出生后10—90天大鼠附睾管上皮主细胞超微结构的观察

本文应用透射电镜观察了生后第10至90天大鼠附睾管上皮主细胞的超微结构。结果表明:第10天上皮细胞处于未分化状态,胞浆内充满了大量游离核糖体,其它细胞器均少见;第19天上皮细胞已开始分化,游离缘出现少量短粗微绒毛和微吞饮内褶,Golgi复合体渐发达且数量增加;第50天上皮细胞已具成年大鼠主细胞的超微结构,微绒毛发达,微吞饮内褶和多泡体丰富,Golgi复合体发达。本文对生后大鼠附睾管主细胞出现上述超微结构变化的意义进行了讨论。     

自发性高血压大鼠视网膜色素上皮超微结构

目的：观察高血压大鼠视网膜色素上皮的超微结构,探讨其与神经视网膜病变的关系。方法：用透射电镜观察５、６、７月龄自发性高血压大鼠及同月龄京都种大鼠视网膜色素上皮细胞超微结构,结果：高血压大鼠视网膜色素线粒体肿胀,内质网扩张,基底部皱褶减少,微绒毛变稀疏,色素颗粒减少,这些退行性改变随鼠龄增大血压的增高而加重。结论：Ｄ 谪血压视网膜病变的早期视网膜色素上皮细胞已有缺血、缺氧所致的退行性改变,提示早期的     

灯盏花素对人视网膜色素上皮细胞和糖尿病大鼠视网膜VEGF表达的影响

目的:观察灯盏花素对高糖环境下人视网膜色素上皮细胞株ARPE-19中VEGF、VEGFR2和pERK蛋白的表达及对2型糖尿病大鼠视网膜VEGF表达的影响。方法:培养ARPE-19细胞,分为对照组、灯盏花素组、高糖组和高糖+灯盏花素组,ELISA检测VEGF的分泌水平,Western blot检测细胞VEGF、VEGFR2和p-ERK的蛋白水平;取正常大鼠,随机分为正常对照组、灯盏花素组、糖尿病模型组和糖尿病灯盏花素治疗组,16周后取各组大鼠眼球,观察大鼠视网膜的病理变化并评分,免疫组化观察VEGF的表达情况。结果:(1)细胞实验结果显示,灯盏花素组VEGF、VEGFR2和p-ERK的蛋白水平与正常对照组比较均减低(P0.05);高糖组细胞上述蛋白水平与正常对照组比较均增加(P0.05);高糖+灯盏花素组上述蛋白水平与高糖组比较表达均减少(P0.05);(2)大鼠视网膜病理学评分显示糖尿病组与正常对照组、糖尿病灯盏花素治疗组相比差异显著,灯盏花素组与正常对照组相比差异无统计学意义。糖尿病大鼠与正常对照组、灯盏花素组相比,视网膜VEGF的表达增加;糖尿病灯盏花素治疗组中VEGF的表达较糖尿病组有所降低(P0.05)。结论:灯盏花素可以降低ARPE-19细胞VEGF、VEGFR2和p-ERK的蛋白表达水平,抑制糖尿病大鼠视网膜VEGF的表达,提示灯盏花素可能通过降低糖尿病大鼠视网膜细胞和组织中VEGF的表达,缓解其糖尿病视网膜病变进程。     

糖尿病大鼠视网膜视细胞超微结构病理变化的研究

目的:探讨糖尿病大鼠视网膜病变（diabetic retinopathy.DR.简称糖网病）,早期视细胞（视感受器）的超微结构病变,为糖网病光感受器功能障碍提供形态学依据。方法:选择清洁级SD大鼠随机分成正常对照组（CON）、糖尿病1月（DM1）、3月（DM3）和6月（DM6）。按60mg/kg腹腔内注射链脉佐菌素（STZ）诱导大鼠糖尿病,每组12只,取大鼠视网膜透射电镜观察。处死前每月测血糖和眼底镜检查1次。结果:病程1个月:视杆、视锥细胞的外节膜盘模糊不清,膜盘间隙略有扩大,椭圆体内中心粒和纤毛仍清晰可见,线粒体规则的排列在周围。病程3个月,视细胞外节膜盘间隙明显扩大,纵横交错,排列紊乱,并发生局灶性断裂,椭圆体内有部分线粒体肿胀、脱嵴。毛细血管扩张,神经纤维中心微管也有水肿。病程6个月:视细胞外节内膜盘断裂溃变,颜色淡,并出现许多泡沫状结构,椭圆体内的线粒体变小,排列不规则,有的脱嵴、水肿,毛细血管已有部分由扩张转为狭窄。结论:糖尿病早期毛细血管由局部扩张转为狭窄,视网膜因缺血、缺氧的加重导致视细胞超微结构的病理改变,并随病程的进展而加重。     

