人真皮成纤维细胞的分离培养及鉴定

Isolation,culture and identification of human fibroblasts

目的对人真皮成纤维细胞(hFbs)进行分离培养及鉴定,为组织工程复合皮肤的构建提供种子细胞。方法利用组织块培养法、消化传代纯化培养hFbs,通过细胞形态的观察及免疫细胞化学、流式细胞术等方法对其进行鉴定。结果经组织块培养法获得的细胞可以传代培养,从P1代到P6代细胞形态保持一致,呈纤维细胞样生长;免疫细胞化学检测可见波形蛋白(vimentin)呈阳性反应,通过流式细胞术检测P3及P6代的细胞vimentin阳性表达量均达到95%以上,角蛋白15(cytokeratin 15)呈阴性表达。结论成功建立体外稳定培养hFbs的方法,为组织工程皮肤的研究提供了理论与实验基础。

Objective To explore a method of the isolation,culture and identification of human fibroblasts（hFbs） in vitro for the provision of seed cells in tissue engineering skin.Methods The cells were cultured and expanded and observed under phase contrast microscope,and they were identified with immunocytochemistry and flow cytometry（anti-vimentin,cytokeratin 15）.Results With phase contrast microscope,the cells were observed to uniformed population of cells and fibroblast-like.Immunocytochemistry and flow cytometry showed all cultural cells were fibroblasts（positive for vimentin,negative for cytokeratin 15）.The cells were sub-cultured 6 generations.Conclusions A method for isolating and culturing hFbs in vitro is established successfully.