早期糖尿病大鼠视网膜蛋白激酶C与内皮素系统关系的研究

目的 探讨糖尿病状态下蛋白激酶C(PKC)与内皮素（ET）系统的改变，并采用特异性PKC抑制剂观察对视网膜内皮素-1的影响。 方法 采用链脲佐菌素（STZ）诱导早期糖尿病大鼠模型，酶联免疫吸附法（ELISA）测定大鼠视网膜内PKC的表达，半定量逆转录聚合酶链反应（RT-PCR）测定视网膜内内皮素ET-1、ET-3、ET-A、ET-受体mRNA的表达，并通过玻璃体内给予PKC抑制剂GF109203X观测对糖尿病大鼠视网膜内ET-1 mRNA表达的影响。 结果 2周糖尿病大鼠视网膜细胞膜PKC活性明显增加（t=3.296, P=0.008），而细胞浆PKC活性无明显改变（t=0.138, P=0.894）；糖尿病大鼠视网膜内ET-1 mRNA 水平较正常组明显升高（P=0.008），而ET-3、ET-A、ET-B受体mRNA水平则无明显改变（P=0.918，P=0.889，P=0.500）；糖尿病大鼠分别予以10-5、10-6、10-7mol/L PKC抑制剂GF109203X玻璃体内注射后,视网膜内ET-1 mRNA水平较对照组下降，呈剂量-反应关系。 结论 早期糖尿病大鼠视 网膜内即存在PKC的激活及ET-1表达的增加，而PKC抑制剂可抑制ET-1的表达。 （中华眼底病杂志，2004，20：168-171）     

过表达TBK1通过调控RIPK1信号通路对糖尿病大鼠视网膜神经节细胞凋亡的抑制作用

目的　探讨TANK结合激酶1（TBK1）对糖尿病大鼠视网膜神经节细胞(RGC)凋亡的作用机制。方法　按60 mg·kg^-1腹腔一次性注射链脲佐菌素制备糖尿病模型。模型制备完成后,随机分成三组:糖尿病(DM)组、TBK1过表达(TBK1-OE)组 (大鼠玻璃体内单次注射TBK1过表达慢病毒载体5 μL,病毒滴度为 1.0×10⁹ IU·mL^-1)、TBK1空载体(TBK1-NC)组(玻璃体内注射等剂量病毒包装的阴性对照液),另取正常大鼠作为对照(CON)组,每组15只。12周后,免疫荧光染色检测大鼠视网膜TBK1蛋白表达定位,Hoechst 33342染色检测大鼠视网膜RGC凋亡,ELISA检测大鼠视网膜肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)含量,Western blot检测大鼠视网膜TBK1、受体相互作用蛋白激酶1（RIPK1）、Caspase-3蛋白相对表达量。结果　TBK1在大鼠RGC中特异表达。与CON组相比,DM组、TBK1-NC组大鼠RGC凋亡率,TNF-α、IL-6、IL-1β含量,RIPK1、Caspase-3蛋白相对表达量均明显增加,TBK1蛋白相对表达量均明显降低(均为P<0.01);TBK1-OE组各指标均增加（均为P<0.01);而与DM组相比,TBK1-OE 组大鼠RGC凋亡率,TNF-α、IL-6、IL-1β含量,RIPK1、Caspase-3蛋白相对表达量均明显降低,TBK1蛋白相对表达量明显增加(均为P<0.01)。而DM组与TBK1-NC组大鼠之间RGC凋亡率,TNF-α、IL-6、IL-1β含量,TBK1、RIPK1、Caspase-3蛋白相对表达量差异均无统计学意义(均为P>0.05)。结论　过表达TBK1对糖尿病大鼠RGC凋亡具有抑制作用,其机制与阻断RIPK1信号通路激活有关。     

Erythropoietin protects retinal neurons and glial cells in early-stage streptozotocin-induced diabetic rats

