Tobacco smoke activated fibrogenic MARCKS/AXL complex promotes macrophage reprogramming and pulmonary fibrosis

Abstract

Abstract Macrophages and tobacco smoke (TS) exposure have been demonstrated to play significant roles in modulating pulmonary fibrosis (PF). However, the mechanisms of how TS exposure modulates pro-fibrotic macrophage polarization and drives lung fibrosis is unclear. In our study, we investigated how TS modulates macrophage polarization and the functional consequences of this polarization. Multicolor flow cytometric data indicated that markers of M2 macrophage polarization were elevated in both human and mouse macrophage cells and tissues upon TS exposure. In addition, multiple primary lung fibroblast cells demonstrated elevated pro-fibrotic markers and aggressive phenotypes upon interacting with TS-exposed macrophage cells in a co-culture system. Elucidation of the signaling pathways activated by TS exposure through a receptor tyrosine kinase array screen revealed AXL receptor as a novel smoke-responsive molecule in macrophage cells. We noted elevated secretion of AXL ligand, Gas6, and AXL activity in TS exposed cells and tissues. Prior work had demonstrated an interaction between MARCKS, a smoke-responsive protein, and AXL in promoting a pro-fibrotic phenotype in lung fibroblasts. Similarly, we observed AXL activity positively correlated with MARCKS phosphorylation in macrophage cells. Pharmacologic and genetic targeting of the MARCKS/AXL signaling complex reduced M2 markers and profibrotic cytokine production in macrophages and reduced fibrotic changes in the co-culture model and in an animal model of smoke-mediated lung fibrosis. In all, our work suggests that the MARCKS/AXL fibrogenic complex is a potential target in attenuating macrophage activity in TS-mediated fibrosis. Supported by grants from NIH/NHLBI (R01HL146802), DOD DCMRP/PRMRP (PR202411), and UCOP TRDRP (28IR-0061 and T31DT1849).