Growth kinetics and transplantation of human retinal progenitor cells
Growth kinetics and transplantation of human retinal progenitor cells
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DOI：
10.1016/j.exer.2009.03.025
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发表时间：
2009-09-01
影响因子：
3.4
通讯作者：
Young, Michael
中科院分区：
文献类型：
--
作者：
Aftab, Unber;Jiang, Caihui;Young, Michael
我们研究了从不同胎龄（G.A.）供体组织中分离的人视网膜祖细胞（hRPC）的生长动力学，确定hRPC是否可以分化为成熟的光感受器，并评估它们在移植后与退化宿主视网膜整合的能力。经 IRB 批准获得眼睛（生后 12-18 周），并通过酶法分离视网膜。细胞在体外扩增，在分离时和每次传代时进行计数，并使用免疫细胞化学和PCR进行表征。将 GFP 阳性 hRPC 与 rd1 和视紫红质 -/- 小鼠的视网膜外植体共培养，或移植到视网膜光凝和视紫红质 -/- 小鼠的 B6 小鼠中。移植后摘取眼睛进行组织学评估。我们的结果表明，16 至 18 周 G.A. 的 hRPC。其体外存活时间最长，细胞数量最多，可增殖至少 6 代。这些细胞表达视网膜干细胞标记物巢蛋白、Ki-67、PAX6 和 Lhx2，并在用血清分化后光感受器标记物染色呈阳性。一些用于移植研究的 GFP 阳性细胞显示出迁移至退化宿主视网膜并表达视紫红质的证据。总之，我们确定了 hRPC 的生长动力学，并表明来自 16-18 周 G.A. 供体组织的细胞。在特定条件下表现出最佳的增殖动力学，并且 hRPC 也可以沿着光感受器谱系进行分化。此外，我们还证明，移植后，其中一些细胞在宿主视网膜内整合并分化以表达视紫红质，从而支持 hRPC 移植在视网膜退行性疾病中的潜在用途。 (c) 2009 Elsevier Ltd. 保留所有权利。
We studied the growth kinetics of human retinal progenitor cells (hRPCs) isolated from donor tissue of different gestational ages (G.A.), determined whether hRPCs can be differentiated into mature photo-receptors and assessed their ability to integrate with degenerating host retina upon transplantation. Eyes (12-18 weeks G.A.) were obtained with IRB approval and retinas were enzymatically dissociated. Cells were expanded in vitro, counted at isolation and at each passage, and characterized using immunocytochernistry and PCR. GFP positive hRPCs were co-cultured with retinal explants from rd1 and rhodopsin -/- mice, or transplanted into B6 mice with retinal photocoagulation and rhodopsin -/- mice. Eyes were harvested for histological evaluation following transplantation. Our results show that hRPCs from 16 to 18 weeks G.A. had the longest survival in vitro and yielded the maximum number of cells, proliferating over at least 6 passages. These cells expressed the retinal stem cell markers nestin, Ki-67, PAX6 and Lhx2, and stained positively for photoreceptor markers upon differentiation with serum. Some of the GFP positive cells used for transplantation studies showed evidence of migration into the degenerative host retina and expressed rhodopsin. In conclusion, we have determined the growth kinetics of hRPCs and have shown that cells from donor tissue of 16-18 weeks G.A. exhibit the best proliferative dynamics under the specified conditions, and that hRPCs can also be differentiated along the photoreceptor lineage. Further, we have also demonstrated that following transplantation, some of these cells integrate within the host retina and differentiate to express rhodopsin, thereby supporting the potential utility of hRPC transplantation in the setting of retinal degenerative disorders. (c) 2009 Elsevier Ltd. All rights reserved.