大鼠角膜缘上皮细胞培养与同体移植的实验研究

Experimental study on the culture and autotransplantation of limbal epithelium cells in rats

目的:观察以明胶为载体培养的角膜缘上皮细胞移植治疗角膜缘干细胞缺乏症的疗效。方法:大鼠角膜缘上皮细胞在铺有明胶载体的细胞培养板上进行培养5d后,角膜上皮细胞移植术前24h用3H胸腺嘧啶核苷标记培养的角膜缘上皮细胞,培养标记的角膜缘上皮细胞对角膜缘干细胞缺乏的大鼠动物模型行角膜缘上皮细胞同体移植术,对移植术后角膜进行观察、病理学检查及同位素检测。结果:大鼠角膜缘上皮细胞可以在明胶载体上培养、增殖、分化为密集角膜上皮细胞层;角膜缘上皮细胞移植术后角膜上皮完整、基质细胞浸润减轻、新生血管减少。病理学检查角膜缘及周边部上皮细胞为多层结构,角膜新生血管消失及基质中炎性细胞浸润减轻。角膜缘上皮细胞移植术后4wk受眼角膜仍可测到3H胸腺嘧啶核苷。结论:角膜缘上皮细胞移植治疗角膜缘干细胞缺乏症可恢复角膜缘干细胞缺乏病变角膜上皮结构的完整性,减少角膜新生血管的形成,维持角膜缘干细胞的屏障功能,为角膜移值提供更好的条件。

AIM： To study treatment of autotransplantation on limbal stem cells deficiency by cultured limbal epithelium cells cultured on gelatin. METHODS： Limbal epithelium cells had been cultured on gelatin in DMEM/HamF12 medium for 5 days,and cultured limbal epithelium cells had been marked with 3HTDR for 5 days,then limbal epithelium cells and gelatin were transplanted on the limbus and sclera of the rat model with limbal stem cells deficiency by autotransplantation.The corneal changes were observed by a slit-lamp every day,the corneaI pathological changes and 3HTDR content were examined. RESULTS： Rat limbal epithelium cells continued to proliferate, differentiate and form multiple cell layers on gelatin.After autotransplantation with cultured epithelium cells and gelatin, the rat epithelium showed corneal phenotype and progressive decrease of new vessels and stromal infiltration in the limbal and peripheral zone.Pathological examination verified that the limbal and peripheral corneal epithelium was composed of multilayer cells;the neovascularization was reduced and stromal inflammatory cells were decreased.The limbal 3HTDR content by isotope was examined four weeks after operation. CONCLUSION： Autotransplantation with cultured limbal epithelium cells can restore the composition of corneal epithelial cell,decrease neo-vascularization,maintain the function of limbal cellular barrier and provide better condition for later keratoplasty.