Chen Xi, Chen Zehua, Li Zhengya, Zhao Chen, Zeng Yuxiao, Zou Ting, Fu Caiyun, Liu Xiaoli, Xu Haiwei, Yin Zheng Qin
Southwest Hospital/Southwest Eye Hospital, Third Military Medical University, Chongqing, 400038, China.
Key Lab of Visual Damage and Regeneration & Restoration of Chongqing, Chongqing, 400038, China.
Stem Cell Res Ther. 2016 Dec 30;7(1):191. doi: 10.1186/s13287-016-0451-8.
Despite diverse pathogenesis, the common pathological change observed in age-related macular degeneration and in most hereditary retinal degeneration (RD) diseases is photoreceptor loss. Photoreceptor replacement by cell transplantation may be a feasible treatment for RD. The major obstacles to clinical translation of stem cell-based cell therapy in RD remain the difficulty of obtaining sufficient quantities of appropriate and safe donor cells and the poor integration of grafted stem cell-derived photoreceptors into the remaining retinal circuitry.
Eye-wall c-kit/stage-specific embryonic antigen 1 (SSEA1) cells were isolated via fluorescence-activated cell sorting, and their self-renewal and differentiation potential were detected by immunochemistry and flow cytometry in vitro. After labeling with quantum nanocrystal dots and transplantation into the subretinal space of rd1 RD mice, differentiation and synapse formation by daughter cells of the eye-wall c-kit/SSEA1 cells were evaluated by immunochemistry and western blotting. Morphological changes of the inner retina of rd1 mice after cell transplantation were demonstrated by immunochemistry. Retinal function of rd1 mice that received cell grafts was tested via flash electroretinograms and the light/dark transition test.
Eye-wall c-kit/SSEA1 cells were self-renewing and clonogenic, and they retained their proliferative potential through more than 20 passages. Additionally, eye-wall c-kit/SSEA1 cells were capable of differentiating into multiple retinal cell types including photoreceptors, bipolar cells, horizontal cells, amacrine cells, Müller cells, and retinal pigment epithelium cells and of transdifferentiating into smooth muscle cells and endothelial cells in vitro. The levels of synaptophysin and postsynaptic density-95 in the retinas of eye-wall c-kit/SSEA1 cell-transplanted rd1 mice were significantly increased at 4 weeks post transplantation. The c-kit/SSEA1 cells were capable of differentiating into functional photoreceptors that formed new synaptic connections with recipient retinas in rd1 mice. Transplantation also partially corrected the abnormalities of inner retina of rd1 mice. At 4 and 8 weeks post transplantation, the rd1 mice that received c-kit/SSEA1 cells showed significant increases in a-wave and b-wave amplitude and the percentage of time spent in the dark area.
Grafted c-kit/SSEA1 cells restored the retinal function of rd1 mice via regulating neural plasticity and forming new graft-to-host synapses.
尽管发病机制多样，但年龄相关性黄斑变性和大多数遗传性视网膜变性（RD）疾病中观察到的共同病理变化是光感受器丧失。通过细胞移植替换光感受器可能是RD的一种可行治疗方法。基于干细胞的细胞疗法临床转化的主要障碍仍然是难以获得足够数量合适且安全的供体细胞，以及移植的干细胞衍生光感受器与剩余视网膜回路的整合不良。
通过荧光激活细胞分选分离眼壁c-kit/阶段特异性胚胎抗原1（SSEA1）细胞，并在体外通过免疫化学和流式细胞术检测其自我更新和分化潜能。用量子纳米晶点标记后移植到rd1 RD小鼠的视网膜下间隙，通过免疫化学和蛋白质印迹评估眼壁c-kit/SSEA1细胞子代的分化和突触形成。通过免疫化学显示细胞移植后rd1小鼠视网膜内层的形态变化。通过闪光视网膜电图和明/暗转换试验测试接受细胞移植的rd1小鼠的视网膜功能。
眼壁c-kit/SSEA1细胞具有自我更新和克隆能力，并且在超过20代中保持其增殖潜能。此外，眼壁c-kit/SSEA1细胞能够分化为多种视网膜细胞类型，包括光感受器、双极细胞、水平细胞、无长突细胞、穆勒细胞和视网膜色素上皮细胞，并且在体外能够转分化为平滑肌细胞和内皮细胞。眼壁c-kit/SSEA1细胞移植的rd1小鼠视网膜中突触素和突触后致密蛋白-95的水平在移植后4周显著增加。c-kit/SSEA1细胞能够分化为功能性光感受器，在rd1小鼠中与受体视网膜形成新的突触连接。移植还部分纠正了rd1小鼠视网膜内层的异常。在移植后4周和8周，接受c-kit/SSEA1细胞的rd1小鼠的a波和b波振幅以及在暗区停留时间的百分比显著增加。
移植的c-kit/SSEA1细胞通过调节神经可塑性和形成新的移植物与宿主突触恢复了rd1小鼠的视网膜功能。
