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Mechanisms of myeloid cell recruitment and biomarker potential in interstitial lung diseases
Mechanisms of myeloid cell recruitment and biomarker potential in interstitial lung diseases
Interstitial lung diseases (ILDs) are fibrotic disorders with chronic inflammation and fibrinogenesis leading to lung scaring and lung function decline. Ultimately, progressive pulmonary fibrosis results in altered pulmonary physiology, abnormal gas exchange, and organ failure. ILDs include known causes and idiopathic causes, as it is the case of idiopathic pulmonary fibrosis (IPF) and non-specific interstitial pneumonia (NSIP). The most detrimental type of ILD is IPF in which anti-fibrotic drugs (nintedanib and pirfenidone) only decrease disease progression. For other ILD types, corticoid treatment helps to decrease exacerbation. Currently, clinical trials are evaluating the applicability of anti-fibrotic drugs for treating non-IPF ILDs. Therefore, mechanistic insights and in-depth cell characterization during tissue injury and remodeling in ILD are of great interest in the respiratory medical field. Circulating immune cell populations have been suggested to play a critical role in ILDs. For instance, mononuclear phagocytes are involved in the initiation, repair and regeneration of pulmonary fibrosis. Moreover, the close interaction between circulating and lung tissue-resident immune cells is critical to contribute to tissue homeostasis or lead to disease. However, precise myeloid phenotypes (e.g. myeloid-derived suppressor cells and monocytes) and their mechanisms of recruitment in ILDs have not yet been explored. In the first results chapter of this thesis, myeloid-derived suppressor cells (MDSC) abundance and function were investigated for the first time in IPF patients. For that, peripheral blood of 170 patients including IPF, non-IPF ILD, chronic obstructive pulmonary diseases (COPD) and controls were collected to characterize and quantify MDSC by flow cytometry. Circulating MDSC in IPF and non-IPF ILD were increased when compared with control. Moreover, cross sectional and longitudinal analysis of the abundance of MDSC inversely correlated with pulmonary function test in IPF only. IPF patients with high number of MDSC showed downregulation of co-stimulatory T cells signals quantified by qRT-PCR. Furthermore, MDSC were able to suppress lymphocytes CD4+ and CD8+ cells proliferation in vitro. Last, CD33 CD11b double positive cells, suggestive of MDSC, were found in neighboring fibrotic niches of the IPF lungs. Taking together, these results show that MDSC are potential biomarker for IPF and are suppressing T cell responses. In the second results chapter, we aimed at analyzing monocyte phenotype and recruitment from the blood to the lung tissue in ILD. Importantly, CX3CR1 expression on immune cells has been demonstrated to increase fibrosis features. For that, flow cytometry analysis of circulating monocytes was performed in 105 subjects (83 ILD, and 22 controls). Monocyte localization and abundance in the lung was assessed by immunofluorescence and flow cytometry analysis. For receptor-ligand function and transmigration pattern, monocytes were isolated from blood and cultured either alone or with endothelial cells. Here, we showed that classical monocytes (CM) were increased, while non-classical monocytes (NCM) were decreased in ILD: NSIP, hypersensitivity pneumonitis (HP) and connective tissue disease associated with ILD (CTD-ILD) compared with controls. Monocytes abundance positively correlated with lung function. Fractalkine levels, the ligand of CX3CR1, were higher in lung tissue than in plasma in ILD and also co-localized with bronchial ciliated cells. Fractalkine enhanced endothelial transmigration of NCM in ILD only. Flow cytometry and immunofluorescence staining showed increased NCM in ILD. NCM-derived cells in the ILD lungs co-stained with CX3CR1, M2-like and phagocytic markers. In summary, we show that epithelial-derived fractalkine drives the migration of NCM-CX3CR1 which provides an interstitial scavenger and phagocytic myeloid cells population in fibrotic ILD lungs.
