injection in mice and evaluate injection-induced retinal detachment (RD) and damage using a dynamic imaging system, electrophysiology, and histology.

Methods

After full dilation of a 2-month-old C57BL/6J mouse pupil, the cornea near the limbus was punctured with a 30 ½-gague disposable beveled needle. A 33 ½-gauge blunt needle was inserted through the corneal perforation into the anterior chamber, avoiding the lens before going deeper into the vitreous cavity, and penetrating the inner retina to reach the subretinal space. The mice were divided into four groups: in group 1, about 80–100% of the retina was filled with subretinally injected solution; in group 2, approximately 50–70% of the retina was filled with injected solution; in group 3, the procedures were stopped before solution injection; and non-injected eyes were used as the negative control in group 4. An optical coherence tomography (OCT) imaging system was used to monitor retinal reattachment during the first three days following the injections. Histological and functional changes were examined by light microscopy and electroretinography (ERG) at five weeks post-injection.

Results

After a short-term training, a 70% success rate with 50% or more coverage (i.e., retinal blebs occupied 50% or more retinal area and filled with the injected solution) with minimal injection-related damages can be achieved. Bleb formation was associated with retinal detachment (RD) between the neuroretina and the retinal pigment epithelium (RPE) layer. Partial RD could be observed at post-injection day 1, and by day 2 most of the retina had reattached. At 5 weeks post-injection, compared to uninjected control group 4, the b-wave amplitudes of ERG decreased 22% in group 1, 16% in group 2, and 7% in group 3; the b-wave amplitudes were statistically different between the uninjected group and the groups with either 50–70% or 80–100% coverage. The subretinal injection-induced RD reattached and became stable at five weeks post-injection, although some photoreceptor damage could still be observed in and around the injection sites, especially in 80–100% coverage group.

Conclusions

Trans-corneal subretinal injection is effective and practical, although subretinal injection-related damages can cause some morphological and functional loss.     

Assessment of Cardiac Function and Myocardial Morphology Using Small Animal Look-locker Inversion Recovery (SALLI) MRI in Rats

Small animal magnetic resonance imaging is an important tool to study cardiac function and changes in myocardial tissue. The high heart rates of small animals (200 to 600 beats/min) have previously limited the role of CMR imaging. Small animal Look-Locker inversion recovery (SALLI) is a T1 mapping sequence for small animals to overcome this problem ¹. T1 maps provide quantitative information about tissue alterations and contrast agent kinetics. It is also possible to detect diffuse myocardial processes such as interstitial fibrosis or edema ^1-6. Furthermore, from a single set of image data, it is possible to examine heart function and myocardial scarring by generating cine and inversion recovery-prepared late gadolinium enhancement-type MR images ¹.The presented video shows step-by-step the procedures to perform small animal CMR imaging. Here it is presented with a healthy Sprague-Dawley rat, however naturally it can be extended to different cardiac small animal models.     

兔眼孔源性视网膜脱离手术复位后视功能以及超微结构的变化

目的对比研究兔眼视网膜脱离后选择不同时期手术复位视功能的变化情况,为临床手术时机的选择及预测术后视功能的恢复情况提供理论与实验依据。方法利用家兔制备孔源性视网膜脱离模型,成模后1 d、7 d1、4 d时经手术达解剖复位,采用多焦视网膜电流图检测复位后视网膜的功能,数据处理应用SPSS软件。制备组织病理学切片。结果多焦视网膜电流图显示1 d、7 d、14 d的RRD手术复位后P1波平均象限反应密度(QAP1,nV/deg2),P1波幅值(AP1,μV),N1波幅值(AN1,μV),P1波潜伏期(TP1,ms),N1波潜伏期(TN1,ms)各项数值差异有显著性(P<0.05)。光镜电镜显示视网膜脱离复位后组织病理学改变。结论视网膜脱离手术复位后视网膜细胞功能的恢复与脱离时间有明显的负相关性;多焦视网膜电流图对于局部视网膜功能的评价有重要意义;组织病理学研究提供了解释视功能变化的重要依据。     

Changes in the Redox State in the Retina and Brain During the Onset of Diabetes in Rats

Diabetic retinopathy is thought to result from chronic changes in the metabolic pathways of the retina. Hyperglycemia leads to increased intracellular glucose concentrations, alterations in glucose degradation and an increase in lactate/pyruvate ratio. We measured lactate content in retina and other ocular and non-ocular tissues from normal and diabetic rats in the early stages of streptozotocin-induced diabetes. The intracellular redox state was calculated from the cytoplasmic [lactate]/[pyruvate] ratio.Elevated lactate concentration were found in retina and cerebral cortex from diabetic rats. These concentrations led to a significant and progressive decrease in the NAD⁺/NADH ratio, suggesting that altered glucose metabolism is an initial step of retinopathy. It is thus possible that tissues such as cerebral cortex have mechanisms that prevent the damaging effect of lactate produced by hyperglycemia and/or alterations of the intracellular redox state