CHEMOKINES AND IMMUNE CELLS IN HIND LIMB ISCHEMIA

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DESCRIPTION (provided by applicant): Muscle regeneration and angiogenesis are important components of limb salvage following traumatic and/or ischemic injury to the extremities. We have demonstrated that knockout mice lacking either monocyte chemotactic protein-1 (MCP-1) or its specific receptor, the CC Chemokine Receptor 2 (CCR2), have impairments in macrophage recruitment and muscle regeneration following ischemic or toxic injury. In addition, CCR2-/- mice exhibit increased adipocyte accumulation in regenerated muscle. Bone marrow (BM) replacement of CCR2 -/- mice with wild type (WT) BM (radiation chimeras) led to increased early macrophage recruitment and normal muscle regeneration (i.e., recapitulated the phenotype of WT mice). Furthermore, replacing the BM of WT mice with CCR2 -/- BM resulted in decreased macrophage recruitment and impaired muscle regeneration (i.e., recapitulated the phenotype of CCR2 -/- mice). This suggests that BM-derived cells modulate the healing responses of muscle and that macrophages are the likely BM-derived cell type that mediates the impaired muscle regeneration in CCR2 -/- mice. Further studies revealed impaired angiogenesis in CCR2 -/- mice in conjunction with decreased tissue vascular endothelial growth factor (VEGF) compared to WT mice. Interestingly, restoration of VEGF to baseline levels was associated with the development of maximal capillary density in both CCR2 -/- and WT mice. While impaired muscle regeneration in CCR2 -/- mice is attributable to a BM-derived cell, the effects of BM-derived vs. host-derived cells have not been studied in regards to angiogenesis. Our long-term goal is to define the influence of inflammation, including the chemokine system, in angiogenesis and skeletal muscle regeneration. The following 3 specific aims will test the overall hypothesis that the recruitment and activation of BM-derived cells, especially monocytes/macrophages, are essential for angiogenesis, a critical component in skeletal muscle regeneration after injury. 1) Determine the contribution of CCR2 expression in BM-derived vs. non-BM- derived cells on angiogenesis in skeletal muscle after injury, 2) Determine the ex vivo and in vivo influence of the MCP-1/CCR2 axis on angiogenesis and 3) Define the effects of selective and complete monocyte/macrophage ablation prior to muscle injury and sustained throughout the time course of tissue repair on inflammation and angiogenesis after injury. The proposed studies are innovative because they will help define the contribution of BM-derived cells to angiogenesis. The significance of this research is that a better understanding of the mechanisms of skeletal muscle regeneration and angiogenesis could lead to the design of novel primary or adjuvant treatments for improved limb salvage and tissue engineering. PUBLICE HEALTH RELEVANCE: Leg and arm wounds, with large muscle defects and high amputation rates, are common in trauma victims and especially in injured soldiers from the Iraqi/Afghanistan war; new treatments to replace the missing muscle are needed to decrease amputation rates and improve limb function. Our research studies the complex relationships between the multiple cells that are needed to make new muscle, including blood vessels. A better understanding of how new blood vessels help muscle recover from injury could lead to new therapies, including tissue engineering strategies, to help patients recover from these devastating injuries.