目的 探讨结核分枝杆菌感染对巨噬细胞极化的影响并分析极化后的巨噬细胞对A549肺癌细胞恶性生物学行为影响.方法 采用佛波酯(PMA)诱导人单核细胞THP-124 h为M0巨噬细胞,继续用IL-4、IL-13刺激M0巨噬细胞48 h向M2极化(M2),用M.tuberculosis H37Rv裂解总蛋白刺激M0巨噬细胞48 h(M0H).WB检测M0、M2、M0H巨噬细胞IL-10、CD163、TNF-α蛋白表达水平;CCK-8、伤口愈合实验、Transwell实验检测M0、M2、M0H巨噬细胞条件培养基(CM)对A549细胞增殖、迁移和侵袭能力的影响.结果 与对照M0组相比,M2极化标志物IL-10和CD163蛋白表达水平在M2组(IL-10:M0 vs M2=0.84±0.29 vs 1.76±0.34;CD163:M0 vs M2=0.84±0.16 vs 1.66±0.21)和M0H组(IL-10:M0 vs M0H=0.84±0.29 vs 1.47±0.29;CD163:M0 vs M0H=0.84±0.16 vs 1.65±0.36)中显著升高(P<0.01);与对照M0组相比,M1极化标志物TNF-α蛋白表达水平在M2组无明显变化(M0 vs M2=0.90±0.13 vs 1.04±0.07,P=0.12),在M0H组显著下降(M0 vs M0H=0.90±0.13 vs 0.65±0.10,P<0.05);与对照M0-CM组相比,M2-CM组和M0H-CM组能够增加A549细胞增殖[48 h:M0-CMvs M2-CM=(100.00±4.75)％vs(129.51±4.59)％,P<0.01;M0-CMvs M0H-CM=(100.00±4.75)％vs(152.04±8.11)％,P<0.01]、横向迁移[24 h:M0-CMvs M2-CM=(17.08±3.16)％vs(30.98±3.28)％,M0 vs M0H=(17.08±3.16)％vs(27.51±0.65)％;48 h:M0-CM vs M2-CM=(52.62±5.45)％vs(70.75±2.33)％;M0-CMvs M0H-CM=(52.62±5.45)％vs(70.91±4.29)％,P<0.01]、纵向迁移(24 h:M0-CM vs M2-CM=100.00±4.93 vs 160.00±18.73,M0 vs M0H=10.00±4.93 vs 195.00±6.24;48 h:M0-CM vs M2-CM=43.11±4.66 vs 103.56±10.62,M0 vs M0H=43.11±4.66 vs 136.56±23.49,P<0.01)和侵袭(24 h:M0-CMvs M2-CM=62.4±4.56 vs 127.2±12.39,M0 vs M0H=62.4±4.56 vs 131.40±10.88;48 h:M0-CMvs M2-CM=86.40±8.96 vs 162.40±15.95,M0 vs M0H=86.40±8.96 vs 177.00±13.11,P<0.01).结论 结核分枝杆菌感染通过诱导巨噬细胞M2极化促进A549细胞增殖、迁移和侵袭.
Mycobacterium tuberculosis promotes proliferation,migration and invasion of A549 lung cancer cells by inducing macrophage M2 polarization
Objective To investigate the effect of Mycobacterium tuberculosis infection on the polarization of mac-rophages and analyze the effect of polarized macrophages on the malignant behavior of A549 lung cancer cells.Methods Human monocytic THP-1 cells were induced into M0 macrophages by Phorbol 12-myristate 13-acetate (PMA)for 24 h,and M0 macrophage was incubated with IL-4 and IL-13 for 48 h to induce M2 macrophage po-larization(M2)or stimulated with M.tuberculosis H37Rv whole-cell lysate(M0H)for 48 h.The expression levels of IL-10,CD163 and TNF-α in M0,M2 and M0H macrophages were detected by WB.The effects of M0,M2 and M0H macrophage conditioned medium(CM)on proliferation,migration and invasion of A549 cells were de-tected by CCK-8,wound healing assay and transwell assay.Results Compared with the control M0 group,the M2 polarisation markers IL-10 and CD163 protein expression levels were significantly increased in the M2 group (IL-10:M0 vs M2=0.84±0.29 vs 1.76±0.34;CD163:M0 vs M2=0.84±0.16 vs 1.66±0.21) and the M0H group (IL-10:M0 vs M0H=0.84±0.29 vs 1.47±0.29;CD163:M0 vs M0H=0.84±0.16 vs 1.65±0.36)(P<0.01);compared to the control M0 group,the M1 polarisation marker TNF-α pro-tein expression level did not change in the M2 group (M0 vs M2=0.90±0.13 vs 1.04±0.07,P=0.12) and decreased significantly in the M0H group (M0 vs M0H=0.90±0.13 vs 0.65±0.10,P<0.05 );com-pared with the control M0-CMgroup,the M2-CMgroup and the M0H-CMgroup were able to increase the prolif-eration of A549 cells[48 h:M0-CMvs M2-CM=(100.00±4.75)％ vs (129.51±4.59)％,P<0.01;M0-CMvs M0H-CM=(100.00±4.75)％vs (152.04±8.11)％,P<0.01],lateral migration[24 h:M0-CMvs M2-CM=(17.08±3.16)％ vs (30.98±3.28)％,M0 vs M0H=(17.08±3.16)％ vs (27.51±0.65)％;48 h:M0-CMvs M2-CM=(52.62±5.45)％ vs (70.75±2.33)％;M0-CM vs M0H-CM=(52.62±5.45)％ vs (70.91±4.29)％,P<0.01],vertical migration (24 h:M0-CM vs M2-CM=100.00±4.93 vs 160.00±18.73,M0 vs M0H=10.00±4.93 vs 195.00±6.24;48 h:M0-CMvs M2-CM=43.11±4.66 vs 103.56±10.62,M0 vs M0H=43.11±4.66 vs 136.56±23.49,P<0.01)and invasion (24 h:M0-CM vs M2-CM=62.4±4.56 vs 127.2±12.39,M0 vs M0H=62.4±4.56 vs 131.40±10.88;48 h:M0-CMvs M2-CM=86.40±8.96 vs 162.40±15.95,M0 vs M0H=86.40±8.96 vs 177.00±13.11,P<0.01).Conclusion Mycobacterium tuberculosis infection promotes the prolifera-tion,migration and invasion of A549 cells by inducing macrophage M2 polarization.
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