胎兔视网膜色素上皮移植的初步研究

目的:观察受体Bruch膜和视网膜色素上皮(retrial pigment epithelial,RPE)受损情况下供体RPE细胞的增殖、分化和凋亡状况。方法:制备有色素胎兔RPE细胞悬液,移植至破坏了Bruch膜和RPE细胞的12只成年新西兰大白兔的视网膜下腔。左眼作为实验组,右眼作为对照,于术后3、7和14d, 采用Ki-67免疫组化和TUNEL染色,并提取RPE细胞DNA作琼脂糖凝胶电泳,观察Ki-67阳性RPE细胞及移植的RPE细胞和受体ONL细胞的凋亡百分率,行统计学分析。结果:术后随着时间的延长,实验组或对照组Ki-67阳性RPE细胞数显著增加(P<0.05);术后14 d实验组或对照组的TUNEL染色ONL核阳性百分率较术后3 d显著减少(P<0.05);术后实验组TUNEL染色阳性和细胞核深染的RPE细胞无显著增加(P>0.05);细胞核深染的RPE细胞百分率明显高于TUNEL阳性的细胞(P<0.01)。对照组未见TUNEL阳性的RPE细胞。术后14d,移植的RPE细胞DNA电泳出现典型的凋亡带。结论:在受体Bruch 膜、RPE受损状态下,供体的RPE细胞具有良好的增殖和分化能力。

Primary study of transplanted embryonic retinal pigment epithelial cells in rabbits

Objective: To observe the proliferation, differentiation and apoptotis of transplanted embryonic retinal pigment epithelial (RPE) cells in rabbits with debrided Bruch's membrane and RPE cells, and investigate the experimental possibility of allogeneic RPE transplants in treating age-related macular degeneration (AMD). Methods: Embryonic RPE cells obtained from pigmented rabbits were transplanted into the subretinal space of 12 adult albino rabbits with damaged Bruch's membrane and RPE. Right eyes as transplanted hosts, left eyes as controls. The transplantation sites were examined on the 3rd, 7th and 14th day after surgery,immunohistochemically stained with Ki-67, terminal deoxy-transferase-mediated dUTP nickend labeling (TUNEL) stained and agarose gel electrophoreticed with DNA of RPE. Statistically analyzed the percentage of Ki-67 positive cells, of TUNEL positive cells of transplanted PRE and of host's outer nuclear layer(ONL). Results: Ki-67 positive RPE cells significantly increased with time in transplanted groups or control (P<0.05). The percentage of TUNEL positive ONL nuclei was not sig nificant difference between transplanted groups and control (P>0.05); but it was significantly different in the 3rd day from the 14th day after surgery (P<0.05). Among the three time points, the percentage of darkly stained RPE cells significantly outnumbered TUNEL positive RPE cells (P<0.01). Fourteen days after surgery, typical DNA ladder appeared on the electrophoresis of transplanted RPE cells. Conclusion: Embryonic RPE cells suspension transplanted in rabbits with debrided Bruch's membrane and RPE cells have the ability of proliferation and differentiation.