Immunoglobulin G4-related sialadenitis (IgG4-RS) is an immune-mediated fibro-inflammatory disease and the pathogenesis is still not fully understood. The aim of this study was to explore the role and mechanism of interleukin-13 (IL-13) in the cellular senescence during the progress of IgG4-RS. We found that the expression of IL-13 and IL-13 receptor α1 (IL-13Rα1) as well as the number of senescent cells were significantly higher in the submandibular glands (SMGs) of IgG4-RS patients. IL-13 directly induced senescence as shown by the elevated activity of senescence-associated β-galactosidase (SA-β-gal), the decreased cell proliferation, and the upregulation of senescence markers (p53 and p16) and senescence-associated secretory phenotype (SASP) factors (IL-1β and IL-6) in SMG-C6 cells. Mechanistically, IL-13 increased the level of phosphorylated signal transducer and activator of transcription 6 (p-STAT6) and mitochondrial-reactive oxygen species (mtROS), while decreased the mitochondrial membrane potential, ATP level, and the expression and activity of superoxide dismutase 2 (SOD2). Notably, the IL-13-induced cellular senescence and mitochondrial dysfunction could be inhibited by pretreatment with either STAT6 inhibitor AS1517499 or mitochondria-targeted ROS scavenger MitoTEMPO. Moreover, IL-13 increased the interaction between p-STAT6 and cAMP-response element binding protein (CREB)-binding protein (CBP) and decreased the transcriptional activity of CREB on SOD2. Taken together, our findings revealed a critical role of IL-13 in the induction of salivary gland epithelial cell senescence through the elevated mitochondrial oxidative stress in a STAT6-CREB-SOD2-dependent pathway in IgG4-RS.
Cellular Senescence/genetics* ; Humans ; Immunoglobulin G/metabolism* ; Interleukin-13/pharmacology* ; Mitochondria/metabolism* ; Sialadenitis/metabolism*

OBJECTIVE: To investigate the effect of interleukin (IL) -13 combined with cold stimulation on synthesis and secretion of mucin (MUC) 5AC in human bronchial epithelial cell line 16HBE and explore the role of transient receptor potential 8 (TRPM8) and anti-apoptotic factor B-cell lymphoblast-2 (Bcl-2) in this process. METHODS: 16HBE cells were stimulated with 10 ng/mL IL-13, 1 mmol/L menthol, or both (1 mmol/L menthol was added after 6 days of IL-13 stimulation), and the changes in the expression of MUC5AC, intracellular Ca RESULTS: The mRNA and protein expressions of MUC5AC increased significantly in 16HBE cells following stimulation with IL-13, menthol, and both ( CONCLUSIONS Menthol combined with IL-13 produces a synergistic effect to promote the synthesis and secretion of MUC5AC in 16HBE cells possibly by activating TRPM8 receptor to upregulate the expression of Bcl-2.
Bronchi ; Epithelial Cells/drug effects* ; Humans ; Interleukin-13 ; Menthol/pharmacology* ; Mucin 5AC

