胚胎干细胞C3B鼠视网膜下腔移植分化研究

Embryonic stem cells transplant into subretinal space of C3B mouse

目的:探讨胚胎干细胞在具有正常视网膜结构的C3B鼠视网膜下腔中的诱导分化情况。方法:胚胎干细胞传1代后进行拟胚体培养。拟胚体消化成单细胞后联合视黄酸注入C3B鼠视网膜下腔,注射后1周、3周、2月处死小鼠取材进行病理切片、电镜检查和免疫组化检测。结果:1周时,可见到注射部位视网膜层水肿增厚。3周和2个月时,3只移植眼内出现畸胎瘤,占所有移植眼的25%,其余眼球注射部位仅见瘢痕组织或眼球萎缩。电镜发现增殖的细胞核异型性明显,具肿瘤细胞特征。免疫组化显示:畸胎瘤部分区域MAP-2强阳性反应;可见团状或集落状细胞GFAP强阳性反应;个别细胞Nestin阳性反应。结论:胚胎干细胞移植入具有正常视网膜结构的C3B鼠视网膜下腔,未能出现ESC向视网膜组织分化或嵌入视网膜组织,相当部分的鼠眼出现了畸胎瘤,其临床安全性和致瘤性是非常值得关注的问题。

AIM： To study the differentiation of embryonic stem cells （ESC） at the subretinai space of C3B mouse whose retina are of normal structure. METHODS ： Embryonic bodies （EB） were gained when embryonic stem cells were propagated 1 generation after anabiosis. The digested EB were transplanted into subretinai space of C3B mouse. The eyes were analysed by photo microscopy, electric microscopy and immune assay at 1st week, 3rd week and 2nd month after injection. RESULTS ： The thickened retina appeared on the place of injection at I st week. Teratoma formation was observed in 3 transplant recipients at 3rd week and 2nd month. The ratio of forming tumor was 25% in all eyes received cell transplant. Eye atrophy or scar at injection position was seen in other eyes. The nucleus of tumour cells had apparent strange type under electrical microscopy. The immune assay showed that part of the tumour were microtubule - associated protein 2 （ MAP -2） positive; part of the tumour were gliai fibrillary acid protein （GFAP） positive; a little cells were nestin positive. CONCLUSION： After transplantation into the subretinai space, ESC were not incorporated into retina or differentiated into retinal cells. The teratoma was formed in part of eyes. The security of application for ESC in eye was considerable.