The purpose of this study was to investigate the efficacy of recombinant human erythropoietin (rhEPO) in preventing and reversing dysfunction of retinal neurons and glial cells in early-stage streptozotocin (STZ)-induced diabetic rats. Two weeks after STZ [60 mg/kg body weight (b.w.), i.p.] injection, diabetic rats had been administered rhEPO (5000 IU/kg of b.w., i.p.) injection three times weekly for 2 weeks. The changes to the electroretinogram, retina ultrastructure, erythropoietin receptors (EPO-Rs) and the content of glutamate in retinas before and after the experiment in test and control groups were compared. The amplitudes of b-wave and oscillatory potentials (OPs) decreased at the end of the experiment in STZ-induced diabetic rats compared with the age-matched controls, whereas the amplitudes of b-wave and OPs showed no decrease in diabetic rats with rhEPO injection. Mitochondrial metamorphosis in ganglion cells occurred in STZ-induced diabetic rats but was not found in those rats with rhEPO treatment. EPO-Rs in retinas and the retinal glutamate of STZ-induced diabetic rats at the end of the experiment had increased obviously compared with rhEPO-injected diabetic rats. The abnormalities of retinal electrophysiological activity, ganglion cells with swollen mitochondria in the retinas, increased retinal glutamate and EPO-R in the retinas occurred early in the process of the disease course in diabetic rats. These aspects can all be improved by intraperitoneal administered rhEPO. The administration of rhEPO may be useful in the neural treatment of diabetic retinopathy at the early stage.     

Protective effect of pomegranate juice on retinal oxidative stress in streptozotocin-induced diabetic rats

AIM: To investigate the effect of pomegranate juice (PJ) intake on overall oxidation status in retinas of diabetic rats. METHODS: Twenty-seven rats were divided into four groups as control (CO), diabetic (DM), control treated with PJ (CO-PJ), and diabetic treated with PJ (DM-PJ).The retina tissues were used to determine 8-hydroxy-2’-deoxyguanosine (8OHdG), malondialdehyde (MDA), reduced glutathione (GSH) levels, and the enzyme activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). RESULTS: The levels of 8OHdG and MDA were significantly increased in the retina of DM group compared to CO group (P=0.001, P<0.001 respectively). Both 8OHdG and MDA levels were decreased in PJ-DM group compared to DM group (P=0.004, P<0.001 respectively). The activities of antioxidant enzymes GSH, SOD, and GDH-Px were significantly decreased in the retina of DM group compared to CO group (P≤0.01). GSH and GSH-Px activities were higher in PJ-DM group compared with DM group (P=0.010, P=0.042, respectively) but SOD activity was not statistically different (P=0.938). CONCLUSION: PJ intake is found to be effective in decreasing oxidative end products, and in increasing the activities of antioxidant enzymes in diabetic retinas of rats, which suggests it may be effective against oxidative stress in diabetic retinas.     

蛋白激酶Cα亚型在视网膜视杆-双极细胞发育过程中表达规律的体外实验研究

目的明确新生小牛视网膜视杆-双极细胞体外发育过程中蛋白激酶Cα亚型(PKCα)的表达规律。方法分离培养新生小牛视网膜神经细胞,培养10、20、30和40 d的神经元用于免疫细胞化学染色,采用PKCα抗体标记视网膜视杆-双极细胞,用图像分析系统定量测量阳性细胞的免疫反应强度。结果培养10 d时,PKCα阳性神经元只有1~2个较短的神经突起;到培养20 d时,阳性细胞的一端为1个细长的神经突起,相对的另一端为丛状的短的树状突起,培养30 d及40 d的神经元保持培养20 d时神经元的形态特征。不同培养阶段的阳性细胞中PKCα免疫反应物质均位于细胞体和神经突起,定量分析显示培养20 d的阳性细胞免疫反应强度最高,培养30 d和40 d时阳性细胞PKCα免疫反应强度下降。结论培养20 d的视杆-双极细胞具备细胞形态和免疫反应特性上的成熟特征,PKCα的表达水平最高,为发育成熟的神经元。培养30 d以后,视杆-双极细胞逐渐衰老。实验结果为选择合适的供体材料进行视网膜移植提供了理论依据。     