Objective To investigate the effect of calcitonin gene-related peptide (CGRP) on the regulation of group 2 innate lymphoid cells (ILC2) in the peripheral blood of patients with allergic rhinitis (AR). Methods Peripheral blood mononuclear cells (PBMCs) were extracted from normal healthy individuals and AR patients, then stimulated with CGRP, interleukin 33 (IL-33) and CGRP combined with IL-33 for 3 days, with blank stimulus as control. The percentage of ILC2 in the four groups was measured by flow cytometry. After being sorted, ILC2 was given to CGRP, IL-33 and CGRP combined with IL-33 stimulation for 3 days, with blank stimulus as control. The percentage of IL-5 and IL-13 positive cells in ILC2 was detected by flow cytometry, and the levels of IL-5 and IL-13 in ILC2 supernatant were measured by ELISA. Results The percentage of ILC2 in the peripheral blood of AR patients was significantly higher than that of the control group. The levels of IL-5⁺ILC2 and IL-13⁺ILC2 were significantly increased by IL-33 single stimulation after culturing PBMCs. After adding IL-33 combined with CGRP stimulation, the levels of IL-5⁺ILC2 and IL-13⁺ILC2 in PBMCs were significantly reduced; after CGRP single stimulation, the levels of IL-5⁺ILC2 and IL-13⁺ILC2 in PBMCs were further decreased. After ILC2 was sorted and cultured, the levels of IL-5⁺ILC2 and IL-13⁺ILC2 showed significant increase after IL-33 single stimulation. The levels of IL-5⁺ILC2 and IL-13⁺ILC2 were decreased by IL-33 and CGRP co-stimulation, and they were further reduced after CGRP single stimulation. Compared to IL-33 single stimulation, IL-5 and IL-13 levels dropped significantly due to the IL-33 and CGRP co-stimulation. The levels of IL-5 and IL-13 were further reduced by CGRP single stimulation. Conclusion CGRP inhibits the proliferation and activation of peripheral blood ILC2 in AR and exert anti-inflammatory effects in AR.
Humans ; Calcitonin Gene-Related Peptide/pharmacology* ; Leukocytes, Mononuclear ; Immunity, Innate ; Interleukin-33/pharmacology* ; Interleukin-13 ; Lymphocytes ; Interleukin-5/pharmacology* ; Rhinitis, Allergic ; Cell Proliferation

OBJECTIVETo explore the effect of nuclear factor-kappa B (NF-κB) decoy oligodeoxynucleotides (ODN) on serum NF-κB, IL-10, IL-13 and pulmonary NF-κB protein expression in rabbits with severe lung contusion.

METHODSForty New Zealand rabbits were randomly divided into severe lung contusion group (group A, n=12), lung contusion with NF-κB scrambled decoy ODN group (group B, n=12), lung contusion with sense NF-κB decoy ODN group (group C, n=12), and normal control group (n=4). After establishment of the contusion injury model, the sense and scrambled NF-κB decoy ODN were infused into the rabbits via the jugular vein accordingly. Serum NF-κB, IL-10, and IL-13 and NF-κB protein expressions in the lung tissue were detected before and at 1, 2, 3, and 4 h after the contusion.

RESULTSTwo hours after sense NF-κB decoy ODN intervention, the expression of NF-κB began to decrease and reached the lowest level at 3 h; pulmonary IL-10 and IL-13 expressions decreased at 1 h after contusion, to the lowest level at 2 and 4 h, respectively. After sense NF-κB decoy ODN intervention, the expression of IL-10 and IL-13 increased and NF-κB protein expression decreased significantly in comparison with those in groups A and B (P<0.01).

CONCLUSIONSense NF-κB decoy ODN can significantly reduce the serum NF-κB expression, increase serum IL-10 and IL-13 levels and decrease pulmonary NF-κB protein expression in the early stages after severe lung contusion in rabbits.

Animals ; Contusions ; blood ; Female ; Interleukin-10 ; blood ; Interleukin-13 ; blood ; Lung ; drug effects ; metabolism ; Lung Injury ; blood ; NF-kappa B ; blood ; Oligodeoxyribonucleotides ; pharmacology ; Rabbits

OBJECTIVETo investigate the dynamic changes of a group of cytokines in phosgene-induced lung injury and the function of different dose of ulinastatin through animal experiment.

METHODS104 male SD rats were randomly assigned into the control group, ulinastatin control group, phosgene treatment groups and different dose of ulinastatin intervention groups, 8 rats each group. Treatment groups were dynamic constant exposure in phosgene, and immediately injected ulinastatin intraperitoneal, and then the experimental animal, the lung tissue biopsy, lung wet/dry ratio, RT-PCR detection, the plasma for detection of Bio-Plex 18 cytokines.

