Asthma is a chronic allergic inflammatory disease of lung. The initiation and progression of asthma is dependent on the cytokines interleukin (IL) -4 and IL-13 acting through related receptor complexes. Disease pathogenesis is effected by intracellular signaling pathways that couple primarily to specific motifs within the intracellular domain of the IL-4 receptor alpha chain (IL-4R alpha), a subunit that is common to the IL-4 and IL-13 receptor complexes. Neutralizing anti-cytokine strategies have proven to be highly successful on dissecting relevant effector pathways in experimental allergic disease, and are now entering clinical trials in human allergic disorders. Although there have been only a few clinical studies on the effects of cytokine modulators in asthma, this line of research and development appears promising.
Asthma* ; Cytokines ; Humans ; Interleukin-13 ; Interleukin-4 ; Interleukins ; Lung ; Receptors, Interleukin-13 ; Receptors, Interleukin-4

Idiopathic interstitial pneumonia (IIP) is characterized by varying degrees of interstitial fibrosis. IL-13 and IL-4 are strong inducers of tissue fibrosis, whereas IFN-gamma has antifibrotic potential. However, the roles of these substances in IIP remain unknown. IL-13, IL-4, and IFN-gamma were measured in the BAL fluid of 16 idiopathic pulmonary fibrosis (IPF) patients, 10 nonspecific interstitial pneumonia (NSIP) patients, and 8 normal controls. The expression of IL-13 and IL-13Ralpha1/alpha2 in lung tissues was analyzed using ELISA and immunohistochemistry. IL-13 levels were significantly higher in IPF patients than the others (P<0.05). IL-4 levels were higher in both IPF and NSIP patients than in normal controls (P<0.05), and IFN-gamma levels were lower in NSIP patients than in normal controls (P=0.047). IL-13 levels correlated inversely with FVC% (r=-0.47, P=0.043) and DLCO% (r=-0.58, P=0.014) in IPF and NSIP patients. IL-13 was strongly expressed in the smooth muscle, bronchial epithelium, alveolar macrophages and endothelium of IPF patients. IL-13Ralpha1, rather than IL-13Ralpha2, was strongly expressed in the smooth muscle, bronchial epithelium, and endothelium of IPF patients. IL-13 and its receptors may contribute to the pathogenesis of fibrosis in IIP and appear to be related to the severity of the disease.
Adult ; Female ; Humans ; Idiopathic Interstitial Pneumonias/diagnosis/*metabolism ; Idiopathic Pulmonary Fibrosis/diagnosis/*metabolism ; Interferon-gamma/analysis ; Interleukin-13/*analysis ; Interleukin-13 Receptor alpha1 Subunit/*metabolism ; Interleukin-13 Receptor alpha2 Subunit/*metabolism ; Interleukin-4/analysis ; Lung/physiopathology ; Male ; Middle Aged

BACKGROUND: Efficacy and safety of tiotropium bromide, a muscarinic receptor antagonist, in treatment of asthma have been reported. However, its effect on airway remodeling in chronic asthma of the elderly has not been clearly verified. The objective of this study was to investigate the effect of tiotropium and expression of muscarinic receptors as its related mechanism in an aged mouse model of chronic asthma with airway remodeling. METHODS: BALB/c female mice age 6 weeks, 9 and 15 months were sensitized and challenged with ovalbumin (OVA) for three months. Tiotropium bromide was administered during the challenge period. Airway hyperresponsiveness (AHR) and pulmonary inflammation were measured. Parameters of airway remodeling, and expression levels of M2 and M3 receptors were examined. RESULTS: Total cell with eosinophils, increased in the OVA groups by age, was decreased significantly after treatment with tiotropium bromide, particularly in the age group of 15 months. AHR and levels of interleukin (IL)-4, IL-5, and IL-13 were decreased, after tiotropium administration. In old aged group of 9- and 15-months-treated groups, hydroxyproline contents and levels of α-smooth muscle actin were attenuated. Tiotropium enhanced the expression of M2 but decreased expression of M3 in all aged groups of OVA. CONCLUSION: Tiotropium bromide had anti-inflammatory and anti-remodeling effects in an aged mouse model of chronic asthma. Its effects seemed to be partly mediated by modulating expression M3 and M2 muscarinic receptors. Tiotropium may be a beneficial treatment option for the elderly with airway remodeling of chronic asthma.
Actins ; Aged ; Airway Remodeling ; Animals ; Asthma* ; Eosinophils ; Female ; Humans ; Hydroxyproline ; Interleukin-13 ; Interleukin-5 ; Interleukins ; Mice* ; Ovalbumin ; Ovum ; Pneumonia ; Receptors, Muscarinic* ; Tiotropium Bromide*

