transforming-growth-factor-beta and Alveolitis--Extrinsic-Allergic

Fibroblasts differ in a variety of phenotypic features, including the expression of Thy-1 a glycophosphatidylinositol-linked glycoprotein. Fibroblasts in idiopathic pulmonary fibrosis (IPF) are Thy-1 negative, whereas most fibroblasts from normal lungs are Thy-1 positive. However, the functional consequences of the absence of Thy-1 are not fully understood. We analyzed the expression of Thy-1 in several primary fibroblasts lines derived from IPF, hypersensitivity pneumonitis (HP), and normal human lungs. We found that a high proportion, independently of their origin, expressed Thy-1 in vitro. We identified a primary culture of HP fibroblasts, which did not express Thy-1, and compared several functional activities between Thy-1 (-) and Thy-1 (+) fibroblasts. Thy-1 (-) fibroblasts were smaller (length: 41.3±20.8 μ versus 83.1±40 μ), showed increased proliferative capacity and enhanced PDGF-induced transmigration through collagen I (59.9% versus 42.2% over control under basal conditions, P<0.01). Likewise, Thy-1 (-) fibroblasts either spontaneously or after TGF-β stimulation demonstrated stronger contraction of collagen matrices (eg, 0.17±0.03 versus 0.6±0.05 cm² after TGF-β stimulation at 24 h; P<0.01). Thy-1 (-) lung fibroblasts stimulated with TGF-β1 expressed MMP-9, an enzyme that is usually not produced by lung fibroblasts. TGFβ-induced MMP-9 expression was reversible upon re-expression of Thy-1 after transfection with full-length Thy-1. β-glycan, a TGF-β receptor antagonist abolished MMP-9 expression. TGF-β1-induced MMP-9 in Thy-1 (-) fibroblasts depended on the activation of ERK1/2 signaling pathway. Finally, we demonstrated that fibroblasts from IPF fibroblastic foci, which do not express Thy-1 exhibit strong staining for immunoreactive MMP-9 protein in vivo. These findings indicate that loss of Thy-1 in human lung fibroblasts induces a fibrogenic phenotype.

Topics: Alveolitis, Extrinsic Allergic; Cell Line; Cell Movement; Cell Proliferation; Collagen; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Female; Fibroblasts; Fibrosis; Gene Transfer Techniques; Humans; Idiopathic Pulmonary Fibrosis; Lung; MAP Kinase Signaling System; Matrix Metalloproteinase 9; Middle Aged; Phenotype; RNA, Messenger; Thy-1 Antigens; Transforming Growth Factor beta; Wound Healing

HP results from the repeated inhalation of environmental antigens; however, the roles of CD4(+)CD25(+) T(reg) cells in HP are unknown. Therefore, we investigated the functions of CD4(+)CD25(+) T(reg) cells in SR-induced murine HP. More severe HP was observed in CD4(+)CD25(+) T(reg) cell-depleted mice than in control mice in terms of histological alterations, inflammatory cell numbers in BALF, and the serum level of SR-specific IgG, which were restored by the adoptive transfer of CD4(+)CD25(+) T(reg) cells. The CD4(+)CD25(+) T(reg) cell-depleted mice also showed elevated levels of IFN-gamma, TGF-beta, and reduced IL-4 production in the lungs. Moreover, IL-10 production of CD4(+)CD25(+) T(reg) cells and direct contact between CD4(+)CD25(+) T(reg) cells and CD4(+) or CD8(+) T cells in BALF resulted in reduced IFN-gamma production. Taken together, CD4(+)CD25(+) T(reg) cells play a protective role in SR-induced HP by suppressing IFN-gamma production by T cells.

