Integrin αVβ5 regulates myoblast proliferation and differentiation in sarcopenia mice treated with FNDC5 gene delivery

Irisin which is encoded by FNDC5 gene, has emerged as a promising therapeutic candidate for alleviating sarcopenia; however, its long-term therapeutic application in muscle regeneration remains insufficiently developed. Our study investigated the potential of FNDC5 gene delivery in both cellular and animal levels via a virus vector. It was indicated that FNDC5 overexpression promoted myoblast proliferation while inhibiting myogenic differentiation, and vice versa. Mechanistically, we confirmed that the product of FNDC5 gene, irisin binds to integrin αVβ5 on the cell membrane of myoblasts, leading to activation of the downstream FAK/SRC complex using coimmunoprecipitation (co-IP) and confocal microscopy. RNA-Seq and qPCR analyses revealed that irisin exposure activated genes related to the extracellular matrix (ECM) and its ligands, thereby promoting the cell cycle and enhancing the proliferation of myoblast. Additionally, treatment with the integrin αVβ5 inhibitor cilengitide enhanced myogenic differentiation but suppressed cell proliferation, highlighting the crucial role of integrin αVβ5 in determining myoblasts’ cell fate. Furthermore, in sarcopenia mouse models, AAV-mediated FNDC5 supplementation activated integrin αVβ5-related signaling as expected, resulting in increased muscle mass and alleviation of sarcopenia symptoms. Our findings demonstrate that integrin αVβ5 acts as a key switch in regulating the proliferation and differentiation of myoblasts under FNDC5 gene delivery. When integrin αVβ5 is active, FNDC5 over-expression enhances cell proliferation rather than differentiation. Conversely, when integrin αVβ5 is inhibited, FNDC5 over-expression promotes myogenic differentiation over proliferation. Moreover, the binding of irisin to integrin αVβ5 induces the expression of ECM proteins in myoblasts and activates the Bgn-TLR4 signaling pathway, which further enhances cell proliferation. This FNDC5-ITGAVB5-FAK-mTOR-Bgn-TLR4 signaling axis was also validated in a sarcopenia mouse model. Integrin αVβ5 is a critical regulator of myoblast proliferation and differentiation in response to FNDC5 gene supplement. These findings provide new insights into FNDC5 biological functions and suggest potential therapeutic strategies for enhancing muscle regeneration and treating sarcopenia.