RGS4 controls airway hyperresponsiveness through GAP-independent mechanisms.

Regulators of G protein signaling (RGS) proteins constrain G protein-coupled receptor (GPCR)-mediated and other responses throughout the body primarily, but not exclusively, through their GTPase-activating protein activity. Asthma is a highly prevalent condition characterized by airway hyper-responsiveness (AHR) to environmental stimuli resulting in part from amplified GPCR-mediated airway smooth muscle contraction. Rgs2 or Rgs5 gene deletion in mice enhances AHR and airway smooth muscle contraction, whereas RGS4 KO mice unexpectedly have decreased AHR because of increased production of the bronchodilator prostaglandin E2 (PGE2) by lung epithelial cells. Here, we found that knockin mice harboring Rgs4 alleles encoding a point mutation (N128A) that sharply curtails RGS4 GTPase-activating protein activity had increased AHR, reduced airway PGE2 levels, and augmented GPCR-induced bronchoconstriction compared with either RGS4 KO mice or WT controls. RGS4 interacted with the p85α subunit of PI3K and inhibited PI3K-dependent PGE2 secretion elicited by transforming growth factor beta in airway epithelial cells. Together, these findings suggest that RGS4 affects asthma severity in part by regulating the airway inflammatory milieu in a G protein-independent manner.

Livin promotes Th2-type immune response in airway allergic diseases.

OBJECTIVES: To investigate the effects of livin on the Th2 immune response in airway allergic diseases (AAD) and explore the interaction among livin, GATA3, IL-4 in peripheral blood CD4+ T cells of AAD patients. METHODS: WT mice and livin KO mice were developed for model of AAD. Th2 cell levels in the lung tissues and spleen were assessed by flow cytometry. Also, it was assessed in the culture after exposing to livin inhibitor (Lp-15); the protein and mRNA levels of livin, GATA3 and IL-4 in peripheral blood CD4+ T cells isolated from patients with or without AAD were measured by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting, respectively. Finally, Co-immunoprecipitation (Co-IP) was employed to identify the interaction between livin and GATA3. RESULTS: Compared with WT mouse, Th2 cell frequency in lung tissues and spleen was significantly decreased in livin KO mouse; after adding Lp-15, the differentiation from Naive CD4+T cells in spleen to Th2 cells was blocked; the protein and mRNA levels of livin, GATA3 and IL-4 in AAD group were higher than that in control group. The levels of livin were positively correlated with IL-4, and GATA3 was also positively correlated with IL-4 and livin. GATA3 was detected in the protein complex co-precipitated with livin antibody, and livin was also detected in the protein complex co-precipitated by GATA3 antibody. CONCLUSION: Livin increases the expression of IL-4 and facilitates naive CD4+ T cells to differentiate into Th2 cells, which triggers airway allergy.

Polysaccharides from Platycodonis Radix ameliorated respiratory syncytial virus-induced epithelial cell apoptosis and inflammation through activation of miR-181a-mediated Hippo and SIRT1 pathways.

Respiratory syncytial virus (RSV) is the leading cause of bronchiolitis in young children, but there are few safe and effective treatments for this disease. Platycodonis Radix is widely used as an antitussive and expectorant drug for preventing various diseases in lower respiratory tract, in which the polysaccharides are one of the main bioactivity constituents. In this study, the protective effects of the P. Radix polysaccharides (PRP) against RSV-induced bronchiolitis in juvenile mice and RSV-induced apoptosis of epithelial HEp-2 cells were investigated. The results showed that PRP obviously decreased the levels of IL-1ß, IL-4, IL-6, TNF-α, IFN-γ and TSLP in lung tissues, and reduced the number of inflammatory cells in bronchoalveolar lavage fluid (BALF) of RSV-infected mice. Furthermore, it reduced the apoptosis of RSV-infected HEp-2 cells and remarkably inhibited the mRNA expressions of RSV L gene, which indicated that PRP affected transcription and replication of RSV in host cells. Compared with that in RSV-infected group, miR-181a-5p in the PRP-treated group presented the highest relative abundance and its expression was violently reduced by approximately 30%. Mechanistically, PRP had the similar effects as miR-181a-5p antagomir on RSV-induced apoptosis and inflammation in HEp-2 cells via upregulating BCL2, MLL3 and SIRT1, which could be reversed by miR-181a-5p mimic. Therefore, it demonstrated that PRP not only protected against RSV-induced lung inflammation in mice but also inhibited apoptosis of RSV-infected HEp-2 cells via suppressing miR-181a-5p and transcriptionally activating Hippo and SIRT1 pathways.

