Interleukins (from IL-1 to IL-38), interferons, transforming growth factor ß, and TNF-α: Receptors, functions, and roles in diseases.

There have been extensive developments on cellular and molecular mechanisms of immune regulation in allergy, asthma, autoimmune diseases, tumor development, organ transplantation, and chronic infections during the last few years. Better understanding the functions, reciprocal regulation, and counterbalance of subsets of immune and inflammatory cells that interact through interleukins, interferons, TNF-α, and TGF-ß offer opportunities for immune interventions and novel treatment modalities in the era of development of biological immune response modifiers particularly targeting these molecules or their receptors. More than 60 cytokines have been designated as interleukins since the initial discoveries of monocyte and lymphocyte interleukins (called IL-1 and IL-2, respectively). Studies of transgenic or gene-deficient mice with altered expression of these cytokines or their receptors and analyses of mutations and polymorphisms in human genes that encode these products have provided essential information about their functions. Here we review recent developments on IL-1 to IL-38, TNF-α, TGF-ß, and interferons. We highlight recent advances during the last few years in this area and extensively discuss their cellular sources, targets, receptors, signaling pathways, and roles in immune regulation in patients with allergy and asthma and other inflammatory diseases.

Differential serum protein markers and the clinical severity of asthma.

BACKGROUND: Asthma is a heterogeneous disease characterized by different clinical phenotypes and the involvement of multiple inflammatory pathways. During airway inflammation, many cytokines and chemokines are released and some are detectable in the sera. OBJECTIVE: Serum chemokines and cytokines, involved in airway inflammation in asthma patients, were investigated. METHODS: A total of 191 asthma patients were classified by hierarchical cluster analysis, including the following parameters: forced expiratory volume in 1 second (FEV1), eosinophil cationic protein (ECP) serum levels, blood eosinophils, Junipers asthma symptom score, and the change in FEV1, ECP serum levels, and blood eosinophils after 3 weeks of asthma therapy. Serum proteins were measured by multiplex analysis. Receiver operating characteristic (ROC) curves were used to evaluate the validity of serum proteins for discriminating between asthma clusters. RESULTS: Classification of asthma patients identified one cluster with high ECP serum levels, increased blood eosinophils, low FEV1 values, and good FEV1 improvement in response to asthma therapy (n=60) and one cluster with low ECP serum levels, low numbers of blood eosinophils, higher FEV1 values, and no FEV1 improvement in response to asthma therapy (n=131). Serum interleukin (IL)-8, eotaxin, vascular endothelial growth factor (VEGF), cutaneous T-cell-attracting chemokine (CTACK), growth-related oncogene (GRO)-α, and hepatocyte growth factor (HGF) were significantly different between the two clusters of asthma patients. ROC analysis for serum proteins calculated a sensitivity of 55.9% and specificity of 75.8% for discriminating between them. CONCLUSION: Serum cytokine and chemokine levels might be predictors for the severity of asthmatic inflammation, asthma control, and response to therapy, and therefore might be useful for treatment optimization.

Vascular endothelial growth factor as a key inducer of angiogenesis in the asthmatic airways.

Asthma is a chronic inflammatory disease of the airways characterized by structural airway changes, which are known as airway remodeling, including smooth muscle hypertrophy, goblet cell hyperplasia, subepithelial fibrosis, and angiogenesis. Vascular remodeling in asthmatic lungs results from increased angiogenesis, which is mainly mediated by vascular endothelial growth factor (VEGF). VEGF is a key regulator of blood vessel growth in the airways of asthma patients by promoting proliferation and differentiation of endothelial cells and inducing vascular leakage and permeability. In addition, VEGF induces allergic inflammation, enhances allergic sensitization, and has a role in Th2 type inflammatory responses. Specific inhibitors of VEGF and blockers of its receptors might be useful to control chronic airway inflammation and vascular remodeling, and might be a new therapeutic approach for chronic inflammatory airway disease like asthma.

Mechanisms of IFN-γ-induced apoptosis of human skin keratinocytes in patients with atopic dermatitis.