RESULTSCompared with the control group, plasma concentrations of IL-1α, IL-6, GM-CSF, TNF-α, INF-γ, MIP-3α, VEGF were increased significantly first (2 h), and gradually decreased with the passage of time , the difference was statistically significant (P < 0.05). Plasma concentrations of IL-4, IL-10 were decreased earlier (2h, 6 h) and increased later (24 h) (P < 0.05). The change of plasma concentration of IL-13 was not obvious earlier (2 h) and still rising later (24h), the difference was statistically significant (P < 0.05). After drug intervention, the levels of pro-inflammatory cytokines declined and the levels of anti-inflammatory cytokines raise by different degrees at different times in ulinastatin intervention groups, the difference was statistically significant. The degree of lung injury was improved than the phosgene treatment groups and better in high dose of ulinastatin intervention group. The expression of IL-10, IL-4, IL-13-mRNA of tissue increased in accordance with plasma results.

CONCLUSIONA group of cytokines are dynamicly changed in phosgene-induced lung injury by time. High dose of ulinastatin can improved phosgene-induced lung injury, regulate the synthesis and release of inflammatory cytokines and inhibit inflammatory react in a dose-dependent manner.

Animals ; Cytokines ; metabolism ; Glycoproteins ; pharmacology ; Granulocyte-Macrophage Colony-Stimulating Factor ; Interleukin-10 ; Interleukin-13 ; Interleukin-1alpha ; Interleukin-4 ; Interleukin-6 ; Lung ; Lung Injury ; chemically induced ; drug therapy ; Male ; Phosgene ; toxicity ; Rats ; Rats, Sprague-Dawley ; Tumor Necrosis Factor-alpha

BACKGROUNDExpression of murine calcium-activated chloride channel family member 3 (mCLCA3) has been reported to be increased in the airway epithelium of asthmatic mice challenged with ovalbumin (OVA). However, its role in asthmatic airway inflammation under no OVA exposure has not yet been clarified.

METHODSmCLCA3 plasmids were transfected into the airways of normal BALB/c mice. mCLCA3 expression and airway inflammation in mouse lung tissue were evaluated. Cell differentials and cytokines in bronchoalveolar lavage fluid (BALF) were analyzed. The expression of mCLCA3 protein and mucus protein mucin-5 subtype AC (MUC5AC) were analyzed by Western blotting. The mRNA levels of mCLCA3, MUC5AC and interleukin-13 (IL-13) were determined quantitatively.

RESULTSmCLCA3 expression was not detected in the control group while strong immunoreactivity was detected in the OVA and mCLCA3 plasmid groups, and was strictly localized to the airway epithelium. The numbers of inflammatory cells in lung tissue and BALF were increased in both mCLCA3 plasmid and OVA groups. The protein and mRNA levels of mCLCA3 and MUC5AC in the lung tissue were significantly increased in the mCLCA3 plasmid and OVA groups compared to the control group. The level of IL-13, but not IL-4, IL-5, IFN-γ, CCL2, CCL5 or CCL11, was significantly increased compared with control group in BALF in the mCLCA3 plasmid and OVA groups. The level of IL-13 in the BALF in the mCLCA3 plasmid group was much higher than that in the OVA group (P < 0.05). The level of mCLCA3 mRNA in lung tissue was positively correlated with the levels of MUC5AC mRNA in lung tissue, IL-13 mRNA in lung tissue, the number of eosinophils in BALF, and the content of IL-13 protein in BALF. The level of IL-13 mRNA in lung tissue was positively correlated with the number of eosinophils in BALF and the level of MUC5AC mRNA in lung tissue.

CONCLUSIONThese findings suggest that increased expression of a single-gene, mCLCA3, could simulate an asthma attack, and its mechanism may involve mCLCA3 overexpression up-regulating IL-13 expression.