沉默信息调节因子相关酶1(SIRT1)调控p38MARK信号通路对糖尿病视网膜病变大鼠视网膜神经节细胞的保护作用及机制

目的 探讨沉默信息调节因子相关酶1(silment information regulator factor related enzymes 1,SIRTl)对糖尿病视网膜病变大鼠视网膜神经节细胞(retinal ganglion cells,RGCs)的保护作用及其机制.方法 取健康清洁级雄性Sprague-Dawley大鼠60只,应用随机数字表法分为正常对照组(正常组)、糖尿病组(病变组)、SIRT1激动剂白藜芦醇治疗组(治疗组).病变组和治疗组大鼠按60 mg·kg-1单次腹腔注射链脲佐菌素以诱导糖尿病大鼠模型;正常组按60 mg· kg-1腹腔注射枸橼酸钠缓冲液.72h后取鼠尾静脉血检测血糖,血糖值＞16.7 mmol·L-1定为糖尿病大鼠.自造模成功后第2天起治疗组每天每只鼠给予白藜芦醇20 g·kg-1灌胃,正常组和病变组每天每只鼠给予亚甲砜灌胃.8周后进行视网膜免疫组织化学染色,TUNEL法检测视网膜RGCs凋亡,Western blot检测SIRT1、p38 MAPK、Caspase-3蛋白的表达.结果 正常组、病变组、治疗组8周后RGCs凋亡指数分别为:(0.848±0.131)％、(19.038±1.327)％、(10.461±1.089)％,三组间差异有统计学意义(F=670.497,P=0.000).进一步两两比较:正常组RGCs凋亡指数与病变组、治疗组间差异均有统计学意义(均为P=0.000);治疗组与病变组间差异亦有统计学意义(P =0.000).与正常组(0.132±0.003)相比,病变组(0.060±0.028)和治疗组(0.073±0.026)大鼠视网膜SIRT1蛋白表达降低,总体差异有统计学意义(F=1310.663,P=0.000).进一步两两比较,病变组和治疗组与正常组间,以及病变组与治疗组间差异均有统计学意义(均为P =0.000).病变组(l.121±0.082,0.266±0.005)和治疗组(0.574±0.012,0.190±0.060)大鼠视网膜p38 MAPK、Caspase-3蛋白表达较正常组(0.402±0.012,0.156±0.006)明显增加,总体差异有统计学意义(F =604.500、1056.709,P=0.000、0.000).进一步两两比较:p38 MAPK、Caspase-3蛋白表达在正常组与病变组间、正常组与治疗组间以及病变组与治疗组间差异均有统计学意义(均为P=0.000).结论 在糖尿病视网膜病变模型中,SIRT1表达上调,抑制RGCs的凋亡,对糖尿病视网膜病变RGCs起保护作用.其抗凋亡作用机制可能与其抑制p38 MAPK的表达相关.p38 MAPK信号通路是糖尿病视网膜病变中SIRT1介导的神经保护作用的重要通路之一.     

Retinal heparanase expression in streptozotocin-induced diabetic rats

Objective: Heparanase, an endoglycosidase, exhibits strong proangiogenic capacity that can induce vascular endothelial growth factor (VEGF) expression in tumour angiogenesis. The purpose of this study was to evaluate heparanase expression and its relationship with VEGF in streptozotocin (STZ)-induced diabetic rats' retinas.Design: Experimental study.Participants: STZ-induced rats and non-diabetic control rats.Methods: Heparanase expression was initially evaluated in cultured human retinal microvascular endothelial cells (HRECs) under high-glucose conditions by Western blot. Diabetes was induced in Sprague-Dawley rats by STZ intraperitoneal injection. Retinal heparanase expression was assayed in rats by immunohistochemistry. Heparanase inhibitor (phosphomannopentaose sulfate) was administrated to high-glucose-treated HRECs and diabetic rats. VEGF levels were evaluated in HRECs and retinas using enzyme-linked immunosorbent assay.Results: Heparanase expression was increased in HRECs under high-glucose conditions compared with controls (p < 0.01). Immunohistochemical studies indicated that heparanase signals were intense in the retinal vascular endothelia of diabetic rats, but faint in those of nondiabetic control rats. Quantitative analysis showed that heparanase protein expression was increased by 3.2-fold in diabetic rats' retinas compared with nondiabetic rats' retinas (p < 0.01). VEGF level was increased, as was heparanase expression, in high-glucose-treated HRECs and in the retinas of diabetic rats, and these increases were significantly decreased by phosphomannopentaose sulfate administration (p < 0.01).Conclusions: Heparanase expression was upregulated and associated with an increase of VEGF expression in STZ-induced diabetic rat retinas. The data suggest that heparanase may be involved in the development of diabetic retinopathy and represents a possible novel target for therapeutic intervention.     

