中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (10): 1780-1784.doi: 10.3969/j.issn.1673-8225.2010.10.014

• 干细胞培养与分化 stem cell culture and differentiation • 上一篇    下一篇

体外分离培养昆明小鼠胚胎干细胞最佳胚龄的筛选：孕2.5 d与3.5 d及4.5 d比较

张海锋1，单 伟2，秦书俭2   

  1. 1徐州医学院解剖教研室，江苏省徐州市  221004； 
    2辽宁医学院解剖教研室，辽宁省锦州市 121001
  • 出版日期:2010-03-05 发布日期:2010-03-05
  • 作者简介:张海锋，男，1980年生，河南省焦作市人，汉族，2004年辽宁医学院毕业，硕士，主要从事组织工程学细胞方面的研究。 pearlbead2004@sina.com
  • 基金资助:

    辽宁省自然科学基金（20032142）；
    徐州医学院课题基金（08KJ19）

Screening of optimal embryonic time for in vitro separation and culture of Kunming mouse embryonic stem cells: Comparison among 2.5, 3.5, and 4.5 pregnant days

Zhang Hai-feng1, Shan Wei2, Qin Shu-jian2   

  1. 1Department of Anatomy, Xuzhou Medical College, Xuzhou   221004, Jiangsu Province, China;
    2Department of Anatomy, Liaoning Medical University, Jinzhou   121001, Liaoning Province, China
  • Online:2010-03-05 Published:2010-03-05
  • About author:Zhang Hai-feng, Master, Department of Anatomy, Xuzhou Medical College, Xuzhou 221004, Jiangsu Province, China pearlbead2004@ sina.com
  • Supported by:

    the Natural Science Foundation of Liaoning Province, No. 20032142*;
    the Project Foundation of Xuzhou Medical College, No. 08KJ19*

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摘要：

背景：昆明小鼠基因型与人类近似，建立昆明小鼠胚胎干细胞系有利于其转基因动物的获得，但目前对于何时收集胚胎尚无明确报道。

目的：探讨体外分离培养昆明小鼠胚胎干细胞的最佳胚龄。

方法：分别从孕2.5 d，3.5 d和4.5 d的昆明系母鼠子宫中分离收集胚胎，于显微镜下观察胚胎的形态、贴壁情况、内细胞团形成情况、胚胎干细胞克隆率、胚胎干细胞亚克隆率等，并进行碱性磷酸酶染色。

结果与结论：孕2.5 d采集的胚胎多为16细胞期胚胎，孕3.5 d采集的胚胎多为桑椹胚，孕4.5 d采集的胚胎多为囊胚。孕2.5 d胚胎和孕3.5 d胚胎的贴壁率、内细胞团形成率、胚胎干细胞克隆率、胚胎干细胞亚克隆率均无明显差异(P > 0.05)；孕4.5 d胚胎上述指标均显著高于前两者，更适合作为胚胎干细胞培养、克隆、分离、传代的材料。

关键词: 昆明小鼠, 胚龄, 内细胞团, 胚胎, 胚胎干细胞

Abstract:

BACKGROUND: Genotype of Kunming mice was similar to human population, thus an establishment of embryonic stem cell line is beneficial for research of transgenic animal. However, the best time to collect embryo has been less reported yet.

OBJECTIVE: To find the best time to collect embryos from Kunming mice.

METHODS: The embryos were collected from mother mice of 2.5, 3.5, and 4.5 pregnant days. Microscope was used to evaluate the growth condition of embryos, embryo attaching rate (A/C), inner cell mass (ICM) growing rate (I/C), embryonic stem cells (ESCs) clone growing rate (P1/C) and ESCs subclone growing rate (P2/C). The cells were then stained with alkaline phosphatase.

RESULTS AND CONCLUSION: Most of the 2.5-pregnant-day embryos were 16-cell-phase embryos. The 3.5-pregnant-day embryos were morulas while the 4.5-pregnant-day embryos were blastulas. There were no significant differences in A/C, I/C, P1/C, P2/C between 2.5 and 3.5 pregnant days (P > 0.05). The 4.5-pregnant-day indicators mentioned above were significantly greater than those two groups; therefore, 4.5-pregnant-day embryos were the best source to culture, clone, isolate and passage ESCs.

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