BACKGROUND: Enhanced apoptosis of keratinocytes is the main cause of eczema and spongiosis in patients with the common inflammatory skin disease atopic dermatitis (AD). OBJECTIVE: The aim of the study was to investigate molecular mechanisms of AD-related apoptosis of keratinocytes. METHODS: Primary keratinocytes isolated from patients with AD and healthy donors were used to study apoptosis by using annexin V/7-aminoactinomycin D staining. Illumina mRNA Expression BeadChips, quantitative RT-PCR, and immunofluorescence were used to study gene expression. In silico analysis of candidate genes was performed on genome-wide single nucleotide polymorphism data. RESULTS: We demonstrate that keratinocytes of patients with AD exhibit increased IFN-γ-induced apoptosis compared with keratinocytes from healthy subjects. Further mRNA expression analyses revealed differential expression of apoptosis-related genes in AD keratinocytes and skin and the upregulation of immune system-related genes in skin biopsy specimens of chronic AD lesions. Three apoptosis-related genes (NOD2, DUSP1, and ADM) and 8 genes overexpressed in AD skin lesions (CCDC109B, CCL5, CCL8, IFI35, LYN, RAB31, IFITM1, and IFITM2) were induced by IFN-γ in primary keratinocytes. The protein expression of IFITM1, CCL5, and CCL8 was verified in AD skin. In line with the functional studies and AD-related mRNA expression changes, in silico analysis of genome-wide single nucleotide polymorphism data revealed evidence of an association between AD and genetic markers close to or within the IFITM cluster or RAB31, DUSP1, and ADM genes. CONCLUSION: Our results demonstrate increased IFN-γ responses in skin of patients with AD and suggest involvement of multiple new apoptosis- and inflammation-related factors in the development of AD.

Inhibition of angiogenesis by IL-32: possible role in asthma.

BACKGROUND: IL-32 is a proinflammatory cytokine involved in various chronic inflammatory diseases. Chronic airway inflammation in asthmatic patients results in structural airway changes, including angiogenesis. Vascular endothelial growth factor (VEGF) is a key inducer of angiogenesis in the airways of asthmatic patients. OBJECTIVE: The aim of the study was to investigate the expression and function of IL-32 in patients with angiogenesis and asthma. METHODS: The expression and regulation of IL-32 in normal human bronchial epithelial (NHBE) cells was analyzed by using RT-PCR, ELISA, Western blotting, immunofluorescent staining, and flow cytometry. After knockdown of IL-32 in NHBE cells by small interfering RNA (siRNA) transfections, VEGF secretion was quantified by means of ELISA. New blood vessel formation was determined with human umbilical vein endothelial cells by culturing with supernatants from IL-32 siRNA-transfected NHBE cells. IL-32 was determined in serum and induced sputum samples of asthmatic patients and healthy control subjects by means of ELISA. RESULTS: IL-32 is expressed in NHBE cells on stimulation with IFN-γ, TNF-α, T(H)1 cells, and rhinovirus. Inhibition of IL-32 expression resulted in significantly increased secretion of the proangiogenic factors VEGF and platelet-derived growth factor by NHBE cells. Human umbilical vein endothelial cells cultured in supernatants from IL-32 siRNA-transfected NHBE cells showed enhanced in vitro angiogenesis. IL-32 is detectable in induced sputum from asthmatic patients. IL-32 serum levels were significantly higher in asthmatic patients compared with those seen in healthy control subjects and correlated with response to asthma treatment. CONCLUSION: IL-32 is induced by IFN-γ, TNF-α, T(H)1 cells, and rhinovirus in bronchial epithelial cells. It inhibits angiogenesis, and its serum levels are associated with a good treatment response in asthmatic patients.

Claudin-1 expression in airway smooth muscle exacerbates airway remodeling in asthmatic subjects.

BACKGROUND: Increased airway smooth muscle (ASM) mass is an essential component of airway remodeling and asthma development, and there is no medication specifically against it. Tight junction (TJ) proteins, which are expressed in endothelial and epithelial cells and affect tissue integrity, might exist in other types of cells and display additional functions in the asthmatic lung. OBJECTIVE: The aim of this study was to investigate the existence, regulation, and function of TJ proteins in ASM in asthmatic patients. METHODS: The expression and function of TJ proteins in primary ASM cell lines, human bronchial biopsy specimens, and a murine model of asthma were analyzed by means of RT-PCR, multispectral imaging flow cytometry, immunohistochemistry, Western blotting, 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester staining, tritiated thymidine incorporation, wound-healing assay, and luminometric bead array. RESULTS: Increased claudin-1 expression was observed in ASM of asthmatic patients, as well as in a murine model of asthma-like airway inflammation. Whereas IL-1ß and TNF-α upregulated claudin-1 expression, it was downregulated by the T(H)2 cytokines IL-4 and IL-13 in primary human ASM cells. Claudin-1 was localized to the nucleus and cytoplasm but not to the cell surface in ASM cells. Claudin-1 played a central role in ASM cell proliferation, as demonstrated by increased ASM cell proliferation seen with overexpression and decreased proliferation seen with small interfering RNA knockdown of claudin-1. Overexpression of claudin-1 induced vascular endothelial growth factor and downregulated IL-6, IL-8, and IFN-γ-induced protein 10 production by ASM cells. Claudin-1 upregulation by IL-1ß or TNF-α was suppressed by dexamethasone but not by rapamycin, FK506, or salbutamol. CONCLUSION: These results demonstrate that claudin-1 might play a role in airway remodeling in asthmatic patients by means of regulation of ASM cell proliferation, angiogenesis, and inflammation.