ILD, lung, fibrosis, monocytes, CX3CR1, fractalkine, MDSC, immunosuppression
Greiffo, Flavia
2019
English
Universitätsbibliothek der Ludwig-Maximilians-Universität München
Greiffo, Flavia (2019): Mechanisms of myeloid cell recruitment and biomarker potential in interstitial lung diseases. Dissertation, LMU München: Faculty of Medicine
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Abstract

Interstitial lung diseases (ILDs) are fibrotic disorders with chronic inflammation and fibrinogenesis leading to lung scaring and lung function decline. Ultimately, progressive pulmonary fibrosis results in altered pulmonary physiology, abnormal gas exchange, and organ failure. ILDs include known causes and idiopathic causes, as it is the case of idiopathic pulmonary fibrosis (IPF) and non-specific interstitial pneumonia (NSIP). The most detrimental type of ILD is IPF in which anti-fibrotic drugs (nintedanib and pirfenidone) only decrease disease progression. For other ILD types, corticoid treatment helps to decrease exacerbation. Currently, clinical trials are evaluating the applicability of anti-fibrotic drugs for treating non-IPF ILDs. Therefore, mechanistic insights and in-depth cell characterization during tissue injury and remodeling in ILD are of great interest in the respiratory medical field. Circulating immune cell populations have been suggested to play a critical role in ILDs. For instance, mononuclear phagocytes are involved in the initiation, repair and regeneration of pulmonary fibrosis. Moreover, the close interaction between circulating and lung tissue-resident immune cells is critical to contribute to tissue homeostasis or lead to disease. However, precise myeloid phenotypes (e.g. myeloid-derived suppressor cells and monocytes) and their mechanisms of recruitment in ILDs have not yet been explored. In the first results chapter of this thesis, myeloid-derived suppressor cells (MDSC) abundance and function were investigated for the first time in IPF patients. For that, peripheral blood of 170 patients including IPF, non-IPF ILD, chronic obstructive pulmonary diseases (COPD) and controls were collected to characterize and quantify MDSC by flow cytometry. Circulating MDSC in IPF and non-IPF ILD were increased when compared with control. Moreover, cross sectional and longitudinal analysis of the abundance of MDSC inversely correlated with pulmonary function test in IPF only. IPF patients with high number of MDSC showed downregulation of co-stimulatory T cells signals quantified by qRT-PCR. Furthermore, MDSC were able to suppress lymphocytes CD4+ and CD8+ cells proliferation in vitro. Last, CD33 CD11b double positive cells, suggestive of MDSC, were found in neighboring fibrotic niches of the IPF lungs. Taking together, these results show that MDSC are potential biomarker for IPF and are suppressing T cell responses. In the second results chapter, we aimed at analyzing monocyte phenotype and recruitment from the blood to the lung tissue in ILD. Importantly, CX3CR1 expression on immune cells has been demonstrated to increase fibrosis features. For that, flow cytometry analysis of circulating monocytes was performed in 105 subjects (83 ILD, and 22 controls). Monocyte localization and abundance in the lung was assessed by immunofluorescence and flow cytometry analysis. For receptor-ligand function and transmigration pattern, monocytes were isolated from blood and cultured either alone or with endothelial cells. Here, we showed that classical monocytes (CM) were increased, while non-classical monocytes (NCM) were decreased in ILD: NSIP, hypersensitivity pneumonitis (HP) and connective tissue disease associated with ILD (CTD-ILD) compared with controls. Monocytes abundance positively correlated with lung function. Fractalkine levels, the ligand of CX3CR1, were higher in lung tissue than in plasma in ILD and also co-localized with bronchial ciliated cells. Fractalkine enhanced endothelial transmigration of NCM in ILD only. Flow cytometry and immunofluorescence staining showed increased NCM in ILD. NCM-derived cells in the ILD lungs co-stained with CX3CR1, M2-like and phagocytic markers. In summary, we show that epithelial-derived fractalkine drives the migration of NCM-CX3CR1 which provides an interstitial scavenger and phagocytic myeloid cells population in fibrotic ILD lungs.