糖尿病早期大鼠视网膜毛细血管及视细胞超微结构变化规律

目的：探讨大糖尿病视网膜病变（糖网病）早期的视网膜毛细血管及视细胞的病变规律。材料与方法：选择健康成年Wistar大鼠,随机分成正常对照组,糖尿病1月,3月和6月组,一次性腹腔内注射链脲佐菌素STZ）诱导大鼠糖尿病,制备视网膜超薄切片,透镜观察并计算机图像分析。结果：病程1月,周细胞和内皮细胞核异染色质聚集靠边;视杆膜盘间隙略扩大。3月,毛细血管细胞异染色质聚集严重,基底膜增厚;膜盘间隙扩大明显,局灶性断裂,溶解。视杆细胞膜固缩,异染色质浓集,视杆内节线粒体肿胀,甚至空泡变性,病程6月,以上改变更加严重,毛细血管狭窄,变形,甚至闭塞。结论：糖网病早期大鼠视 膜毛细血管病变的同时,变出现视感受器超微结构的改变,随病程的进展病变逐渐加重。     

人视网膜色素上皮细胞诱导型一氧化氮合酶和活性氧在高糖状态下的表达

背景：高血糖导致的自由基损伤是糖尿病视网膜病变发病机制的中心环节。 目的：观察高糖对体外培养的人视网膜色素上皮细胞的氧化损伤作用以及高糖对人视网膜色素上皮细胞诱导型一氧化氮合酶和活性氧表达的影响。 方法：将培养人视网膜色素上皮细胞,分为对照组、高糖组和甘露醇组,分别用含5.5 mmol/L葡萄糖,33 mmol/L葡萄糖及5.5 mmol/L葡萄糖和27.5 mmol/L甘露醇的DMEM培养液培养。采用相差倒置显微镜观察细胞生长形态,采用免疫荧光染色研究诱导型一氧化氮合酶和3-硝基酪氨酸蛋白表达的变化,用氯甲基二氯二氢荧光素二乙酯荧光染色检测视网膜色素上皮细胞中活性氧的产生量。 结果与结论：与对照组相比,应用含33 mmol/L葡萄糖的DMEM培养基处理视网膜色素上皮细胞48 h可见细胞胞体变薄,形态表现多样,不规则细胞增多;高糖培养的视网膜色素上皮细胞诱导型一氧化氮合酶和3-硝基酪氨酸蛋白表达增加,活性氧产生明显增多。说明高浓度葡萄糖培养可造成人视网膜色素上皮细胞氧化损伤,使细胞形态发生变化,并导致细胞中3-硝基酪氨酸产生增多。     

慢性镉中毒小鼠肾脏的超微结构观察

透射电镜下观察10只慢性镉中毒小鼠肾脏的超微结构变化，结果表明肾小体和近曲小管上皮细胞的超微结构均受损伤，肾小体，毛细血管内皮细胞肿胀，增厚，血管系膜增生，血管球基膜增厚或分裂成层，基膜和内皮细胞之间或基膜的分裂之间有电子致密物沉积，近曲小管上皮细胞，胞核变形，固缩，线粒体肿胀，嵴断裂，脱落，细胞游离面微绒毛肿胀，变形，排列紊乱，细胞基底面质膜内褶减少或消失，这些超微结构的改变与某些类型肾小球肾炎的病理特征相似。     

糖尿病患者角结膜上皮稳定性的改变

目的了解糖尿病患者病程、血糖控制状况以及糖尿病视网膜病变与角结膜上皮改变的关系。方法选择60例（120眼）2型糖尿病患者作为病例组，54名（108眼）非糖尿病患者作为对照者，性别年龄成组匹配。分别进行角膜知觉检查、角膜荧光素染色（FL）、结膜印迹细胞学检查（CIC）检查、糖尿病眼底检查、眼底照相或荧光造影，观察杯状细胞及结膜化生情况。结果糖尿病组角膜知觉下降（P〈0．05），角膜荧光素染色阳性增加（P〈0．05），杯状细胞丢失，结膜上皮细胞鳞状化生级别增加。糖尿病病程、血糖控制状况以及糖尿病视网膜病变严重程度与结膜上皮细胞鳞状化生级别、杯状细胞密度和角膜FL累积分间有显著的相关性（均P〈0．01）。结论糖尿病患者角结膜上皮的损害，且糖尿病病程长、血糖控制状况差和糖尿病视网膜病变严重者，损害程度亦重。     