Allergens ; Animals ; Asthma ; Chloride Channels ; Female ; Inflammation ; chemically induced ; metabolism ; Interleukin-13 ; metabolism ; Interleukin-4 ; genetics ; metabolism ; Interleukin-5 ; genetics ; metabolism ; Mice ; Mice, Inbred BALB C ; Mucin 5AC ; genetics ; metabolism ; Ovalbumin ; pharmacology

OBJECTIVETo investigate the effects of polyinosinic-polycytidylic acid (polyI:C) on the production of thymic stromal lymphopoietin (TSLP) and airway inflammation in mice with exacerbated asthma induced by respiratory syncytial virus (RSV).

METHODSThirty-two female BALB/c mice were randomly divided into 4 groups, namely the PBS control group, OVA group, OVA/RSV group, and OVA/RSV/polyI:C group. In the latter 3 groups, the mice were sensitized by OVA and stimulated with nebulized OVA. RSV was inoculated into the nasal cavity of the sensitized mice and polyI:C (1 mg/kg) was intramuscularly administered. The airway response to metacholine was examined, and the serum levels of IL-4, IL-5, IL-13, and IFN-γ and TSLP in the supernatants of bronchoalveolar lavage fluid (BALF) were detected using ELISA. The total BALF cells, eosinophils, lymphocytes and neutrophils were counted. The lung specimens were collected to observe the inflammation with HE staining, and immunohistochemistry was employed to determine TSLP production in the airway epithelial cells.

RESULTSThe mice in RSV/OVA/polyI:C group showed a significantly lower airway responsiveness to metacholine than those in OVA/RSV group (P<0.01). Compared with OVA/RSV group, RSV/OVA/polyI:C group showed significantly lower serum levels of IL-4, IL-5, IL-13 and TSLP in BALF (P<0.05), with also lower total BALF cells, eosinophils and lymphocytes (P<0.05) and lessened infiltration of the airway inflammatory cells. Immunohistochemistry of TSLP also demonstrated a lower production of TSLP in the airway epithelial cells in RSV/OVA/polyI:C group than in OVA/RSV group.

CONCLUSIONSpolyI:C can inhibit the increase in TSLP production in the airway epithelial cells after RSV infection and relieve airway inflammation in mice with RSV-induced asthma exacerbation.

Animals ; Asthma ; blood ; metabolism ; virology ; Bronchoalveolar Lavage Fluid ; Cytokines ; secretion ; Female ; Inflammation ; pathology ; Interleukin-13 ; blood ; Interleukin-4 ; blood ; Interleukin-5 ; blood ; Mice ; Mice, Inbred BALB C ; Poly I-C ; pharmacology ; Respiratory Syncytial Virus Infections ; blood ; metabolism ; Respiratory Syncytial Viruses

OBJECTIVETo study the effect of 1,25-(OH)2D3 on lipopolysaccharide (LPS)-induced expression of interleukin-13 (IL-13) and interleukin-17 (IL-17) in cord blood CD4(+)T cells, providing theoretical basis for clinical reasonable application of vitamin D and prevention of asthma and allergic diseases.

METHODSMononuclear cells (MNCs) were isolated from umbilical cord blood (50 mL) of 12 normal eutocia term newborns by gravity centrifugation. CD4(+)T cells were isolated using magnetic beads, which was cultured with following three kinds of stimulus for 72 hours: natural state (blank group), LPS (10 μg/mL)stimulation alone and LPS(10 μg/mL)+1,25-(OH)2D3 (10(-8) mmol/L)stimulation. Levels of IL-13 and IL-17 in the culture supernatant and mRNA expressions in cord blood CD4(+)T cells were detected using ELISA and real Time-PCR respectively.

RESULTSCompared with the blank group, levels of IL-13 and IL-17 in the culture supernatant and mRNA expression of IL-13 and IL-17 in the cord blood CD4(+)T cells increased in the LPS stimulation alone group (P<0.01). When co-stimulation of 1,25-(OH)2D3 with LPS, levels of IL-13 and IL-17 in the culture supernatant and mRNA expression of IL-13 and IL-17 in the cord blood CD4(+)T cells decreased compared with LPS-stimulated alone group (P<0.05), but remained higher than the blank group (P<0.01).