DRP1-Mediated Mitochondrial Fission Regulates Lung Epithelial Response to Allergen.

Mitochondria regulate a myriad of cellular functions. Dysregulation of mitochondrial control within airway epithelial cells has been implicated in the pro-inflammatory response to allergens in asthma patients. Because of their multifaceted nature, mitochondrial structure must be tightly regulated through fission and fusion. Dynamin Related Protein 1 (DRP1) is a key driver of mitochondrial fission. During allergic asthma, airway epithelial mitochondria appear smaller and structurally altered. The role of DRP1-mediated mitochondrial fission, however, has not been fully elucidated in epithelial response to allergens. We used a Human Bronchial Epithelial Cell line (HBECs), primary Mouse Tracheal Epithelial Cells (MTECs), and conditional DRP1 ablation in lung epithelial cells to investigate the impact of mitochondrial fission on the pro-inflammatory response to house dust mite (HDM) in vitro and in vivo. Our data suggest that, following HDM challenge, mitochondrial fission is rapidly upregulated in airway epithelial cells and precedes production of pro-inflammatory cytokines and chemokines. Further, deletion of Drp1 in lung epithelial cells leads to decreased fission and enhanced pro-inflammatory signaling in response to HDM in vitro, as well as enhanced airway hyper-responsiveness (AHR), inflammation, differential mucin transcription, and epithelial cell death in vivo. Mitochondrial fission, therefore, regulates the lung epithelial pro-inflammatory response to HDM.

Mechanisms of Immunotoxicity: Stressors and Evaluators.

The immune system defends the body against certain tumor cells and against foreign agents such as fungi, parasites, bacteria, and viruses. One of its main roles is to distinguish endogenous components from non-self-components. An unproperly functioning immune system is prone to primary immune deficiencies caused by either primary immune deficiencies such as genetic defects or secondary immune deficiencies such as physical, chemical, and in some instances, psychological stressors. In the manuscript, we will provide a brief overview of the immune system and immunotoxicology. We will also describe the biochemical mechanisms of immunotoxicants and how to evaluate immunotoxicity.

Contribution of Regulatory T Cell Methylation Modifications to the Pathogenesis of Allergic Airway Diseases.

Regulatory T (Treg) cells are a subtype of CD4+ T cells that play a significant role in the protection from autoimmunity and the maintenance of immune tolerance via immune regulation. Epigenetic modifications of Treg cells (i.e., cytosine methylation at the promoter region of the transcription factor, Forkhead Box P3) have been found to be closely associated with allergic diseases, including allergic rhinitis, asthma, and food allergies. In this study, we highlighted the recent evidence on the contribution of epigenetic modifications in Treg cells to the pathogenesis of allergic diseases. Moreover, we also discussed directions for future clinical treatment approaches, with a particular emphasis on Treg cell-targeted therapies for allergic disorders.

GLCCI1 reduces collagen deposition and airway hyper-responsiveness in a mouse asthma model through binding with WD repeat domain 45B.

Asthma is a serious public health problem worldwide, without effective therapeutic methods. Our previous study indicated that glucocorticoid-induced transcript 1 gene (GLCCI1) knockout reduces the sensitivity to glucocorticoid in asthmatic mouse. Here, we explored the role and action mechanism of GLCCI1 in asthma development. In ovalbumin-sensitized mice, airway resistance and tissue damage increased, the production of inflammatory cytokines were up-regulated, GLCCI1 expression was reduced and autophagy was activated. Increasing of GLCCI1 inhibited human and mouse airway epithelial cell (AEC) autophagy, while decreasing of GLCCI1 promoted autophagy. Furthermore, we found that GLCCI1 bound with WD repeat domain 45B (WDR45B) and inhibited its expression. Increasing of WDR45B partly reversed the inhibition of GLCCI1 to autophagy-related proteins expression and autophagosome formation in vitro. Increasing of WDR45B in vivo reversed the improvement of GLCCI1 on airway remodelling in asthma and the inhibition to autophagy level in lung tissues. Overall, our data showed that GLCCI1 improved airway remodelling in ovalbumin-sensitized mice through inhibiting autophagy via combination with WDR45B and inhibiting its expression. Our results proved a new idea for asthma treatment.