Therapy for inflammatory bowel disease (IBD) has changed, with several new agents being evaluated. The era of anti-tumor necrosis factor (anti-TNF) antibody therapy saw remarkable progress in IBD therapy. Some patients, however, do not respond to anti-TNF treatment, or their response decreases over time. This phenomenon highlights the need to identify new molecular targets for therapy in IBD. The targets of new therapeutic molecules in IBD must aim to restore immune dysregulation by the inhibition of proinflammatory cytokines (TNF-α, interleukin [IL]-6, IL-13, IL-17, IL-18, and IL-21) and augmentation of the effect of anti-inflammatory cytokines (IL-10, IL-11, and transforming growth factor β) and to pursue new anti-inflammatory targets, such as regulatory T-cell therapy, Smad7 antisense, Janus-activated kinase inhibition, Toll-like receptor stimulation, leukocyte adhesion, and blockade of T-cell homing via integrins and mucosal addressin cellular adhesion molecule-1. In addition, potential molecular targets could restore mucosal barrier function and stimulate mucosal healing. Despite these potential targets, the value and clinical significance of most new molecules remain unclear, and clinical efficacy and safety must be better defined before their implementation in clinical practice. This article aims to review the promising and emerging molecular targets that could be clinically meaningful for novel therapeutic approaches.
Biological Products* ; Colitis, Ulcerative ; Crohn Disease ; Cytokines ; Humans ; Inflammatory Bowel Diseases* ; Integrins ; Interleukin-11 ; Interleukin-13 ; Interleukin-17 ; Interleukin-18 ; Interleukins ; Leukocytes ; Necrosis ; Phosphotransferases ; T-Lymphocytes ; Toll-Like Receptors ; Transforming Growth Factors ; Treatment Outcome

PURPOSE: Asthma is a pulmonary chronic inflammatory disease characterized by airway obstruction and hyperresponsiveness. Pattern recognition receptors are known to play a key role in the development of allergic diseases as well as host defenses against microbial infection. Receptor interacting protein 2 (RIP2), a serine/threonine kinase, is an adaptor molecule of NOD1 and NOD2, and genetic variation in this receptor is known to be associated with the severity of allergic asthma in children. In this study, we examined the role of RIP2 in the development of allergic airway inflammation in a mouse model. METHODS: Airway inflammation was induced in mice through intranasal administration of ovalbumin (OVA) after 2 intraperitoneal immunizations with OVA. Lung inflammation and mucus hypersecretion were examined histologically and total cell infiltration in bronchoalveolar (BAL) fluids was determined. Levels of the Th2-related cytokines, IL-5 and IL-13, in lung extracts were measured by ELISA. Serum antigen-specific IgE and IgG1 levels were also assessed. RESULTS: OVA-induced lung inflammation and mucus hypersecretion were not different between WT and RIP2-deficient mice. The IL-5 and IL-13 levels in the bronchoalveolar (BAL) fluids were also not impaired in RIP2-deficient mice compared to WT mice. Moreover, RIP2 deficiency did not affect serum OVA-specific IgG1 and IgE levels. CONCLUSIONS: Our results suggest that RIP2 is not associated with the development of allergic airway inflammation.
Administration, Intranasal ; Airway Obstruction ; Animals ; Asthma ; Child ; Cytokines ; Enzyme-Linked Immunosorbent Assay ; Genetic Variation ; Humans ; Immunization ; Immunoglobulin E ; Immunoglobulin G ; Inflammation* ; Interleukin-13 ; Interleukin-5 ; Lung ; Methods ; Mice* ; Mucus ; Ovalbumin ; Ovum ; Phosphotransferases ; Pneumonia ; Receptors, Pattern Recognition

OBJECTIVETo study the effects of IL-13 on the differentiation and expression of transcription factor c-fos of human erythroleukemia cell line (HEL) cells.

METHODSReverse transcription polymerase chain reaction (RT-PCR) was used to observe the mRNA expression of IL-13 receptor a 1, GP i b, vWF and c-fos, and Western blot and cytometry were used to analyse their protein expression.

RESULTSIL-13 receptor a 1 was expressed on HEL cells. IL-13 (100 ng/ml ) up-regulated the mRNA expression of GP II b and vWF. The ratio of luminous absorption (LA) of GP I b to p-actin bands ( AB) was 1. 303 in control group, whereas was 2. 912 in experiment group; being 2. 23-fold higher than that in control group (P < 0. 05). The ratio of LA to AB for vWF was 0.217 in control group, and 0. 506 in experiment group; indicating a 2. 33-fold increase in experiment group (P <0. 05). The protein expression of GP I b and vWF was significantly increased in experiment group, compared with that in control group. IL-13 inducing the increased expression of c-fos mRNA and protein of HEL cells peaked at 30 min and 60 min, respectively. The ratio of LA to AB for c-fos was also increased at 30 min and 60 min (P <0. 05).

