CAJ | 학술논문

目的观察肌肉微粒与人脐带间充质干细胞(hUCMSCs)共培养体系中干细胞成肌分化的可行性及其分化能力.方法体外分离、鉴定并扩增hUCMSCs,0.05％胰酶消化后制备干细胞悬液.手术切取新西兰白兔后肢骨骼肌,剪碎成＜1 mm3的肌肉微粒.按照1×105∶1的比例(n:mg)行干细胞与肌肉微粒的间接或直接共培养,分别通过Tranwell迁移实验、噻唑蓝(MTT)检测及免疫组织化学与流式细胞分析等方法观察共培养体系中hUCMSCs的迁移能力、增殖能力及成肌细胞标志物desmin的表达.结果间接共培养体系建立12h后,hUCMSCs向肌肉微粒方向迁移率达(39.73±5.92)％,直接共培养体系建立5d后干细胞增殖及成肌转化达到高峰,干细胞增殖率可达原来的6倍,其分布以肌肉微粒为中心,呈向心性聚集排列,流式细胞术检测到desmin的阳性表达率为(21.57±2.39)％,与对照组间差异均有统计学意义(P＜0.05).结论通过与肌肉微粒共培养的方法,可以实现对hUCMSCs的成肌分化诱导,该方法是一种非化学性的简便、快速、有效的干细胞成肌肉诱导方法.
목적 관찰기육미립여인제대간충질간세포(hUCMSCs)공배양체계중간세포성기분화적가행성급기분화능력.방법 체외분리、감정병확증hUCMSCs,0.05％이매소화후제비간세포현액.수술절취신서란백토후지골격기,전쇄성＜1 mm3적기육미립.안조1×105∶1적비례(n:mg)행간세포여기육미립적간접혹직접공배양,분별통과Tranwell천이실험、새서람(MTT)검측급면역조직화학여류식세포분석등방법관찰공배양체계중hUCMSCs적천이능력、증식능력급성기세포표지물desmin적표체.결과 간접공배양체계건립12h후,hUCMSCs향기육미립방향천이솔체(39.73±5.92)％,직접공배양체계건립5d후간세포증식급성기전화체도고봉,간세포증식솔가체원래적6배,기분포이기육미립위중심,정향심성취집배렬,류식세포술검측도desmin적양성표체솔위(21.57±2.39)％,여대조조간차이균유통계학의의(P＜0.05).결론 통과여기육미립공배양적방법,가이실현대hUCMSCs적성기분화유도,해방법시일충비화학성적간편、쾌속、유효적간세포성기육유도방법.
Objective To investigate the feasibility and ability of the muscular differentiation of human umbilical cord stem cells (hUCMSCs) in the in vitro co-culture system of hUCMSCs with minced rabbit skeletal muscle fragments.Methods hUCMSCs were isolated,identified and proliferated in vitro.In a proportion of 1 × 105 to 1 (n:mg) of hUCMSCs to the mince muscle,direct co-culture system was established by adding the minced muscle fragments to the inoculated hUCMSCs in plates,and the indirect coculture system was established by using a transwell with the hUCMSCs and the mince muscle fragments in the upper and the lower chamber,respectively.Migration,proliferation,and differentiation of the hUCMSCs were evaluated by using the inverted microscopy,MTT assays,flow cytometry,and immunohistochemistry.Results Twelve h after the establishment of the indirect co-culture system,the migration rate of the hUCMSCs to the minced muscle fragments reached to (39.73 ± 5.92) ％.Five days after the establishment of the direct co-culture system,hUCMSCs centripetally accumulated to the minced muscle fragments and increased 6 folds to that of their initial number,and their muscular differentiation rate reached (21.57 ±2.39) ％ under flow cytometry inspections.Conclusion Co-culture of hUCMSCs with the minced muscle fragments is a sampler non-chemical method,which could successfully induce the muscular differentiation of hUCMSCs in a short time.