CAJ | 학술논문

目的研究人脐带华尔通胶来源间充质干细胞的特性及向软骨细胞、骨细胞的分化能力,为椎间盘再生研究寻找新的细胞来源.方法取人的正常分娩或剖腹产胎儿的脐带,酶消化法从Wharton胶中分离干细胞,应用复合胶原酶NB4、dispaseⅡ和透明质酸的组合混合酶消化,检测人脐带来源的MSCs的细胞表面标记;分别应用成软骨诱导液、成骨诱导液诱导人脐带来源的MSCs向软骨细胞、骨细胞分化.用免疫细胞化学方法对分化和未分化的细胞进行鉴定.结果人脐带分离培养的贴壁细胞,体外生长形态类似于成纤维细胞,可以维持在未分化状态稳定增殖,这类细胞MSCs的表面标记CD44、CD105、CD90和D73呈现高表达,而不表达CD45、CD34、CD14、CD19及HLA-DR.向软骨细胞、骨细胞诱导结果提示P3代细胞有向终末软骨细胞、骨细胞分化能力,具有干细胞的特性.结论人脐带华尔通胶含有丰富的MSCs,易于培养扩增.人脐带来源的MSCs能分化为软骨细胞、骨细胞,这类细胞可能成为椎间盘移植的一个干细胞来源.
목적 연구인제대화이통효래원간충질간세포적특성급향연골세포、골세포적분화능력,위추간반재생연구심조신적세포래원.방법 취인적정상분면혹부복산태인적제대,매소화법종Wharton효중분리간세포,응용복합효원매NB4、dispaseⅡ화투명질산적조합혼합매소화,검측인제대래원적MSCs적세포표면표기;분별응용성연골유도액、성골유도액유도인제대래원적MSCs향연골세포、골세포분화.용면역세포화학방법대분화화미분화적세포진행감정.결과 인제대분리배양적첩벽세포,체외생장형태유사우성섬유세포,가이유지재미분화상태은정증식,저류세포MSCs적표면표기CD44、CD105、CD90화D73정현고표체,이불표체CD45、CD34、CD14、CD19급HLA-DR.향연골세포、골세포유도결과제시P3대세포유향종말연골세포、골세포분화능력,구유간세포적특성.결론 인제대화이통효함유봉부적MSCs,역우배양확증.인제대래원적MSCs능분화위연골세포、골세포,저류세포가능성위추간반이식적일개간세포래원.
Objective To investigate the isolation and expansion of mesenchymal stem cells (MSCS) from human umibilical cord Wharton's jelly and their biological identities , and explore the possibility of inducing human umbilical cord-derived MSCS to differentiate into chondrogenic and osteogenic cells. Methods The hUCMSCs were isolated form human umbilical cord by tissue adherence and digested with collagenaseNB4, dispase Ⅱ and hyaluronidase. The morphology, proliferation and immunophenotype of the 3rd passage cells were analyzed, and then the chondrogenic and osteogenic differentiation was tested and evaluated by specific staining methods. cells were induced to chondrogenic and osteogenic differentiation in vitro. Results The isolation of hUCMSCs by digestion with collagenaseNB4, dispase Ⅱ and hyaluronidase was efficient. After seeded for 24 hours, the adherent cells showed spindle shape and fibroblast cell-like shape and the size of hUCMSCs was homogeneous. Flow cytometry analysis revealed that the hUCMSCs were positive for CD44,CD105, CD90, CD73, but were negative for CD45, CD34, CD14, CD19 and HLA-DR.These cells could be induced to differentiate into chondrogenic and osteogenic cells under proper inducing conditions. The hUCMSCs retained the appearance and phenotype even after being expanded more than 40 passages in vitro. Conclusions The human MSCs could be isolated from human umbilical cord Wharton's jelly ,and it was easy to propagate these MSCs. An in vitro method for isolation and purification of hUCMSCs from human umbilical cord has been established. The cultured cells were composed of only undifferentiated cells and their biological properties were stable. The hUCMSCs are expected to be a new type of stem cells of tissue engineering.