Huang Xiao Yong, Yin Zheng Qin, Tan Xiao Ling
Southwest Eye Hospital, Third Military Medical University, Chongqing, People's Republic of China.
Cell Tissue Res. 2008 Sep;333(3):381-93. doi: 10.1007/s00441-008-0653-1. Epub 2008 Jul 8.
Photoreceptor loss causes irreversible blindness in many retinal diseases. Repair of such damage by cell transplantation is one of the most feasible types of central nervous system treatment. Retinal stem cells (RSC) are a substrate for cell-replacement therapy, and previous studies have shown that RSCs from different developmental stages have distinct properties in proliferative capacity and differentiation potential. The tailbud stage is of special interest in retinogenesis, because RSCs commence differentiation after this period. However, no information about the characteristics of RSCs from the tailbud stage is available. In this study, the characteristics of cell cultures from the rat optic cup (referred to as optic-cup-derived RSCs; OC-RSCs) at embryonic day 12.5 (tailbud stage) were analyzed. OC-RSCs grew either as monolayers or as neurospheres in the presence of basic fibroblast growth factor and could be dissociated into a single cell suspension. Using the MTT assay, immunochemistry, cytogenetic analysis, and flow cytometry, we found that OC-RSCs were easily enriched to 92% by three passages, had a normal diploid karyotype, and exhibited no obvious differences in proliferative rate during eight passages (doubling time: 36 h). OC-RSCs produced retinal specific cells after the addition of serum to the medium, but the differentiation potential was affected by serum concentration. Preliminary results showed that transplanted OC-RSCs were incorporated into the degenerated retina of RCS rats and differentiated into rhodopsin-positive cells. Thus, OC-RSCs, after suitable enrichment, provide a population of stem cells with distinct growth and differentiation properties that make them suitable for research into RSC differentiation and transplantation.
在许多视网膜疾病中，光感受器丧失会导致不可逆的失明。通过细胞移植修复此类损伤是中枢神经系统治疗中最可行的方法之一。视网膜干细胞（RSC）是细胞替代疗法的一种底物，先前的研究表明，来自不同发育阶段的RSC在增殖能力和分化潜能方面具有不同的特性。尾芽期在视网膜发生过程中具有特殊意义，因为在此阶段之后RSC开始分化。然而，关于尾芽期RSC特性的信息尚无报道。在本研究中，我们分析了胚胎第12.5天（尾芽期）大鼠视杯细胞培养物（称为视杯来源的RSC；OC-RSC）的特性。在碱性成纤维细胞生长因子存在的情况下，OC-RSC可以单层生长或形成神经球，并且可以解离成单细胞悬液。通过MTT法、免疫化学、细胞遗传学分析和流式细胞术，我们发现经过三次传代后，OC-RSC很容易富集到92%，具有正常的二倍体核型，并且在八次传代过程中增殖率没有明显差异（倍增时间：36小时）。向培养基中添加血清后，OC-RSC产生视网膜特异性细胞，但分化潜能受血清浓度影响。初步结果表明，移植的OC-RSC被整合到RCS大鼠的退化视网膜中，并分化为视紫红质阳性细胞。因此，经过适当富集的OC-RSC提供了一群具有独特生长和分化特性的干细胞，使其适合用于RSC分化和移植的研究。
