脂肪干细胞对角膜基质细胞增殖凋亡的调节及其机制研究  

Effect of rabbit adipose-derived stem cell on proliferation and apoptosis of corneal stromal cells

作　　者：郑青青[1] 沈江[2] 沈婷[1] 李秋实[2] 洪朝阳[2] 李文伟[3] 

机构地区：[1]浙江省人民医院、杭州医学院附属人民医院眼科,杭州310014 [2]温州医科大学眼视光学院 [3]浙江省立同德医院眼科

出　　处：《浙江医学》2017年第5期340-344,共5页Zhejiang Medical Journal

基　　金：浙江省科技厅重大专项(2013C03048-1)

摘　　要：目的探讨在体外共培养中,兔脂肪干细胞(ADSCs)对兔角膜基质细胞(CSCs)的影响及其作用机制,为角膜基质损伤治疗提供实验依据。方法取新西兰大白兔腹股沟皮下脂肪组织,用I型胶原酶消化分离出ADSCs进行培养,贴壁细胞至3代,评价其多向分化能力。另取新西兰大白兔角膜基质层组织,用II型胶原酶消化分离出CSCs,传代至2代后备用。将CSCs及ADSCs(实验组)共培养3d后,与单纯CSCs(对照组)采用CCK-8法检测CSCs增殖情况,流式细胞术检测细胞凋亡情况,Western blot法检测细胞金属蛋白酶(MMPs)、胶原相关蛋白水平,评估在体外共培养中ADSCs对CSCs的影响。结果与对照组比较,CCK-8法检测结果提示实验组CSCs数量显著增高(P<0.05),流式细胞术检测结果提示实验组CSCs细胞凋亡减少,Western blot法检测结果提示实验组MMPs表达水平降低,胶原相关蛋白表达水平增加。结论 ADSCs不仅可以促进CSCs增殖并抑制其凋亡,还可以促进细胞外基质的合成,在角膜基质损伤治疗中具有较好的应用前景。Objective To investigate the effects of rabbit adipose-derived stem cells(ADSCs)on proliferation and apoptosis of rabbit corneal stromal cells(CSCs).Methods ADSCs were isolated fromsubcutaneous adipose tissue in groin area of New Zealand white rabbit with the method of collagenase type I digestion.The isolated ADSCs were cultured and passed to3generation and its differentiation ability was evaluated.CSCs were obtained from the corneas of New Zealand rabbits with the method of collagenase II digestion.After initial expansion,the CSCs were cultured up to passage2and then prepared for experiment.The CSCs were cultured alone(control group)or co-cultured with ADSCs(indirect co-culture group).After incubationfor72h the proliferative capacity of CSCs was evaluated by CCK-8assay,apoptosis of CSCs was determined by flow cytometry,the expression of matrix metalloproteinases(MMPs)and collagens was detected by Western blot,and the invasion activity ofcorneal stromal cells was also examined in vitro.Results Compared with the control group,ADSCs significantly promotedproliferation and invasion of CSCs in indirect co-culture group(P<0.05).The group with ADSCs showed low percentage of apoptotic cells.CSCs secreted lower levels of MMPs and higher levels of collagens in indirect co-culture group.Conclusion ADSCs can promote plasticity and EMC synthesis of corneal stromal cells,and suppress the apoptosis at the same time.It suggests that application of ADSCs may be a promising approach for corneal injury therapy.

关 键 词：脂肪干细胞 角膜基质细胞 损伤 增殖 基质金属蛋白酶 

分 类 号：R772.2[医药卫生—眼科]