人胚胎干细胞来源角膜上皮样细胞用于治疗兔角膜缘干细胞缺乏  

Corneal Epithelioid Cells Derived from Human Embryonic Stem Cells Were Used to Treat Limbal Stem Cell Deficiency in Rabbits

作　　者：杨哲 曾湘纹 李钊[2] 王丽强[2] YANG Zhe;ZENG Xiangwen;LI Zhao;WANG Liqiang(Graduate School,Chinese People's Liberation Army Medical School,Beijing 100853,China)

机构地区：[1]解放军医学院研究生院,北京100853 [2]解放军总医院眼科医学部

出　　处：《联勤军事医学》2023年第7期553-558,共6页Military Medicine of Joint Logistics

基　　金：国家重点研发计划项目(2017YFA0103204)。

摘　　要：目的 通过小分子化合物诱导人胚胎干细胞(human embryonic stem cells, hESCs)分化为角膜上皮样细胞,观察角膜上皮样细胞治疗兔角膜缘干细胞缺乏(limbal stem cell deficiency, LSCD)的效果。方法 利用拟胚体联合小分子化合物方法,将hESCs H9细胞系向角膜上皮样细胞分化,将角膜上皮样细胞在去上皮羊膜片上培养构建重组细胞膜片。选取造模成功的12只LSCD兔,按照随机数表法分为对照组和实验组,对照组接受去上皮的的羊膜片治疗,实验组接受角膜上皮样细胞膜片移植治疗。术后通过裂隙灯、前节照相等方法,对角膜透明度、新生血管、荧光素钠染色情况进行评分,并于移植后4周观察角膜病理切片结构。结果 成功将hESCs诱导为角膜上皮样细胞,细胞形状和结构类似角膜上皮细胞。病理结果显示,实验组的角膜形成复层上皮结构,上皮下炎症细胞浸润轻,治疗效果明显优于对照组。进行细胞移植治疗后发现,实验组的角膜透明度高、新生血管少,角膜评分明显优于对照组(P<0.05)。诱导后的角膜上皮样细胞的角膜上皮标记物ΔNP63和CK12的表达与hESCs H9相比差异具有统计学意义(P<0.05)。结论 采用拟胚体联合小分子化合物可以成功诱导出角膜上皮样细胞,角膜上皮样细胞膜片移植到兔体内后可有效治疗LSCD。Objective To investigate the therapeutic effects of human embryonic stem cells(hESCs)on rabbit limbal stem cell deficiency(LSCD)by inducing small molecule compounds to differentiate into corneal epithelioid cells.Methods hESCs H9 cell lines were differentiated into corneal epithelioid cells by embryoid body combined small molecule compound method,and the corneal epithelioid cells were cultured on the denuded amniotic membrane to construct recombinant cell membrane.Twelve LSCD rabbits with successful modeling were selected and divided into control group and experimental group according to random number table method,the control group received deepithelialized amniotic membrane treatment,and the observation group received corneal epithelioid cell membrane transplantation.After surgery,the corneal transparency,neovascular and fluorescein sodium staining were scored by slit lamp observation and anterior segment photography,and the corneal pathological section structure was observed 4 weeks after transplantation.Results hESCs were successfully induced into corneal epithelial cells,which were similar in shape and structure to corneal epithelial cells.The pathological results showed that the cornea of the experimental group formed a multilayer epithelial structure,and the subepithelial inflammatory cells infiltrated lightly,and the therapeutic effect was obviously better than that of the control group.After cell transplantation,it was found that the corneal score of the experimental group was significantly better than that of the control group with higher corneal transparency and less new blood vessels(P<0.05).The expression of corneal epithelial markers△NP63 and CK12 in induced corneal epithelioid cells was statistically different from that of hESCs H9(P<0.05).Conclusion Corneal epithelioid cells can be successfully induced by embryoid bodies combined with small molecule compounds,and corneal epithelioid cells membrane grafted into rabbits can effectively treat LSCD.

关 键 词：人胚胎干细胞 角膜上皮样细胞 角膜缘干细胞缺乏 小分子化合物 

分 类 号：R772.2[医药卫生—眼科]