CONCLUSIONIL-13 prompts the differentiation of HEL cells and up-regulates the expression of c-fos.

Cell Differentiation ; drug effects ; Cell Line, Tumor ; Flow Cytometry ; Humans ; Interleukin-13 ; pharmacology ; Leukemia, Erythroblastic, Acute ; metabolism ; Platelet Membrane Glycoprotein IIb ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-fos ; biosynthesis ; genetics ; RNA, Messenger ; biosynthesis ; Receptors, Interleukin-13 ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Up-Regulation ; von Willebrand Factor ; biosynthesis ; genetics

IL-4 and IL-13 are closely related cytokines that are produced by Th2 cells. However, IL-4 and IL-13 have different effects on the development of asthma phenotypes. Here, we evaluated downstream molecular mechanisms involved in the development of Th2 type asthma phenotypes. A murine model of Th2 asthma was used that involved intraperitoneal sensitization with an allergen (ovalbumin) plus alum and then challenge with ovalbumin alone. Asthma phenotypes, including airway-hyperresponsiveness (AHR), lung inflammation, and immunologic parameters were evaluated after allergen challenge in mice deficient in candidate genes. The present study showed that methacholine AHR and lung inflammation developed in allergen-challenged IL-4-deficient mice but not in allergen-challenged IL-13-deficient mice. In addition, the production of OVA-specific IgG2a and IFN-gamma-inducible protein (IP)-10 was also impaired in the absence of IL-13, but not of IL-4. Lung-targeted IFN-gamma over-expression in the airways enhanced methacholine AHR and non-eosinophilic inflammation; in addition, these asthma phenotypes were impaired in allergen-challenged IFN-gamma-deficient mice. Moreover, AHR, non-eosinophilic inflammation, and IFN-gamma expression were impaired in allergen-challenged IL-12Rbeta2- and STAT4-deficient mice; however, AHR and non-eosinophilic inflammation were not impaired in allergen-challenged IL-4Ralpha-deficient mice, and these phenomena were accompanied by the enhanced expression of IL-12 and IFN-gamma. The present data suggest that IL-13-mediated asthma phenotypes, such as AHR and non-eosinophilic inflammation, in the Th2 type asthma are dependent on the IL-12-STAT4-IFN-gamma axis, and that these asthma phenotypes are independent of IL-4Ralpha-mediated signaling.
Allergens/immunology ; Animals ; Asthma/complications/*immunology/pathology/physiopathology ; Bronchial Hyperreactivity/complications/immunology/pathology ; Disease Models, Animal ; Interferon-gamma/*immunology ; Interleukin-12/*immunology ; Interleukin-12 Receptor beta 2 Subunit/metabolism ; Interleukin-13/deficiency/*immunology ; Interleukin-4/deficiency ; Methacholine Chloride ; Mice ; Mice, Transgenic ; Models, Immunological ; Organ Specificity ; Pneumonia/complications/immunology/pathology ; Receptors, Cell Surface/metabolism ; STAT4 Transcription Factor/*metabolism ; Signal Transduction/*immunology ; Th2 Cells/*immunology

BACKGROUND/AIMS: Inhaled corticosteroids are the most effective treatment currently available for asthma, but their beneficial effect against airway remodeling is limited. The tyrosine kinase inhibitor nilotinib has inhibitory activity against c-kit and the platelet-derived growth factor receptor. We compared the effects of fluticasone and nilotinib on airway remodeling in a chronic asthma model. We also examined whether co-treatment with nilotinib and fluticasone had any synergistic effect in preventing airway remodeling. METHODS: We developed a mouse model of airway remodeling, including smooth muscle thickening, in which ovalbumin (OVA)-sensitized female BALB/c-mice were repeatedly exposed to intranasal OVA administration twice per week for 3 months. Mice were treated with fluticasone and/or nilotinib intranasally during the OVA challenge. RESULTS: Mice chronically exposed to OVA developed eosinophilic airway inflammation and showed features of airway remodeling, including thickening of the peribronchial smooth muscle layer. Both fluticasone and nilotinib attenuated airway smooth muscle thickening. However, only nilotinib suppressed fibrotic changes, demonstrating inhibition of collagen deposition. Fluticasone reduced pro-inflammatory cells, such as eosinophils, and several cytokines, such as interleukin 4 (IL-4), IL-5, and IL-13, induced by repeated OVA challenges. On the other hand, nilotinib reduced transforming growth factor β1 levels in bronchoalveolar lavage fluid and inhibited fibroblast proliferation significantly. CONCLUSIONS: These results suggest that fluticasone and nilotinib suppressed airway remodeling in this chronic asthma model through anti-inflammatory and anti-fibrotic pathways, respectively.
Adrenal Cortex Hormones ; Airway Remodeling ; Animals ; Asthma* ; Bronchoalveolar Lavage Fluid ; Collagen ; Cytokines ; Eosinophils ; Female ; Fibroblasts ; Fluticasone* ; Hand ; Humans ; Inflammation ; Interleukin-13 ; Interleukin-4 ; Interleukin-5 ; Mice ; Muscle, Smooth ; Ovalbumin ; Ovum ; Protein-Tyrosine Kinases ; Receptors, Platelet-Derived Growth Factor ; Transforming Growth Factors

