半乳糖凝集素-3通过PI3K/Akt/mTOR通路促进巨噬细胞自噬分化

doi:10.6040/j.issn.1671-7554.0.2025.0658

摘要/Abstract

摘要： 目的探讨半乳糖凝集素-3(galectin-3, Gal-3)通过磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶点(phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin, PI3K/Akt/mTOR)通路调控巨噬细胞自噬及极化的作用,为炎性疾病的靶向治疗提供新的理论依据。方法采用RAW264.7细胞,实验分为4组:对照组(不作处理)、模型组(100 nmol/L雷帕霉素诱导自噬6 h)、Gal-3过表达组(Gal-3 OE组,转染pcDNA3.1-Gal-3质粒)及敲低组(Gal-3 KD组,感染shRNA-Gal-3 慢病毒)。采用免疫荧光法检测微管相关蛋白1轻链3B(microtubule-associated protein 1 light chain 3B, LC3B)水平、Western blotting法检测自噬标志物p62及PI3K/Akt/mTOR通路相关蛋白;流式细胞术评估巨噬细胞极化(CD86/CD206)及凋亡情况;酶联免疫吸附试验检测炎症因子［(肿瘤坏死因子α(tumor necrosis factor-α, TNF-α)、白细胞介素6(interleukin-6,IL-6)、白细胞介素10(interleukin-10,IL-10)、转化生长因子β(transforming growth factor-β, TGF-β)］水平;RT-qPCR法检测自噬相关基因精氨酸酶1(arginase 1,Arg-1)和诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)表达;荧光染色法测定活性氧(reactive oxygen species, ROS)水平。结果与模型组相比,Gal-3 OE组p-PI3K、p-Akt、p-mTOR水平下调(P<0.01),LC3B升高(P<0.05),差异有统计学意义;CD206阳性细胞比例、IL-10、TGF-β及Arg-1 mRNA表达升高(P<0.01),iNOSmRNA表达降低(P<0.01),差异有统计学意义;同时ROS水平与细胞凋亡率升高(P<0.05)。Gal-3 KD组则呈现相反效应。结论 Gal-3通过抑制PI3K/Akt/mTOR通路参与巨噬细胞自噬及M2型极化,伴随ROS依赖性凋亡调控。

关键词: 半乳糖凝集素-3, 磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶点, 自噬, 活性氧, 凋亡

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

中图分类号:

R392.5

陈杨,冯莹,卢晓,郑蓉. 半乳糖凝集素-3通过PI3K/Akt/mTOR通路促进巨噬细胞自噬分化[J]. 山东大学学报 (医学版), 2026, 64(4): 14-22.

CHEN Yang, FENG Ying, LU Xiao, ZHENG Rong. Galectin-3 promotes autophagy and polarization of macrophages via the PI3K/Akt/mTOR pathway[J]. Journal of Shandong University (Health Sciences), 2026, 64(4): 14-22.

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