Notch通路参与调控成年小鼠c-kit+视网膜前体细胞分裂和分化修复NMDA视网膜损伤  

Notch pathway participates in regulation of c-kit+retinal precursor cell division and differentiation in adult mice for repair of NMDA-induced retinal injury

作　　者：陈曦[1] 李珊珊 赵露[1] 尤冉 王艳玲[1] CHEN Xi;LI Shanshan;ZHAO Lu;YOU Ran;WANG Yanling(Department of Ophthalmology,Beijing Friendship Hospital Affiliated to Capital Medical University,Beijing,100050,China)

机构地区：[1]首都医科大学附属北京友谊医院眼科,北京100050

出　　处：《第三军医大学学报》2021年第1期59-67,共9页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(81870686);北京市自然科学基金青年项目(7184201);首都卫生发展科研专项(首发2018-1-2021)。

摘　　要：目的分析成年小鼠视网膜中c-kit+内源性前体细胞的生物学特性及其在N-甲基-D-天冬氨酸(N-methyl-D-aspartic acid,NMDA)视网膜损伤修复中的作用和机制。方法选取4周龄成年C57BL/6J小鼠,采用NMDA诱导视网膜损伤。免疫荧光检测c-kit+视网膜前体细胞(retinal precursor cells,RPCs)与c-kit配体干细胞因子(stem cell factor,SCF)阳性细胞的关系以及NMDA损伤后1、2、4周视网膜中c-kit+RPCs细胞数量及其分裂状态。荧光定量PCR对比NMDA损伤后不同阶段视网膜中Hes1、Hes5、S100b和Pou4f1的表达变化。结果在成年小鼠视网膜内核层存在SCF+Müller细胞,包围在c-kit+RPCs周围。c-kit+RPCs仍可保持低水平的细胞分裂。在NMDA损伤后2周,c-kit+RPCs细胞数量明显升高(P<0.001),处于对称分裂和不对称分裂期的细胞比例也显著升高(P=0.016),伴随c-kit+GAD65/67+细胞/c-kit+细胞比例升高(P<0.001)。在NMDA损伤后2周,Notch通路因子Hes1和Hes5明显上调(P<0.001),S100b和Pou4f1表达也出现升高(P<0.001)。结论NMDA损伤后2周c-kit+RPCs同时存在对称分裂和不对称分裂两种模式,可向GAD65/67+神经元分化,这一过程可能受到Notch通路的调控。ObjectiveTo analyze the characteristics of c-kit+endogenous retinal precursor cells(RPCs)in the retina of adult mice and their roles in the repair of N-methyl-D-aspartic acid(NMDA)-induced retinal damage.MethodsMouse models of NMDA-induced retinal injury were established in 4-week-old adult C57BL/6J mice.Immunofluorescence assay was used to analyze the relationship between c-kit+RPCs and c-kit stem cell factor(SCF)-positive cells and determine the number of c-kit+RPCs and their division activity at 1,2 and 4 weeks after NMDA injury.The mRNA expression levels of Hes1,Hes5,S100b and Pou4f1 in the injured retinal tissues were analyzed using real-time PCR.ResultsSCF+Müller cells surrounding the c-kit+RPCs were observed in the inner nuclear layer(INL)of the retina of adult mice.C-kit+RPCs still maintained a low level of cell division activity.Two weeks after NMDA-induced retinal injury,the number of c-kit+RPCs significantly increased in the retina(P<0.001)with markedly increased proportions of dividing cells(P=0.016)and c-kit+GAD65/67+cells/c-kit+cells(P<0.001).Two weeks after retinal injury,the Notch pathway components Hes1 and Hes5 were up-regulated(P<0.001)and the expression levels of S100b and Pou4f1 were augmented significantly in the retina(P<0.001).ConclusionTwo weeks after NMDA-induced retinal injury,c-kit+RPCs in the retina undergo both symmetrical and asymmetrical cell division and then differentiate into GAD65/67+neurons possibly under regulation by the Notch pathway.

关 键 词：视网膜 c-kit+视网膜前体细胞 N-甲基-D-天冬氨酸 细胞分裂 NOTCH通路 

分 类 号：R322.9[医药卫生—人体解剖和组织胚胎学] R329.25[医药卫生—基础医学]