Hematopoietic stem cell cytokine control of developmental vascularization

在这个新的提案中，我们调查了我们的新发现，即一种独特的造血干细胞组合 细胞细胞因子(即干细胞因子-SCF、白介素3-IL-3和基质衍生因子-1α-SDF-1) 控制人内皮细胞(EC)管形态发生和EC-周细胞管共组装 确定了3D胶原蛋白基质中的无血清条件(以及在不存在佛波酯的情况下)。 有趣的是，血管内皮生长因子本身没有影响，需要这些造血因子的协同作用。 为了对EC的萌发起作用，它的主要形态发生影响。我们提出这样的假设： SCF、IL-3和SDF-1是血管形态发生的关键调节因子，它们以协同的方式发挥作用，并 是已知的发育血管形成介质作用的必要辅助因子(例如，血管内皮生长因子和 BMP-4)。C-Kit(SCF受体)和CXCR4(SDF-1受体)抑制剂联合应用或 阻断抗SCF和IL-3抗体导致发育中的鹌鹑胚胎血管出血表型 这与血管形成和重塑中的缺陷有关。此外，我们还表明，这三个因素协同作用 激活Src、B-Raf和Erk激酶，诱导已知的整合素和Ak2的表达 血管形态发生的调节剂。我们将阐明信号协同作用的分子基础。 内皮细胞中的三种细胞因子，这些因子如何与血管内皮生长因子、骨形态发生蛋白-4和周细胞相互作用来控制内皮细胞的萌发 以及它们在体外和体内的管子共组装和成熟事件中如何影响内皮细胞和周细胞。 我们提出了三个具体目标： 具体目标1.阐明SCF、IL-3和SDF-1协同作用的信号机制 在3D细胞外基质中刺激血管生成EC管组装。 具体目标2.确定SCF、IL-3和SDF-1如何影响血管内皮细胞生长因子和骨形态发生蛋白-4信号转导 EC在3D细胞外基质中的萌发反应。 具体目标#3.确定SCF、IL-3和SDF-1如何调节周细胞诱导的EC 3D细胞外基质中的萌发反应和血管生成EC-周细胞管共组装。

In this new proposal, we investigate our novel finding that a unique combination of hematopoietic stem cell cytokines (i.e. stem cell factor-SCF; interleukin 3- IL-3; and stromal-derived factor-1 alpha- SDF-1¿) control human endothelial cell (EC) tube morphogenesis and EC-pericyte tube coassembly under defined serum-free conditions (and in the absence of phorbol ester) in 3D collagen matrices. Interestingly, VEGF has no influence by itself and requires the synergistic action of these hematopoietic factors in order to exert an effect on EC sprouting, its major morphogenic influence. We propose the hypothesis that SCF, IL-3 and SDF-1¿ are critical regulators of vascular morphogenesis which act in a synergistic manner and are necessary cofactors for the action of known mediators of developmental vascularization (e.g. VEGF and BMP-4). Combined administration of c-Kit (the SCF receptor) and CXCR4 (the SDF-1¿ receptor) inhibitors or blocking antibodies to SCF and IL-3 leads to vascular hemorrhage phenotypes in developing quail embryos that relates to defects in vessel formation and remodeling. Also, we show that the three factors synergize to activate Src, B-Raf and Erk kinases and induce the expression of the ¿2¿1 integrin and Pak2 which are known regulators of vascular morphogenesis. We will elucidate the molecular basis for the signaling synergism of the three cytokines in ECs, how these factors interface with VEGF, BMP-4, and pericytes to control EC sprouting and how they affect both ECs and pericytes during tube coassembly and maturation events in vitro and in vivo. We propose three specific aims which are; Specific Aim #1. To elucidate the signaling mechanisms by which SCF, IL-3 and SDF-1¿ synergize to stimulate vasculogenic EC tube assembly in 3D extracellular matrices. Specific Aim #2. To determine how SCF, IL-3 and SDF-1¿ affect VEGF and BMP-4 signaling to control EC sprouting responses in 3D extracellular matrices. Specific Aim #3. To determine how SCF, IL-3 and SDF-1¿ act to regulate pericyte-induced EC sprouting responses and vasculogenic EC-pericyte tube coassembly in 3D extracellular matrices.