Interactions between myoblasts and macrophages under high glucose milieus result in inflammatory response and impaired insulin sensitivity

Abstract

BACKGROUND Skeletal muscle handles about 80% of insulin-stimulated glucose uptake and become the major organ occurring insulin resistance (IR). Many studies have confirmed the interactions between macrophages and skeletal muscle regulated the inflammation and regeneration of skeletal muscle. However, despite of the decades of research, whether macrophages infiltration and polarization in skeletal muscle under high glucose (HG) milieus results in the development of IR is yet to be elucidated. C2C12 myoblasts are well-established and excellent model to study myogenic regulation and its responses to stimulation. Further exploration of macrophages' role in myoblasts IR and the dynamics of their infiltration and polarization is warranted. AIM To evaluate interactions between myoblasts and macrophages under HG, and its effects on inflammation and IR in skeletal muscle. METHODS We detected the polarization status of macrophages infiltrated to skeletal muscles of IR mice by hematoxylin and eosin and immunohistochemical staining. Then, we developed an in vitro co-culture system to study the interactions between myoblasts and macrophages under HG milieus. The effects of myoblasts on macrophages were explored through morphological observation, CCK-8 assay, Flow Cytometry, and enzyme-linked immunosorbent assay. The mediation of macrophages to myogenesis and insulin sensitivity were detected by morphological observation, CCK-8 assay, Immunofluorescence, and 2-NBDG assay. RESULTS The F4/80 and co-localization of F4/80 and CD86 increased, and the myofiber size decreased in IR group (P < 0.01, g = 6.26). Compared to Mc group, F4/80+CD86+CD206- cells, tumor necrosis factor-α (TNFα), inerleukin-1β (IL-1β) and IL-6 decreased, and IL-10 increased in McM group (P < 0.01, g > 0.8). In McM + HG group, F4/80+CD86+CD206- cells, monocyte chemoattractant protein 1, TNFα, IL-1β and IL-6 were increased, and F4/80+CD206+CD86- cells and IL-10 were decreased compared with Mc + HG group and McM group (P < 0.01, g > 0.8). Compered to M group, myotube area, myotube number and E-MHC were increased in MMc group (P < 0.01, g > 0.8). In MMc + HG group, myotube area, myotube number, E-MHC, GLUT4 and glucose uptake were decreased compared with M + HG group and MMc group (P < 0.01, g > 0.8). CONCLUSION Interactions between myoblasts and macrophages under HG milieus results in inflammation and IR, which support that the macrophage may serve as a promising therapeutic target for skeletal muscle atrophy and IR.