Mechanical stretching of lung epithelial cells induces mesenchymal transition of epithelial cells by promoting macrophages polarization_Chinese Journal of Respiratory and Critical Care Medicine

Authors：

ZHENG Yongxin , YANG Yuanyuan , HUANG Yongbo , YANG Baoxin , CHEN Kangxie , MAO Pu , ZHANG Haibo ,  LI Yimin

Department of Respiratory and Critical Care Medicine, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Institute of Respiratory Health, State Key Laboratory of Respiratory Diseases, Guangzhou, Guangdong 510120, P. R. China;

Corresponding author：

LI Yimin, Email: dryiminli@163.com

Keywords：

Acute respiratory distress syndrome; Macrophage; Platelet-derived growth factor; Epithelial-mesenchymal transition; Mechanical stretch

DOI：

10.7507/1671-6205.202105077

Video：

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Objective To study the effect of mechanical stretch on the microenvironment of BEAS-2B on macrophage polarization and the role of polarized macrophages in the epithelial-mesenchymal transition (EMT) of BEAS-2B. Methods Using enzyme linked immunosorbent assay to detect the changes in the levels of cytokines such as interferon-γ, granulocyte-macrophage colony stimulating factor, tumor necrosis factor-α, interleukin (IL)-4, IL-6, IL-10 in the supernatant of lung epithelial cells cultured statically and mechanically stretched. The M0 macrophages (derived from THP-1) were stimulated by stretch/static conditioned medium of BEAS-2B. The surface markers of M1 (CD197) /M2 (CD206) macrophages were detected by flow cytometer. Stretch/static conditioned medium were used to stimulate the co-culture system of macrophages and BEAS-2B in the presence or absence of platelet-derived growth factor receptor inhibitor (PDGFRi), then the protein expression level of EMT makers was examined by Western blot. Results Exposure of BEAS-2B to mechanical stretch resulted in significantly higher production of the pro-M1/M2 polarized factor. The EMT of the co-culture system of M0 and BEAS-2B could be induced by stretch conditioned medium, epithelial marker cytokeratin (CK)-8 and E-cadherin were decreased, while mesenchymal marker α-smooth muscle actin, N-cadherin and vimentin were increased in stretch conditioned medium group. The expression of platelet-derived growth factor (PDGF) was significantly higher in stretch conditioned medium group. The PDGFRi can block the EMT in stretch conditioned medium group. Conclusions The lung epithelial cell supernatant induced by mechanical stretch can promote the polarization of macrophages to M1 and M2. Polarized macrophages promote EMT in human lung epithelial cells via PDGF, and blocking PDGF might attenuate the VILI-associated lung fibrosis.

Citation： ZHENG Yongxin, YANG Yuanyuan, HUANG Yongbo, YANG Baoxin, CHEN Kangxie, MAO Pu, ZHANG Haibo, LI Yimin. Mechanical stretching of lung epithelial cells induces mesenchymal transition of epithelial cells by promoting macrophages polarization. Chinese Journal of Respiratory and Critical Care Medicine, 2021, 20(12): 870-876. doi: 10.7507/1671-6205.202105077 Copy