Topics: Alveolitis, Extrinsic Allergic; Animals; CD8-Positive T-Lymphocytes; Immunoglobulin G; Interferon-gamma; Interleukin-10; Male; Mice; Mice, Knockout; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Hypersensitivity pneumonitis (HP) is an immunologically-mediated lung disease caused by repeated inhalation of dispersed antigen. Various cytokines have been reported to be involved in the immunopathogenesis of HP Recently, some reports suggested an association between the genetic control of cytokine production and disease susceptibility. To evaluate whether cytokine gene polymorphisms are associated with HP, we performed a case-control association study involving 61 patients with HP, consisting of summer-type HP (SHP) and bird fancier's lung (BFL, also named bird fancier's disease), as well as 101 healthy controls. Polymorphisms of the genes for tumor necrosis factor (TNF)-alpha (-308G/A, -857C/T, -863C/A, -1031T/C), interleukin (IL)-10 (-592C/A, -819C/T, -1082G/A), transforming growth factor (TGF)-beta1 (-509C/T, +869T/C), and IL-6 (-634C/G) were examined by restriction fragment length polymorphism analysis. There were no significant differences in allele frequency and genotype distribution among control, SHP, and BFL group. When HP group was divided into acute or chronic, no significant differences were detected between any groups. LPS-stimulated IL-6 secretion by whole blood cells was similar between subjects with GG genotype and non-GG genotype in IL-6 -634C/G polymorphism. In conclusion, the association between HP susceptibility and cytokine polymorphisms studied was not demonstrated.

Topics: Alveolitis, Extrinsic Allergic; Bird Fancier's Lung; Case-Control Studies; Cytokines; Gene Frequency; Genetic Predisposition to Disease; Genotype; Humans; Interleukin-10; Interleukin-6; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Statistics, Nonparametric; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha

The expression of alphaEbeta7 integrin has been related to the selective retention of lymphocytes in mucosal tissues of gut, urogenital tract and lung. To identify potential disease-associated alphaEbeta7 expression patterns on cells accounting for lymphocytic alveolitis in interstitial lung disease (ILD), alphaE expression on CD4+ and CD8+ T cell subsets was evaluated by dual-colour flow cytometry in peripheral blood and bronchoalveolar lavage fluid (BALF) of patients with idiopathic pulmonary fibrosis (IPF; n = 18), hypersensitivity pneumonitis (HP; n = 20) and sarcoidosis (n = 44) in comparison with healthy controls (n = 15). In both healthy individuals and all patient groups the proportion of alphaE-bearing T cells in peripheral blood was < 2%, whereas the vast majority of alveolar CD8+ T cells consistently co-expressed alphaE. Absolute alveolar CD8+alphaE+ cell numbers/ml were up to 30-fold increased in HP patients. Proportions of alphaE-bearing CD4+ cells in BALF were significantly elevated in IPF (74.0 +/- 2.7%) and HP (70.0 +/- 2.4%) compared with normals (30.0 +/- 1.8%) (mean +/- s.e.m.; P < 0.01). In sarcoidosis, the alphaE expression on BALF CD4+ cells displayed subgroup dependency: proportions significantly lower than normal were noted in chest radiographic stage I (14.3 +/- 1.5%), but increased proportions in stages II (50.0 +/- 3.8%) and III (64.0 +/- 4.8%). Correlations between common markers of T cell activation or BALF transforming growth factor-beta (TGF-beta ) bioactivity and alphaE expression were not noted. We conclude that the vast majority of alveolar CD8+ T cells consistently express alphaEbeta7 and that distinct patterns of alphaEbeta7 expression on alveolar CD4+ lymphocytes in sarcoidosis are related to the diverse manifestations of the sarcoid inflammatory process in the lung.