氨基胍对STZ糖尿病大鼠视网膜微血管病变的影响

目的:观察STZ糖尿病大鼠视网膜微血管病理改变及氨基胍对微血管病变的影响.方法:制备STZ糖尿病大鼠动物模型,随机分为正常对照(CON)组、糖尿病3mo(DM3)组,6mo(DM6)组,氨基胍3mo(AG3)组和6mo(AG6)组.每组大鼠各10只,分别饲养3,6mo.取大鼠视网膜进行视网膜微血管形态及超微结构观察.结果:DM组视网膜周细胞数目随病程逐渐减少,细胞核染色质浓缩,边集,分布于核膜下;胞质内线粒体肿胀,变性,空泡化,胞质消失,基底膜增厚.AG治疗组则明显改善.结论:氨基胍对STZ糖尿病大鼠视网膜微血管病变有防治作用.     

Nogo受体在早期糖尿病大鼠视网膜神经节细胞凋亡中的作用

目的:探讨Nogo受体(nogo receptor,NgR)在糖尿病(diabetes mellitus,DM)大鼠视网膜神经节细胞凋亡中的作用及机制。方法:链脲佐菌素诱导SD大鼠DM模型,实验分4组,对照组;DM组;siNgR组:DM大鼠玻璃体内给予NgR反义核苷酸序列;siRNA空白组:DM大鼠玻璃体内给予阴性核苷酸序列。1mo后TUNEL染色检测视网膜神经节细胞(retinal ganglion cell,RGC)凋亡,试剂盒检测视网膜丙二醛(malondialdehyde,MDA)含量,Western-blot检测视网膜NgR及caspase-3含量。结果:对照组、DM组、siRNA空白组及siNgR组MDA含量分别为3.74±0.49,7.21±0.96,6.41±1.34及4.02±0.53nmol/mg prot。与对照组相比,DM组及siRNA空白组视网膜RGC凋亡增加,NgR及caspase-3表达上调,MDA含量升高(P<0.05),而siNgR组视网膜RGC数量、NgR及caspase-3表达、MDA含量较对照组均无明显变化(P>0.05)。结论:NgR表达增加是糖尿病RGC凋亡的重要原因之一,其机制可能与NgR诱导视网膜氧化应激、上调caspase-3表达有关。     

Diabetes induces expression of aquaporin-0 in the retinal nerve fibers of spontaneously diabetic Torii rats

Diabetes redistributes the expression of glial aquaporin (AQP) water channels in the retina. However, it is not known whether diabetes also affects retinal AQP-0 expression. This study examined the effects of the development of diabetes on the expression of retinal AQP-0 in spontaneously diabetic Torii (SDT) rats. Male SDT rats at 10 and 40 weeks of age and age-matched male Sprague-Dawley (SD) rats were used. The localization of AQP-0 was assessed immunohistochemically using sagittal cryosections of the rats’ retinas and optic nerves. Fold changes in AQP-0 gene expression relative to controls were assessed by real-time RT-PCR. All SDT rats spontaneously developed diabetes by 40 weeks of age (the mean hemoglobin (Hb) A1c levels were 2.8 ± 0.2% and 11.2 ± 1.0% at 10 and 40 weeks, respectively). SD rats did not develop diabetes (the HbA1c levels were 2.7 ± 0.2% and 2.6 ± 0.3% at 10 and 40 weeks, respectively). In the retinas of SD rats and in those of SDT rats at 10 weeks of age, immunoreactivity for AQP-0 was confined predominantly to the inner nuclear layer and to the border between the inner plexiform layer and the ganglion cell layer (GCL), where AQP-0 colocalized with protein kinase C-α. AQP-0 immunoreactivity was also observed in the GCL to a lesser degree, which colocalized with the neuronal nuclei. In the 40-week-old SDT rat retinas, additional AQP-0 immunoreactivity was observed in the GCL and colocalized with neurofilaments, indicating expression of AQP-0 in ganglion cell axons. However, the axonal AQP-0 immunoreactivity was restricted to the retinal nerve fibers, whereas the optic nerve axons were devoid of AQP-0. Retinal blood vessels did not express AQP-0. AQP-0 gene expression was 3.4-fold higher in SDT rat retinas than in SD rat retinas at 40 weeks of age.AQP-0 was predominantly expressed in the bipolar cells of the non-diabetic rat retinas, whereas it was also expressed in the retinal nerve fibers of diabetic rat retinas. The disrupted water transport between astrocytes and retinal nerve fibers may be associated with the known accelerated apoptosis of retinal ganglion cells induced by diabetes.     

