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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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In vivo quantification and characterization of MP populations 5 days after TK-I/R injury.A) Flow cytometric identification and quantification of myeloid and MP cell populations in TK-injured muscle. Total myeloid cells (CD45+CD11b+), resident MPs (CD11b+F4/80+PKH2.6-), transplanted MPs (CD11b+F4/80+PKH2.6+).Values expressed as mean ±SEM, n = 3, B) Expression of CD206 and Ly-6C surface proteins on F4/80+MP populations in control and MP-treated muscles. Values expressed as mean ±SEM, n = 3.
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pone.0145550.g007: In vivo quantification and characterization of MP populations 5 days after TK-I/R injury.A) Flow cytometric identification and quantification of myeloid and MP cell populations in TK-injured muscle. Total myeloid cells (CD45+CD11b+), resident MPs (CD11b+F4/80+PKH2.6-), transplanted MPs (CD11b+F4/80+PKH2.6+).Values expressed as mean ±SEM, n = 3, B) Expression of CD206 and Ly-6C surface proteins on F4/80+MP populations in control and MP-treated muscles. Values expressed as mean ±SEM, n = 3.

Mentions: MPs were shown to undergo phenotypic switching at the site of TK-I/R muscle injury [31] from an M1-like pro-inflammatory to an M2-like anti-inflammatory and pro-regenerative population[32]. Ly-6C and CD206 are two primary markers used to discriminate between pro-inflammatory (M1) MPs and anti-inflammatory (M2) MPs. We wanted to assess whether transplanted MPs were able to undergo phenotypic switching after i.m. delivery as well as to determine whether we can see any differences in the expression of Ly-6C and CD206 proteins on the surface of MP populations, both transplanted and resident, at 5 days after reperfusion injury (Fig 7). We did not see any differences in the total number of CD45+CD11b+ myeloid cells or CD11b+F4/80+ MPs among treatment groups on day 5 after TK-I/R injury (Fig 7A). Likewise, there were no differences in CD206 expression on resident F4/80+PKH- MPs, with CD206 marker elevated on all three groups consistent with an ongoing regeneration process (Fig 7B). Both groups of transplanted MPs (M0 and M1 (LPS/IFN-γ) PKH+ MPs) transitioned to higher expression level of CD206 marker following in vivo delivery (Fig 7B). No Ly-6C expression was detected on MP populations, which is consistent with existing literature on MP phenotypes in muscle regeneration [31, 32].Although we anticipated to see higher levels of CD206 expression in the M1 (LPS/IFN-γ) treatment group on both PKH+ and PKH- populations of MPs, no such differences were apparent at 5 days post-reperfusion injury, suggesting that if differences in MP transition exist among treatment groups, they occur between 1 and 5 days post-reperfusion.


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)

In vivo quantification and characterization of MP populations 5 days after TK-I/R injury.A) Flow cytometric identification and quantification of myeloid and MP cell populations in TK-injured muscle. Total myeloid cells (CD45+CD11b+), resident MPs (CD11b+F4/80+PKH2.6-), transplanted MPs (CD11b+F4/80+PKH2.6+).Values expressed as mean ±SEM, n = 3, B) Expression of CD206 and Ly-6C surface proteins on F4/80+MP populations in control and MP-treated muscles. Values expressed as mean ±SEM, n = 3.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0145550.g007: In vivo quantification and characterization of MP populations 5 days after TK-I/R injury.A) Flow cytometric identification and quantification of myeloid and MP cell populations in TK-injured muscle. Total myeloid cells (CD45+CD11b+), resident MPs (CD11b+F4/80+PKH2.6-), transplanted MPs (CD11b+F4/80+PKH2.6+).Values expressed as mean ±SEM, n = 3, B) Expression of CD206 and Ly-6C surface proteins on F4/80+MP populations in control and MP-treated muscles. Values expressed as mean ±SEM, n = 3.
Mentions: MPs were shown to undergo phenotypic switching at the site of TK-I/R muscle injury [31] from an M1-like pro-inflammatory to an M2-like anti-inflammatory and pro-regenerative population[32]. Ly-6C and CD206 are two primary markers used to discriminate between pro-inflammatory (M1) MPs and anti-inflammatory (M2) MPs. We wanted to assess whether transplanted MPs were able to undergo phenotypic switching after i.m. delivery as well as to determine whether we can see any differences in the expression of Ly-6C and CD206 proteins on the surface of MP populations, both transplanted and resident, at 5 days after reperfusion injury (Fig 7). We did not see any differences in the total number of CD45+CD11b+ myeloid cells or CD11b+F4/80+ MPs among treatment groups on day 5 after TK-I/R injury (Fig 7A). Likewise, there were no differences in CD206 expression on resident F4/80+PKH- MPs, with CD206 marker elevated on all three groups consistent with an ongoing regeneration process (Fig 7B). Both groups of transplanted MPs (M0 and M1 (LPS/IFN-γ) PKH+ MPs) transitioned to higher expression level of CD206 marker following in vivo delivery (Fig 7B). No Ly-6C expression was detected on MP populations, which is consistent with existing literature on MP phenotypes in muscle regeneration [31, 32].Although we anticipated to see higher levels of CD206 expression in the M1 (LPS/IFN-γ) treatment group on both PKH+ and PKH- populations of MPs, no such differences were apparent at 5 days post-reperfusion injury, suggesting that if differences in MP transition exist among treatment groups, they occur between 1 and 5 days post-reperfusion.

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