Topics: Adolescent; Adult; Aged; Alveolitis, Extrinsic Allergic; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Female; Humans; Integrins; Male; Middle Aged; Pulmonary Fibrosis; Sarcoidosis; Transforming Growth Factor beta

T lymphocytes expressing the alpha E beta 7 integrin are localized and selectively retained in mucosal tissues. To investigate a potential relationship between alpha E beta 7 expression and pulmonary inflammation, the distribution of alpha E beta 7-bearing CD4+ and CD8+ T cells in peripheral blood and bronchoalveolar lavage (BAL) fluids obtained from patients with allergic asthma, sarcoidosis, hypersensitivity pneumonitis, and idiopathic pulmonary fibrosis (IPF) was determined. In contrast to the distribution in peripheral blood, BAL fluid from these patients contained high number of cells expressing alpha E beta 7 with markedly different expression patterns on CD4 or CD8 cells as well as among the various diseases. Despite similar numbers of activated CD4 cells, alpha E beta 7+CD4+ T cells ranged from 15% in asthmatics to 70% in IPF. In contrast, even in normal individuals, 60% to 90% of BAL fluid CD8+ T cells express alpha E beta 7, suggesting differential induction mechanisms on CD4 and CD8 cells. In vitro experiments revealed that a substantial proportion of peripheral blood CD+ T cells express alpha E beta 7 after stimulation with anti-CD3 antibodies, and up to 80% positive cells were found after the addition of TGF-beta. In contrast, less than 10% of CD4 cells express this particular integrin after in vitro stimulation, and the presence of TGF-beta only increased the number to 30%. Supernatants from in vitro-activated BAL cells as well as concentrated BAL fluid from patients with high alpha E beta 7 expression had no further enhancing effect. However, crosslinking of alpha 4 beta 1-, but not beta 2-integrins, significantly increased the number of alpha E beta 7 expressing CD4+ and CD8+ T cells, even in the absence of TGF-beta. These data indicate that in addition to TGF-beta, the interaction of particular T-cell subsets with specific endothelial cell and extracellular matrix proteins may upregulate alpha E beta 7 integrin expression and thereby contribute to the selective accumulation of these cells in inflammatory lung diseases.

Topics: Adult; Alveolitis, Extrinsic Allergic; Antibodies; Asthma; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cross-Linking Reagents; Eosinophils; Female; HLA-DR Antigens; Humans; Integrin alpha4beta1; Integrins; Leukocyte Common Antigens; Lung Diseases, Interstitial; Lymphocyte Activation; Lymphocyte Subsets; Macrophages, Alveolar; Male; Middle Aged; Neutrophils; Pulmonary Fibrosis; Receptors, Interleukin-2; Receptors, Lymphocyte Homing; Sarcoidosis; T-Lymphocytes; Transforming Growth Factor beta

Mice of the C57BL/6 strain were instilled with optimal doses (150 micrograms/day for 3 days/wk) of the thermophilic actinomycete Faeni rectivirgula (also known as Saccharopolyspora rectivirgula or Micropolyspora faeni) to induce a hypersensitivity pneumonitis inflammation that mimics the human disease affecting certain occupational groups. This mouse model was characterized by a very significant alveolitis (3-fold increase in bronchoalveolar lavage [BAL] cell number at 48 h and a 10-fold increase at 3 wk). Also, total lung transforming growth factor (TGF-beta) was shown to be elevated in treated mice as early as 1 wk after the first instillation and increased gradually to 2.5 micrograms/lung at 3 wk (approximately 0.3 microgram/lung in saline-instilled controls). Intranasal instillation with F. rectivirgula was also associated with very significant increases in lung fibroblast collagen synthesis, starting at 2 wk. BAL macrophages from mice instilled with F. rectivirgula were found to release significantly more TGF-beta upon in vitro stimulation with zymosan beads than did BAL macrophages from saline-instilled mice. Finally, we show that supernatants from activated BAL macrophages of mice given F. rectivirgula increased quite significantly collagen synthesis in normal mouse lung fibroblasts. This increase could be abrogated by treating conditioned medium with a rabbit antibody against TGF-beta. Collectively, these data suggest that TGF-beta is generated in the course of experimental mouse hypersensitivity pneumonitis and contributes significantly to collagen synthesis.

Topics: Alveolitis, Extrinsic Allergic; Animals; Bronchoalveolar Lavage Fluid; Collagen; Disease Models, Animal; Fibroblasts; Lung; Macrophages, Alveolar; Mice; Mice, Inbred C57BL; Saccharopolyspora; Transforming Growth Factor beta