Evaluation of inhibitory effects of neuroprotective treatment on diabetic retinopathy

为探讨尼普拉地洛和拉坦前列素对视网膜神经节细胞（retina ganglion cells，RGCs）的抗凋亡作用，采用去血清法和链脲佐菌素（streptozotocin，STZ）诱导糖尿病大鼠视网膜细胞凋亡。通过激活的caspase-3免疫组织化学或末端dUTP缺口末端标记染色定量细胞凋亡。尼帕地洛和拉坦前列素酸抑制RGC-5细胞凋亡，具有剂量依赖性（p=0.001），而去硝基尼帕地洛没有。抑制剂研究表明，尼普拉地洛的抗凋亡作用是蛋白激酶A和G依赖性的，而拉坦前列素酸是MAPK依赖性的。CREB显性负突变体的转染也阻断了尼普拉洛的拯救作用。当滴注5天时，与对照组相比，4周和12周糖尿病视网膜的视网膜神经节细胞层中的凋亡细胞均减少（分别为p<0.05和p<0.0001）。去硝基尼普拉洛滴眼液未显示任何影响。拉坦前列素滴眼液可显著增加大鼠视网膜MAPR的磷酸化水平，提示尼普拉地洛和拉坦前列素通过不同的细胞内信号通路保护糖尿病诱导的RGCs凋亡

In order o investigate if nipradilol and latanoprost have anti-apoptotic effects in retinal ganglion cells (RGCs), apoptosis was induced in RGC-5 cells by withdrawing serum and in rat retina by inducing diabetes with streptozotocin (STZ) injection. Apoptosis was quantified by activated caspase-3 immunohistochemistry on retinal cryosection or terminal dUTP nick end labeling staining. Nipradilol and latanoprost acid suppressed apoptosis of RGC-5 cells with dose dependency (p=0.001), whereas desnitro-nipradilol did not. Inhibitor studies showed that the anti-apoptotic effects of nipradilol were protein kinase A and G-dependent, whereas those of latanoprost acid were MAPK-dependent. Transfection of dominant negative mutants of CREB also blocked the nipradilol rescue effect. When instilled for 5 days, both nipradilol and latanoprost reduced apoptotic cells in retinal ganglion cell layer of 4-week and 12-week diabetic retina than controls (p<0.05 and p<0.0001, respectively). Desnitro-nipradilol eye drops did not show any effects. Latanoprost eye drops significantly increased MAPR phosphorylation in rat retina .These results indicated that nipradilol and latanoprost rescue RGCs from apoptosis induced by diabetes through distinct intracellular signaling pathways