GDF11减轻巨噬细胞极化参与流感病毒感染免疫调控的研究-东南大学学报医学版

GDF11减轻巨噬细胞极化参与流感病毒感染免疫调控的研究

作者：彭璐冯锦

单位：华中科技大学同济医学院附属武汉金银潭医院儿科传染性疾病科, 湖北武汉 430000

分类号：R511.7;R503

出版年·卷·期（页码）：2025·44·第三期（404-411）

摘要：

目的: 探讨生长分化因子11(GDF11)通过调节mTOR途径减轻巨噬细胞极化参与流感病毒感染免疫调控的影响。方法: 将细胞分为阴性对照组(NC组)、模型组(Model组)、GDF11^+/+组、GDF11^-/-组和GDF11^-/-+MHY1485组。采用流式细胞术检测各组RAW264.7细胞的iNOS、CD86表达水平;采用ELISA试剂盒检测各组RAW264.7细胞培养基中炎症因子及免疫球蛋白水平;采用实时定量聚合酶链反应(RT-qPCR)、蛋白质印迹法及细胞免疫荧光法检测各组RAW264.7细胞中GDF11、mTOR信号通路蛋白、炎症因子及免疫球蛋白表达水平。结果: 与NC组相比,流感病毒RP8感染细胞后,可促进RAW264.7细胞M1极化(P<0.05),下调GDF11分子水平(P<0.05),上调TNF-α、IL-6和IL-1β炎症因子水平,同时下调IgA、IgG和IgM免疫球蛋白水平(P<0.05),促进mTOR、AMPK、AKT蛋白磷酸化(P<0.05);与Model组相比,过表达GDF11蛋白可抑制RAW264.7细胞M1极化(P<0.05),下调炎症因子TNF-α、IL-6和IL-1β水平,上调免疫球蛋白IgA、IgG和IgM水平(P<0.05),抑制mTOR、AMPK、AKT蛋白磷酸化(P<0.05);而敲除GDF11蛋白则显著促进RAW264.7细胞M1极化(P<0.05),上调炎症因子水平,下调免疫球蛋白水平(P<0.05),并促进mTOR、AMPK和AKT蛋白磷酸化(P<0.05);mTOR蛋白抑制剂MHY1485可抑制RAW264.7细胞M1极化(P<0.05),下调炎症因子TNF-α、IL-6和IL-1β水平,上调免疫球蛋白IgA、IgG和IgM水平(P<0.05),抑制mTOR、AMPK、AKT蛋白磷酸化(P<0.05)。结论: GDF11可通过抑制mTOR信号通路,从而减轻流感病毒感染引起的RAW264.7细胞M1极化水平,进而参与流感病毒感染免疫调控反应。

Objective: To investigate the effects of growth differentiation factor 11(GDF11) on the modulation of macrophage polarization through the mTOR pathway in the context of immune regulation during influenza virus infection. Methods: Cells were divided into five groups: negative control(NC), Model, GDF11^+/+, GDF11^-/-, and GDF11^-/-+MHY1485 group. Flow cytometry was employed to assess the expression levels of iNOS and CD86 in RAW264.7 cells across the groups. ELISA kits were utilized to measure levels of inflammatory cytokines and immunoglobulins in the culture supernatants of RAW264.7 cells. The expression levels of GDF11, mTOR pathway proteins, inflammatory cytokines, and immunoglobulins were analyzed using real-time quantitative polymerase chain reaction(RT-qPCR) and Western blot, and immunofluorescence assays. Results: Compared with the NC group, influenza virus RP8 infection promoted M1 polarization of RAW264.7 cells(P<0.05), downregulated GDF11 levels(P<0.05), upregulated inflammatory cytokines(TNF-α, IL-6, and IL-1β) levels(P<0.05), and downregulated immunoglobulins(IgA, IgG, and IgM)levels(P<0.05), while enhancing phosphorylation of mTOR, AMPK, and AKT proteins(P<0.05). In comparison with the Model group, overexpression of GDF11 significantly inhibited M1 polarization of RAW264.7 cells(P<0.05), decreased levels of inflammatory cytokines(TNF-α, IL-6, and IL-1β)(P<0.05), increased levels of immunoglobulins(IgA, IgG, and IgM)(P<0.05), and suppressed phosphorylation of mTOR, AMPK, and AKT proteins(P<0.05). Knockout of GDF11 protein significantly promoted M1 polarization in RAW264.7 cells(P<0.05), upregulated the levels of inflammatory factors, downregulated the levels of immunoglobulins(P<0.05), and enhanced the phosphorylation of mTOR, AMPK, and AKT proteins(P<0.05). The mTOR inhibitor MHY1485 effectively suppressed M1 polarization in RAW264.7 cells(P<0.05), reduced levels of inflammatory cytokines(TNF-α, IL-6, and IL-1β) (P<0.05), increased levels of immunoglobulins(IgA, IgG, and IgM)(P<0.05), and inhibited phosphorylation of mTOR, AMPK, and AKT proteins(P<0.05). Conclusion: GDF11 can mitigate M1 polarization of RAW264.7 cells induced by influenza virus infection through the inhibition of the mTOR signaling pathway, thereby contributing to immune regulatory responses during influenza virus infection.