Interleukins, from 1 to 37, and interferon-γ: receptors, functions, and roles in diseases.

Advancing our understanding of mechanisms of immune regulation in allergy, asthma, autoimmune diseases, tumor development, organ transplantation, and chronic infections could lead to effective and targeted therapies. Subsets of immune and inflammatory cells interact via ILs and IFNs; reciprocal regulation and counter balance among T(h) and regulatory T cells, as well as subsets of B cells, offer opportunities for immune interventions. Here, we review current knowledge about ILs 1 to 37 and IFN-γ. Our understanding of the effects of ILs has greatly increased since the discoveries of monocyte IL (called IL-1) and lymphocyte IL (called IL-2); more than 40 cytokines are now designated as ILs. Studies of transgenic or knockout mice with altered expression of these cytokines or their receptors and analyses of mutations and polymorphisms in human genes that encode these products have provided important information about IL and IFN functions. We discuss their signaling pathways, cellular sources, targets, roles in immune regulation and cellular networks, roles in allergy and asthma, and roles in defense against infections.

TNF-like weak inducer of apoptosis (TWEAK) and TNF-α cooperate in the induction of keratinocyte apoptosis.

BACKGROUND: Activation of skin keratinocytes followed by their apoptotic death leads to eczema and spongiosis formations in patients with atopic dermatitis (AD). TNF-like weak inducer of apoptosis (TWEAK) binds to its receptor, fibroblast growth factor-inducible 14 (Fn14), and controls many cellular activities, including proliferation, migration, differentiation, apoptosis, angiogenesis, and inflammation. OBJECTIVE: The aim of the study was to investigate the role of TWEAK and Fn14 in the formation of eczema in patients with AD. METHODS: Primary keratinocytes were isolated from nonlesional skin from patients with AD and psoriasis and from normal skin of healthy donors. Apoptosis analysis was performed by using annexin V/7-aminoactinomycin D and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling staining. The expression and regulation of TWEAK, TNF-α, Fn14, TNF receptor (TNFR) 1, and TNFR2 were measured by means of RT-PCR, flow cytometric analysis, and ELISA. TWEAK and Fn14 expression of lesional AD and psoriatic skin and normal control skin was analyzed by using immunohistochemistry and immunofluorescence. RESULTS: TWEAK and TNF-α cooperate in the induction of apoptosis in primary keratinocytes obtained from patients with AD, patients with psoriasis, and healthy subjects and in artificial skin equivalents. TNFR1 and Fn14 were the main receptors involved. TWEAK upregulates TNF-α expression in primary keratinocytes, whereas TNF-α did not affect the expression of TWEAK and its receptors. High TWEAK expression was observed in AD lesions but not in psoriatic lesions or normal skin. Fn14 was highly expressed in the lesional skin of patients with AD and patients with psoriasis and in healthy control skin. CONCLUSION: The high expression of TWEAK in lesional AD skin contributes to the difference in keratinocyte apoptosis and lesional formation between AD and psoriasis.

IL-32 is expressed by human primary keratinocytes and modulates keratinocyte apoptosis in atopic dermatitis.