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2.	Matthay MA, Zemans RL, Zimmerman GA, et al. Acute respiratory distress syndrome. Nat Rev Dis Primers, 2019, 5(1): 18.
3.	Ichikado K, Muranaka H, Gushima Y, et al. Fibroproliferative changes on high-resolution CT in the acute respiratory distress syndrome predict mortality and ventilator dependency: a prospective observational cohort study. BMJ Open, 2012, 2(2): e000545.
4.	张容, 毛璞, 傅威, 等. 机械牵张对人肺上皮细胞转分化的影响. 中华危重病急救医学, 2013, 25(8): 455-459.
5.	Dehai C, Bo P, Qiang T, et al. Enhanced invasion of lung adenocarcinoma cells after co-culture with THP-1-derived macrophages via the induction of EMT by IL-6. Immunol Lett, 2014, 160(1): 1-10.
6.	Wang L, Zhang Y, Zhang NN, et al. Potential role of M2 macrophage polarization in ventilator-induced lung fibrosis. Int Immunopharmacol, 2019, 75: 105795.
7.	Jaguin M, Fardel O, Lecureur V. AhR-dependent secretion of PDGF-BB by human classically activated macrophages exposed to DEP extracts stimulates lung fibroblast proliferation. Toxicol Appl Pharmacol, 2015, 285(3): 170-178.
8.	Das S, Majid M, Baker AB. Syndecan-4 enhances PDGF-BB activity in diabetic wound healing. Acta Biomater, 2016, 42: 56-65.
9.	Kovalenko M, Gazit A, Böhmer A, et al. Selective platelet-derived growth factor receptor kinase blockers reverse sis-transformation. Cancer Res, 1994, 54(23): 6106-6114.
10.	Maess MB, Wittig B, Cignarella A, et al. Reduced PMA enhances the responsiveness of transfected THP-1 macrophages to polarizing stimuli. J Immunol Methods, 2014, 402(1-2): 76-81.
11.	Nikonova A, Khaitov M, Jackson DJ, et al. M1-like macrophages are potent producers of anti-viral interferons and M1-associated marker-positive lung macrophages are decreased during rhinovirus-induced asthma exacerbations. EBioMedicine, 2020, 54: 102734.
12.	Wang J, Xu L, Xiang Z, et al. Microcystin-LR ameliorates pulmonary fibrosis via modulating CD206(+) M2-like macrophage polarization. Cell Death Dis, 2020, 11(2): 136.
13.	Singh M, Yelle N, Venugopal C, et al. EMT: Mechanisms and therapeutic implications. Pharmacol Ther, 2018, 182: 80-94.
14.	Cabrera-Benitez NE, Laffey JG, Parotto M, et al. Mechanical ventilation-associated lung fibrosis in acute respiratory distress syndrome: a significant contributor to poor outcome. Anesthesiology, 2014, 121(1): 189-198.
15.	Cabrera-Benitez NE, Parotto M, Post M, et al. Mechanical stress induces lung fibrosis by epithelial-mesenchymal transition. Crit Care Med, 2012, 40(2): 510-517.
16.	Murray PJ. Macrophage polarization. Annu Rev Physiol, 2017, 79: 541-566.
17.	Gharib SA, Johnston LK, Huizar I, et al. MMP28 promotes macrophage polarization toward M2 cells and augments pulmonary fibrosis. J Leukoc Biol, 2014, 95(1): 9-18.
18.	Sunderkotter C, Goebeler M, Schulze-Osthoff K, et al. Macrophage-derived angiogenesis factors. Pharmacol Ther, 1991, 51(2): 195-216.
19.	Neri S, Miyashita T, Hashimoto H, et al. Fibroblast-led cancer cell invasion is activated by epithelial-mesenchymal transition through platelet-derived growth factor BB secretion of lung adenocarcinoma. Cancer Lett, 2017, 395: 20-30.
20.	Devarajan E, Song YH, Krishnappa S, et al. Epithelial-mesenchymal transition in breast cancer lines is mediated through PDGF-D released by tissue-resident stem cells. Int J Cancer, 2012, 131(5): 1023-1031.
21.	Aono Y, Kishi M, Yokota Y, et al. Role of platelet-derived growth factor/platelet-derived growth factor receptor axis in the trafficking of circulating fibrocytes in pulmonary fibrosis. Am J Respir Cell Mol Biol, 2014, 51(6): 793-801.
22.	Kishi M, Aono Y, Sato S, et al. Blockade of platelet-derived growth factor receptor-beta, not receptor-alpha ameliorates bleomycin-induced pulmonary fibrosis in mice. PLoS One, 2018, 13(12): e0209786.
23.	Deng X, Jin K, Li Y, et al. Platelet-derived growth factor and transforming growth factor beta1 regulate ARDS-associated lung fibrosis through distinct signaling pathways. Cell Physiol Biochem, 2015, 36(3): 937-946.
24.	Glim JE, Niessen FB, Everts V, et al. Platelet derived growth factor-CC secreted by M2 macrophages induces alpha-smooth muscle actin expression by dermal and gingival fibroblasts. Immunobiology, 2013, 218(6): 924-929.

