摘要
目的:体外培养人胚胎生殖干细胞(EG),在不添加细胞因子的培养条件下,观察细胞生长情况。方法:取5～10周人胚胎的生殖腺嵴和肠背系膜,进行组织块培养,采用组织化学及免疫细胞化学技术对培养的细胞进行鉴定。结果:培养4d后,在成纤维细胞的上面出现EG细胞集落;培养2周后,显示细胞呈圆形,胞质染成深蓝色;细胞染色体均为正常的二倍体核型;碱性磷酸酶活性强阳性;并检测到SSEA-1。结论:体外培养人胚胎生殖腺嵴,利用源于自身胚胎组织的成纤维细胞作为饲养层,可观察到EG细胞集落的形成;培养的细胞初步鉴定为人胚胎干细胞。
Objective: To culture human embryonic germ stem cell in vitro and to observe the cell growth without any cytokines in presence. Methods: Human embryonic (5-10 weeks postfertilization ) genital ridges and mesenteries were cultured. Histochemistry and immunocytochemistry were used to identify the cultures. Results: 12 h after culture, fibroblasts appeared around the tissue fragments. Culture till 4 d, embryonic germline cells cluster was observed on the fibroblasts, the colony appeared spherical, and the cells arranged compactly. Two weeks later, the cell was round in shape and the cytoplasm was stained oxford blue with Giemsa. All of the karyotypes of the primordial germ cells, which were under conventional chromosome test, were diploid without abnormity. Gomoris calcium cobalt for alkaline phosphatase was strongly positive. Immunocytochemistry studies showed that the cultured human embryonic germ cells were stained deep brown by using anti-SSEA-1 monoclonal antibody. Conclusion: Tissue from genital ridges and mesenteries can be cultured in vitro, with the fibroblasts in its own embryonic tissue serving as the feeder layer. The formation of the embryonic stem cell colony can be observed, and the cells cultured by this method is confirmed to be embryonic stem cells.
出处
《解剖学杂志》
CAS
CSCD
北大核心
2005年第3期295-297,F004,共4页
Chinese Journal of Anatomy
基金
江苏省高校自然科学研究计划(Q2134042)
