Vascular Heterogeneity Determination by Marrow Progenit*

描述(由申请人提供)： 这一建议的广泛、长期的目标是确定参与选择性动员和招募骨髓来源的内皮和造血干/祖细胞的血管生成途径，从而决定器官特异性血管系统的异质性。特别是，我们计划确定血管内皮生长因子受体VEGFR2(Flk-1，KDR)、VEGFR1(Flt-1)和VEGFR3(Flt-4)的表达在再生过程中协调BM来源的前体细胞增殖、动员和整合到器官特异性新生血管中的机制，包括肺再生和BM血管生成重建。我们已经证明，血管生成因子家族促进CD133+VEGFR2+内皮祖细胞(EPC)从骨髓向新生血管募集。我们还证明了功能性的VEGFR1表达在造血干细胞和祖细胞(HSPC)的亚群上，支持从BM动员这些细胞。血管内皮细胞生长因子受体1+血管内皮祖细胞共同募集到新生血管中，可促进血管内皮细胞生长因子受体2+内皮祖细胞进入功能性新生血管。骨髓来源的祖细胞的动员是一个动态的过程，需要从独特的骨髓细胞定位中招募这些细胞。血管生成因子诱导金属蛋白酶-9(MMP9)的表达，进而促进可溶性Kit配体(SKitL)的释放。增加生物可利用的sKitL促进HSPC的循环和增殖，为动员进入循环奠定基础。骨髓中还含有CD133+VEGFR3+淋巴源性内皮祖细胞，可能有助于淋巴管生成。在这些研究的基础上，我们假设再生肺和骨髓提供了一个有利于血管生成的微环境，允许招募和整合具有血管功能的骨髓来源的前体细胞。器官特异性血管生成因子促进VEGFR2+和VEGFR3+EPC向新生血管的动员和募集。在器官再生的初始阶段，VEGFR1+造血细胞的共募集促进了血管前体细胞的功能整合，并决定了血管的异质性。通过研究以下特定目标，我们提出了这一假设：我们计划确定组织血管重建过程中骨髓来源的前体细胞的时间、空间和区域招募模式，并比较它们在血管生成潜力降低的转基因小鼠中的整合模式，包括VEGF164/164、VEGF189/189、PIGF-/-和Id1+/-ID3-/-小鼠。明确VEGFR1、VEGFR2和VEGFR3信号在调节祖细胞动员、归巢和募集到肺和骨髓血管系统中的作用。探讨骨髓来源的CD133+VEGFR2+、CD133+VEGFR3+内皮祖细胞和VEGFR1+HSPC在器官再生过程中对血管重建的生理意义和贡献。这些研究将为将骨髓来源的细胞用于治疗性细胞治疗以促进器官(即肺、骨髓)的血运重建奠定基础。

DESCRIPTION (provided by applicant): The broad, long-term objective of this proposal is to identify angiogenic pathways that are involved in selective mobilization and recruitment of bone marrow (BM)-derived endothelial and hematopoietic stem and progenitor cells thereby dictating heterogeneity of organ-specific vasculature. In particular, we plan to determine the mechanism by which the expression of Vascular Endothelial Growth Factor-receptors, VEGFR2 (Flk-1, KDR), VEGFR1 (Flt-1) and VEGFR3 (Flt-4) orchestrate proliferation, mobilization and incorporation of BM-derived progenitors into organ-specific neo-vasculature during regenerating processes, including lung regeneration and BM hemangiogenic reconstitution. We have shown that VEGF family of angiogenic factors promote recruitment of CD133+VEGFR2+ endothelial progenitor cells (EPCs) from BM to the angiogenic neo-vessels. We have also demonstrated that functional VEGFR1 is expressed on the subsets of hematopoietic stem and progenitors cells (HSPCs) supporting mobilization of these cells from BM. Co-recruitment of angio-competent VEGFR1+HSPCs to the neo-angiogenic vessels facilitate incorporation of VEGFR2+EPCs into functional neo-vessels. Mobilization of BM-derived progenitor cellsl is a dynamic process and requires recruitment of these cells from unique BM niches. Angiogenic factors, induce expression of metalloproteinase-9 (MMP-9), which in turn promote the release of soluble Kit-ligand (sKitL). Increase in bio-available sKitL enhance cycling and proliferation of HSPCs, setting up the stage for mobilization to the circulation. BM also contains a population of CD133+VEGFR3+ lymphatic EPCs that could possibly contribute to lymphangiogenesis. Based on these studies, we hypothesize that regenerating lung and BM provide for a pro-hemangiogenic microenvironment that is permissive for recruitment and incorporation of angio-competent BM-derived progenitors. Organ-specific angiogenic factors promote mobilization and recruitment of VEGFR2+ and VEGFR3+ EPCs to the neo-vessels. Co-recruitment of the VEGFR1+ hematopoietic cells facilitate functional incorporation of vascular progenitors and dictate vascular heterogeneity in the initial phases of organ regeneration. This hypothesis through studying the following specific aims: We plan to determine temporal, spatial and regional recruitment patterns of BM-derived progenitors during tissue revascularization-remodeling and compare their incorporation pattern in transgenic mice with diminished hemangiogenic potential including, VEGF164/164, VEGF189/189, PIGF-/-, and Id1+/-Id3-/- mice. Define the role of VEGFR1, VEGFR2 and VEGFR3 signaling in the regulation of mobilization, homing and recruitment of progenitors to the pulmonary and BM vasculature. Assess the physiological significance and contribution of BM-derived CD133+VEGFR2+, CD133+VEGFR3+EPCs and VEGFR1+HSPCs to revascularization during organ regeneration. These studies will lay the foundation for using BM-marrow derived cells for therapeutic cell therapy to enhance organ (i.e. lung, marrow) revascularization.