Chinese Journal of Tissue Engineering Research ›› 2011, Vol. 15 ›› Issue (27): 4969-4974.doi: 10.3969/j.issn.1673-8225.2011.27.007

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Effects of sequential inductive systems with feeder cells from human aorta-gonad-mesonephros region, fetal liver and bone marrow on the differentiation of mouse embryonic stem cells into hematopoietic stem cells

Cai Yun1, 2, Zhang Xu-chao2, 3, Chen Hui-qin1, 2, Huang Shao-liang2   

  1. 1Department of Pediatrics, Third Affiliated Hospital, Sun Yat-sen University, Guangzhou  510630, Guangdong Province, China
    2Center for Stem Cell Research, Second Affiliated Hospital, Sun Yat-sen University, Guangzhou  510120, Guangdong Province, China
    3Research Center of Medical Sciences, Guangdong Provincial People’s Hospital, Guangzhou  510080, Guangdong Province, China
  • Received:2011-01-23 Revised:2011-03-09 Online:2011-07-02 Published:2011-07-02
  • Contact: Zhang Xu-chao, Doctor, Associate researcher, Center for Stem Cell Research, Second Affiliated Hospital, Sun Yat-sen University, Guangzhou 510120, Guangdong Province, China; Research Center of Medical Sciences, Guangdong Provincial People’s Hospital, Guangzhou 510080, Guangdong Province, China zhangxuchao@263.net Correspondence to: Huang Shao-liang, Professor, Doctoral supervisor, Center for Stem Cell Research, Second Affiliated Hospital, Sun Yat-sen University, Guangzhou 510120, Guangdong Province, China hshl@gzsums.edu.cn
  • About author:Cai Yun☆, Doctor, Attending physician, Department of Pediatrics, Third Affiliated Hospital, Sun Yat-sen University, Guangzhou 510630, Guangdong Province, China; Center for Stem Cell Research, Second Affiliated Hospital, Sun Yat-sen University, Guangzhou 510120, Guangdong Province, China wcyf@163.com
  • Supported by:

    the National Natural Science Foundation of China, No. 30300377*; the Scientific Research Project of Guangdong Province, No. 2008B030301046*

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Abstract:

BACKGROUND: Previous studies have prepared human aorta-gonad-mesonephros (AGM) region stromal cell line and fetal liver stromal cell line, and found that AGM can promote directional differentiation of mouse embryonic stem cells (ESCs) into hemopoietic stem cells (HSCs).
OBJECTIVE: To simulate the spatial and temporal hematopoietic microenvironment changes in embryonic development, investigate the supportive effects of sequential inductive systems with feeder cells from human AGM region and fetal liver and bone marrow on the differentiation of mouse ESCs into HSCs, and design more effective conditions for HSCs output.
METHODS: E14 mouse ESCs were induced into embryoid body firstly. Then the cells from embryoid body were further co-cultured with human AGM region, fetal liver and bone marrow stromal cells in Transwell non-contact system in sequential orders. The induced embryoid body cells were divided into four groups according to different culture stages: embryoid body control group; embryoid body/AGM group; embryoid body/AGM+ fetal liver group; embryoid body /AGM+ fetal liver + bone marrow group. On day 6 of co-cultured, cells derived from embryoid body in all groups were collected for Sca-1+c-Kit+ cells analysis by flow cytometry, colony forming unit assay and observed.
RESULTS AND CONCLUSION: ①Primitive hematopoietic cells were detected in embryoid body/AGM+fetal liver group and embryoid body/AGM+fetal liver+bone marrow group. ②Sca-1+c-Kit+ cell proportion of embryoid body/AGM group was much higher than pre-treated embryoid body cells (P < 0.05). ③Embryoid body/AGM group, embryoid body/AGM+fetal liver group and embryoid body/AGM+fetal liver+bone marrow group had more colony forming unit amount than embryoid body control group (P < 0.05). And the cells collected from embryoid body /AGM+ fetal liver group and embryoid body/AGM+ fetal liver + bone marrow group had better ability of forming multiple lineage hematopoietic colonies than that from embryoid body/AGM group. It suggested that microenvironment of AGM+ fetal liver or AGM+ fetal liver + bone marrow stromal cells can significantly expand much more primitive hematopoietic stem cells than AGM stromal cells only.

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