参考文献：

[1] DIN G U, WU C, TARIQ Z, et al.Unlocking influenza B:exploring molecular biology and reverse genetics for epidemic control and vaccine innovation[J].Virol J, 2024, 21(1):196.
[2] IULIANO A D, ROGUSKI K M, CHANG H H, et al.Estimates of global seasonal influenza-associated respiratory mortality:a modelling study[J].Lancet, 2018, 391(10127):1285-1300.
[3] CLARK T W, TREGONING J S, LISTER H, et al.Recent advances in the influenza virus vaccine landscape:a comprehensive overview of technologies and trials[J].Clin Microbiol Rev, 2024, 37(4):e0002524.
[4] STUMPFF J P 2nd, KIM S Y, MCFADDEN M I, et al.Pleural macrophages translocate to the lung during infection to promote improved influenza outcomes[J].Proc Natl Acad Sci U S A, 2023, 120(51):e2300474120.
[5] TAKASHITA E.Influenza polymerase inhibitors:mechanisms of action and resistance[J].Cold Spring Harb Perspect Med, 2021, 11(5):a038687.
[6] LUDWIG S, HRINCIUS E R, BOERGELING Y.The two sides of the same coin-influenza virus and intracellular signal transduction[J].Cold Spring Harb Perspect Med, 2021, 11(1):a038513.
[7] ZHU B, GU G, REN J, et al.Schwann cell-derived exosomes and methylprednisolone composite patch for spinal cord injury repair[J].ACS Nano, 2023, 17(22):22928-22943.
[8] HE L, JHONG J H, CHEN Q, et al.Global characterization of macrophage polarization mechanisms and identification of M2-type polarization inhibitors[J].Cell Rep, 2021, 37(5):109955.
[9] WALKER R G, POGGIOLI T, KATSIMPARDI L, et al.Biochemistry and biology of GDF11 and myostatin:similarities, differences, and questions for future investigation[J].Circ Res, 2016, 118(7):1125-1142.
[10] 肖雯, 田源, 蒋宇, 等.GDF11调控NF-κB通路影响巨噬细胞M1极化介导的炎症反应[J].中国医科大学学报, 2024, 53(4):342-348, 354.
[11] DUAN F, WANG X, WANG H, et al.GDF11 ameliorates severe acute pancreatitis through modulating macrophage M1 and M2 polarization by targeting the TGFβR1/SMAD-2 pathway[J].Int Immunopharmacol, 2022, 108:108777.
[12] SU Y, SUN X, LIU X, et al.hUC-EVs-ATO reduce the severity of acute GVHD by resetting inflammatory macrophages toward the M2 phenotype[J].J Hematol Oncol, 2022, 15(1):99.
[13] 唐坤, 吴黎虹, 曾陈芳, 等.Sesn2通过激活mTOR信号通路调节LPS诱导的巨噬细胞M1极化和炎症反应[J].中国病理生理杂志, 2024, 40(4):689-696.
[14] LUO Y, YANG Z, ZHAO X, et al.Immune regulation enhances osteogenesis and angiogenesis using an injectable thiolated hyaluronic acid hydrogel with lithium-doped nano-hydroxyapatite (Li-nHA) delivery for osteonecrosis[J].Mater Today Bio, 2024, 25:100976.
[15] XIA C, XU W, AI X, et al.Autophagy and exosome coordinately enhance macrophage M1 polarization and recruitment in influenza a virus infection[J].Front Immunol, 2022, 13:722053.
[16] ZHANG Y, LI J, QIU Z, et al.Insights into the mechanism of action of pterostilbene against influenza A virus-induced acute lung injury[J].Phytomedicine, 2024, 129:155534.
[17] ATRI C, GUERFALI F Z, LAOUINI D.Role of human macrophage polarization in inflammation during infectious diseases[J].Int J Mol Sci, 2018, 19(6):1801.
[18] GUNASSEKARAN G R, POONGKAVITHAI VADEVOO S M, BAEK M C, et al.M1 macrophage exosomes engineered to foster M1 polarization and target the IL-4 receptor inhibit tumor growth by reprogramming tumor-associated macrophages into M1-like macrophages[J].Biomaterials, 2021, 278:121137.
[19] JIANG G, LI S, YU K, et al.A 3D-printed PRP-GelMA hydrogel promotes osteochondral regeneration through M2 macrophage polarization in a rabbit model[J].Acta Biomater, 2021, 128:150-162.
[20] LI M, CHU X, WANG D, et al.Tuning the surface potential to reprogram immune microenvironment for bone regeneration[J].Biomaterials, 2022, 282:121408.
[21] 常立功, 黄培林.mTOR信号通路与大肠癌演进相关性的研究进展[J].东南大学学报(医学版), 2013, 32(2):243-247.

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