The effects of vascular endothelial growth factor （VEGF） on neural differentiation of human embryonic stem cells （hESCs） in vitro and the possible mechanism were observed. The hESCs lines, TJMU1 and TJMU2, were es...The effects of vascular endothelial growth factor （VEGF） on neural differentiation of human embryonic stem cells （hESCs） in vitro and the possible mechanism were observed. The hESCs lines, TJMU1 and TJMU2, were established and stored by our laboratory, hESCs differentiated into neuronal cells through embryonic body formation. In this induction process, hESCs were divided into three groups： group A, routine induction; group B, routine induction＋10 ng/mL VEGF; group C, routine in- duction＋10 ng/mL VEGF＋10 ng/mL VEGFR2/Fc. OCT4, Nestin and GFAP in each group were de- tected by RT-PCR, and the cells expressing Nestin and GFAP were counted by immunofluorescence. The percentage of Nestin positive cells in group B was significantly higher than in groups A and C, while the percentage of GFAP positive cells in group B was significantly lower than in groups A and C （P〈0.01）. There was no significant difference between groups A and C （P〉0.05）. It was concluded that VEGF, via VEGFR2, stimulated the neural differentiation of hESCs in vitro.展开更多
Background Hematopoietic stem cells （HSCs） give rise to all blood and immune cells and are used in clinical transplantation protocols to treat a wide variety of refractory diseases, but the amplification of HSCs has...Background Hematopoietic stem cells （HSCs） give rise to all blood and immune cells and are used in clinical transplantation protocols to treat a wide variety of refractory diseases, but the amplification of HSCs has been difficult to achieve in vitro. In the present study, the expansive effects of aorta-gonad-mesonephros （AGM） region derived stromal cells on HSCs were explored, attempting to improve the efficiency of HSC transplantation in clinical practice.Methods The murine stromal cells were isolated from the AGM region of 12 days postcoitum （dpc） murine embryos and bone marrow（BM） of 6 weeks old mice, respectively. After identification with flow cytometry and immunocytochemistry, the stromal cells were co-cultured with ESCs-derived, cytokines-induced HSCs. The maintenance and expansion of ESCs-derived HSCs were evaluated by detecting the population of CD34^＋ and CD34^＋ Sca-1^＋ cells with flow cytometry and the blast colony-forming cells （BL-CFCs）, high proliferative potential colony-forming cells （HPP-CFCs） by using semi-solid medium colonial culture. Finally, the homing and hematopoietic reconstruction abilities of HSCs were evaluated using a murine model of HSC transplantation in vivo.Results AGM and BM-derived stromal cells were morphologically and phenotypically similar, and had the features of stromal cells. When co-cultured with AGM or BM stromal cells, more primitive progenitor cells （ HPP-CFCs ） could be detected in ESCs derived hematopoietic precursor cells, but BL-CFC＇ s expansion could be detected only when co-cultured with AGM-derived stromal cells. The population of CD34^＋ hematopoietic stem/ progenitor cells were expanded 3 times, but no significant expansion in the population of CD34^＋ Sca-1^＋ cells was noted when co-cultured with BM stromal cells. While both CD34^＋ hematopoietic stem/progenitor cells and CD34^＋ Sca-1^＋ cells were expanded 4 to 5 times respectively derived stromal cells, like BM-derived stromal cells, homing to BM in vivo. when co-cultured with AGM stromal cells. AGM regioncould promote hematopoietic reconstruction and HSCs＇homing to BM in vivo.Conclusions AGM-derived stromal cells in comparison with the BM-derived stromal cells could not only support the expansion of HSCs but also maintain the self-renewal and multi-lineage differentiation more effectively. They are promising in HSC transplantation.展开更多
文摘The effects of vascular endothelial growth factor （VEGF） on neural differentiation of human embryonic stem cells （hESCs） in vitro and the possible mechanism were observed. The hESCs lines, TJMU1 and TJMU2, were established and stored by our laboratory, hESCs differentiated into neuronal cells through embryonic body formation. In this induction process, hESCs were divided into three groups： group A, routine induction; group B, routine induction＋10 ng/mL VEGF; group C, routine in- duction＋10 ng/mL VEGF＋10 ng/mL VEGFR2/Fc. OCT4, Nestin and GFAP in each group were de- tected by RT-PCR, and the cells expressing Nestin and GFAP were counted by immunofluorescence. The percentage of Nestin positive cells in group B was significantly higher than in groups A and C, while the percentage of GFAP positive cells in group B was significantly lower than in groups A and C （P〈0.01）. There was no significant difference between groups A and C （P〉0.05）. It was concluded that VEGF, via VEGFR2, stimulated the neural differentiation of hESCs in vitro.
文摘Background Hematopoietic stem cells （HSCs） give rise to all blood and immune cells and are used in clinical transplantation protocols to treat a wide variety of refractory diseases, but the amplification of HSCs has been difficult to achieve in vitro. In the present study, the expansive effects of aorta-gonad-mesonephros （AGM） region derived stromal cells on HSCs were explored, attempting to improve the efficiency of HSC transplantation in clinical practice.Methods The murine stromal cells were isolated from the AGM region of 12 days postcoitum （dpc） murine embryos and bone marrow（BM） of 6 weeks old mice, respectively. After identification with flow cytometry and immunocytochemistry, the stromal cells were co-cultured with ESCs-derived, cytokines-induced HSCs. The maintenance and expansion of ESCs-derived HSCs were evaluated by detecting the population of CD34^＋ and CD34^＋ Sca-1^＋ cells with flow cytometry and the blast colony-forming cells （BL-CFCs）, high proliferative potential colony-forming cells （HPP-CFCs） by using semi-solid medium colonial culture. Finally, the homing and hematopoietic reconstruction abilities of HSCs were evaluated using a murine model of HSC transplantation in vivo.Results AGM and BM-derived stromal cells were morphologically and phenotypically similar, and had the features of stromal cells. When co-cultured with AGM or BM stromal cells, more primitive progenitor cells （ HPP-CFCs ） could be detected in ESCs derived hematopoietic precursor cells, but BL-CFC＇ s expansion could be detected only when co-cultured with AGM-derived stromal cells. The population of CD34^＋ hematopoietic stem/ progenitor cells were expanded 3 times, but no significant expansion in the population of CD34^＋ Sca-1^＋ cells was noted when co-cultured with BM stromal cells. While both CD34^＋ hematopoietic stem/progenitor cells and CD34^＋ Sca-1^＋ cells were expanded 4 to 5 times respectively derived stromal cells, like BM-derived stromal cells, homing to BM in vivo. when co-cultured with AGM stromal cells. AGM regioncould promote hematopoietic reconstruction and HSCs＇homing to BM in vivo.Conclusions AGM-derived stromal cells in comparison with the BM-derived stromal cells could not only support the expansion of HSCs but also maintain the self-renewal and multi-lineage differentiation more effectively. They are promising in HSC transplantation.
