Artesunate alleviated DSS induced colitis via inhibiting M1 polarization through directly targeting INSR/mTORC1/HIF-1α pathway.

Abstract

INTRODUCTION: Artesunate (ARS) has demonstrated therapeutic potential in experimental ulcerative colitis (UC) through immunomodulatory activities. However, the role of macrophages and the underlying mechanisms remain unclear. METHODS: In this study, a DSS-induced UC model was established, along with macrophage depletion, polarized macrophage reinfusion, and Transwell coculture systems. The typical colitis characteristics, including weight change, disease activity index, inflammation, and tissue damage, were systematically assessed. Key proteins in the mTORC1/HIF-1α pathway were analyzed. SPR-MS was employed to identify ARS targets in M1 macrophages. RESULTS: This study indicated that ARS markedly suppressed DSS-enhanced M1 polarization and macrophage infiltration. In a Caco2-BMDM coculture system, ARS reduced macrophage migration and inflammation. Macrophage depletion significantly attenuated the anti-colitis effect of ARS. Compared with M1 reinfusion, ARS-pretreated M1 cells alleviated UC symptoms. ARS also inhibited LPS- or DSS-induced M1 polarization and proinflammatory cytokine upregulation in vivo and in vitro, linked to mTORC1/HIF-1α signaling, as confirmed by the agonist Leucine (LEU). Moreover, ARS protected intestinal barrier function by preventing tight junction loss and permeability increases via suppression of M1 polarization. SPR-MS and molecular docking revealed that ARS directly activated INSR, inhibiting mTORC1/HIF-1α-driven glycolytic M1 polarization. CONCLUSIONS: Overall, macrophages are essential for ARS-mediated protection in UC. ARS alleviates UC by reprogramming macrophage polarization via the INSR/mTORC1/HIF-1α axis, providing new mechanistic insights and potential clinical applications.