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The Development of Macrophage-Mediated Cell Therapy to Improve Skeletal Muscle Function after Injury.

Rybalko V, Hsieh PL, Merscham-Banda M, Suggs LJ, Farrar RP - PLoS ONE (2015)

Bottom Line: A variety of MP phenotypes have been identified and characterized in vitro as well as in vivo.We detected a 15% improvement in specific tension and force normalized to mass after M1 (LPS/IFN-γ) MP transplantation 24 hours post-reperfusion.Interestingly, we found that M0 bone marrow-derived unpolarized MPs significantly impaired muscle function highlighting the complexity of temporally coordinated skeletal muscle regenerative program.

View Article: PubMed Central - PubMed

Affiliation: Department of Kinesiology, The University of Texas at Austin, 1 University Station D3700, Austin, TX 78712, United States of America.

ABSTRACT
Skeletal muscle regeneration following acute injury is a multi-step process involving complex changes in tissue microenvironment. Macrophages (MPs) are one of the key cell types involved in orchestration and modulation of the repair process. Multiple studies highlight the essential role of MPs in the control of the myogenic program and inflammatory response during skeletal muscle regeneration. A variety of MP phenotypes have been identified and characterized in vitro as well as in vivo. As such, MPs hold great promise for cell-based therapies in the field of regenerative medicine. In this study we used bone-marrow derived in vitro LPS/IFN-y-induced M1 MPs to enhance functional muscle recovery after tourniquet-induced ischemia/reperfusion injury (TK-I/R). We detected a 15% improvement in specific tension and force normalized to mass after M1 (LPS/IFN-γ) MP transplantation 24 hours post-reperfusion. Interestingly, we found that M0 bone marrow-derived unpolarized MPs significantly impaired muscle function highlighting the complexity of temporally coordinated skeletal muscle regenerative program. Furthermore, we show that delivery of M1 (LPS/IFN-γ) MPs early in regeneration accelerates myofiber repair, decreases fibrotic tissue deposition and increases whole muscle IGF-I expression.

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Related in: MedlinePlus

Assessment of tissue revascularization and immune cell infiltrate in saline or MP-treated TK-I/R injured muscles 14 days post-reperfusion.Representative images of D14 regenerating skeletal muscle stained with anti-CD31antibody identifying vascular endothelium and anti-CD45 staining for immune cells, scale bar 100μm. Quantification graphs of the number of CD31+ cell per myofiber (capillary/fiber ratio) and CD45+ cells per field of view (FOV). M0 group n = 3, M1 and saline groups n = 4–5, 3–5 images/muscle, values presented as mean ± SEM, (*) p<0.05 relative to M0 MP group
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pone.0145550.g009: Assessment of tissue revascularization and immune cell infiltrate in saline or MP-treated TK-I/R injured muscles 14 days post-reperfusion.Representative images of D14 regenerating skeletal muscle stained with anti-CD31antibody identifying vascular endothelium and anti-CD45 staining for immune cells, scale bar 100μm. Quantification graphs of the number of CD31+ cell per myofiber (capillary/fiber ratio) and CD45+ cells per field of view (FOV). M0 group n = 3, M1 and saline groups n = 4–5, 3–5 images/muscle, values presented as mean ± SEM, (*) p<0.05 relative to M0 MP group

Mentions: Microvascular cell dysfunction accounts for most of the inflammatory response associated with I/R injury and is a rate-limiting step in I/R pathogenesis [49]. Restoration of adequate microvascular supply to regenerating muscle is crucial for functional muscle recovery and performance. We, therefore, evaluated skeletal muscle revascularization in our TK-I/R injured skeletal muscles at 14 days after acute saline or MP treatment. By using CD31 specific antibodies we were able to identify endothelial cells and quantify capillary per myofiber ratio in regenerating muscles. Our results show significantly lower capillary per myofiber ratio in M0 MP treated muscles, without significant differences between saline and M1 MP treated groups (Fig 9). It is worth noting, that uninjured skeletal muscle tissue has a capillary per myofiber ratio value around 2 (data not shown). Both saline and M1 MP treated muscles showed about 80% revascularization, consistent with our histological evaluation showing an ongoing regeneration process in all three groups. Moreover, we utilized anti-CD45 staining to determine the extent of the immune cell infiltrate into regenerating muscles at 14 day after TK-I/R injury. Our data show significantly higher immune cell infiltrate into M0 MP injected muscles suggesting that skeletal muscle regeneration after M0 MP treatment may be delayed (Fig 9).


