塞来昔布对糖尿病大鼠视网膜血管内皮生长因子表达的影响

目的 观察塞来昔布对糖尿病大鼠视网膜血管内皮生长因子（VEGF）表达的影响。 方法 将36只大鼠用链脲佐菌素（STZ）腹腔注射制成糖尿病大鼠模型，随机分成糖尿病组(n=18)和塞来昔布灌胃组(n=18)。塞来昔布灌胃组大鼠经口灌胃塞来昔布50 mg/kg；糖尿病组经口灌胃等体积生理盐水。另18只正常大鼠为正常对照组。3个月后处死全部大鼠，应用免疫组织化学技术检测视网膜VEGF蛋白表达，逆转录多聚酶链式反应（RT-PCR）方法检测视网膜VEGF-mRNA和环氧化酶(COX)-2-mRNA的表达。 结果 正常对照组视网膜VEGF-mRNA和COX-2-mRNA表达量少，VEGF免疫组织化学反应弱阳性，VEGF蛋白表达量低；糖尿病组较正常对照组视网膜VEGF-mRNA和COX-2-mRNA表达均上调(P＜0.05)，VEGF免疫组织化学反应呈强阳性，VEGF蛋白表达增高(P＜0.01)；塞来昔布灌胃组较糖尿病组视网膜VEGF mRNA表达显著下降(P＜0.05)，COX-2- mRNA的表达无显著降低(P>0.05)，VEGF免疫组织化学反应阳性减弱，VEGF蛋白表达降低(P＜0.01)。 结论 塞来昔布可通过抑制COX-2的活性进一步抑制STZ诱导的糖尿病大鼠视网膜VEGF-mRNA及蛋白表达。(中华眼底病杂志,2007,23:265-268)     

Ets-1和VEGF在实验性糖尿病视网膜中的共表达

目的：阐明VEGF和Ets-1在糖尿病大鼠视网膜内的表达特点和分布特点。 方法：腹腔注射STZ诱导糖尿病大鼠模型。STZ注射后4wk处死动物，分离各组视网膜总蛋白进行Western　blot分析。将眼球固定后制成14μm厚的冰冻切片进行Ets-1和VEGF免疫荧光双标检测。 结果：Ets-1和VEGF在糖尿病大鼠视网膜内表达均显著增强，而且Ets-1和VEGF在视网膜内的分布特点相似，几乎在视网膜各层均共表达。 结论：Ets-1可能对VEGF诱导的糖尿病视网膜病变有促进作用。     

造血干细胞对糖尿病小鼠视网膜血管内皮细胞影响的研究

目的探讨造血干细胞(HSCs)倍增对糖尿病小鼠视网膜微血管内皮细胞的影响。方法应用小鼠干细胞生长因子(SCF)200μg·kg~(-1)·d~(-1)联合重组人粒细胞集落刺激因子(rhG-csf)50μg·kg~(-1)·d~(-1),连续5d 对糖尿病小鼠和非糖尿病小鼠分别进行自体 HSCs 动员,以生理盐水皮下注射作为对照。应用流式细胞仪以 CD34-/low 和 Scal+作为标记鉴定外周血 HSCs。应用免疫组化方法以 CD31和5-溴脱氧尿嘧啶(BrdU)标记新生的视网膜血管内皮细胞,PAS 染色标记成熟的视网膜血管内皮细胞。半定量 RT-PCR 检测血管内皮生长因子(VEGF)、可溶性血管内皮生长因子受体2(VEGFR-2)及血管生成素2(ang-2)在视网膜的表达情况。结果自体干细胞动员可以使正常小鼠和糖尿病小鼠外周血中产生 CD34-/low 和 Scal+的细胞数目倍增。在免疫组化切片上可观察到糖尿病小鼠 CD31标记的新生血管数目增多,内皮细胞更生速度加快。在 BrdU 与 CD31的免疫组化双重染色中,可观察到新生毛细血管内皮细胞两种抗原双重表达的现象。自体干细胞动员可使小鼠视网膜 VEGFR-2含量显著增高,明显下调糖尿病小鼠 VEGF 和 ang-2的表达量。结论自体 HSCs 动员可以通过外周血干细胞倍增,环境诱导分化为血管内皮细胞,双向调节 VEGF 和 ang-2等血管渗漏性相关因子,实现治疗性血管重建。     

