CXCR4-dependent tissue remodeling after experimental stroke

CXCR4趋化因子受体在造血干细胞和神经干细胞中表达。该受体引导白细胞和来源于这些干细胞的前体细胞的迁移。最近，一种CXCR4拮抗剂(plerixafor，AMD3100)被批准用于造血干细胞动员。深入研究了CXCR4在肿瘤、炎症和再生过程中的功能。在此背景下，据报道，CXCR4介导的趋化因子CXCL12信号将干细胞和炎症细胞导向受损组织。因此，CXCR4被认为与组织重塑、新生血管和肿瘤形成有关。然而，对这些过程的适当研究需要毫不含糊地识别从表达CXCR4的干细胞发展而来的组织。此外，CXCR4的可控失活也是可取的。由于CXCL12和CXCR4缺陷小鼠在围产期死亡，有条件的方法是必要的。申请者因此产生了携带CXCR4基因敲除/Creer敲除等位基因(Cxcr4CreER)和已建立的Cre报告基因的小鼠。在这些小鼠中，他莫昔芬诱导荧光蛋白TdTomato在具有活性CXCR4启动子的细胞中永久表达。当Cxcr4Creer等位基因与Cxcr4LoxP等位基因结合时，CXCR4-Creer允许CXCR4消融。对Cxcr4Creer/WT和Cxcr4Creer/loxP小鼠的全面鉴定表明，对TdTomato阳性细胞的分析允许在存在和不存在功能性CXCR4的情况下追踪表达CXCR4的干细胞的谱系(Cxcr4Creer/WT)。申请人计划使用这个模型来评估CXCR4在实验性中风后重塑过程中的功能(单核细胞、中性粒细胞和未成熟神经元的招募，受损区域的胶质反应和新生血管)。在第二个焦点中，干细胞将被体内标记CXCR4-Creer，在体外繁殖和再移植过程中分离和鉴定。该项目将阐明CXCR4是否是治疗中风的合适药物靶点的问题，并严格测试CXCR4在缺血损伤重塑期间指导干细胞的概念。

CXCR4 chemokine receptors are expressed in hematopoietic and neural stem cells. The receptor guides migration of leukocytes and precursor cells that are derived from these stem cells. Consistently, a CXCR4 antagonist (plerixafor, AMD3100) was recently approved for hematopoietic stem cell mobilization. Functions of CXCR4 in neoplastic, inflammatory and regenerative processes are intensively investigated. In this context, it was reported that the CXCR4-mediated signal of the chemokine CXCL12 directs stem cells and inflammatory cells towards lesioned tissues. Accordingly, CXCR4 is thought to be involved in tissue remodeling, neovascularization and tumor formation. However, proper investigation of these processes requires non-ambiguous identification of tissues that develop from Cxcr4-expressing stem cells. In addition, controllable inactivation of CXCR4 is desirable. Since CXCL12- and CXCR4-deficient mice die perinatally, a conditional approach is necessary. The applicant thus generated mice harboring a Cxcr4-knockout/CreER-knockin allele (Cxcr4CreER) and an established Cre-reporter. In these mice, tamoxifen induces permanent expression of the fluorescent protein TdTomato in cells with an active Cxcr4 promoter. When the Cxcr4CreER allele is combined with a Cxcr4LoxP allele, Cxcr4-CreER permits Cxcr4 ablation. Thorough characterization of Cxcr4CreER/WT and Cxcr4CreER/LoxP mice showed that analysis of TdTomato-positive cells permits to trace the lineage of Cxcr4-expressing stem cells in the presence (Cxcr4CreER/WT) and in the absence (Cxcr4CreER/LoxP) of functional CXCR4. The applicant plans to use this model to assess the function of CXCR4 in remodeling processes after experimental stroke (recruitment of monocytes, neutrophils and immature neurons, glial reaction and neovascularization in the lesioned area). In the second focus, stem cells will be in vivo-labeled with Cxcr4-CreER, isolated and characterized during in vitro propagation and reimplantation. The project will shed light on the question if CXCR4 is a suitable drug target in stroke and rigorously test the concept that CXCR4 directs stem cells during remodeling of ischemic lesions.