IL-17A Regulation Of Alveolar Macrophage Polarization Promotes Early Inflammation During Silica-induced Pulmonary Fibrosis

Background Silicosis is a systemic disease characterized by diffuse fibrosis of the lungs caused by the prolonged inhalation of free silica（SiO₂）dust.The development of silicosis includes lung inflammation and fibrosis stages.Alveolar macrophages（AMs）are the main target cells for silicosis.During the progression of pulmonary fibrosis,the function of AMs is closely related to their polarization of AMs,which is regulated by a variety of cytokines.IL-17A,as a helper Th17 cells-secreted pro-inflammatory cytokine,plays an important role in the development and progression of pulmonary disorders,but the role of IL-17A in the inflammatory and pulmonary fibrosis process of silicosis and its relationship to the polarisation of AMs is still not clear.Objective1.To explore the relationship between IL-17A expression and AMs polarization,lung tissue inflammation and fibrosis during SiO₂exposure-induced lung fibrosis.2.To clarify the mechanism by which IL-17A regulates polarization of AMs in SiO₂-induced pulmonary fibrosis to promote early inflammation in pulmonary fibrosis using IL-17A knockout mice.MethodsTo investigate the relationship between IL-17A expression and lung tissue inflammation,fibrosis and AMs polarization in SiO₂-exposed mice,8-week-old wild type（WT）C57BL/6J female mice were randomly divided into WT+Saline group and（WT+SiO₂）group（6 mice each group）.Mice were treated with 100μL saline or 100μL20 mg/m L SiO₂by intratracheal instillation,respectively.IL-17A knockout(IL-17A^-/-)mice was used to verify whether IL-17A regulates AMs polarization during SiO₂-induced pulmonary fibrosis.8-week-old female IL-17A^-/-mice were randomly divided into IL-17A^-/-+Saline and IL-17A^-/-+SiO₂group（6 mice each group）.Mice were treated with 100μL saline or 100μL 20 mg/m L SiO₂by intratracheal instillation,respectively.All mice continued to be fed after treatment and alveolar lavage fluid（BALF）and lung tissue were collected at 3 and 28 days to determine IL-17A content and expression levels,to observe lung inflammation and fibrosis,and to detect changes in AMs polarization.Results1.The correlation of IL-17A expression and lung tissue inflammation,fibrosis and polarization of AMs in SiO₂-exposed miceCompared to the WT+Saline group,the expression levels of IL-17A in the lung tissue and BALF of mice in the SiO₂group increased significantly after 3 days of exposure,and the infiltration of inflammatory cells in the lung increased,while collagen fibril deposition,hydroxyproline（HYP）levels and the expression of molecules related to pulmonary fibrosis in the lung tissue were not significantly changed.There was a significant increase in the proportion of M1 AMs（P<0.05）and no significant difference in the proportion of M2 AMs（P>0.05）in BALF.On day 28 after SiO₂exposure,the differences in IL-17A expression levels and lung inflammation in lung tissue and BALF were not significant,but there was a significant increase in collagen deposition,an increase in hydroxyproline（HYP）levels,a down-regulation in the expression of the lung fibrosis-related molecules E-calcadherin（E-cad）and an up-regulation in the expression of vimentin,α-smooth muscle actin（α-SMA）and collagen I（COL-Ⅰ）in lung tissue.The expression of vimentin,α-smooth muscle actin（α-SMA）and collagenⅠ（COL-Ⅰ）was upregulated;the difference in the proportion of M1 subtype AMs in BALF was not significant（P>0.05）,while the proportion of M2 subtype AMs increased significantly（P<0.05）.Correlation analysis showed that IL-17A levels showed significant correlation with early lung inflammation,late lung fibrosis,and the proportion of early M1 subtype AMs.2.IL-17A regulates early polarization of AMs to M1 type in SiO₂exposure to promote early inflammation in pulmonary fibrosisCompared to the WT+SiO₂group,On day 3 after SiO₂exposure,IL-17A expression levels in lung tissues and BALF of mice in the IL-17A^-/-+SiO₂group were significantly decreased,lung inflammation was significantly reduced,lung tissue collagen fibrils,HYP levels and expression of molecules related to lung fibrosis were not significantly altered;the proportion of M1 subtype AMs was significantly decreased（P<0.05）and M2 subtype in the proportion of AMs were not significant（P>0.05）.After28 days of SiO₂exposure,the level of IL-17A expression in lung tissue and BALF decreased,lung inflammation was significantly alleviated,lung tissue collagen deposition was significantly reduced,HYP)level decreased,the expression of lung fibrosis-related molecule E-cad was upregulated,and the expression of vimentin,α-SMA and COL-Ⅰwas downregulated;the differences in the proportion of M1 and M2subtype AMs in BALF were not significant（P>0.05）.Conclusions1.IL-17A expression was significantly associated with early lung tissue inflammation,late fibrosis,and polarization of AMs to M1 type in SiO₂-exposed mice.2.IL-17A promotes early inflammation and mediates late lung fibrosis by regulating the polarization of AMs to M1 type in the early stages of SiO₂exposure.