视网膜消化铺片的免疫荧光染色探讨

目的：探讨视网膜消化铺片联合免疫荧光染色技术的可行性。方法：正常Wistar大鼠和链尿佐菌素（Streptozotocin,STZ)诱导的糖尿病2周大鼠各5只。经心脏灌注后，取右眼视网膜作胰酶消化铺片，并在铺片上进行血管细胞粘附分子－1（Vascular cell adhesion molecule-1,VCAM-1)单克隆抗体免疫荧光染色。结果；大鼠视网膜消化铺片可以清晰看到视网膜血管走行、分布、正常大鼠未见VCAM－1阳性表达，糖尿病2周组视网膜血管壁见VCAM－1阳性细胞表达。结论：视网膜血管铺片联合免疫荧光染色技术是可行的。     

视网膜母细胞瘤和正常视网膜组织中基质调控蛋白表达及其意义(英文)

目的:研究视网膜母细胞瘤(RB)和正常视网膜组织中EMMPRIN、MMP1、MMP9和TIMP2表达水平及其临床病理意义和相互联系。方法:收集30例RB眼球摘除手术标本和15例正常视网膜组织标本。常规制作石蜡包埋切片,EMMPRIN、MMP1、MMP9和TIMP2染色方法均为Envision免疫组织化学法。结果:RB组织中EMMPRIN、MMP1、MMP9表达阳性率均高于正常视网膜组织(P<0.01),TIMP2则低于正常视网膜组织(P<0.01)。临床分期I期、分化型、未侵犯视神经及生存期≥2aRB病例EMMPRIN、MMP1、MMP9表达阳性率低于临床分期III期、未分化型、侵犯视神经和生存<2a病例(P<0.05或P<0.01);分化型、未侵犯视神经及生存期≥2aRB病例TIMP2表达阳性率明显高于未分化型、侵犯视神经和生存<2a病例(P<0.05或P<0.01);RB中,EMMPRIN、MMP1、MMP9表达呈高度一致性(P<0.05),TIMP2与EMMPRIN、MMP1、MMP9表达水平呈高度不一致性(P<0.05)。结论:EMMPRIN、MMP1、MMP9和TIMP2的表达水平可能是反映RB进展、侵袭能力及预后的重要标记物,它们之间存在着内在的相互调控关系。     

大肠杆菌K5多糖对糖尿病视网膜病变小鼠视网膜新生血管形成作用的研究

目的 探究大肠杆菌K5多糖对糖尿病视网膜病变小鼠视网膜新生血管形成的影响.方法 发酵培养大肠杆菌K5菌株,对培养液进行分离纯化制备K5多糖,并分析K5多糖的纯度;雄性ICR小鼠被随机分为正常纽、模型组和干预组,其中正常组是健康小鼠,而模型组和干预组均使用链脲佐菌素建立糖尿病小鼠模型,干预组于造模后使用K5多糖干预4周,造模后8周各组取小鼠眼球组织,行HE染色及视网膜铺片ADPase染色;采用Real-time PCR和Western blotting检测各组小鼠眼球组织血管内皮生长因子(vascular endothelial gTowth factor,VEGF)及基质金属蛋白酶-2(matrix metalloproteinase,MMP-2)的表达情况.结果 HPLC结果显示制备的K5多糖纯度在95％以上:HE染色及ADPase染色结果显示正常组小鼠视网膜只见少量血管,模型组小鼠视网膜血管数目显著高于正常组,而干预组小鼠视网膜血管数目较模型组显著下降(P＜ 0.05);Real-time PCR和Western blot结果显示,模型组小鼠眼球组织VEGF和MMP-2的mRNA及蛋白表达水平显著高于正常组(均为P＜0.05),而干预组VEGF和MMP-2的mRNA及蛋白表达水平显著低于模型组(均为P＜0.05).结论 大肠杆菌K5多糖能够抑制糖尿病视网膜病变小鼠视网膜新生血管形成,其作用机制可能与抑制VEGF及MMP-2的表达有关.     