Requirement of Gαi1 and Gαi3 in interleukin-4-induced signaling, macrophage M2 polarization and allergic asthma response.

IL-4 induces Akt activation in macrophages, required for full M2 (alternative) polarization. We examined the roles of Gαi1 and Gαi3 in M2 polarization using multiple genetic methods. Methods and Results: In MEFs and primary murine BMDMs, Gαi1/3 shRNA, knockout or dominant negative mutations attenuated IL-4-induced IL4Rα endocytosis, Gab1 recruitment as well as Akt activation, leaving STAT6 signaling unaffected. Following IL-4 stimulation, Gαi1/3 proteins associated with the intracellular domain of IL-4Rα and the APPL1 adaptor, to mediate IL-4Rα endosomal traffic and Gab1-Akt activation in BMDMs. In contrast, gene silencing of Gαi1/3 with shRNA or knockout resulted in BMDMs that were refractory to IL-4-induced M2 polarization. Conversely, Gαi1/3-overexpressed BMDMs displayed preferred M2 response with IL-4 stimulation. In primary human macrophages IL-4-induced Akt activation and Th2 genes expression were inhibited with Gαi1/3 silencing, but augmented with Gαi1/3 overexpression. In Gαi1/3 double knockout (DKO) mice, M2 polarization, by injection of IL-4 complex or chitin, was potently inhibited. Moreover, in a murine model of asthma, ovalbumin-induced airway inflammation and hyperresponsiveness were largely impaired in Gαi1/3 DKO mice. Conclusion: These findings highlight novel and essential roles for Gαi1/3 in regulating IL-4-induced signaling, macrophage M2 polarization and allergic asthma response.

Fetomaternal immune cross talk modifies T-cell priming through sustained changes to DC function.

BACKGROUND: Prenatal exposure to infections can modify immune development. These environmental disturbances during early life potentially alter the incidence of inflammatory disorders as well as priming of immune responses. Infection with the helminth Schistosoma mansoni is widely studied for its ability to alter immune responsiveness and is associated with variations in coinfection, allergy, and vaccine efficacy in endemic populations. OBJECTIVE: Exposure to maternal schistosomiasis during early life, even without transmission of infection, can result in priming effects on offspring immune responses to bystander antigenic challenges as related to allergic responsiveness and vaccination, with this article seeking to further clarify the effects and underlying immunologic imprinting. METHODS: Here, we have combined a model of chronic maternal schistosomiasis infection with a thorough analysis of subsequent offspring immune responses to allergy and vaccination models, including viral challenge and steady-state changes to immune cell compartments. RESULTS: We have demonstrated that maternal schistosomiasis alters CD4+ responses during allergic sensitization and challenge in a skewed IL-4/B-cell-dominant response to antigenic challenge associated with limited inflammatory response. Beyond that, we have uncovered previously unidentified alterations to CD8+ T-cell responses during immunization that are dependent on vaccine formulation and have functional impact on the efficacy of vaccination against viral infection in a murine hepatitis B virus model. CONCLUSION: In addition to steady-state modifications to CD4+ T-cell polarization and B-cell priming, we have traced these modified CD8+ responses to an altered dendritic cell phenotype sustained into adulthood, providing evidence for complex priming effects imparted by infection via fetomaternal cross talk.

The Cxxc1 subunit of the Trithorax complex directs epigenetic licensing of CD4+ T cell differentiation.

Different dynamics of gene expression are observed during cell differentiation. In T cells, genes that are turned on early or turned off and stay off have been thoroughly studied. However, genes that are initially turned off but then turned on again after stimulation has ceased have not been defined; they are obviously important, especially in the context of acute versus chronic inflammation. Using the Th1/Th2 differentiation paradigm, we found that the Cxxc1 subunit of the Trithorax complex directs transcription of genes initially down-regulated by TCR stimulation but up-regulated again in a later phase. The late up-regulation of these genes was impaired either by prolonged TCR stimulation or Cxxc1 deficiency, which led to decreased expression of Trib3 and Klf2 in Th1 and Th2 cells, respectively. Loss of Cxxc1 resulted in enhanced pathogenicity in allergic airway inflammation in vivo. Thus, Cxxc1 plays essential roles in the establishment of a proper CD4+ T cell immune system via epigenetic control of a specific set of genes.