BACKGROUND: Keratinocyte (KC) apoptosis is an important mechanism of eczema and spongiosis in patients with atopic dermatitis (AD) and is mediated by IFN-gamma, which is secreted by T(H)1 cells. IL-32 is a proinflammatory cytokine that is involved in the inflammatory processes of rheumatoid arthritis, chronic obstructive pulmonary disease, and Crohn disease. Recently, it was shown that upregulation of IL-32 induces apoptosis. OBJECTIVE: The aim of the study was to investigate the expression and function of IL-32 in patients with AD. METHODS: The expression of IL-32 in KCs was analyzed by means of RT-PCR, ELISA, and flow cytometry. Transfections of small interfering RNA were performed in primary KCs, and apoptosis was analyzed by means of terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling, annexin-V, and 7-amino actinomycin D stainings. Immunofluorescence stainings were used to detect IL-32 in skin biopsy specimens, and serum levels of IL-32 were analyzed by means of ELISA. RESULTS: We report that IL-32 is expressed in human primary KCs on stimulation with IFN-gamma, TNF-alpha, and T(H)1 cells in contrast to T(H)2, regulatory T (Treg), or T(H)17 cells, which showed no effect. Transfection of primary KCs and artificial skin equivalents with small interfering RNA to IL-32, which resulted in a clear decrease in IL-32 expression, significantly reduced KC apoptosis. Immunofluorescence staining demonstrated that IL-32 was expressed in AD lesional skin, whereas it was present in neither skin biopsy specimens from healthy donors nor in lesional skin from patients with psoriasis. Serum levels of IL-32 from patients with AD correlated with disease severity, but increased serum levels of IL-32 were also detected in asthmatic patients. CONCLUSION: The present study demonstrates KCs as a source of IL-32, which modulates KC apoptosis and contributes to the pathophysiology of AD.

Dual nature of T cell-epithelium interaction in chronic rhinosinusitis.

BACKGROUND: T-cell infiltration of submucosa, release of proinflammatory cytokines leading to epithelial activation, and contributions to inflammation are observed in chronic rhinosinusitis (CRS). OBJECTIVES: Molecular mechanisms and kinetics of T-cell interaction with sinus epithelium leading to activation followed by subsequent apoptosis of epithelial cells were the focus of the current study. METHODS: Primary human sinus epithelial cells and T cells generated from sinus tissues of healthy individuals and patients with CRS with or without allergy and sinus tissue biopsies were characterized in terms of activation (surface marker expression, cytokine production via real-time PCR, confocal microscopy, ELISA) and apoptosis (annexin V/7-amino-actinomycin D staining, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay, receptor expression by flow cytometry, confocal microscopy) of epithelial cells. RESULTS: Primary human sinus epithelial cells isolated from patients with CRS were at an activated state with upregulated expression of HLA-DR, IFN-gamma-inducible protein 10, monokine induced by IFN-gamma, and TNF-related apoptosis-inducing ligand (TRAIL) compared with healthy individuals. The expressions of these chemokines, HLA-DR, TRAIL, and TNF receptor 2 were significantly induced by IFN-gamma, whereas TRAIL receptor 4 was downregulated. Epithelial cells started to undergo apoptosis 48 hours after IFN-gamma stimulation when the transcription of proinflammatory cytokines and chemokines decreased to initial levels. The essential factors for sinus epithelial apoptosis were T(H)1 cells and IFN-gamma. Epithelial apoptosis was enhanced by Fas-Fas-ligand and TRAIL-TRAIL receptor 2 interactions. Remarkable apoptosis of epithelial cells and shedding was observed in CRS in situ. CONCLUSION: Epithelial cell interaction with activated T cells is a biphasic phenomenon in CRS. Initially activated T cells lead to activation and induction of proinflammatory functions of epithelial cells, and thereafter their apoptotic death, resulting in no more contribution to inflammation, takes place.

Differentiation and functional analysis of human T(H)17 cells.

BACKGROUND: T(H)17 cells are of pathologic relevance in autoimmune disorders and presumably also in allergy and asthma. Regulatory T (Treg) cells, in contrast, suppress inflammatory and allergen-driven responses. Despite these disparate functions, both T-cell subsets have been shown to be dependent on TGF-beta for their development. OBJECTIVE: The aim of the study was to analyze the differentiation and function of human T(H)17 cells in comparison with other T(H) cell subsets. METHODS: Naive human CD4(+) T cells were differentiated in vitro, and gene expression was analyzed by means of quantitative real-time PCR, ELISA, and immunofluorescence. The function of T(H) cell subsets was assessed by monitoring the response of primary bronchial epithelial cells in coculture experiments. RESULTS: In vitro differentiated T(H)17 cells differ from Treg and other T(H) cells in their potency to induce IL-6 and IL-1beta expression in primary bronchial epithelial cells. TGF-beta, IL-1beta, IL-6, and IL-23 are necessary during T(H)17 cell differentiation to acquire these functions, including IL-17 production. In contrast, TGF-beta alone is necessary and sufficient to induce the transcription factor RORC2. This transcription factor, previously thought to be specific for T(H)17 cells, is also expressed in Treg cells, CD25(+) cells, cytotoxic T cells, and natural killer T cells. CONCLUSION: This study demonstrates mechanisms of differentiation to human T(H)17 cells, a subset that effectively and uniquely modulates the function of primary bronchial epithelial cells.