人毛囊外根鞘细胞的分离及培养

Isolation And Cultivation In Vitro Of Human Hair Follicle Outer Root Sheath Cells

目的建立毛囊外根鞘细胞的分离和培养方法。方法利用中性蛋白酶分离得到单个毛囊,采用组织块法和消化法进行细胞培养,倒置显微镜、扫描及透射电镜观察细胞形态,培养各代细胞做生长曲线,RT-PCR检测干细胞相对特异性标识分子K15和细胞分化标识分子外皮蛋白的mRNA表达情况。结果组织块法和消化法均能培养出毛囊外根鞘细胞,并可传代培养,组织块法细胞生长慢,不易汇合,消化法细胞增殖快。各代细胞中K15和外皮蛋白mRNA均有表达。结论消化法适合毛囊外根鞘细胞的体外培养,该方法的建立为进一步分离纯化培养毛囊干细胞奠定了基础。

Objective To establish a method for isolation and cultivation in vitro of human hair follicle outer root sheath cells. Methods The single hair follicle were obtained by incubating with dispase, then harvest the outer root sheath cells from enzymic digestion and explant culture. Cells were observed by phase contrast microscope, scanning and transmission electron microscope, meanwhile growth curve was made, mRNA expression of stem cell specific marker-cytokeratin 15(K15) and cell differentiation marker-involucrin was detected by RT-PCR. Results Human hair follicle outer root sheath cells can be obtained from enzymic digestion and explant culture. The cells from explant culture method grow slowly and not easy to confluence, the cells from enzymic digestion grow rapidly, from passage0 to passage5, the cells express K15 and involucrin mRNA. Conclusion Enzymic digestion method is feasible in cultivation of human hair follicle outer root sheath cells in vitro.