Increased IL-6 and Potential IL-6 trans-signalling in the airways after an allergen challenge.

BACKGROUND: In asthma, IL-6 is a potential cause of enhanced inflammation, tissue damage and airway dysfunction. IL-6 signalling is regulated by its receptor, which is composed of two proteins, IL-6R and GP130. In addition to their membrane form, these two proteins may be found as extracellular soluble forms. The interaction of IL-6 with soluble IL-6R (sIL-6R) can trigger IL-6 trans-signalling in cells lacking IL-6R. Conversely, the soluble form of GP130 (sGP130) competes with its membrane form to inhibit IL-6 trans-signalling. OBJECTIVES: We aimed to analyse IL-6 trans-signalling proteins in the airways of subjects after an allergen challenge. METHODS: We used a model of segmental bronchoprovocation with an allergen (SBP-Ag) in human subjects with allergy. Before and 48 h after SBP-Ag, bronchoalveolar lavages (BALs) allowed for the analysis of proteins in BAL fluids (BALFs) by ELISA, and membrane proteins on the surface of BAL cells by flow cytometry. In addition, we performed RNA sequencing (RNA-seq) and used proteomic data to further inform on the expression of the IL-6R subunits by eosinophils, bronchial epithelial cells and lung fibroblasts. Finally, we measured the effect of IL-6 trans-signalling on bronchial fibroblasts, in vitro. RESULTS: IL-6, sIL-6R, sGP130 and the molar ratio of sIL-6R/sGP130 increased in the airways after SBP-Ag, suggesting the potential for enhanced IL-6 trans-signalling activity. BAL lymphocytes, monocytes and eosinophils displayed IL-6R on their surface and were all possible providers of sIL-6R, whereas GP130 was highly expressed in bronchial epithelial cells and lung fibroblasts. Finally, bronchial fibroblasts activated by IL-6 trans-signalling produced enhanced amounts of the chemokine, MCP-1 (CCL2). CONCLUSION AND CLINICAL RELEVANCE: After a bronchial allergen challenge, we found augmentation of the elements of IL-6 trans-signalling. Allergen-induced IL-6 trans-signalling activity can activate fibroblasts to produce chemokines that can further enhance inflammation and lung dysfunction.

Induction of Airway Hypersensitivity to Ovalbumin and Dust Mite Allergens as Mouse Models of Allergic Asthma.

Mouse models of allergic asthma have been utilized to establish the role of T helper type 2 (Th2) cells in driving lung inflammation, airway hyperresponsiveness, and obstruction. Here, we present the allergic asthma models, in which mice are hypersensitized to ovalbumin (OVA) and house dust mite (HDM). These models mimic the major characteristics of human asthma including the eosinophilic inflammation and hyperactivity of the airway, overproduction of Th2 cytokines in the lung, and elevated total and allergen-specific immunoglobulin E (IgE) in serum.

IL-4Rα signaling in CD4+CD25+FoxP3+ T regulatory cells restrains airway inflammation via limiting local tissue IL-33.

Impaired tolerance to innocuous particles during allergic asthma has been linked to increased plasticity of FoxP3+ regulatory T cells (Tregs) reprogramming into pathogenic effector cells, thus exacerbating airway disease. However, failure of tolerance mechanisms is driven by Th2 inflammatory signals. Therefore, the in vivo role of canonical IL-4 receptor α (IL-4Rα) signaling, an essential driver of Th2-type airway responses to allergens, on the regulatory function of FoxP3+ Tregs in allergic asthma was explored. Here, we used transgenic Foxp3cre IL-4Rα-/lox and littermate control mice to investigate the role of IL-4 and IL-13 signaling via Tregs in house dust mite-induced (HDM-induced) allergic airway disease. We sensitized mice intratracheally on day 0, challenged them on days 6-10, and analyzed airway hyperresponsiveness (AHR), airway inflammation, mucus production, and cellular profile on day 14. In the absence of IL-4Rα responsiveness on FoxP3+ Tregs, exacerbated AHR and airway inflammation were shown in HDM-sensitized mice. Interestingly, reduced induction of FoxP3+ Tregs accompanied increased IL-33 alarmin production and type 2 innate lymphoid cell activation in the lung, exacerbating airway hyperreactivity and lung eosinophilia. Taken together, our findings indicate that IL-4Rα-unresponsive FoxP3+ Tregs result in exaggerated innate Th2-type, IL-33-dependent airway inflammation and a break in tolerance during allergic asthma.

