兔角膜内皮细胞移植治疗角膜内皮损伤

目的 研究兔角膜内皮细胞（CECs）移植的可行性和移植后细胞的形态和功能。方法 体外培养兔CECs并用Brdu标记后接种在Gelatin膜载体上;采用α-氰基丙烯酸脘基酯作为组织黏合剂,将接种有兔CECs的载体与机械去除了内皮细胞的自体兔角膜植片相黏合,原位缝合对21只新西兰白兔的右眼进行CECs移植;另外17只兔的右眼作为对照仅移植没有接种任何细胞的载体膜。术后1、2、4、8、12周观察兔眼角膜移植片的透明情况、眼压和植片厚度,并利用共焦显微镜进行移植细胞的形态观察和计数。观察12周后处死动物,角膜标本分别进行组织切片、免疫组织化学染色和电镜观察。结果 CECs在Gelatin膜载体上生长良好,体外培养3—5d后细胞汇合,细胞密度可以达到约2700个／mm^2。术后2周,所有植片均发生了水肿,并逐渐浑浊。CECs组植片在术后4周左右逐渐水肿减退,开始恢复透明;而对照组角膜植片则持续水肿,逐渐浑浊。术后移植细胞的密度随时间推移逐渐降低,术后12周时,移植细胞的平均密度为（2023±330）个／mm^2。Brdu单克隆抗体免疫组织化学染色显示,植片上的细胞为移植细胞。结论 Gelatin膜作为内皮细胞培养载体并进行内皮细胞移植是一种可行的方法。

Corneal endothelial cell transplantation for cornea endothelium cell destruction in rabbits

OBJECTIVE: To evaluate the feasibility of using cultured corneal endothelial cell (CECs) transplantation for cornea endothelium cell destruction with Gelatin membrane as the carrier in rabbits. METHODS: The cultured CECs were labeled by Brdu and subcultured in vitro on glutaraldehyde-fixed Gelatin membranes and then the membranes were glued by alpha-cyanoacrylate alkyl to 7.00 mm autologous rabbit corneal bottons whose endothelium were mechanically removed previously. The buttons were sutured in place. With this method the right eyes of 21 rabbits were transplanted with CECs, and the right eyes of another 17 rabbits were transplanted with non-cells carrier as controls. The rabbits were bred and observed by slit microscopy and confocal microscopy at 1, 2, 4, 8, and 12 weeks after surgeries. Also, introcular pressure and corneal thickness were measured by Perkin's tonometer and ultrasonic pachymeter. After 12 weeks, all the animals were sacrificed and the grafts were examined by light microscopy and electronic microscopy. RESULTS: CECs grew well on the gelatine memberane, and formed confluent monolayers in 3-5 days; the cell density reached as high as 2700 cells/mm2. After 2 weeks of operation, all corneal buttons were edema and began to be opaque. The control eyes remained opaque throughout the observation period. In eyes with CECs transplanted, the grafts began to be clear and thin 4 weeks after operation. The cell density of grafts decreased along with time, and the mean cell density of CECs transplantation buttons was (2023.3 +/- 330.3) cells/mm2 12 weeks after operation. The transplanted cells were stained with the anti-Brdu monoclonal antibody. CONCLUSION: It is feasible to culture and translate CECs with the Gelatin membrane.