细胞骨架相关蛋白2(CKAP2)在db/db小鼠视网膜组织中的表达及其与HIF-1α/VEGF信号通路的关系

目的 观察细胞骨架相关蛋白2(CKAP2)在db/db小鼠视网膜组织中的表达及分布特点,分析其与缺氧诱导因子1α(HIF-1α)/血管内皮生长因子(VEGF)信号通路的关系.方法 选取雄性db/db糖尿病小鼠(db/db组)及同窝正常野生型wt/wt小鼠(wt/wt组)各15只.采用HE染色观察两组小鼠视网膜结构变化,...     

脯氨酰羟化酶在糖尿病大鼠视网膜的表达及其意义

目的 观察探讨糖尿病(DM)大鼠视网膜组织中脯氨酰羟化酶(PHD-2)的表达及意义.方法 雄性Wistar大鼠108只,随机分为DM模型组和正常对照组,分别为60、48只.对DM模型组大鼠进行一次性腹腔注射1％链脲霉素枸橼酸(STZ)溶液建模,正常对照组腹腔注射等量的构橼酸缓冲液.造模后1、3、6个月,荧光显微镜下观察大鼠视网膜血管分布形态.造模后1～6个月,采用伊凡思蓝(EB)灌注视网膜铺片检测视网膜组织内EB浓度;免疫组织化学染色法观察大鼠视网膜PHD-2阳性染色的分布特点;蛋白质免疫印迹法(Western blot)检测大鼠视网膜中PHD-2、缺氧诱导因子-1α(HIF-1α)及血管内皮生长因子(VEGF)的表达.结果 荧光显微镜观察结果显示,正常对照组大鼠视网膜血管走行规则,浅深层血管形态清晰;DM模型组大鼠视网膜血管渗漏持续存在.造模后1～6个月,DM模型组大鼠视网膜血管外组织内EB含量较正常对照组明显增加,差异有统计学意义(t=3.810,2.722,2.845,2.342,2.456,3.823;P＜0.05).免疫组织化学染色结果显示,DM模型组及正常对照组大鼠视网膜中均可见PHD-2阳性染色,主要分布于视网膜内核层、神经节细胞层及血管壁.Western blot检测结果显示,DM模型组大鼠视网膜中PHD-2表达在造模后1、2个月时较正常对照组下降(t=16.230,16.390;P＜0.05);HIF-1a表达在造模后1、2、3个月时较正常对照组明显升高(t=27.073,36.709,10.176; P＜0.05);VEGF表达在造模后1～6个月均较正常对照组升高(t=13.547,31.984,21.897,8.912,9.019,14.046;P＜0.05).结论 PHD-2在DM大鼠视网膜组织中有丰富表达.PHD-2可能在糖尿病视网膜病变发生发展中有一定的调控作用,其作用途径及机制与VEGF作用通路有关.     

MMP-2和VEGF在视网膜新生血管的表达

目的检测基质金属蛋白酶-2(matrix metalloproteinases-2,MMP-2)和血管内皮生长因子(vascular endothelial growthfactor,VEGF)在视网膜新生血管中的表达,探讨两者的相互关系。方法 取7 d龄C57BL/6J小鼠40只,随机分为高氧组和对照组,每组各20只。建立高氧诱导的视网膜新生血管小鼠模型。采用ADP酶组织化学染色、HE染色和免疫组化方法分别观察2组视网膜血管的变化、计数突破视网膜内界膜的内皮细胞核数目和检测MMP-2、VEGF的表达。结果 高氧组视网膜铺片可见大量视网膜新生血管形成,对照组未见新生血管形成;高氧组突破视网膜内界膜内皮细胞核数目为14(14.230±5.388),与对照组数目0(0.110±0.386)相比差异有统计学意义(t=23.537,P<0.001);对照组和高氧组视网膜组织中VEGF的积分光密度值分别为36.81±14.60、60.85±24.55,差异有统计学意义(t=3.348,P<0.01);对照组和高氧组视网膜组织中MMP-2的积分光密度值分别为16.33±4.13、21.12±6.29,差异有统计学意义(t=3.160,P<0.01)。高氧组视网膜组织中MMP-2和VEGF的表达呈正相关(r=0.633,P<0.01)。结论 在ROP小鼠模型的视网膜组织中,VEGF、MMP-2的表达同步升高,二者的高表达可能与视网膜新生血管形成密切相关。     

