缺血损伤后，免疫炎症通路的激活，包括TNFα（肿瘤坏死因子α）信号通路，对于再灌注和外周肌肉组织修复至关重要。然而，TNFα驱动的炎症级联反应机制，导致前期通向缺血部位的免疫细胞介导的血管新生仍然未知。通过激光多普勒成像和光片荧光显微镜监测了永久性股动脉结扎后的肌肉组织再灌注情况，研究了TNFα介导的信号传导和CCL20（C-C动机化学因子配体20）-CCR6（C-C趋化因子受体6）轴在血管新生中的作用，包括骨髓移植、流式细胞术以及生物化学和分子生物学技术。发现TNFα介导的肿瘤坏死因子受体TNFR（肿瘤坏死因子受体）1的激活是缺血损伤后肌肉组织再灌注所必需的，而不是TNFR2。骨髓源CCR6+中性粒细胞被鉴定为一种新发现的TNFα诱导的前血管生成性中性粒细胞群体，其表达VEGFA（血管内皮生长因子A）上调。机械上，缺血后TNFR1的激活诱导了血管细胞中CCL20的表达，并促进了CCR6受体在中性粒细胞的细胞表面的转位，从而导致VEGFA表达的前血管生成性中性粒细胞在缺血部位对CCL20的依赖性招募。此外，糖尿病小鼠缺血外周肌肉组织再灌注不良与前血管生成性中性粒细胞数量减少和CCL20表达降低相关。重组CCL20的给药增加了前血管生成性中性粒细胞的招募，并改善了糖尿病缺血骨骼肌组织再灌注，而联合用药的含义则通过顺序处理含氟那汀，进一步巩固了结果。我们证明了通过TNFα介导的CCL20-CCR6轴的特异性激活可将表达VEGFA的前血管生成性中性粒细胞招募至缺血损伤部位，以启动肌肉组织再灌注。这些发现为组织再灌注提供了一种有吸引力的选择，尤其在糖尿病情况下。

Activation of immune-inflammatory pathways involving TNFα (tumor necrosis factor alpha) signaling is critical for revascularization and peripheral muscle tissue repair after ischemic injury. However, mechanisms of TNFα-driven inflammatory cascades directing recruitment of proangiogenic immune cells to sites of ischemia are unknown.Muscle tissue revascularization after permanent femoral artery ligation was monitored in mutant mice by laser Doppler imaging and light sheet fluorescence microscopy. TNFα-mediated signaling and the role of the CCL20 (C-C motif chemokine ligand 20)-CCR6 (C-C chemokine receptor 6) axis for formation of new vessels was studied in vitro and in vivo using bone marrow transplantation, flow cytometry, as well as biochemical and molecular biological techniques.TNFα-mediated activation of tumor necrosis factor receptor TNFR (tumor necrosis factor receptor) 1 but not TNFR2 was found to be required for postischemic muscle tissue revascularization. Bone marrow-derived CCR6+ neutrophil granulocytes were identified as a previously undescribed TNFα-induced population of proangiogenic neutrophils, characterized by increased expression of VEGFA (vascular endothelial growth factor A). Mechanistically, postischemic activation of TNFR1 induced expression of the CCL20 in vascular cells and promoted translocation of the CCL20 receptor CCR6 to the cell surface of neutrophils, essentially conditioning VEGFA-expressing proangiogenic neutrophils for CCL20-dependent recruitment to sites of ischemia. Moreover, impaired revascularization of ischemic peripheral muscle tissue in diabetic mice was associated with reduced numbers of proangiogenic neutrophils and diminished CCL20 expression. Administration of recombinant CCL20 enhanced recruitment of proangiogenic neutrophils and improved revascularization of diabetic ischemic skeletal muscles, which was sustained by sequential treatment with fluvastatin.We demonstrate that site-specific activation of the CCL20-CCR6 axis via TNFα recruits proangiogenic VEGFA-expressing neutrophils to sites of ischemic injury for initiation of muscle tissue revascularization. The findings provide an attractive option for tissue revascularization, particularly under diabetic conditions.