Chen Hui-Qin, Zhang Xu-Chao, Tang Xin-Yi, Wu Bei-Yan, Huang Shao-Liang
Center for Stem Cell Research, The Second Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510120, China.
Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2006 Oct;14(5):999-1003.
This study was aimed to investigate the hematopoietic growth factors expressed in human aorta-gonad-mesonephros (AGM)-derived stromal cells in vitro in order to provide the basic data for elucidating the role of AGM -derived-stromal cells in embryo-hematopoiesis and its hematopoietic suppoitive effect. RT-PCR was used to analyze the expression of IL-6, SCF, Flt3-L, oncostatin M (OSM), IL-3, TPO, M-CSF and LIF in human aorta-gonad-mesonephros-derived stromal cells (hAGMS1-S5) at mRNA level. IL-6, SCF and Flt3-L levels were detected in the supernatant of hAGMS1-S5 stromal cells by ELISA assay. Umbilical cord blood CD34(+) cells were cocultured with hAGMS1-S5 feeder cells, and hematopoietic cells were collected at day 14 for colony analysis in methylcellulose semisolid medium. The results showed that human aorta-gonad-mesonephros-derived stromal cells S1-S5 expressed IL-6, SCF, Flt3-L and OSM mRNA, but did not express IL-3, TPO, M-CSF and LIF mRNA. In the supernatant of hAGMS1-S5 cells, IL-6, SCF and Flt3-L could be detected by ELISA assay at different levels, while there was no significant difference between groups of hAGMS1-S5 (P > 0.05). When cocultured with umbilical cord blood CD34(+) cells, hAGMS1-S5 could support the expansion of CFU-GM, BFU-E, and CFU-Mix. The supportive effects of hAGM S1-S5 were significantly different (P < 0.05), hAGM S3 and S4 were better than hAGM S1, S2, and S5. It is concluded that detection of hematopoietic growth factors expressed in human aorta-gonad-mesonephros-derived stromal cells provided a solid foundation to elucidate the mechanism of hematopoiesis and the hematopoietic supportive effect of these stromal cells.
本研究旨在探讨人主动脉-性腺-中肾（AGM）来源的基质细胞在体外表达的造血生长因子，以便为阐明AGM来源的基质细胞在胚胎造血中的作用及其造血支持效应提供基础数据。采用逆转录聚合酶链反应（RT-PCR）分析人主动脉-性腺-中肾来源的基质细胞（hAGMS1-S5）中白细胞介素-6（IL-6）、干细胞因子（SCF）、Flt3配体（Flt3-L）、制瘤素M（OSM）、IL-3、血小板生成素（TPO）、巨噬细胞集落刺激因子（M-CSF）和白血病抑制因子（LIF）在mRNA水平的表达。通过酶联免疫吸附测定（ELISA）检测hAGMS1-S5基质细胞上清液中的IL-6、SCF和Flt3-L水平。将脐血CD34(+)细胞与hAGMS1-S5饲养细胞共培养，在第14天收集造血细胞，在甲基纤维素半固体培养基中进行集落分析。结果显示，人主动脉-性腺-中肾来源的基质细胞S1-S5表达IL-6、SCF、Flt3-L和OSM mRNA，但不表达IL-3、TPO、M-CSF和LIF mRNA。在hAGMS1-S5细胞的上清液中，ELISA检测可在不同水平检测到IL-6、SCF和Flt3-L，而hAGMS1-S5各组之间无显著差异（P>0.05）。当与脐血CD34(+)细胞共培养时，hAGMS1-S5可支持粒-单系集落形成单位（CFU-GM）、红系爆式集落形成单位（BFU-E）和混合集落形成单位（CFU-Mix）的扩增。hAGM S1-S5的支持作用有显著差异（P<0.05），hAGM S3和S4优于hAGM S1、S2和S5。结论是，检测人主动脉-性腺-中肾来源的基质细胞中表达造血生长因子为阐明造血机制及这些基质细胞的造血支持效应提供了坚实基础。