胰岛素控制血糖对糖尿病大鼠早期视网膜血管的影响

目的:探讨胰岛素控制血糖对糖尿病早期大鼠视网膜血管形态改变和血管内皮细胞生长因子(Vascular endothelial growth factor,VEGF)表达的影响。方法:实验动物Wistar大鼠分正常对照组、糖尿病组、糖尿病血糖严格控制组和糖尿病血糖非严格控制组。链尿佐菌素(Streptozotocin,STZ)诱导糖尿病动物模型,动物成模3周后糖尿病血糖严格控制组及血糖非严格控制组大鼠予胰岛素治疗,20d后处死,行Fluorescein isothiocyanate-dextran(FITC-Dextran)灌注视网膜血管铺片观察大鼠视网膜血管形态变化,并行VEGF免疫荧光染色观察大鼠视网膜VEGF的表达改变。结果:FITC-Dextran灌注示糖尿病血糖非严格控制组大鼠视网膜浅层血管多处荧光素聚集,不均匀分布,深层毛细血管局部扩张、迂曲;血糖严格控制组视网膜血管管径较均匀一致,无局部扩张或狭窄。糖尿病血糖非严格控制组VEGF高表达,血糖严格控制组较糖尿病组、血糖非严格控制组视网膜VEGF表达下降,统计学比较具有显著性差异(P<0.05)。结论:胰岛素治疗严格控制血糖至正常水平可以降低糖尿病早期大鼠视网膜VEGF的表达、减轻视网膜血管损害;血糖波动会加重糖尿病视网膜病变的发展。眼科学报2007;23:205-211.     

Streptozotocin induced diabetic retinopathy in rat and the expression of vascular endothelial growth factor and its receptor

AIM: To investigate the characteristics and criterion of graft rejection in mice model.METHODS: C57BL/6 or BALB/c mice corneal grafts were grafted onto BALB/c hosts. Each group was divided into two subgroups according to the corneal opacity scores 12d after transplantation. The characteristics of opacity and neovascularization were observed. Mice of the 12^th, 50^th day after transplantation, the grafts biopsy of mice in allogeneic group 1, which opacity score exceed 3, were prepared for histological observation and those restore transparent were endothelial stained.RESULTS: There was no difference of corneal opacity score on the 7^th and 12^th day after operation; the histological results had no disparity between syngeneic group and allogeneic group. On the 12^th day after surgery, the turbidity curve was apparent in grafts with opacity score ＜2. Mononuclear cells were shown in grafts with opacity score reached 3 in allogeneic group 1. Different rejection performance was observed in tissue sections on the 50^th day after surgery.CONCLUSION:Grafts, opacity score exceeds 3 from the 7^th to the 12^th day after operation could not be judged as a rejection. We should pay more attention to the variation of grafts opacity since 12d after corneal transplantation.     

Resveratrol blocks diabetes‐induced early vascular lesions and vascular endothelial growth factor induction in mouse retinas

Purpose: Vessel leakage and loss of pericytes are early signs of diabetic retinopathy (DR), which leads to vision loss. Upregulation of the vascular endothelial growth factor (VEGF) during diabetes plays a key role in mediating these vascular lesions. The aim of this study is to investigate the effects of resveratrol, a natural plant‐derived phytoalexin, on vascular damage and VEGF induction in mouse retinas of early diabetes. Methods: Diabetes was induced in C57BL/6 mice by five consecutive‐intraperitoneal injections of 55 mg/kg of streptozotocin (STZ). Animals injected with buffer only were used as controls. Beginning 1 month after the fifth injection of STZ or buffer, 20 mg/kg of resveratrol was administered by oral gavage daily for 4 weeks to diabetic and control mice, and all mice were killed 2 months after the injections. We assessed vessel leakage, pericyte loss and VEGF protein expression in mouse retinas of 2‐month diabetes compared with controls with or without resveratrol treatment. Results: Diabetes led to increase vessel leakage, pericyte loss and VEGF protein level in the mouse retinas compared with controls; however, these changes were effectively blocked by resveratrol treatment. Conclusion: Our data suggest that resveratrol is effective to decrease vascular lesions and VEGF induction in mouse retinas of early diabetes.     