Interleukin-4 (IL-4) is a pleiotropic cytokine which plays a pivotal role in shaping immune responses. IL-4 mediates important proinflammatory functions in asthma, including the IgE isotype switch. IL-4 exerts its biological effects through binding to its receptor (IL-4R) complex, with the alpha chain as the high affinity binding subunit. Soluble IL-4R lacks the transmemberane and cytoplasmic domains, so it cannot induce cellular activation. By acting as a decoy to circulating IL-4 and neutralize its activity, its high specificity and affinity make it ideal as an IL-4 antagonist. Some companies have embarked the clinical research for asthma treatment with the sIL-4R and the result revealed well therapeutic effect. With RNA extracted from human monocyte as the template, The sIL-4R cDNA encoding the extracellular domain of IL-4R a chain was obtained by RT-PCR. Compared with the sIL-4R encoding sequence in GenBank, the nucleotide sequencing analysis indicated that there was a A-->G mutation at 148bp and the mutation caused Ile-->Val at 50th amino acid. According to the references, numerous polymorphisms have been identified in the IL-4R gene and the Ile50Val was the only known extracellular variant of human IL-4R. Then the recombinant vector pPIC9K/sIL-4R was constructed, linearized and introduced into Pichia pastoris GS115 by electroporation. The recombinant sIL-4R was identified by SDS-PAGE, Western blot and Ligand binding blot. The SDS-PAGE and Western blot analysis showed that the apparent molecular weight of expressed sIL-4R was about 30kD. And the Ligand binding blot analysis indicated the expressed sIL-4R had the biological activity. The sIL-4R, which had the biological activity, was successfully secretorily expressed in the Pichia pastoris (GS 115).
Genetic Vectors ; genetics ; Humans ; Interleukin-4 Receptor alpha Subunit ; biosynthesis ; genetics ; Pichia ; genetics ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics

BACKGROUND: Interleukin (IL)-4 is a pleiotropic cytokine that plays an important role in the pathogenesis of the allergic inflammation and asthma. Upon IL-4 receptor (IL-4R) engagement, a variety of signaling mediators, such as JAK kinases and STAT-6 are activated, leading to induction of IL-4 target gene expression including CD23 and germline C epsilon transcription. The function of a membrane-proximal domain of IL-4Ra, termed ID-1, remains to be characterized to date. OBJECTIVE: To assess whether the ID-1 domain mediates the induction of IL-4 target gene expression in a STAT-6-dependent manner. METHODS: The intracellular region of IL-4Ralpha was translationally fused to the extracellular region of IL-2Rbeta to provide ligand specificity to IL-2. Acidic amino acids and serine residues in the ID-1 domain of the chimeric receptor were substituted by site-directed mutagenesis. These receptor cDNAs were stably transfected to M12.4.1 murine B lymphoma cells. Following IL-2 stimulation, wild type and mutant clones for the ID-1 motif were subjected to FACS. RNA blotting and elecroporetic mobility shift assays to address the levels of CD23, germline C epsilon and STAT-6 inductions, respectively. RESULTS: ID-1 mutant clones were defective in gene induction of CD23 and germline C epsilon in response to IL-2 stimulation, as compared with wildtype clones. Moreover, IL-2-mediated STAT-6 activation was abolished in ID-1 mutant clones. CONCLUSION: These results demonstrate that the ID-1 domain of IL-4Ra is essential to induce IL-4 target gene expression through a STAT-6-dependent pathway.
Amino Acids, Acidic ; Asthma ; Clone Cells ; DNA, Complementary ; Electrophoretic Mobility Shift Assay ; Gene Expression ; Inflammation ; Interleukin-2 ; Interleukin-4 Receptor alpha Subunit* ; Interleukin-4* ; Interleukins ; Janus Kinases ; Lymphoma ; Mutagenesis, Site-Directed ; Receptors, Interleukin-4 ; RNA ; Sensitivity and Specificity ; Serine