玻璃体腔注射人脐带间充质干细胞诱导分化的神经干细胞对糖尿病大鼠视网膜脑源性神经营养因子表达及视网膜神经节细胞计数的影响

摘要

Objective To investigate the effect of intravitreal injection of neural stem cells (NSC)derived from human umbilical cord mesenchymal stem cells (hUCMSC) on the expression of brain-derived neurotrophic factor (BDNF) and the number of retinal ganglion cells (RGC).Methods Fifty-two adult male Sprague-Dawley rats were randomly divided into normal group (group A) and diabetes mellitus group which received intraperitoneal injection of streptozocin to make diabetic rat models.One month after the diabetic rat models were confirmed successfully,diabetic rats were randomly divided into diabetic group (group B),hUCMSC group (group C) and hUCMSC-induced NSC group (group D).And thirteen diabetic rats were included in each group.Immuno-cytochemistry was applied to observe BDNF and thymosin-1 (Thy-1) staining in the retina.Then mean integrated absorbance of the staining region on the retina slices were analyzed by Image-Pro Plus 6.0.The number of Thy-1 labeled RGC was record.Results BDNF and Thy-1 were positive on the retina slices from group A.The staining intensity from group B became weak and the expression of BDNF and Thy-1 gradually decrease with time (P＜0.05),and those from group C and group D were positively (P＜0.05),especially in group D (P＜0.05).The BDNF expression and Thy-1 labeled RGC were the same between group B and C (P＞0.05) at 2 weeks after injection,but were significant different for other time points (P＜0.05).Significant positive correlation between the expression of BDNF and the number of RGC were found by the Pearson correlation analysis (r=0.964,P＜0.05).Conclusion Intravitreal injection of hUCMSC-derived NSC to diabetic rat may protect the retina by promoting the expression of BDNF and increasing the number of RGC.%目的 观察玻璃体腔注射人脐带间充质干细胞(hUCMSC)诱导分化的神经干细胞(NSC)对糖尿病大鼠视网膜脑源性神经营养因子(BDNF)表达及视网膜神经节细胞计数(RGC)的影响.方法 成年雄性Sprague-Dawley大鼠52只,随机数字表法将其分为正常对照组(A组)、假干预组(B组)、玻璃体腔注射hUCMSC组(C组)、玻璃体腔注射NSC组(D组),每组13只大鼠.B、C、D组大鼠腹腔注射链尿佐菌素建立糖尿病模型;并于建模后1个月,分别对其左眼行玻璃体腔注射无菌磷酸盐缓冲液、hUCMSC、NSC各2μl.干预后2、4、6、8周,免疫组织化学染色观察视网膜BDNF和胸腺肽-1(Thy-1)阳性染色情况,图像分析软件分析视网膜染色区BDNF平均累积吸光度[A,旧称光密度(OD)](M)值,并计数Thy-1阳性染色RGC.对比分析视网膜阳性染色区BDNF平均IA值与Thy-1阳性RGC计数的关系.结果 免疫组织化学染色观察发现,A组大鼠视网膜BDNF和Thy-1呈阳性染色;B组大鼠视网膜BDNF和Thy-1阳性染色随干预后时间延长而逐渐减少;C、D组大鼠视网膜BDNF和Thy-1阳性染色随干预后时间延长而逐渐增加,D组表现更为明显.除干预后2周,B、C组大鼠视网膜染色区BDNF平均IA值、Thy-1阳性RGC计数间差异无统计学意义外(P＞0.05);其余各时间点各组大鼠视网膜染色区BDNF平均IA值、Thy-1阳性RGC计数比较,差异均有统计学意义(P＜0.05).相关性分析结果显示,糖尿病大鼠视网膜染色区BDNF平均IA值与Thy-1阳性RGC计数呈明显正相关(r=0.964,P=0.000).结论 玻璃体腔注射hUCMSC诱导分化的NSC能促进糖尿病大鼠视网膜BDNF表达,提高RGC计数.