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Heterogeneous macrophage activation in injured skeletal muscle

ABSTRACT: Macrophages are the predominant inflammatory cells during skeletal muscle injury repair. They are required for the complete repair of acutely injured skeletal muscle, but detrimentally contribute to the pathological progression of dystrophic muscle in Duchenne muscular dystrophy (DMD) patients and DMD mouse model mdx5cv mice. The cellular and molecular mechanism underlying this difference has yet to be fully elucidated. To this end, the present study compared the single-cell transcriptomes of intramuscular monocytes/macrophages from uninjured and acutely injured quadriceps of 14 weeks-old wild-typed (WT) mice, and from quadriceps and diaphragm of age-matched mdx5cv mice, using single cell-based RNA sequencing (scRNA-seq) analysis. Our results identified multiple functionally diverse monocyte/macrophage clusters co-existing in each injured muscle sample. Both Ly6Chi and Ly6Clo intramuscular monocytes/macrophages were heterogenous, containing different clusters among different injury conditions. These clusters did not conform to strict M1/M2 activation, and were involved differentially in inflammation, extracellular matrix (ECM) remodeling, and myogenesis during skeletal muscle injury repair. These findings uncovered the heterogeneity of monocyte/macrophage activation in both acutely injured and dystrophic skeletal muscle, shedding light on the molecular and cellular mechanisms underlying the differential roles of monocytes/macrophages in acute skeletal muscle injury and muscular dystrophy in DMD.

ORGANISM(S): Mus musculus

PROVIDER: GSE270031 | GEO | 2025/06/14

REPOSITORIES: GEO

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Project description:Background: Niemann-Pick disease type A (NPDA), a disease caused by mutations in acid sphingomyelinase (ASM), involves severe neurodegeneration and early death. Intracellular lipid accumulation and plasma membrane alterations are implicated in the pathology. ASM is also linked to the mechanism of plasma membrane repair, so we investigated the impact of ASM deficiency in skeletal muscle, a tissue that undergoes frequent cycles of injury and repair in vivo. Methods: Utilizing the NPDA/B mouse model ASM−/− and wild type (WT) littermates, we performed excitation- contraction coupling/Ca2+ mobilization and sarcolemma injury/repair assays with isolated flexor digitorum brevis fibers, proteomic analyses with quadriceps femoris, flexor digitorum brevis, and tibialis posterior muscle and in vivo tests of the contractile force (maximal isometric torque) of the quadriceps femoris muscle before and after eccentric contraction-induced muscle injury. Results: ASM−/− flexor digitorum brevis fibers showed impaired excitation-contraction coupling compared to WT, a defect expressed as reduced tetanic [Ca2+]i in response to electrical stimulation and early failure in sustaining [Ca2+]i during repeated tetanic contractions. When injured mechanically by needle passage, ASM−/− flexor digitorum brevis fibers showed susceptibility to injury similar to WT, but a reduced ability to reseal the sarcolemma. Proteomic analyses revealed changes in a small group of skeletal muscle proteins as a consequence of ASM deficiency, with downregulation of calsequestrin occurring in the three different muscles analyzed. In vivo, the loss in maximal isometric torque of WT quadriceps femoris was similar immediately after and 2 min after injury. The loss in ASM−/− mice immediately after injury was similar to WT, but was markedly larger at 2 min after injury. Conclusions: Skeletal muscle fibers from ASM−/− mice have an impairment in intracellular Ca2+ handling that results in reduced Ca2+ mobilization and a more rapid decline in peak Ca2+ transients during repeated contraction-relaxation cycles. Isolated fibers show reduced ability to repair damage to the sarcolemma, and this is associated with an exaggerated deficit in force during recovery from an in vivo eccentric contraction- induced muscle injury. Our findings uncover the possibility that skeletal muscle functional defects may play a role in the pathology of NPDA/B disease.

Project description:Massage therapy is commonly used for the physical rehabilitation of skeletal muscle to ameliorate pain and promote recovery from injury. While there is some evidence that massage may relieve pain in injured muscle, the cellular effects remain unknown. To assess the effects of massage, we administered either massage therapy or no treatment to separate quadriceps of eleven young, male participants after exercised-induced muscle damage. Muscle biopsies were acquired from the quadriceps (vastus lateralis) at baseline (rest), immediately after 10 minutes of massage treatment (0h), and after a 2.5 hour period of recovery. We found that massage activated the mechanotransduction signaling pathways FAK and Erk1/2, potentiated mitochondrial biogenesis signaling (nuclear PGC-1α), and mitigated the rise in NFkB (p65) nuclear accumulation caused by muscle trauma. Moreover, in spite of having no effect on muscle metabolites (glycogen, lactate), massage attenuated the production of the inflammatory cytokines TNF-α and IL-6 and reduced HSP27 phosphorylation, effectively mitigating cellular stress resulting from myofiber injury. In summary, massage therapy appears to be clinically beneficial when administered to acutely damaged skeletal muscle by reducing inflammation and promoting mitochondrial biogenesis. 55 muscle biopsies were used in this study as material to evaluate the effects of massage on gene expression. These biopsies were taken from the vastus lateralis of eleven healthy recreationally active males with full ethical approval at McMaster's university. Each individual had an initial biopsy at rest (baseline), followed by exercist testing for peak aerobic capacity. On a second visit, the individual performed a bout of exhaustive aerobic exercise, prior to recieving a randomized massage treatment to one leg. Immediatley following the exercise, subjects were allowed to recover for 10 minutes, while massage oil was lightly applied to both quadriceps. Then a single leg was randomized to recieve massage treatment for 10 minutes. Subjects then rested for 10 minutes, and one additional biopsies were carried out (on each leg). Two and half hours later (3 hours after the cessation of the exercise bout), 2 more biopsies were taken (one from each leg). This amounted to 5 biopsies from each individual.

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Heterogeneous macrophage activation in injured skeletal muscle

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