Abstract: Objective To explore the role of galectin-3(Gal-3)in regulating autophagy and polarization of macrophages through the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR)pathway, thus providing a new theoretical basis for targeted therapy of inflammatory diseases. Methods RAW264.7 cells were divided into four groups: control group(untreated), model group(treated with 100 nmol/L rapamycin for 6 hours to induce autophagy), Gal-3 overexpression group(Gal-3 OE, transfected with pcDNA3.1-Gal-3 plasmid), and Gal-3 knockdown group(Gal-3 KD, infected with shRNA-Gal-3 lentivirus). Immunofluorescence was used to detect the level of microtubule-associated protein 1 light chain 3B(LC3B), Western blotting to assess autophagy markers p62 and related proteins in the PI3K/Akt/mTOR pathway, flow cytometry to evaluate macrophage polarization(CD86/CD206)and apoptosis, enzyme-linked immunosorbent assay to measure inflammatory factors(tumor necrosis factor-α, interleukin-6, interleukin-10, transforming growth factor-β), RT-qPCR to detect the expression of autophagy-related genes arginase 1 and inducible nitric oxide synthase, and fluorescent staining to determine reactive oxygen species levels. Results Compared with the model group, the Gal-3 OE group showed downregulated levels of p-PI3K, p-Akt, and p-mTOR(P<0.01), increased LC3B(P<0.05), with statistical significance. The proportion of CD206-positive cells and the expressions of IL-10, TGF-β, and Arg-1 mRNA increased(P<0.01), while iNOS mRNA expression decreased(P<0.01), with statistical significance. Simultaneously, ROS levels and apoptosis rate increased(P<0.05). The Gal-3 KD group exhibited opposite effects. Conclusion Gal-3 participates in macrophage autophagy and M2-type polarization by inhibiting the PI3K/Akt/mTOR pathway, accompanied by ROS-dependent apoptosis regulation.

Key words: Galectin-3, Phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin pathway, Autophagy, Reactive oxygen species, Apoptosis