Intraocular expression of thymosin β4 in proliferative diabetic retinopathy

Purpose: To examine an association between thymosin β4 as potentially angioproliferative factor and proliferative diabetic retinopathy. Methods: The clinical study part included 62 patients with proliferative diabetic retinopathy (PDR) (study group) and 24 patients with non‐diabetic pre‐retinal membranes (control group). All patients underwent pars plana vitrectomy. We examined the thymosin β4 concentration in vitreous and plasma; and the expression of thymosin β4, glial fibrillary acidic protein (GFAP) and CD31 (PECAM‐1 or Platelet Endothelial Cell Adhesion Molecule) and the levels of thymosin β4 mRNA and vascular endothelial growth factor (VEGF) mRNA in the excised membranes. The experimental study part consisted of 24 Sprague–Dawley rats with streptozotocin‐induced diabetes mellitus and 24 age‐matched control animals without diabetes. We determined the mRNA concentrations of thymosin β4, VEGF and GFAP in the rat retinas. Results: In the clinical study part, the vitreal and plasma thymosin β4 concentrations were significantly higher in the study group than control group (p = 0.04 and p = 0.01, respectively), and were significantly (p = 0.028) correlated with each other. Co‐expression of thymosin β4 and CD31 was observed in the diabetic fibrovascular membranes. Thymosin β4 mRNA and VEGF mRNA levels were significantly (p < 0.01) higher in diabetic membranes than in non‐diabetic membranes. In the experimental study part, the diabetic retinas showed co‐localization of thymosin β4 and GFAP. The mRNA levels of thymosin β4, VEGF and GFAP were significantly (p < 0.01) higher in diabetic rats than in control animals. Conclusions: Thymosin β4 was produced in intraocular fibrovascular membranes of patients with PDR and in rats with experimental diabetes mellitus. Thymosin β4 may play a role in diabetic retinal neovascularization.     

Inhibition of experimental choroidal neovascularization in mice by anti-VEGFA/VEGFR2 or non-specific siRNA

Choroidal neovascularization (CNV) is one of the severe pathological consequences of the end-stage of age-related macular degeneration (AMD). Several lines of evidence implicate increased levels of vascular endothelial growth factor (VEGF) in the retinas of AMD patients. Current available agents for the inhibition of VEGF protein such as bevacizumab show significant promise for the treatment of exudative AMD. However, this compound still has limited efficacy and requires multiple administrations; thus, it is associated with a variety of ocular complications, including endophthalmitis and retinal detachment. In this study, we used anti-VEGFA/VEGFR2 or non-specific siRNA and evaluated their suppression of laser-induced choroidal neovascularization (CNV) in mice. Male adult C57BL/6J mice were used in the study. The mice were subjected to laser rupture of Bruch’s membrane to induce CNV and then randomized to five groups with six mice per group. The five groups were blank control, vehicle control (5% glucose solution, GS), VEGFA.siRNA, VEGFR2.siRNA, and non-specific siRNA. Two days after laser photocoagulation, each group, with the exception of the blank control group, received 1 μl of the appropriate agent by intravitreal injection to both eyes. Seven days later, after taking fundus photography and fundus fluorescein angiography (FFA), the mice were killed for tissue sampling. Six eyes from three mice in each group were used for choroidal flatmounts to examine CNV. Six eyes from three mice in each group were subjected to RNA extraction for VEGF mRNA quantification by qRT-PCR. Retinal tissue from 2 mice without laser treatment was harvested as the assay reference. The incidence of burns which showed fluorescein leakage was 80.0% in blank control, 75.0% in GS, 55.0% in VEGFA.siRNA, 40.0% in VEGFR2.siRNA and 30.0% in non-specific siRNA group. The flatmounted specimens showed that the retinal pigment epithelium (RPE) was visualized as a uniform hexagonal array in nonlasered areas. On day 7 after laser burn, well-defined isolectin-B4 labeled CNV networks were shown within the burn spots of the 2 control groups. The inhibitory effects of the 3 siRNAs on CNV formation were statistically significant as compared to the 2 control groups. Compared to the control groups, the ocular expression of VEGF mRNA was decreased significantly in the 3 siRNA treated groups. The areas of CNV correlated with the expression of VEGF mRNA. Anti-VEGFA/VEGFR2 or non-specific siRNA can inhibit CNV and attenuate VEGF mRNA expression in a laser-induced mouse model of CNV. We conclude that the siRNAs may be an option for treating patients with exudative AMD, and more studies are needed to test the possible side-effects of the treatment.     