1. Bellani G, Laffey JG, Pham T, et al. Epidemiology, patterns of care, and mortality for patients with acute respiratory distress syndrome in intensive care units in 50 countries. JAMA, 2016, 315(8): 788-800.
2. Matthay MA, Zemans RL, Zimmerman GA, et al. Acute respiratory distress syndrome. Nat Rev Dis Primers, 2019, 5(1): 18.
3. Ichikado K, Muranaka H, Gushima Y, et al. Fibroproliferative changes on high-resolution CT in the acute respiratory distress syndrome predict mortality and ventilator dependency: a prospective observational cohort study. BMJ Open, 2012, 2(2): e000545.
4. 张容, 毛璞, 傅威, 等. 机械牵张对人肺上皮细胞转分化的影响. 中华危重病急救医学, 2013, 25(8): 455-459.
5. Dehai C, Bo P, Qiang T, et al. Enhanced invasion of lung adenocarcinoma cells after co-culture with THP-1-derived macrophages via the induction of EMT by IL-6. Immunol Lett, 2014, 160(1): 1-10.
6. Wang L, Zhang Y, Zhang NN, et al. Potential role of M2 macrophage polarization in ventilator-induced lung fibrosis. Int Immunopharmacol, 2019, 75: 105795.
7. Jaguin M, Fardel O, Lecureur V. AhR-dependent secretion of PDGF-BB by human classically activated macrophages exposed to DEP extracts stimulates lung fibroblast proliferation. Toxicol Appl Pharmacol, 2015, 285(3): 170-178.
8. Das S, Majid M, Baker AB. Syndecan-4 enhances PDGF-BB activity in diabetic wound healing. Acta Biomater, 2016, 42: 56-65.
9. Kovalenko M, Gazit A, Böhmer A, et al. Selective platelet-derived growth factor receptor kinase blockers reverse sis-transformation. Cancer Res, 1994, 54(23): 6106-6114.
10. Maess MB, Wittig B, Cignarella A, et al. Reduced PMA enhances the responsiveness of transfected THP-1 macrophages to polarizing stimuli. J Immunol Methods, 2014, 402(1-2): 76-81.
11. Nikonova A, Khaitov M, Jackson DJ, et al. M1-like macrophages are potent producers of anti-viral interferons and M1-associated marker-positive lung macrophages are decreased during rhinovirus-induced asthma exacerbations. EBioMedicine, 2020, 54: 102734.
12. Wang J, Xu L, Xiang Z, et al. Microcystin-LR ameliorates pulmonary fibrosis via modulating CD206(+) M2-like macrophage polarization. Cell Death Dis, 2020, 11(2): 136.
13. Singh M, Yelle N, Venugopal C, et al. EMT: Mechanisms and therapeutic implications. Pharmacol Ther, 2018, 182: 80-94.
14. Cabrera-Benitez NE, Laffey JG, Parotto M, et al. Mechanical ventilation-associated lung fibrosis in acute respiratory distress syndrome: a significant contributor to poor outcome. Anesthesiology, 2014, 121(1): 189-198.
15. Cabrera-Benitez NE, Parotto M, Post M, et al. Mechanical stress induces lung fibrosis by epithelial-mesenchymal transition. Crit Care Med, 2012, 40(2): 510-517.
16. Murray PJ. Macrophage polarization. Annu Rev Physiol, 2017, 79: 541-566.
17. Gharib SA, Johnston LK, Huizar I, et al. MMP28 promotes macrophage polarization toward M2 cells and augments pulmonary fibrosis. J Leukoc Biol, 2014, 95(1): 9-18.
18. Sunderkotter C, Goebeler M, Schulze-Osthoff K, et al. Macrophage-derived angiogenesis factors. Pharmacol Ther, 1991, 51(2): 195-216.
19. Neri S, Miyashita T, Hashimoto H, et al. Fibroblast-led cancer cell invasion is activated by epithelial-mesenchymal transition through platelet-derived growth factor BB secretion of lung adenocarcinoma. Cancer Lett, 2017, 395: 20-30.
20. Devarajan E, Song YH, Krishnappa S, et al. Epithelial-mesenchymal transition in breast cancer lines is mediated through PDGF-D released by tissue-resident stem cells. Int J Cancer, 2012, 131(5): 1023-1031.
21. Aono Y, Kishi M, Yokota Y, et al. Role of platelet-derived growth factor/platelet-derived growth factor receptor axis in the trafficking of circulating fibrocytes in pulmonary fibrosis. Am J Respir Cell Mol Biol, 2014, 51(6): 793-801.
22. Kishi M, Aono Y, Sato S, et al. Blockade of platelet-derived growth factor receptor-beta, not receptor-alpha ameliorates bleomycin-induced pulmonary fibrosis in mice. PLoS One, 2018, 13(12): e0209786.
23. Deng X, Jin K, Li Y, et al. Platelet-derived growth factor and transforming growth factor beta1 regulate ARDS-associated lung fibrosis through distinct signaling pathways. Cell Physiol Biochem, 2015, 36(3): 937-946.
24. Glim JE, Niessen FB, Everts V, et al. Platelet derived growth factor-CC secreted by M2 macrophages induces alpha-smooth muscle actin expression by dermal and gingival fibroblasts. Immunobiology, 2013, 218(6): 924-929.

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Mechanical stretching of lung epithelial cells induces mesenchymal transition of epithelial cells by promoting macrophages polarization

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