角膜缘niche细胞与基质细胞维持角膜缘干细胞功能的对比研究  

Differences between limbal niche cells and limbal stromal cells in maintenance of corneal limbal stem cell function

作　　者：黎颖莉 席兴华 Tomoyuki-Inoue Fumihiko-Takamatsu Naoyuki-Maeda Yuichi-Ohashi Kohji-Nishida LI Yingli;XI Xinghua;Tomoyuki-Inoue;Fumihiko-Takamatsu;Naoyuki-Maeda;Yuichi-Ohashi;Kohji-Nishida(Department of Ophthalmology,Shenzhen Hospital,Southern Medical University,the Third of Clinical Medicine,Southern Medical University,Shenzhen 518100,Guangdong Province,China;Department of Ophthalmology,Ehime University Medical School,Matsuyama 790-8577,Ehime,Japan;Department of Ophthalmology,Osaka University Medical School,Suita 790-8577,Osaka,Japan)

机构地区：[1]南方医科大学深圳医院眼科,南方医科大学第三临床医学院,广东省深圳市518100 [2]爱媛大学医学院眼科教研室,日本爱媛县松山市790-8577 [3]大阪大学医学院眼科教研室,日本大阪府吹田市565-0871

出　　处：《眼科新进展》2020年第6期505-509,共5页Recent Advances in Ophthalmology

基　　金：国家自然科学基金资助(编号81800804);广东省自然科学基金资助(编号2017A030310108);深圳市医疗卫生“三名工程”高层次医学团队(编号SZSM201612022)。

摘　　要：目的比较角膜缘niche细胞(limbal niche cells,LNCs)与角膜缘基质细胞(limbal stromal cells,LSCs)在维持角膜缘干细胞功能上的不同特性。方法将LNCs和LSCs分别从6个角膜缘组织分离,并在相同的条件下培养、传代。LNCs与LSCs经丝裂霉素C(mitomycin C,MMC)处理后分为LNCs组与LSCs组作为饲养细胞分别与角膜缘干细胞共培养,比较两组角膜缘干细胞克隆形成率(colony-forming efficiency,CFE)、上皮细胞复层化以及细胞标志物和部分基因的表达。结果LNCs组角膜缘干细胞CFE(6.57±1.54)%高于LSCs组(1.43±0.47)%。LNCs组细胞复层上皮数(4~5层)多于LSCs组(2~3层)。角膜缘干细胞克隆与免疫荧光染色及mRNA半定量分析结果显示,LNCs组比LSCs组表达了更多干细胞标志物ΔNp63,能更有效地维持角膜缘干细胞的细胞特性。逆转录PCR分析结果显示,LNCs组与LSCs组都分泌了一些维持角膜缘干细胞生长的生长因子,但LNCs组比LSCs组高表达上皮型钙黏蛋白(E-cadherin),低表达营养神经素3(NT3),能更好地支持角膜上皮增殖。结论LNCs比LSCs能更好地支持角膜缘干细胞的生长及维持其干细胞特性。Objective To investigate and compare the different characteristics of limbal niche cells(LNCs)and limbal stromal cells(LSCs)in the maintenance of corneal limbal stem cell function.Methods LNCs and LSCs were obtained from six limbal tissues,cultured and passaged under the same conditions.LNCs and LSCs received treatment of mitomycin C(MMC),and then divided into LNCs group and LSCs group with co-culture of limbal stem cells.The colony-forming efficiency(CFE),epithelial cell stratification,expression of cell markers and some genes were compared between the two groups.Results The CFE of limbal stem cells was(6.57±1.54)%in LNCs group,which was higher than(1.43±0.47)%in LSCs group.The epithelium stratified into 4-5 layers in the LNCs group was more than that stratified into 2-3 layers in the LSCs group.Limbal cell cloning,immunofluorescent staining and mRNA semi-quantitative analysis results showed that LNCs group showed a higher intensity of limbal stem cell markerΔNp63 than that in the LSCs group,and can effectively maintain the characteristics of limbal stem cells.Moreover,reverse transcription polymerase chain reaction analysis revealed that both LNCs group and LSCs group can secrete growth factors to keep growth of limbal stem cells,but LNCs showed a higher expression level of E-cadherin and a lower expression level of NT3 than those in the LSCs group,and better efficacy to support corneal epithelial proliferation.Conclusion LNCs have the distinct ability to support growth of limbal stem cells and maintain their characteristics compared with LSCs.

关 键 词：角膜缘干细胞 niche细胞 角膜基质细胞 克隆形成率 

分 类 号：R772[医药卫生—眼科]