糖网病发病机理的研究进展

糖尿病视网膜病变是致盲的常见原因 ,因而最引人注意。多年来 ,国内外学者对糖网病进行了大量实验性和应用性研究 ,取得了一定的进展。本文在简要回顾糖网病病理变化的基础上 ,对其发病机制 ,诸如高血糖与糖网病、糖网病与糖代谢异常的学说以及血管生成因子的作用等作一综述。     

糖尿病早期大鼠视网膜视细胞超微结构改变与醛糖还原酶的相关性及氨基胍的保护作用

目的 :探讨糖尿病早期大鼠视网膜视细胞超微结构改变与醛糖还原酶 (Aldosereductase ,AR)含量变化的相关性及氨基胍 (Aminoguanidine,AG)的保护作用。方法 :选择健康成年雄性SD大鼠 ,随机分成正常对照组、糖尿病组和糖尿病AG治疗组。一次性腹腔注射链脲佐菌素 (Streptozotin ,STZ)诱发糖尿病模型 ,分别于第 3 0天和第 90天测定视网膜组织AR含量 ;并在第 90天时取视网膜行透射电镜观察视细胞超微结构。结果 :糖尿病早期大鼠视网膜AR含量随糖尿病时间的增长而增多 ;糖尿病 90天大鼠视网膜的视细胞外节膜盘排列紊乱 ,间隙扩大 ,内节线粒体水肿 ,有的空泡化 ;AG治疗组大鼠视网膜AR含量明显低于糖尿病未治疗组 ,糖尿病 90天大鼠视网膜视细胞外节膜盘少量排列紊乱 ,内节线粒体正常。结论 :糖尿病早期视网膜视细胞超微结构改变与AR含量变化有关 ,AG可通过抑制AR活性而保护视细胞     

rAAV2-NTF2对糖尿病大鼠血视网膜屏障的保护作用

目的： 观察2型重组腺相关病毒－核运输因子2（rAAV2-NTF2）对糖尿病大鼠血视网膜屏障功能损伤的拮抗作用。方法: 伊凡思蓝（EB）灌注铺片观察糖尿病大鼠视网膜血管分布和形态。亚克隆构建pSNAV－NTF2载体, 腺相关病毒包装形成rAAV2-NTF2。成年雄性Wistar大鼠,以右眼为实验眼,玻璃体腔注射4 μL rAAV2-NTF2,左眼为对照眼,玻璃体腔注射rAAV2-增强型绿色荧光蛋白（EGFP）4 μL。1个月后取4只大鼠左眼,固定后取视网膜制备全视网膜铺片,荧光显微镜下观察EGFP表达情况。再行STZ腹腔注射诱导糖尿病,同时设单纯糖尿病组（diabetes,D组）和正常组（normal,N组）。糖尿病1个月后以45 mg/kg剂量静脉注射EB,循环2 h后1%多聚甲醛枸橼酸缓冲液灌注并摘除眼球取视网膜,置于甲酰胺中浸泡提取染料,用分光光度计测量甲酰胺中染料的浓度。结果: 正常组大鼠可见在低背景荧光下视网膜血管对EB有很好的屏障作用,糖尿病2周大鼠视网膜血管仅见背景荧光增强,糖尿病1月后血管出现异常节段性扩张,局部血管周围EB渗漏。糖尿病1个月大鼠视网膜内EB浓度是正常组的4.67倍,P<0.01,提示糖尿病大鼠血-视网膜屏障受损。玻璃体腔注射rAAV2-NTF2,视网膜内EB浓度为(8.30±4.16)μg/g视网膜干重,较对侧眼(22.13±8.43)μg/g视网膜干重明显减少,可以部分改善血-视网膜屏障的破坏,降低EB-白蛋白渗漏（P<0.05）。结论: 玻璃体腔注射rAAV2-NTF2可减轻糖尿病大鼠视网膜血-视网膜屏障功能的破坏,为糖尿病视网膜病变的基因治疗提供了实验依据。     