糖尿病视网膜病变中氧化应激反应的研究及应用

目的通过研究抗氧化剂Vitamin E在糖尿病动物模型-大鼠视网膜中对氧化应激反应的作用,探寻糖尿病视网膜病变的发病机理.方法将大鼠随机分为正常组、糖尿病组、维生素E治疗1组(剂量0.12g·kg-1)、2组(剂量0.25g·kg-1).饲养8个月后,观察其视网膜蛋白激酶C(protein kinase C,PKC)的活性和无细胞血管及影细胞的变化.结果治疗1组与糖尿病组大鼠视网膜PKC的活性及无细胞血管、影细胞无明显差异(P>0.05),治疗2组与糖尿病组大鼠视网膜PKC的活性及无细胞血管、影细胞有明显差异(P<0.05).结论抗氧化剂Vitamin E能抑制糖尿病视网膜病变的氧化应激反应进而减轻其早期病理损害,糖尿病视网膜病变的发病机理与氧化应激反应的关系有待于进一步被证实.     

链脲佐菌素诱导的糖尿病大鼠早期视网膜NO的变化

目的利用链脲佐菌素(STZ)诱导的1型糖尿病大鼠模型,研究糖尿病早期视网膜一氧化氮(NO)浓度的变化。方法健康成年雄性SD大鼠腹腔注射STZ60mg/kg,7d后血糖>13.9mmol/L入选为糖尿病组,与正常对照组在无特定病原体(SPF)环境下饲养,每周监测体重与血糖。8周后氯胺酮全身麻醉下摘取眼球,分离视网膜,制成组织匀浆。应用硝酸还原酶法测定组织NO浓度。结果糖尿病组与对照组在8周末时的平均体重差异有极显著统计学意义(t=11,P<0.01)。两组在各观察时段的平均血糖水平差异均有显著统计学意义(P<0.05)。两组视网膜样品的蛋白浓度差异无显著统计学意义(t=0.765,P>0.05)。糖尿病组视网膜平均NO浓度为(0.352±0.256)μmol/gprot,对照组为(0.031±0.017)μmol/gprot,两组间差异有显著统计学意义(t=2.79,P<0.05)。结论STZ诱导的糖尿病大鼠早期视网膜NO浓度增加,是引起糖尿病早期视网膜血管功能性改变的重要因素。     

Vascular endothelial growth factor expression and secretion by retinal pigment epithelial cells in high glucose and hypoxia is protein kinase C-dependent

Retinal pigment epithelial (RPE) cells express vascular endothelial growth factor (VEGF) in response to high glucose or hypoxia. We hypothesised that VEGF expression and secretion by RPE cells in high glucose and hypoxia are regulated by protein kinase C (PKC). Primary cultured RPE cells from Sprague-Dawley rats were growth-arrested for 48 hr in 0.5% FBS in 5.6 or 30 mm D-glucose. Cells were exposed to hypoxic conditions (<1% O(2), 5% CO(2)) for the last 15-18 hr of growth-arrest. PKC -alpha, -beta(1), -delta, -epsilon, and -zeta were expressed by RPE cells and exposure to high glucose for 48 hr had no effect on expression as demonstrated by Western immunoblotting. High glucose, hypoxia or VEGF stimulated translocation of a number of the PKC isozymes to the membrane or particulate fractions implying activation. In response to high glucose or acute phorbol myristate acetate (PMA) stimulation, VEGF mRNA analysed by RT-PCR was increased. Intracellular VEGF protein identified by immunoblotting and confocal immunofluorescence imaging was significantly increased by high glucose, hypoxia or acute PMA stimulation. Calphostin C or a specific inhibitor of PKC-zeta prevented high glucose-stimulated VEGF expression in high glucose. VEGF secretion, as measured by ELISA in the culture medium, was enhanced in hypoxia but not in high glucose. Following exposure of RPE cells to PMA for 24 hr, PKC-delta was significantly down regulated, whereas PKC-alpha, -beta, -epsilon and -zeta remained unchanged. Secretion of VEGF in normal or high glucose, or hypoxia was significantly reduced following treatment with PMA for 24 hr but not with the PKC-zeta inhibitor. We conclude that in high glucose and hypoxia PKC isozymes are activated and are necessary for VEGF expression. Secretion of VEGF is enhanced in hypoxia and appears to be regulated by PKC-delta. RPE cells may contribute to the pathogenesis of retinopathy caused by high glucose and hypoxia through the expression and secretion of VEGF that are regulated by PKC isozymes.     

