Liu Yu-Xiao, Ji Lei, Yue Wen, Yan Zhi-Feng, Wang Jing, Xi Jia-Fei, Zhang Rui, Nan Xue, Bai Ci-Xian, Chen Lin, Wang Yun-Fang, Pei Xue-Tao
Beijing Institution of Transfusion Medicine, People's Republic of China.
Cloning Stem Cells. 2009 Mar;11(1):51-60. doi: 10.1089/clo.2008.0049.
Here, we have now developed a new inducing system to promote the differentiation of human stem cells (hESCs) toward hematopoietic lineages by the treatment with cells extract of human fetal liver tissue (hFLT). The embryoid bodies (EBs) obtained from human H1 embryonic stem cells were exposed to buffer, hFLT cells extract, heated hFLT cell extract, and cell extract of human liver cells lines-LO2. Then, the feature of EBs in different groups was characterized by real-time RT-PCR and colony-forming assays. The results showed the treatment by hFLT cells extract could activate the hematopoietic genes expression and improve the capacity for hematopoietic progenitor development of hEBs. After that, we cocultured hFLT extract treated hEBs on the hFLSCs (human fetal liver stromal cells) feeder to differentiate them into hematopoietic cells. As a control, untreated hEBs were cocultured on hFLSCs feeder with cytokines. The feature of induced cells from hEBs was characterized by flow cytometry, Wright-Giemsa staining, and colony-forming assays. The results demonstrated that hFLT cells extract was capable of inducing hEBs into hematopoietic cells and combing it with hFLSCs feeder could largely promote hematopoietic differentiation of hESCs. This method may supply a new way to substitute the cytokines required in hematopoietic induction of hESCs.
在此,我们现已开发出一种新的诱导系统,通过用人胎儿肝脏组织(hFLT)的细胞提取物处理,来促进人类干细胞(hESCs)向造血谱系分化。将从人H1胚胎干细胞获得的胚状体(EBs)分别暴露于缓冲液、hFLT细胞提取物、加热的hFLT细胞提取物以及人肝细胞系-LO2的细胞提取物中。然后,通过实时RT-PCR和集落形成试验对不同组EBs的特征进行表征。结果表明,hFLT细胞提取物处理可激活造血基因表达,并提高hEBs的造血祖细胞发育能力。之后,我们将经hFLT提取物处理的hEBs与人胎儿肝脏基质细胞(hFLSCs)饲养层共培养,使其分化为造血细胞。作为对照,未处理的hEBs与细胞因子一起在hFLSCs饲养层上共培养。通过流式细胞术、瑞氏-吉姆萨染色和集落形成试验对hEBs诱导细胞的特征进行表征。结果表明,hFLT细胞提取物能够将hEBs诱导为造血细胞,并且将其与hFLSCs饲养层相结合可极大地促进hESCs的造血分化。该方法可能为替代hESCs造血诱导所需的细胞因子提供一种新途径。