Functional role and microfluidic gene chip analysis of proangiogenic chemokines in the course of ischemia-dependent endothelial progenitor cell (EPC) vasculogenesis

血管内皮祖细胞(EPC)被证明对缺血区的血运重建有很大的贡献。内皮祖细胞的发现实质上改变了我们对成人组织血管生成的看法，表明骨髓来源的细胞可以促进新血管的形成。然而，经过10年的积极研究，关于EPC的出现还没有达成共识。人们普遍认为，内皮祖细胞通过释放其携带的旁分泌血管生成因子来促进血管生成和血管生成。其中一个血管生成因子，也是我感兴趣的焦点是多效性细胞因子巨噬细胞移动抑制因子(MIF)。最近发现，MIF具有趋化素样功能，并与CXC趋化因子受体CXCR4非同源相互作用，CXCR4是调节祖细胞迁移的主要受体，也表达于内皮祖细胞，已有研究表明，MIF参与了缺血依赖的EPC募集。基于这些结果，我现在想要阐明促血管生成趋化因子，如MIF和CXCL12在低氧依赖诱导内皮细胞和内皮细胞后的功能作用。此外，还将进行趋化因子-细胞因子-多重分析、静态黏附实验和动态迁移(趋化)实验，以确定MIF/CXCR4轴和CXCL12/CXCR4轴的作用。为了回答有关异质内皮祖细胞性质的基本问题，将开发一种创新的单细胞分析方法，利用单细胞流式细胞仪分选和基于商业可用的微流控技术，同时对给定细胞群体中的许多单个细胞进行48个基因靶点的定量PCR基因表达分析。该项目的最后一步涉及另一种创新的方法，即用于组织转移的转基因游离瓣，以扩大我的研究组合的临床范围。

Endothelial progenitor cells (EPCs) have been shown to amply contribute to the revascularization of ischemic areas. The discovery of EPCs has substantively altered our view of adult tissue angiogenesis, indicating that bone marrow-derived cells can contribute to new blood vessel formation. However, after 10 years of vigorous research no consensus about the appearance of EPCs has been reached yet. It is widely accepted that EPCs promote angiogenesis and vasculogenesis by virtue of the release of paracrine angiogenic factors which they carry with them as cargo. One of these angiogenic factors and my focus of interest is the pleiotropic cytokine macrophage migration inhibitory factor (MIF). Based on the recent discovery that MIF exhibits chemokine-like functions and interacts as a non-cognate interaction of the CXC chemokine receptor CXCR4, a major receptor regulating progenitor cell migration that is also expressed on EPCs, it has been shown that MIF contributes to ischemia-dependent recruitment of EPCs. Based on these results I now want to clarify the functional role of proangiogenic chemokines, such as MIF and CXCL12 after hypoxia dependent induction in endothelial cells and EPCs. Furthermore chemokine-cytokine-multiplex assays, static adhesion experiments and dynamic transmigration assays (chemotaxis) will be performed to functional underlie the role of the MIF/CXCR4 axis and CXCL12/CXCR4 axis vice versa. To answer fundamental questions regarding the nature of the heterogeneous EPCs, an innovative approach to single cell analysis utilizing single cell FACS sorting and commercially available microfluidic-based technology to perform quantitative PCR gene expression analysis across 48 gene targets simultaneously for many individual cells within a given cell population will be developed. A last step of the program deals with another innovative approach of genetically modified free flaps for tissue transfer to expand the clinical spectrum of my research portfolio.