M2型丙酮酸激酶在糖尿病视网膜病变中的作用

糖尿病视网膜病变(DR)是一种糖尿病中最常见的高度特异性微血管并发症,是全球20~65岁人群视力障碍和失明的主要原因,主要表现为视网膜内微血管的异常及新生血管的形成。糖酵解过程是从葡萄糖分解开始到丙酮酸生成的过程,可以为机体迅速提供能量,内皮细胞多数通过糖酵解产生的ATP来维持其功能,包括维持紧密连接和屏障作用。丙酮酸激酶(PK)的M2亚型(PKM2)作为糖酵解的关键酶,在机体的大多数组织中均有表达。内皮细胞和光感受器细胞作为视网膜中的重要细胞成分,在DR的发生发展过程中发挥重要作用。研究表明,PKM2通过代谢和非代谢方式调节内皮细胞和光感受器的功能,在DR发展中发挥重要作用。因此,本文将重点通过内皮细胞和光感受器细胞两个方面来综述PKM2在糖尿病视网膜病变中的研究进展,从而为DR的诊断和治疗提供新思路。     

糖尿病视网膜神经变性机制的研究进展

糖尿病视网膜病变(diabetic retinopathy,DR)一直被认为是微血管病变。然而,大量的研究已经证实,DR不仅会引起视网膜的血管病变,也会引起视网膜神经退行性改变。越来越多的证据也表明,在DR早期未发生视网膜血管病变之前就已经出现了视网膜神经变性,且视网膜神经变性可能参与了微血管异常的发生发展。目前,糖尿病视网膜神经变性的机制尚不十分明确,现就近年来糖尿病视网膜神经变性机制的研究进展做一综述。     

高血糖对糖尿病大鼠视网膜神经节细胞的影响

目的 通过观察大鼠视网膜病理改变,从组织病理学角度揭示高血糖对大鼠视网膜神经节细胞的影响。方法 将100只大鼠分为对照组10只和糖尿病组90只。糖尿病组用链脲佐菌素造模,9个月后通过组织化学法观察高血糖对观察组大鼠视网膜内层神经节细胞的影响。结果 糖尿病组大鼠视网膜内层厚度降低、视网膜神经节细胞数目明显减少、视网膜神经节细胞肿胀平均周长、面积升高,视网膜神经节细胞黑度值降低,与对照组比较差别有显著意义(P＜0．01)。结论 高血糖能使糖尿病大鼠视网膜变薄,神经节细胞肿胀、数目减少,从而对糖尿病大鼠视网膜内层造成损害。     

Raf-1激酶抑制蛋白(RKIP)对糖尿病大鼠视网膜神经损伤的保护作用

目的　探讨Raf-1激酶抑制蛋白(raf-1 kinase inhibitory protein,RKIP)通过p38-MAPK通路对糖尿病(DM)大鼠视网膜神经损伤的保护作用。方法　雄性SD大鼠48只,采用随机数字表法将大鼠分为对照组、空白组、糖尿病组、RKIP组,每组12只;空白组、糖尿病组和RKIP组大鼠腹腔注射链脲佐菌素建立DM模型。RKIP组和空白组于模型建成第2周玻璃体内分别注射携带RKIP基因片段重组慢病毒(LV-RKIP)和等量空白病毒,对照组和糖尿病组注射等量生理盐水。注射10周后利用Western blot和免疫荧光化学检测各组RKIP、p38丝裂原激活蛋白激酶(p38-mitogen-activated protein kinase,p38-MAPK) 、胶质细胞谷氨酸转运体(L-glutamate/L-asparate transporter,GLAST)和谷氨酰胺合成酶(glutamine synthetase,GS)在视网膜中的表达,HE染色检测各组视网膜神经节细胞(retinal ganglion cell,RGC)密度。结果　与对照组相比,糖尿病组p38-MAPK表达升高,RKIP、GLAST、GS表达下降,RGC密度减少(均为P<0.01);与糖尿病组相比,空白组各指标差异均无统计学意义(均为P>0.05),RKIP组p38-MAPK表达降低,RKIP、GLAST、GS表达升高,RGC密度增多(均为P<0.01)。结论　糖尿病视网膜病变早期注射RKIP可降低视网膜细胞中p38-MAPK表达量,提高GLAST、GS表达活性,改善Müller细胞功能,减少RGC损害,提示RKIP对糖尿病视网膜病变中视网膜神经细胞具有保护作用。     