CONCLUSIONSLPS can promote expression of IL-13 and IL-17 in cord blood CD4(+)T cells. 1,25-(OH)2D3 inhibits the expression of IL-13 and IL-17, suggesting that vitamin D intake may provide protective effects in the development of atopy-predisposing immune responses in early life.

Asthma ; drug therapy ; immunology ; CD4-Positive T-Lymphocytes ; immunology ; Calcitriol ; pharmacology ; Female ; Fetal Blood ; immunology ; Humans ; Infant, Newborn ; Interleukin-13 ; blood ; genetics ; Interleukin-17 ; blood ; genetics ; Lipopolysaccharides ; pharmacology ; Male ; RNA, Messenger ; blood

PURPOSE: The aim of this study was to investigate whether pathologic changes in zonula occludens-1 (ZO-1) are induced by interleukin-13 (IL-13) in the experimental minimal-change nephrotic syndrome (MCNS) model and to determine whether montelukast, a leukotriene receptor antagonist, has an effect on ZO-1 restoration in cultured human podocytes. MATERIALS AND METHODS: Human podocytes cultured on bovine serum albumin-coated plates were treated with different doses of IL-13 and montelukast and then examined for distribution using confocal microscopy and for ZO-1 protein levels using Western blotting. RESULTS: ZO-1 was internalized and shown to accumulate in the cytoplasm of human podocytes in an IL-13 dose-dependent manner. High doses (50 and 100 ng/mL) of IL-13 decreased the levels of ZO-1 protein at 12 and 24 h (both p<0.01; n=3), which were significantly reversed by a high dose (0.5 microM) montelukast treatment (p<0.01; n=3). CONCLUSION: Our results suggest that IL-13 alters the expression of ZO-1, and such alterations in the content and distribution of ZO-1 may be relevant in the pathogenesis of proteinuria in the MCNS model.
Acetates/*pharmacology ; Blotting, Western ; Dose-Response Relationship, Drug ; Humans ; Interleukin-13/*pharmacology ; Leukotriene Antagonists/*pharmacology ; Microscopy, Confocal ; Podocytes/*drug effects/metabolism ; Proteinuria/pathology ; Quinolines/*pharmacology ; Tight Junctions ; Zonula Occludens-1 Protein/*metabolism

PURPOSE: The aim of this study was to investigate whether pathologic changes in zonula occludens-1 (ZO-1) are induced by interleukin-13 (IL-13) in the experimental minimal-change nephrotic syndrome (MCNS) model and to determine whether montelukast, a leukotriene receptor antagonist, has an effect on ZO-1 restoration in cultured human podocytes. MATERIALS AND METHODS: Human podocytes cultured on bovine serum albumin-coated plates were treated with different doses of IL-13 and montelukast and then examined for distribution using confocal microscopy and for ZO-1 protein levels using Western blotting. RESULTS: ZO-1 was internalized and shown to accumulate in the cytoplasm of human podocytes in an IL-13 dose-dependent manner. High doses (50 and 100 ng/mL) of IL-13 decreased the levels of ZO-1 protein at 12 and 24 h (both p<0.01; n=3), which were significantly reversed by a high dose (0.5 microM) montelukast treatment (p<0.01; n=3). CONCLUSION: Our results suggest that IL-13 alters the expression of ZO-1, and such alterations in the content and distribution of ZO-1 may be relevant in the pathogenesis of proteinuria in the MCNS model.
Acetates/*pharmacology ; Blotting, Western ; Dose-Response Relationship, Drug ; Humans ; Interleukin-13/*pharmacology ; Leukotriene Antagonists/*pharmacology ; Microscopy, Confocal ; Podocytes/*drug effects/metabolism ; Proteinuria/pathology ; Quinolines/*pharmacology ; Tight Junctions ; Zonula Occludens-1 Protein/*metabolism