内皮生长因子和胎盘生长因子对小鼠胎肝造血干细胞髓系分化的作用

Myeloid Differentiation of Murine Fetal Liver Hematopoiesis Could Be Regulated by VEGF and PlGF

【目的】观察VEGF(内皮生长因子)和PlGF(胎盘生长因子)对小鼠胎肝造血干细胞髓系分化的影响,并探讨其调控机制。【方法】体外培养E13胎肝造血细胞,培养造血集落CFU-GM,观察VEGF和PlGF对胎肝造血干细胞髓系分化的作用,应用Western-Blot检测不同处理条件对胎肝造血细胞Akt蛋白以及p-Akt蛋白表达水平的影响,用MTT法来评估不同浓度的LY294002(PI3K/Akt抑制剂)对胎肝造血细胞增殖的影响,通过CFU-GM观察LY294002对VEGF和PlGF调控胎肝造血作用的影响。【结果】VEGF和PlGF均可促进胎肝造血干细胞髓系分化,集落数分别为对照组的141%(P<0.01)和123%(P<0.05);PlGF可上调胎肝造血细胞p-Akt蛋白表达,其表达水平为对照组的143%(P<0.05);VEGF及其与PlGF联合应用对胎肝造血细胞p-Akt蛋白表达水平无明显影响。LY294002可抑制胎肝造血细胞的增殖,并具有明显的剂量依赖关系(r=0.9647,P<0.01),在LY294002存在的条件下加入VEGF或PlGF,其CFU-GM产率与单独应用LY294002相比无明显差异(P>0.05)。【结论】VEGF和PlGF均能促进胎肝造血干细胞的髓系分化;PlGF可通过上调胎肝造血细胞Akt的磷酸化水平调节胎肝造血细胞的髓系分化;LY294002可通过抑制PI3K/Akt信号通路抑制VEGF和PlGF对胎肝造血干祖细胞髓系分化的调控作用。

【Objective】To investigate the effects of VEGF and PlGF on the myeloid differentiation of murine fetal liver hematopoietic stem cells（FL HSCs） and related mechanism.【Methods】E13 murine FL hematopoietic cells were isolated and cultured in vitro to evaluate the effects of VEGF and PlGF on the myeloid differentiation of murine FL HSCs.The influence on proliferation of hematopoietic progenitor cells was detected by the colony forming unit（CFU） formation assay.The expression of Akt and p-Akt in hematopoietic cells were analyzed by Western blotting.The influence of LY294002（inhibitor of PI3K/Akt） on proliferation of FL hematopoietic cells was detected by MTT assay.【Results】Application of VEGF and PlGF could increase the yield of CFU-GM to 141%（P 0.01）and 123%（P 0.05） respectively compared with the control.PlGF upregulated the p-Akt expression level in FL hematopoietic cells,the p-Akt expression level was increased to 143% compare with the control（P 0.05）.Application of VEGF separate or combined with PlGF affected the expression level of p-Akt scarcely compared with the control group.The proliferation of FL hematopoietic cells could be inhibited by LY294002 in a dose-dependent manner（r = 0.9647,P 0.01）.No apparente difference could be observed on the yield of CFU-GM by application of LY294002 separate or combined with VEGF and PlGF（P 0.05）.【Conclusion】The myeloid differentiation of the fetal liver hematopoietic cells could be promoted by VEGF and PlGF.PlGF promotes the myeloid differentiation by up-regulating the phosphorylation of Akt.