不同体细胞来源人诱导多能干细胞向视网膜类器官分化能力的评价  

Evaluation of the retinal differentiation ability of human induced pluripotent stem cells reprogrammed from different somatic cells

作　　者：谢冰冰[1] 关远远 高冠杰 郑丹丹 徐平 宋小景 钟秀风[1] Xie Bingbing;Guan Yuanyuan;Gao Guanjie;Zheng Dandan;Xu Ping;Song Xiaojing;Zhong Xiufeng(State Key Laboratory of Ophthalmology,Zhongshan Ophthalmic Center,Sun Yat-sen University,Guangzhou 510060,China)

机构地区：[1]中山大学中山眼科中心眼科学国家重点实验室,广州510060

出　　处：《中华实验眼科杂志》2020年第10期821-827,共7页Chinese Journal Of Experimental Ophthalmology

基　　金：广东省科技计划项目(2017B020230003);广州市科技计划项目(201803010078);科技部国家重点研发计划项目(2017YFA0104101、2016YFC1101103);国家自然科学基金项目(81570874、81970842);中山大学百人计划项目(PT1001010);中山大学中山眼科中心眼科学国家重点实验室PI经费支持项目。

摘　　要：目的评价不同体细胞来源的人诱导多能干细胞(hiPSCs)向3D视网膜类器官分化的能力。方法采用血液细胞重编程获得的hiPSCs系绿色荧光蛋白(GFP)标记的BC1细胞(BC1-GFP)及Gibco、尿液细胞重编程获得的hiPSCs系UE017进行视网膜诱导分化。光学显微镜下记录视网膜形态的发生和发展过程,采用免疫荧光染色法检测获得的视网膜中各种细胞亚类的特异性分子标志物表达情况,对不同细胞系分化视网膜的效率进行分析和比较。结果血液和尿液细胞来源的hiPSCs均可成功诱导分化为3D视网膜类器官,包括神经视网膜和视网膜色素上皮细胞。视网膜类器官能够模拟体内视网膜的发育过程,逐渐分化出所有神经视网膜亚类细胞,包括视网膜神经节细胞、光感受器细胞、无长突细胞、水平细胞、双极细胞和Müller细胞,甚至形成板层状结构。2种体细胞来源的hiPSCs在分化为视网膜类器官的效率上存在差异,血液来源的细胞比尿液来源的细胞分化为视网膜类器官的效率更高。结论血液和尿液体细胞来源的hiPSCs均可诱导出3D视网膜类器官,获得的视网膜类器官中包括所有视网膜细胞亚类,但2种体细胞来源的hiPSCs分化成视网膜类器官的效率不同。Objective To evaluate the retinal differentiation ability of human induced pluripotent stem cells(hiPSCs)from various somatic cell sources.Methods The hiPSCs lines BC1-green fluorescent protein(GFP)and Gibco obtained by blood cell reprogramming and the hiPSCs line UE017 obtained by urine cell reprogramming were used to induce retinal differentiation.The morphogenesis and development of retina were recorded with an optical microscope,and the expression of specific molecular markers of various cell subclasses in the retina was detected by immunofluorescence,and the efficiency of retinal differentiation of different cell lines was analyzed and compared.Results All three hiPSC lines derived from blood and urine cells were able to be induced into three-dimensional(3D)retinal organoids,including neuroretina and retinal pigment epithelial cells.Retinal organoids simulated the development process of retina in vivo and gradually differentiated into all cell subtypes of retina,including retinal ganglion cells,photoreceptor cells,amacrine cells,horizontal cells,bipolar cells,Müller cells,and even formed lamellar structures.However,in terms of the efficiency of acquiring retinal organoids,the hiPSCs derived from blood were more efficient than those derived from urine.Conclusions hiPSCs from both blood and urine somatic cells can differentiate into 3D retinal organoids,including all subtypes of retinal cells.The differentiation efficiency among lines is different.

关 键 词：人诱导多能干细胞 诱导分化 视网膜类器官 视网膜亚类细胞 

分 类 号：R774.1[医药卫生—眼科]