Changes in gene expression in the retina of diabetic Long Evans rats after one, four, and twelve weeks of diabetes

Purpose. Patients with diabetic retinopathy may experience severe vision loss due to macular edema and neovascularization secondary to vascular abnormalities. However, before these abnormalities become apparent, there are functional deficits in contrast sensitivity, color perception, and dark adaptation. The goals of this study are to evaluate early changes (up to 3 months) in retinal gene expression, selected visual cycle proteins, and optokinetic tracking (OKT) in streptozotocin (STZ)-induced diabetic rats.Methods. Retinal gene expression in diabetic Long Evans rats was measured by whole genome microarray 7 days, 4 weeks, and three months after onset of hyperglycemia. Select gene and protein changes were probed by PCR and immunohistochemistry respectively, and OKT thresholds were measured using a virtual optokinetics system. Results. Microarray analysis showed that the most consistently affected molecular and cellular functions were cell-to-cell signaling and interaction, cell death, cellular growth and proliferation, molecular transport, and cellular movement. Further analysis revealed reduced expression of several genes encoding visual cycle proteins including lecithin:retinol acyltransferase (LRAT), retinal pigment epithelium (RPE)-specific protein 65kDa (RPE65) and RPE retinal G protein coupled receptor (RGR). Immunohistochemistry revealed a decrease in RPE65 in the RPE layer of diabetic rats. These molecular changes occurred simultaneously with a decrease in OKT thresholds by 4 weeks of diabetes. Conclusions. The data presented here are further evidence that inner retinal cells are affected by hyperglycemia prior to vasculopathy suggesting that glial and neuronal dysfunction may underlie some of the early visual deficits in diabetics.研究目的 糖尿病视网膜病变患者可因血管异常继发的黄斑水肿与新生血管形成出现严重视力丧失。然而在这些血管异常显现之前，患者的对比敏感度、色觉与暗适应功能已存在缺损。本研究旨在评估链脲佐菌素（streptozotocin, STZ）诱导的糖尿病大鼠视网膜基因表达、筛选出的视觉循环蛋白以及视动追踪（optokinetic tracking, OKT）的早期变化（最长至3个月）。 研究方法 对高血糖造模后7天、4周及3个月的糖尿病朗伊文斯（Long Evans）大鼠进行全基因组微阵列检测，以分析其视网膜基因表达水平。分别采用聚合酶链式反应（PCR）与免疫组织化学法探究目标基因与蛋白的表达变化，并通过虚拟视动系统检测OKT阈值。 研究结果 微阵列分析结果显示，受影响最持续的分子与细胞功能包括细胞间信号传导与相互作用、细胞死亡、细胞生长与增殖、分子转运以及细胞迁移。进一步分析发现，多种编码视觉循环蛋白的基因表达水平下调，其中包括卵磷脂：视黄酰基转移酶（lecithin:retinol acyltransferase, LRAT）、视网膜色素上皮（RPE）特异性65kDa蛋白（RPE65）以及RPE视网膜G蛋白偶联受体（RGR）。免疫组织化学检测结果显示，糖尿病大鼠视网膜色素上皮层内的RPE65表达量降低。上述分子变化与糖尿病造模4周时OKT阈值的下降同步出现。 研究结论 本研究提供的实验数据进一步证实，在血管病变发生之前，视网膜内层细胞已受高血糖影响，这提示神经胶质细胞与神经元功能障碍可能是糖尿病患者早期视觉缺损的部分诱因。