非酶糖化抑制剂对STZ-糖尿病大鼠视网膜微血管超微结构的影响

目的　观察病程 2 4wk STZ-糖尿病大鼠视网膜微血管超微结构的病理改变及非酶糖化仰制剂 -氨基胍对超微结构改变的影响。方法　制备 STZ-糖尿病大鼠动物模型 ,随机分为氨基胍 (AG)组、糖尿病 (DM)组各 6只。设正常对照组 6只。 2 4wk时处死 ,取大鼠视网膜进行电镜观察。结果　 DM组视网膜微血管内皮细胞、周细胞核异染色质聚集、靠边 ;胞质内线粒体肿胀、变性 ,胞浆消失 ;基底膜增厚 ;毛细血管壁断裂。 AG组上述损害明显轻于 DM组。结论　非酶糖化抑制剂对 STZ-糖尿病大鼠视网膜微血管超微结构的损害有明显的防治作用     

Latanoprost rescues retinal neuro-glial cells from apoptosis by inhibiting caspase-3, which is mediated by p44/p42 mitogen-activated protein kinase

The purpose of this study was to investigate whether latanoprost, a prostaglandin F2alpha analogue, has a direct anti-apoptotic effect both in retinal neuro-glial cells in culture and in diabetic retina. R28 cells, immortalized retinal neuroglial progenitor cells, were induced apoptosis by 24h serum deprivation. Serum withdrawal made up to 15% of R28 cells pyknotic and activated caspase-3 immunoreactive, and latanoprost acid suppressed apoptosis with dose dependency at an optimum concentration of 1.0 microM (P<0.001). UO126, a mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase kinase (MEK) 1 and 2 inhibitor reversed this effect. Streptozotocin induced one- or three-month diabetic rats received balanced-salt-solution (BSS) in the left eye and latanoprost eye drops in the right for 5 days. Retinal wholemount was subjected to terminal dUTP nick end labeling (TUNEL) staining, whereas eyeballs were enucleated for cleaved caspase-3 immunofluorescence. Retinal homogenates were probed for phospho- or total p44/p42 MAPK and Akt. One- and three-month diabetic retina had 30.2+/-15.3 and 23.6+/-9.0 TUNEL positive cells per 0.5 cm(2), respectively, whereas control retina had few TUNEL positive cells. Latanoprost instillation significantly reduced these cells (10.0+/-3.1 and 11.3+/-3.1 cells per 0.5 cm(2) for 1M and 3M, respectively, P<0.01), whereas BSS did not. Latanoprost also significantly reduced cleaved caspase-3 immunoreactive cells in ganglion cell and inner nuclear layers (P<0.05). Latanoprost increased phosphorylated to total protein ratio of p44/p42 MAPK (P<0.05), but not of Akt. Taken together, the present findings suggest that latanoprost rescues retinal neurons and/or glial cells from apoptosis, which is probably mediated by p44/p42 MAPK through caspase-3 inhibition.     

丙酮酸对糖尿病大鼠视网膜氧化损伤及超微结构的影响

目的:研究糖尿病大鼠视网膜病变过程中视网膜组织学和氧化应激的变化,以及丙酮酸的对抗作用。方法:将80只Wistar大鼠分成3组:对照组(20只),模型组(30只),治疗组(30只)。模型组和治疗组用STZ诱导糖尿病,治疗组在大鼠饲料和饮水中添加2%丙酮酸。观察大鼠的血糖、体质量变化,并在造模后12wk观察3组大鼠视网膜组织中GSH-Px、MDA和Na+-K+-ATP酶水平及其超微结构改变。结果:模型组和对照组相比,体质量显著下降,视网膜中GSH-Px和ATP酶活性显著下降,MDA水平显著升高,视网膜超微结构有显著改变;治疗组和模型组相比,血糖没有显著改变,视网膜组织中GSH-Px和ATP水平升高,MDA水平下降,视网膜超微结构病变相对较轻。结论:丙酮酸可以减轻氧化应激反应,改善视网膜的能量代谢,延缓视网膜病变的发展。