Vasoprotective Actions of Autologous Cell Transfer

描述(由申请人提供)：本建议的目的是在急性血管损伤的动物模型中表征和增强培养修饰的单个核细胞(CMMC)和内皮生长细胞(EOCs)的血管保护作用。最近在嵌合动物模型中的研究表明，骨髓来源的细胞参与了对血管损伤的细胞反应。在适当条件下体外培养的循环单个核细胞可能呈现内皮细胞表型。我们的目标是为了治疗目的而适应和修饰这些自体细胞。CMMC是通过使用特定的条件培养外周血单个核细胞(PBMC)向内皮表型转化而产生的。培养7天后，大多数早期CMMC表达CD14(内毒素受体和单核细胞标志)，尽管这个异质性群体也包括内皮生长细胞(EOCs)和脱落的成熟循环内皮细胞(CD14-)的前体细胞。在以后的时间点，这些培养物由同质的EOC群体(CD14-)组成。我们实验室的工作已经证明，在直接血管损伤模型中，局部输送的早期CMMC和EOCs具有强大的血管保护作用。我们的第一个工作假设是，自体早期的CMMC和EOCs以内皮依赖的方式在受损的血管系统中发挥血管保护作用。我们的第二个工作假设是，基因改造将进一步增强EOCs的血管保护作用。我们的四个具体目标是：具体目标1：比较早期CMMC和EOCs局部注射的血管保护作用。具体目标2：明确早期CMMC和EOC传递的血管保护作用的机制。具体目标3：优化转基因EOCs的表达，增强其局部血管保护作用。具体目标4：优化转基因EOCs的系统转基因表达，并确定其系统血管保护潜力。

DESCRIPTION (provided by applicant): The objective of this proposal is to characterize and enhance the vasoprotective effects of culture-modified mononuclear cells (CMMCs) and endothelial outgrowth cells (EOCs) in an animal model of acute vascular injury. Recent studies in chimeric animal models have demonstrated that bone marrow-derived cells participate in the cellular response to vascular injury. Circulating mononuclear cells, cultured in vitro under appropriate conditions, may assume an endothelial phenotype. Our goal is to adapt and modify these autologous cells for therapeutic purposes. CMMCs are generated by culturing peripheral blood mononuclear cells (PBMCs) towards an endothelial phenotype using defined conditions. Following 7 days in culture, the majority of early CMMCs express CD14 (the LPS receptor and a monocyte marker) although this heterogenous population also includes precursors to endothelial outgrowth cells (EOCs) and sloughed mature circulating endothelial cells (both CD14-). At later time points, these cultures consist of homogenous populations of EOCs (CD14-). Work in our laboratory has demonstrated potent vasoprotective effects of locally delivered early CMMCs and EOCs in a model of direct vascular injury. Our first working hypothesis is that autologous early CMMCs and EOCs exert vasoprotective actions in the injured vasculature in an endothelial dependent fashion. Our second working hypothesis is that genetic modification will further enhance the vasoprotective effects of EOCs. Our four specific aims are: Specific Aim 1: To compare the vasoprotective effects associated with local delivery of early CMMCs and EOCs. Specific Aim 2: To define the mechanisms responsible for the vasoprotective effects of early CMMC and EOC delivery. Specific Aim 3: To optimize transgene expression from genetically modified EOCs and to enhance their local vasoprotective actions. Specific Aim 4: To optimize systemic transgene expression from genetically modified EOCs and to determine their systemic vasoprotective potential.