核运输因子2在糖尿病大鼠视网膜的时空表达及其意义

目的:探讨糖尿病大鼠视网膜中核运输因子2(NTF2)的时空表达变化及意义。方法:伊凡思蓝(EB)灌注铺片观察糖尿病大鼠视网膜血管分布和形态。逆转录-聚合酶链式反应(RT-PCR)检测与糖尿病大鼠不同时点鼠龄匹配的对照组(N2w,N1m,N3m,N6m)和糖尿病成模后2周、1月、3月、6月(D2w,D1m,D3m,D6m)大鼠视网膜中NTF2、血管内皮生长因子(VEGF)mRNA的表达。免疫组化法检测NTF2蛋白在视网膜中的表达和分布位置。结果:正常组大鼠可见在低背景荧光下视网膜血管对EB有很好的屏障作用,成模1月后糖尿病大鼠视网膜血管仅见背景荧光增强,成模3月后血管出现异常节段性扩张,局部血管周围EB渗漏。与糖尿病大鼠年龄匹配的正常大鼠视网膜NTF2和VEGF并未随时间延长而变化,mRNA表达稳定。糖尿病成模2周后开始,NTF2mRNA有轻度增高,并随病程的延长保持较高水平,病程达6个月时,NTF2表达开始回落,与正常大鼠基本一致。NTF2蛋白在正常大鼠及糖尿病大鼠视网膜中免疫组化均可以检出,主要分布在视网膜内层,以节细胞层、内核层为主。结论:NTF2mRNA水平在糖尿病大鼠视网膜中升高,主要分布在视网膜内层。NTF2很可能在糖尿病视网膜病变中起着一定的调控作用,其作用途径及机制可能与VEGF的作用通路有一定的联系。     

黄腐酸钠对糖尿病大鼠视网膜病变的作用

目的 :探讨黄腐酸钠对糖尿病大鼠视网膜病变 (DR)的治疗作用及其机制。方法 :对佐菌素诱导的糖尿病大鼠早期给予 0 .5 %黄腐酸钠 3 0mg/kg皮下注射 ,1次 /日 ,共 6个月。利用光镜和透射电镜观察其视网膜组织的形态改变 ,同时检测血浆纤溶酶原激活剂 (t PA)、纤溶酶原激活剂抑制物 (PAI 1)活性及循环血中粒细胞表面抗原CD11a、CD11b。结果 :(1)黄腐酸钠治疗组视网膜组织光镜下及超微结构变化较病变组大鼠有明显改善 ,毛细血管基底膜的增厚明显减轻 (P <0 .0 1)。 (2 )治疗组t PA、PAI 1活性的改变较病变组得以纠正 (P <0 .0 1)而接近于正常对照组水平 (P <0 .0 5 )。基底膜厚度与血浆t PA活性呈显著负相关。 (3 )治疗组CD11a、CD11b的表达受到抑制 (P <0 .0 5 )。结论 :黄腐酸钠对DR的进展有一定的抑制作用。此作用可能与维持血浆t PA、PAI 1平衡而改善纤溶活性 ,减少白细胞粘附有关。     

Cellular and subcellular expression of monocarboxylate transporters in the pigment epithelium and retina of the rat

The cellular and subcellular expression of the monocarboxylate transporters MCT1, MCT2 and MCT4 [corresponding to MCT3 of Price N. T. et al. (1998) Biochem. J. 329, 321-328] were investigated in the pigment epithelium and outer retina of rats. Immunofluorescence and postembedding immunogold analyses revealed strong MCT1 labelling in the apical membrane of the pigment epithelial and no detectable signal in the basolateral membrane. In contrast, antibodies to the glucose transporter GLUT1 produced intense labelling in both membranes. Neither MCT1 nor GLUT1 was enriched in intracellular compartments. The monocarboxylate transporter MCT4 was very weakly expressed in the retinal pigment epithelium of adult animals, but occurred at higher concentrations at this site in 14-day-old rats. However, even at the latter stage, the immunolabelling of MCT4 was weak compared to that of MCT1. In the neural retina, the data were consistent with a predominant glial localization of MCT1. Specifically, immunogold particles signalling MCT1 occurred in Müller cell microvilli and in the velate processes between the photoreceptors. No labelling was obtained with antibodies to MCT2. Taken together with previous biochemical analyses, the present findings indicate that MCT1 is involved in the outward transport of lactate through the retinal pigment epithelial cells, and in the transfer of lactate between Müller cells and photoreceptors.     