Alterations in the polysialylated neural cell adhesion molecule and retinal ganglion cell density in mice with diabetic retinopathy

AIM: To investigate the impact of polysialylated neural cell adhesion molecule (PSA-NCAM) on the survival of retinal ganglion cells (RGCs) in the experimentally induced diabetes in mice. METHODS: Diabetes was induced in 2.5 months old Swiss Webster mice by intraperitoneal injection of streptozotocin (STZ, 90 mg/kg) once daily for two consecutive days. Examination of the proteins of interest in the retinas from diabetic mice at 2mo after diabetes induction was performed using immunohistochemistry and Western blot analysis. RGCs were counted in the wholemounted retinas, and Brn3a marker was used. RESULTS: Examination of retinas from diabetic mice at 2mo after diabetes induction revealed a considerable reduction in RGC density. Our experiments also demonstrated a redistribution of PSA-NCAM in the retina of diabetic animals. PSA-NCAM immunoreactivity was diminished in the inner part of the retina where RGCs were located. In contrast, an enhanced PSA-NCAM immunoreactivity was detected in the outer layers of the retina. PSA-NCAM signal was co-localized with glial fibrillary acidic protein immunoreactivity in the Müller cell branches. Previous studies have shown that matrix metalloproteinase-9 (MMP-9) is responsible for the reduction in PSA-NCAM levels in neuronal cells. The reduced levels of PSA-NCAM in inner layers (nerve fiber layer, ganglion cell layer) were accompanied by the increased expression of MMP-9. In contrast, in the outer retinal layers, the expression of MMP-9 was much less pronounced. CONCLUSION: MMP-9 induces PSA-NCAM shedding in the inner part of the retina and the decreased level of PSA-NCAM in the inner part of the retina might be, at least in part, responsible for the loss of RGCs in diabetic mice.     

CD146/Soluble CD146 Pathway Is a Novel Biomarker of Angiogenesis and Inflammation in Proliferative Diabetic Retinopathy

PurposeInflammation, angiogenesis and fibrosis are pathological hallmarks of proliferative diabetic retinopathy (PDR). The CD146/sCD146 pathway displays proinflammatory and proangiogenic properties. We investigated the role of this pathway in the pathophysiology of PDR.MethodsVitreous samples from 41 PDR and 27 nondiabetic patients, epiretinal fibrovascular membranes from 18 PDR patients, rat retinas, human retinal microvascular endothelial cells (HRMECs) and human retinal Müller glial cells were studied by ELISA, Western blot analysis, immunohistochemistry and immunofluorescence microscopy analysis. Blood-retinal barrier breakdown was assessed with fluorescein isothiocyanate-conjugated dextran.ResultssCD146 and VEGF levels were significantly higher in vitreous samples from PDR patients than in nondiabetic patients. In epiretinal membranes, immunohistochemical analysis revealed CD146 expression in leukocytes, vascular endothelial cells and myofibroblasts. Significant positive correlations were detected between numbers of blood vessels expressing CD31, reflecting angiogenic activity of PDR, and numbers of blood vessels and stromal cells expressing CD146. Western blot analysis showed significant increase of CD146 in diabetic rat retinas. sCD146 induced upregulation of phospho-ERK1/2, NF-κB, VEGF and MMP-9 in Müller cells. The hypoxia mimetic agent cobalt chloride, VEGF and TNF-α induced upregulation of sCD146 in HRMECs. The MMP inhibitor ONO-4817 attenuated TNF-α-induced upregulation of sCD146 in HRMECs. Intravitreal administration of sCD146 in normal rats significantly increased retinal vascular permeability and induced significant upregulation of phospho-ERK1/2, intercellular adhesion molecule-1 and VEGF in the retina. sCD146 induced migration of HRMECs.ConclusionsThese results suggest that the CD146/sCD146 pathway is involved in the initiation and progression of PDR.     

