Impact of Aging on Stem Cell Repair in Atherosclerosis

动脉粥样硬化是一种异常的、持续的修复性炎症过程， 血管壁受伤。事实上，心血管系统持续暴露于多种 其影响随着时间的推移而累积的侮辱。参与修复的两种细胞来源 过程存在：（1）局部的，分化的，血管内皮细胞和平滑肌细胞， 邻近血管节段;和（2）通过以下途径从骨髓募集干细胞/血管祖细胞： 外周循环我们最近发现，长期给予全骨髓（BM） 细胞显着减少动脉粥样硬化病变的形成在建立的小鼠模型， 动脉粥样硬化-ApoE“/”C57/B6小鼠喂养高脂胆固醇，西式饮食-即使在没有 ApoE基因表达恢复和血浆胆固醇水平正常化（>1200 mg/dl）。 此外，来自年轻而非老年ApoE-/-小鼠的BM细胞能够使血管再生， 动脉粥样硬化的预防这些数据强调了干细胞/血管祖细胞在 血管愈合和动脉粥样硬化形成，并为使用干细胞/祖细胞疗法作为 用于动脉粥样硬化的新的预防和/或治疗策略， 骨髓及其老化是心血管系统长期完整性的瓶颈。 在这个项目中，我们提出了一个假设，即经过一生的修复动脉粥样硬化， 供应维持血管内稳态所需的特定类型的血管祖细胞（VPC）。 心血管系统在某种程度上耗尽或这些细胞功能受损。如果VPC“备用 部件”可以外包，修复过程可以在适当的时间间隔和动脉粥样硬化 后果被推迟，也许是无限期的。具体目标1：确定定量组成 (FACS分析），后代功能特征和基因表达表型（微阵列分析）， 全骨髓细胞或谱系阴性侧群（linSP）细胞，其从年轻人与老年人获得 （野生型和ApoE）小鼠;具体目标2：为了确定感受态VPC被包括在所述细胞系中， 骨髓中的SP分数，其传达抗动脉粥样硬化的功效，通过测试 未分级的骨髓细胞、富集的linSP细胞和去除linSP级分的骨髓细胞 从年轻与老年（野生型和ApoE +/-）小鼠中获得的抑制血浆中升高的 化学细胞因子和生长因子以及预防ApoE /小鼠中动脉粥样硬化病变形成; 具体目的3：确定年轻apoE -#和野生型BM的linSP部分内的相对生物学活性。 a）候选VPC（CVPC，在衰老小鼠中耗尽或功能受损的细胞）的功效 使用目的1中鉴定的标志物分离，B）未分级分离的linSP细胞减去CVPC（USP-CVPC），和c） 未分级的linSP细胞（USP）在抑制升高的IL-6、VEGF和其他 具体目的4：确定ApoE /-小鼠中的炎症标志物和预防动脉粥样硬化; O_4-整合素、VCAM-1和E-选择素在介导血管祖细胞移植至血管内皮细胞中的作用 apoE-/-小鼠慢性血管壁损伤。作为推论，细胞移植的相对贡献 相对于非细胞自主机制的血管修复将被表征。

Atherosclerosis represents an aberrant, continuous reparative inflammatory process in response to repeated injuries to the vessel wall. Indeed, the cardiovascular system is continuously exposed to a multitude of insults whose impact cumulates with the passage of time. Two sources of cells participating in the repair process exist: (1) local, differentiated, vascular endothelial and smooth muscle cells that migrate from adjacent vessel segments; and (2) recruited stern cells/vascular progenitor cells from the bone marrow, via peripheral circulation. We have recently discovered that chronic administration of whole bone marrow (BM) cells significantly reduced atherosclerotic lesion formation in an established mouse model of atherosclerosis--ApoE "/" C57/B6 mice fed high fatcholesterol, Western-type diet--even in the absence of restoration of ApoE gene expression and normalization of plasma cholesterol levels (>1200 mg/dl). Furthermore, BM cells from young, but not old, ApoE-/- mice were capable of vascular rejuvenation and atherosclerosis prevention. These data underscore the importance of stem cells/vascular progenitor cells in vascular healing and atherogenesis and provide support for the use of stern/progenitor cell therapy as a novel preventative and/or treatment strategy for atherosclerosis, particularly for individuals whose bone marrow, and its obsolescence, represents the bottleneck for long-term integrity of the cardiovascular system. In this project, we propose to test the hypothesis that after a lifetime of repairing atherosclerotic arteries, the supply of the specific type(s) of vascular progenitor cells (VPCs) needed to maintain the homeostasis of the cardiovascular system is somehow exhausted or these cells are functionally impaired. If the VPC "spare parts" can be outsourced, the repair process can be boosted at appropriate intervals and atherosclerotic consequences delayed, perhaps indefinitely. Specific Aim 1: To determine the quantitative composition (FACS analysis), progeny functional characteristics, and gene expression phenotype (microarray analysis) of whole bone marrow cells or lineage negative side population (linSP) cells obtained from young versus old (wild-type, and ApoE "#) mice; Specific Aim 2: To establish that competent VPCs are encompassed in the lin SP fraction in the marrow, which convey the anti-atherosclerotic efficacy, by testing the effects of unfractionated bone marrow cells, enriched linSP cells and bone marrow cells deprived of linSP fraction obtained from young versus old (wild-type and ApoE "/') mice in suppressing elevated plasma levels of chemo-cytokines and growth factors and in preventing atherosclerotic lesion formation in ApoE / mice; Specific Aim 3: To determine within the linSP fraction of young apoE -# and wild-type BM the relative efficacy of a) candidate VPCs (CVPC, cells that are either depleted or functionally impaired in aging mice) isolated using markers identified in Aim 1, b) unfractionated linSP cells minus CVPCs (USP-CVPC), and c) unfractionated linSP cells (USP) in suppressing elevated plasma levels of IL-6, VEGF and other inflammatory markers and in preventing atherosclerosis in ApoE /- mice; and Specific Aim 4: To determine the role of o_4-integrin, VCAM-1, and E-selectin in mediating the engraftment of vascular progenitor cells to chronically injured vessel wall in apoE-/- mice. As a corollary, the relative contribution of cell engraftment versus a non-cell autonomous mechanism to vascular repair will be characterized.