The Development of Macrophage-Mediated Cell Therapy to Improve Skeletal Muscle Function after Injury.

Rybalko V, Hsieh PL, Merscham-Banda M, Suggs LJ, Farrar RP - PLoS ONE (2015)

Assessment of tissue revascularization and immune cell infiltrate in saline or MP-treated TK-I/R injured muscles 14 days post-reperfusion.Representative images of D14 regenerating skeletal muscle stained with anti-CD31antibody identifying vascular endothelium and anti-CD45 staining for immune cells, scale bar 100μm. Quantification graphs of the number of CD31+ cell per myofiber (capillary/fiber ratio) and CD45+ cells per field of view (FOV). M0 group n = 3, M1 and saline groups n = 4–5, 3–5 images/muscle, values presented as mean ± SEM, (*) p<0.05 relative to M0 MP group
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4696731&req=5

pone.0145550.g009: Assessment of tissue revascularization and immune cell infiltrate in saline or MP-treated TK-I/R injured muscles 14 days post-reperfusion.Representative images of D14 regenerating skeletal muscle stained with anti-CD31antibody identifying vascular endothelium and anti-CD45 staining for immune cells, scale bar 100μm. Quantification graphs of the number of CD31+ cell per myofiber (capillary/fiber ratio) and CD45+ cells per field of view (FOV). M0 group n = 3, M1 and saline groups n = 4–5, 3–5 images/muscle, values presented as mean ± SEM, (*) p<0.05 relative to M0 MP group
Mentions: Microvascular cell dysfunction accounts for most of the inflammatory response associated with I/R injury and is a rate-limiting step in I/R pathogenesis [49]. Restoration of adequate microvascular supply to regenerating muscle is crucial for functional muscle recovery and performance. We, therefore, evaluated skeletal muscle revascularization in our TK-I/R injured skeletal muscles at 14 days after acute saline or MP treatment. By using CD31 specific antibodies we were able to identify endothelial cells and quantify capillary per myofiber ratio in regenerating muscles. Our results show significantly lower capillary per myofiber ratio in M0 MP treated muscles, without significant differences between saline and M1 MP treated groups (Fig 9). It is worth noting, that uninjured skeletal muscle tissue has a capillary per myofiber ratio value around 2 (data not shown). Both saline and M1 MP treated muscles showed about 80% revascularization, consistent with our histological evaluation showing an ongoing regeneration process in all three groups. Moreover, we utilized anti-CD45 staining to determine the extent of the immune cell infiltrate into regenerating muscles at 14 day after TK-I/R injury. Our data show significantly higher immune cell infiltrate into M0 MP injected muscles suggesting that skeletal muscle regeneration after M0 MP treatment may be delayed (Fig 9).

Bottom Line: A variety of MP phenotypes have been identified and characterized in vitro as well as in vivo.We detected a 15% improvement in specific tension and force normalized to mass after M1 (LPS/IFN-γ) MP transplantation 24 hours post-reperfusion.Interestingly, we found that M0 bone marrow-derived unpolarized MPs significantly impaired muscle function highlighting the complexity of temporally coordinated skeletal muscle regenerative program.

View Article: PubMed Central - PubMed

Affiliation: Department of Kinesiology, The University of Texas at Austin, 1 University Station D3700, Austin, TX 78712, United States of America.

ABSTRACT
Skeletal muscle regeneration following acute injury is a multi-step process involving complex changes in tissue microenvironment. Macrophages (MPs) are one of the key cell types involved in orchestration and modulation of the repair process. Multiple studies highlight the essential role of MPs in the control of the myogenic program and inflammatory response during skeletal muscle regeneration. A variety of MP phenotypes have been identified and characterized in vitro as well as in vivo. As such, MPs hold great promise for cell-based therapies in the field of regenerative medicine. In this study we used bone-marrow derived in vitro LPS/IFN-y-induced M1 MPs to enhance functional muscle recovery after tourniquet-induced ischemia/reperfusion injury (TK-I/R). We detected a 15% improvement in specific tension and force normalized to mass after M1 (LPS/IFN-γ) MP transplantation 24 hours post-reperfusion. Interestingly, we found that M0 bone marrow-derived unpolarized MPs significantly impaired muscle function highlighting the complexity of temporally coordinated skeletal muscle regenerative program. Furthermore, we show that delivery of M1 (LPS/IFN-γ) MPs early in regeneration accelerates myofiber repair, decreases fibrotic tissue deposition and increases whole muscle IGF-I expression.

Show MeSH
Related in: MedlinePlus