Tonicity response element binding protein associated with neuronal cell death in the experimental diabetic retinopathy

AIM:To studythe contribution of tonicity response element binding protein (TonEBP) in retinal ganglion cell (RGC) death of diabetic retinopathy (DR).METHODS:Diabetes was induced in C57BL/6 mice by five consecutive intraperitoneal injections of 55 mg/kg streptozotocin (STZ). Control mice received vehicle (phosphate-buffered saline). All mice were killed 2mo after injections, and the extent of cell death and the protein expression levels of TonEBP and aldose reductase (AR) were examined.RESULTS:The TonEBP and AR protein levels and the death of RGC were significantly increased in the retinas of diabetic mice compared with controls 2mo after the induction of diabetes. Terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL)-positive signals co-localized with TonEBP immunoreactive RGC. These changes were increased in the diabetic retinas compared with controls.CONCLUSION:The present data show that AR and TonEBP are upregulated in the DR and TonEBP may contribute to apoptosis of RGC in the DR.     

血管内皮生长因子在糖尿病大鼠视网膜中的表达

目的 :观察血管内皮生长因子 (VEGF)蛋白在链脲佐菌素 (STZ)诱导的糖尿病大鼠视网膜中的表达情况 ,以探讨其在早期糖尿病视网膜病变 (DR)中的作用。方法 :选择健康成年Wistar大鼠 4 0只 ,随机分成正常对照组 (CON)、糖尿病 1个月组 (DM1)、糖尿病 3个月组 (DM3)及糖尿病 6个月 (DM6 )组 ,每组 10只。大鼠腹腔内注射STZ诱发大鼠糖尿病。制备大鼠视网膜石蜡切片行免疫组化ABC法染色 ,光镜观察。结果 :VEGF于CON和DM1组只表达于内核层及节细胞层部分细胞 ;病程 3个月时 ,尚可见视网膜血管的阳性反应 ;6个月时 ,阳性反应范围又扩大至视杆内节和色素上皮细胞。结论 :随病程进展 ,VEGF在视网膜中的表达呈逐渐增强的趋势 ,提示它在早期DR的发生、发展中起重要作用     

糖尿病小鼠视网膜中JNK3 mRNA表达的动态变化

背景 糖尿病视网膜病变（DR）是糖尿病常见的眼部并发症,其发病机制与多种因素有关,c-jun N末端激酶（JNK）作为一种凋亡基因,在糖尿病的病理过程中发挥着重要作用,其研究已成为近些年的热点之一,而JNK3作为JNK的亚基因之一,在DR中的研究目前较少.目的 定量检测JNK3在糖尿病小鼠视网膜中的表达,探讨JNK3在DR中的作用.方法 选取SPF级6～8周龄C57BL/6雄性小鼠48只,适应性喂养1周后按照随机数字表法分为糖尿病组和正常对照组.30只糖尿病组小鼠用一次性腹腔内注射柠檬酸钠缓冲液溶解的质量分数1％链脲佐菌素（STZ）的方法诱导糖尿病模型,18只对照组小鼠腹腔内注射等容量柠檬酸钠缓冲液.分别于造模后2、4、8周摘除小鼠左眼眼球,提取视网膜组织,采用实时定量PCR法检测JNK3 mRNA在小鼠视网膜中表达的动态变化.结果 造模后2、4、8周,糖尿病组小鼠血糖水平明显高于正常对照组,差异均有统计学意义（t=-5.675、-5.498、-5.347,P＜0.01）.糖尿病组和正常对照组在造模后不同时间点小鼠视网膜中JNK3 mRNA相对表达量（A值）的差异均有统计学意义（F分组=102.345,P＜0.05;F时间 =131.679,P＜0.05）;其中造模后4周和8周糖尿病组小鼠视网膜中JNK3 mRNA相对表达量分别为3.21±0.14和5.43±0.37,均明显高于正常对照组的2.54 ±0.42和2.26±0.67,差异均有统计学意义（t=4.073、23.399,P＜0.05）;糖尿病组内比较可见,造模后8周小鼠视网膜中JNK3 mRNA相对表达量明显高于2周和4周值,差异均有统计学意义（t=10.756、16.857,P＜0.05）.结论 JNK3在糖尿病小鼠视网膜中表达量上调,其早期表达量随时间延长而增加.JNK3可能参与早期DR的发生和发展.