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Updated: Feb 16, 2026
Non-thermal Infrared Light Treatment of Ischemia/Reperfusion Injury and Subsequent Analysis of Macrophage Differentiation
Published on: December 30, 2021
Recruitment and therapeutic application of macrophages in skeletal muscles after hind limb ischemia
Pei-Ling Hsieh1, Viktoriya Rybalko2, Aaron B Baker2
1Department of Kinesiology, The University of Texas at Austin, Austin, Tex.
Objective:
Peripheral arterial disease can cause not only ischemia but also skeletal muscle damage. It has been known that macrophages (MPs) play an important role in coordinating muscle repair; however, phenotype transition of monocyte-MP in ischemic muscle has not been well defined. Hence, the purpose of this study was to examine the temporal recruitment of MPs and to explore their therapeutic effect on ischemic muscle regeneration.
Methods:
Unilateral femoral artery excision was performed on C57BL/6 mice. Myeloid cells were isolated from the ischemic muscles, characterized using flow cytometry. Bone marrow-derived MPs were injected (2 × 106 cells) into the ischemic gastrocnemius muscle 24 hours after injury. Blood flow recovery was measured using laser speckle imaging. Functional outcome was evaluated by assessing the contractile force of ischemic muscles. Histologic analysis included quantification of myofiber size, collagen deposition, number of inflammatory and MyoD-expressing cells, and capillary density.
Results:
Neutrophils and inflammatory monocytes-MPs were present at day 1 after injury. The mature MPs then remained elevated as the dominant population from day 5 to day 21 with the observation of regenerating fibers. Functional measurements revealed that the force production was significantly enhanced after treatment with proinflammatory M1 MPs (94.9% vs 77.9%; P < .05), and this was consistent with increased myofiber size, capillary- fiber ratio, and perfusion (78.6% vs 39.9%; P < .05). Moreover, the percentage of MyoD-expressing nuclei was significantly higher at day 4, indicating that M1 MPs may hasten muscle repair. Whereas early delivery of anti-inflammatory M2 MPs improved myofiber size, this was accompanied by persistent fibrosis suggesting ongoing tissue remodeling, and lower force production was observed.
Conclusions:
We demonstrated the dynamics of myeloid cells in skeletal muscle after ischemic insult, and the administration of exogenous M1 MPs in a temporally coordinated manner successfully improved angiogenesis and skeletal muscle regeneration. Our results suggested that cell therapy using MPs may be a promising adjunctive therapeutic approach for peripheral arterial disease.
Insights
Macrophages (MPs) are crucial for muscle repair after peripheral arterial disease. This study shows M1 MPs enhance skeletal muscle regeneration and blood flow recovery, suggesting cell therapy potential for peripheral arterial disease.
Area of Science:
- Muscle regeneration and repair
- Immunology and inflammation
- Vascular biology and disease
Background:
- Peripheral arterial disease (PAD) causes ischemia and skeletal muscle damage.
- Macrophages (MPs) are vital for muscle repair, but their role in ischemic muscle regeneration is not fully understood.
- Understanding monocyte-to-macrophage phenotype transition is key to exploring therapeutic strategies.
Purpose of the Study:
- To investigate the temporal recruitment dynamics of macrophages in ischemic skeletal muscle.
- To explore the therapeutic potential of exogenous macrophage administration on ischemic muscle regeneration.
- To define the specific roles of different macrophage phenotypes in muscle repair.
Main Methods:
- Created an ischemic muscle model in C57BL/6 mice via femoral artery excision.
- Isolated and characterized myeloid cells using flow cytometry.
- Administered bone marrow-derived macrophages (M1 and M2 phenotypes) to ischemic muscle and assessed functional and histological outcomes, including blood flow, muscle force, myofiber size, and capillary density.
Main Results:
- Neutrophils and inflammatory monocytes-macrophages (MPs) were recruited early (day 1), with mature MPs dominating from day 5 to 21.
- Proinflammatory M1 MPs significantly enhanced muscle force production, myofiber size, capillary density, and perfusion.
- Anti-inflammatory M2 MPs improved myofiber size but led to fibrosis and reduced force production.
Conclusions:
- Demonstrated the dynamic recruitment of myeloid cells in ischemic skeletal muscle.
- Exogenous M1 MP administration, when timed correctly, effectively improved angiogenesis and skeletal muscle regeneration.
- Macrophage-based cell therapy presents a promising adjunctive treatment strategy for peripheral arterial disease.
