糖尿病大鼠视网膜毛细血管细胞凋亡及其凋亡相关基因的表达

目的 确定早期糖尿病大鼠视网膜毛细血管细胞凋亡并观测凋亡相关基因（Bax、bcl-2）在早期糖尿病大鼠视网膜的表达。方法 选择健康成年Wistar大鼠40只,随机分成正常对照组（contrast group,CON）和糖尿病（diabetes mellitus)1个月（DM1）、3个月（DM3）及6个月（DM6）组。腹腔内注射链脲佐菌素（streptozotocin,STZ)诱发大鼠糖尿病。制备大鼠视网膜血管辅片及切片,分别行TUNEL法和免疫组化ABC法染色,并对ABC法染色结果进行计算机图像分析。结果 TUNEL法标记的阳性周细胞核出现于DM3和DM6组,而内皮细胞见于DM6组,CON和DM1组未见阳性反应。TUNEL阳性反应细胞核染色质分布不均,表现为环形核、新月形核等典型凋亡细胞特征。ABC法染色：于CON及DM各组,Bax和bcl-2二基因的蛋白均在视网膜血管表达。随病程进展,其阳性反应强度逐渐增加。此外,bcl-2阳性反应在DM3组于色素上皮细胞出现。在DM6组进一步扩大到视杆内节和节细胞。而Bax在DM6组亦出现于细胞。结论 早期糖尿病大鼠视网膜毛细血管周细胞丧失的性质为凋亡,内皮细胞亦存在凋亡;Bax和bcl-2在早期糖尿病大鼠视网膜的表达增加。

Apoptosis of retinal capillary cells in diabetic rats and expression of apoptosis-related genes

OBJECTIVE: To determine the apoptosis of retinal capillary cells in early diabetic rats and observe the expression of apoptosis-related genes (bcl-2, Bax) in retinas of early diabetic rats. METHODS: 40 healthy adult Wistar rats were chosen and randomly divided into four groups: normal control (CON), diabetes mellitus one month (DM1), three months (DM3) and six months (DM6). Diabetic mellitus in rats was induced by streptozotocin (STZ) intraperitoneally. Retinal preparations of blood vessels and paraffin sections were stained by TdT-mediated dUTP nick end labeling (TUNEL) and immunohistochemical (ABC) method respectively and analyzed by computer-picture analytic system on the results of ABC method. RESULTS: Positive pericyte nuclei labeled by TUNEL were emerged in DM3 and DM6, but positive endothelial cell nuclei only in DM6. No positive reaction was seen in CON and DM1. Chromatin of TUNEL positive nuclei was distributed unequally, showing apoptotic characteristics of ring-like and crescent nuclei. In ABC staining method, two genes' (bcl-2, Bax) proteins were expressed in retinal vessels. Along with the progression of diabetes, the intensity of positive reaction was increased in degrees. Moreover, pigment epithelial cells in DM3 showed Bcl-2 positive reaction that again extended further to inner rod segments and ganglion cells in DM6, while in DM6, Bax was expressed in ganglion cells. CONCLUSIONS: The nature of pericyte dropout in early diabetic rats is apoptosis that also exists in endothelial cells; the expression of Bax and bcl-2 in retinas of early diabetic rats is enhancing.