人牙周膜干细胞、脐带间充质干细胞对体外培养角膜缘干细胞的影响  

Effects of periodontal ligament stem cells and human umbilical cord mesenchymal stem cells on microenvironment of corneal limbal stem cells cultured in vitro

作　　者：王慧娴[1,2] 高晓唯[1,2] 蔡岩[2] 李文静[2] 

机构地区：[1]石河子大学医学院,新疆维吾尔自治区石河子市832003 [2]解放军第474医院眼科中心,新疆维吾尔自治区乌鲁木齐市830012

出　　处：《眼科新进展》2016年第11期1015-1020,共6页Recent Advances in Ophthalmology

基　　金：新疆自治区科技支疆项目(编号:201491171)~~

摘　　要：目的比较人牙周膜干细胞(human periodontal ligament stem cells,HPDLSCs)、人脐带间充质干细胞(human umbilical cord mesenchymal stem cells,HUCMSCs)作为饲养层培养角膜缘干细胞(limbal stem cells,LSCs)的优越性,从而筛选出扩增LSCs的最佳饲养层。方法体外分离培养HPDLSCs、HUCMSCs,诱导细胞成脂、成骨分化,并检测细胞表面标志物的表达;将兔LSCs与HPDLSCs、HUCMSCs、NIH-3T3共培养和无饲养层单独培养,比较各组克隆形成率,免疫荧光比较各组LSCs克隆团ABCB5、IPO13、CK3/12的表达情况。结果体外培养HPDLSCs、HUCMSCs均可向脂肪、成骨细胞分化,两种细胞均高表达间充质来源的表面标志CD90,不表达CD45;三组饲养层与兔LSCs共培养均可形成小克隆团,HPDLSCs组、HUCMSCs组、NIH-3T3组、无饲养层组克隆形成率分别为(4.90±0.96)%、(4.10±0.56)%、(4.67±0.76)%、(0.83±0.35)%,四组间总体比较差异有统计学意义(F=22.047,P=0.00);HPDLSCs组、HUCMSCs组与NIH-3T3组的克隆形成率差异均无统计学意义(均为P>0.05);HPDLSCs组、HUCMSCs组、NIH-3T3组与无饲养层组间差异均有统计学意义(均为P<0.01)。免疫荧光化学检测三种饲养层LSCs标志物ABCB5、IPO13呈高表达,CK3/12呈低表达。结论 HPDLSCs、HUCMSCs均可作为替代饲养层用以培养LSCs。Objective To find optimal feeding layer of amplification with limbal stem cells, compare the superiority of the human periodontal ligament stem cells （ HP- DLSCs） and human umbilical cord mesenchymal stem cells （HUCMSCs） as a breeding cultivation of corneal limbus stem cell layer, then find the more efficient feeder layer. Methods HPDLSCs and HUCMSCs were cultured in vitro, then the cells was induced and differentiated into fat, osteoblast cells, and the expression of cell surface markers were detected. The rabbit corneal limbal stem cells co-cultured with HPDLSCs, HUCM- SCs,NIH-3T3 and no feeder layer,then each clone formation rate was counted, LSCs markers ABCB5, IPO13, CK3/12 expression was compared by immunofluorescence between groups. Results HPDLSCs and HUCMSCs cultured in vitro could differentiate into fat, osteoblast cells and the two cells both expressed high surface of mesenchymal origin marks CD00, not expressed CD45. These groups could form cloning, the clone for- mation rate were （4.90 ±0.96）% , （4. 10 ±0.55）% , （4.57 ±0.75）% , （0.83 ±0.35 ）% in HPDLSCs, HUCMSCs, NIH-3T3 and no feeder layer group, overall difference between the four groups was statistically significant （F = 22.047 ,P 〈0.01 ）. There was no statistical significance of HPDLSCs, HUCMSCs groups compared with NIH-3T3 group （all P 〉 0. 05）,but there were significance differences of HPDLSCs,HUCMSCs, NIH-3T3 groups compared with no feeder layer group （all P 〈 0.01 ）. Immunofluorescence showed three layer of LSCs markers ABCBS, IPO13 were highly expressed, CK3/12 showed lower expression. Conclusion HPDLSCs, HUCMSCs both can be used as ideal substitute feeder layer to cultivate LSCs.

关 键 词：人牙周膜干细胞 人脐带间充质干细胞 角膜缘干细胞 饲养层 

分 类 号：R772.2[医药卫生—眼科]