Table_1_Overexpression of Mechano-Growth Factor Modulates Inflammatory Cytokine Expression and Macrophage Resolution in Skeletal Muscle Injury.PDF

In muscle regeneration, infiltrating myeloid cells, such as macrophages mediate muscle inflammation by releasing key soluble factors. One such factor, insulin-like growth factor 1 (IGF-1), suppresses inflammatory cytokine expression and mediates macrophage polarization to anti-inflammatory phenotype during muscle injury. Previously the IGF-1Ea isoform was shown to be anti-inflammatory. Another isoform of IGF-1, mechano-growth factor (MGF), is structurally and functionally distinct from IGF-1Ea, but its role in muscle inflammation has not yet been characterized. In this study, we hypothesized that MGF expression in muscle injury modulates muscle inflammation. We first investigated changes of transcription and expression of MGF in response to skeletal muscle injury induced by cardiotoxin (CTX) in vivo. At 1–2 days post-injury, Mgf expression was significantly upregulated and positively correlated with that of inflammatory cytokines. Immunostaining revealed that infiltration of neutrophils and macrophages coincided with Mgf upregulation. Furthermore, infiltrating neutrophils and macrophages expressed Mgf, suggesting their contribution to MGF upregulation in muscle injury. Macrophages seem to be the predominant source of MGF in muscle injury, whereas neutrophil depletion did not affect muscle Mgf expression. Given the association of MGF and macrophages, we then studied whether MGF could affect macrophage infiltration and polarization. To test this, we overexpressed MGF in CTX-injured muscles and evaluated inflammatory marker expression, macrophage populations, and muscle regeneration outcomes. MGF overexpression delayed the resolution of macrophages, particularly the pro-inflammatory phenotype. This coincided with upregulation of inflammatory markers. Annexin V-based flow cytometry revealed that MGF overexpression likely delays macrophage resolution by limiting macrophage apoptosis. Although MGF overexpression did not obviously affect muscle regeneration outcomes, the findings are novel and provide insights on the physiological roles of MGF in muscle regeneration.

在肌肉再生过程中，浸润的髓系细胞（myeloid cells）——如巨噬细胞（macrophages）——通过释放关键可溶性因子介导肌肉炎症反应。其中一类关键因子为胰岛素样生长因子1（insulin-like growth factor 1，IGF-1），其可抑制炎症细胞因子的表达，并在肌肉损伤过程中介导巨噬细胞极化为抗炎表型。既往研究表明，IGF-1Ea亚型具有抗炎活性。胰岛素样生长因子1的另一亚型——机械生长因子（mechano-growth factor，MGF）——在结构与功能上均与IGF-1Ea存在差异，但目前尚未明确其在肌肉炎症中的作用。本研究假设，肌肉损伤过程中MGF的表达可调控肌肉炎症反应。我们首先在体内探究了心脏毒素（cardiotoxin，CTX）诱导的骨骼肌损伤后，MGF的转录与表达水平变化情况。损伤后1~2天，MGF的表达显著上调，并与炎症细胞因子的表达呈正相关。免疫染色结果显示，中性粒细胞（neutrophils）与巨噬细胞的浸润与MGF的上调同步出现。此外，浸润的中性粒细胞与巨噬细胞可表达MGF，提示二者参与了肌肉损伤过程中MGF的上调过程。在肌肉损伤中，巨噬细胞似乎是MGF的主要来源，而清除中性粒细胞并不会影响肌肉组织中MGF的表达。鉴于MGF与巨噬细胞的关联，我们进一步探究了MGF是否可影响巨噬细胞的浸润与极化过程。为验证这一假设，我们在CTX损伤的肌肉组织中过表达MGF，并对炎症标志物表达、巨噬细胞群以及肌肉再生结局进行了评估。MGF过表达延缓了巨噬细胞的清除过程，尤其是促炎表型巨噬细胞的清除。该现象与炎症标志物的上调同时发生。基于膜联蛋白V（Annexin V）的流式细胞术结果显示，MGF过表达可能通过抑制巨噬细胞凋亡来延缓其清除过程。尽管MGF过表达并未显著影响肌肉再生结局，但本研究发现具有创新性，为阐明MGF在肌肉再生中的生理功能提供了新的见解。