Effects of centrophenoxine on lipofuscin in the retinal pigment epithelium of old mice

The effects of centrophenoxine on the retinal pigment epithelium (RPE) of 17 month old female mice have been studied. Animals were injected subcutaneously for 3 months (60 injections) with the drug (0.1 mg/g of body wt) daily in 0.1 M phosphate buffered saline at pH 7.0. The morphological changes in the pigment layers of the retina of both eyes were studied by light and electron microscopy and the lipofuscin pigment was demonstrated by its autofluorescence and ultrastructural characteristics. There was a significant reduction of the lipofuscin pigment in the treated animals, but the melanin pigment remained unchanged. The lipofuscin granules also appeared less osmiophilic and showed a greater preponderance of membranes and vacuoles. Although the precise mechanism of action of the drug is not clear, an increased protective function of the pigment epithelium by the drug has been suggested.     

Survivin contributes to the progression of diabetic retinopathy through HIF-1α pathway

Hypoxia plays a critical role in the formation of diabetic retinopathy (DR). Hypoxia inducible factor-1α (HIF-1α) and survivin are upregulated following hypoxia, and contribute to the angiogenesis in cancer. The goal of our study was to investigate if the expression of HIF-1α and survivin is related to the retinal vascular change of rats with different stages of diabetes mellitus (DM). Wistar rats were randomly divided into DM for 1 month (DM1) group, DM3, DM6 group and normal control group. Streptozotocin (STZ) was used to induce diabetic rat models. Retinal vascular digest preparation was taken to observe the change of retinal microvessels. The expression of HIF-1α and survivin was determined by real-time PCR and Western blot. In DM3 group, capillaries became circuitous and irregular. Acellular capillary were detected in DM6 group. In the same time, the number of pericytes significantly decreased in DM3 and DM6 groups. The expressions of HIF-1α were more increased in the DM3 and DM6 groups. However, the expressions of survivin only were upregulated in DM6 group .These data suggest that HIF-1α induced the upregulation of survivin in the early stage of DR. Survivin contributed to the development and progression of DR through the HIF-1α pathway and become an important progression marker of DR.     

Time course and development of light adaptation processes in the outer zebrafish retina

Retinomotor movements are morphological changes in the outer retina in response to changing light conditions. They can be separated into two components: Migration of pigment granules within the microvilli of the retinal pigment epithelium (RPE) and positional changes in photoreceptor cells. These positional changes optimize exposure of the cone and rod photoreceptors to light. The aim of this study was to analyze both the time course of retinomotor movements in the adult zebrafish and the maturation of these processes in the developing fish. We show that retinomotor movements are used as a dark/light adaptation mechanism in zebrafish. In adult zebrafish, melanin granules of the RPE migrate with constant speed and reach the fully light adapted (LA) state approximately after 1 h. In contrast, about two thirds of double cone outer segment movements are finished in 5 min, and are fully completed in 10 to 20 min. During development there are three crucial stages leading to mature retinomotor movements in response to light: at 5 dpf (days post fertilization) the migration of pigment granules begins, at 20 dpf the pigment granules condense in the apical part of the RPE microvilli, and at 28 dpf, concomitant with the functional maturation of rods, the double cones contract as in adult retinas.     

泛素蛋白酶系统在四氧嘧啶诱导的糖尿病大鼠视网膜中的表达

目的:建立泛素蛋白酶系统(UPS)在正常和8周糖尿病大鼠视网膜的表达图谱,探讨其在糖尿病视网膜病变(DR)发生发展中可能的分子机制。方法:在限制片段差异显示PCR(RFDD-PCR)技术建立正常和8周糖尿病大鼠视网膜基因表达谱的基础上,对差异片段进行生物信息学分析,筛选UPS DR相关基因,并以免疫组织化学、半定量RT-PCR技术进行验证。结果:RFDD-PCR结果显示,泛素蛋白酶系统DR相关基因UBS3A、PSMD8和PSMD11在糖尿病组表达上调。RT-PCR结果显示,糖尿病组UBS3A表达比正常组明显增高,PSMD8和PSMD11则仅在糖尿病组中表达。免疫组织化学结果显示,正常组UBE3A阳性免疫反应物见于内丛状层,外丛状层和锥、杆体细胞层,PSMD8和PSMD11未见阳性细胞;糖尿病组UBE3A、PSMD8和PSMD11阳性细胞明显增多,见于节细胞层、内核层和外核层。结论:UPS与DR发生发展有关。