Environmental Factor-Mediated Transgenerational Inheritance of Igf2r Hypomethylation and Pulmonary Allergic Response via Targeting Dendritic Cells.

The developmental origin of allergic diseases has been suggested, but the molecular basis remains enigmatic. Exposure to environmental factors, such as di-(2-ethylhexyl) phthalate (DEHP; a common plasticizer), is suggested to be associated with increased childhood allergic asthma, but the causal relationship and its underlying mechanism remain unknown. This study explored the transgenerational mechanism of DEHP on allergic asthma and dendritic cell (DC) homeostasis through epigenetic modification. In a murine model, ancestral exposure of C57BL/6 mice to low-dose DEHP led to trans-generational promoter hypomethylation of the insulin-like growth factor 2 receptor (Igf2r), concomitant with enhanced Igf2r expression and increased apoptosis prominently in CD8α+ DCs upon ligand stimulation, with consequent reduction in their IL-12 secretion and subsequent T cell-derived IFN-γ, thereby promoting a default Th2-associated pulmonary allergic response. Increased apoptosis was also noted in circulating IGF2Rhigh human DCs. Further, in human placenta, the methylation level at the orthologous IGF2R promoter region was shown to be inversely correlated with the level of maternal DEHP intake. These results support the importance of ancestral phthalate exposure in conferring the trans-generational risk of allergic phenotypes, featuring hypo-methylation of the IGF2R gene and dysregulated DC homeostasis.

The Impact of Interleukin (IL)-33 Gene Polymorphisms and Environmental Factors on Risk of Asthma in the Iranian Population.

BACKGROUND: Airway epithelial cells secrete Interleukin-33 in response to the different allergens. Several single nucleotide polymorphisms (SNP) of this cytokine have been reported to be involved in the development of asthma. We conducted this study to evaluate the impact of the two most common SNPs of the IL-33 gene (rs1342326 and rs3939286) and environmental factors on the susceptibility to asthma in the Iranian population. SUBJECTS AND METHODS: In this study, we enrolled 126 asthmatics patients and 300 age, sex-matched controls. Genotyping was performed by real-time PCR using the TaqMan SNP genotyping assay. Moreover, total serum IgE level, eosinophil count, and skin prick test were accomplished and complete history was taken from all the participants. RESULTS: The frequencies of mutant genotypes in both SNPs were significantly higher in asthmatics than controls. C/C genotype of rs1342326 [OR (95% CI) 2.50 (1.33-4.69)] and A/A genotype of rs3939286 [OR (95% CI) 2.18 (1.05-4.52)] were associated with higher risk of asthma development. While A/C+C/C genotype of rs1342326 was more prevalent in mild asthma [OR (95% CI) 2.36 (1.14-4.89)], G/A+A/A genotype of rs3939286 was associated with increased risk of moderate and severe asthma [OR (95% CI) 2.53 (1.30-4.94)]. CONCLUSION: This study revealed that both IL-33 SNPs were associated with an increased risk of asthma. The rs1342326 was associated with atopic, mild and adult-onset asthma and a higher level of eosinophils in peripheral blood. However, rs3939286 was more frequent in moderate and severe asthma. Moreover, rs3939286 was associated with non-atopic and childhood-onset asthma.

Particulate matters induce acute exacerbation of allergic airway inflammation via the TLR2/NF-κB/NLRP3 signaling pathway.

BACKGROUND: Exposure to particulate matters (PMs) can lead to an acute exacerbation of allergic airway diseases, increasing the severity of symptoms and mortality. However, little is known about the underlying molecular mechanism. This study aimed to investigate the effects of PMs on acute exacerbation of allergic airway inflammation and seek potential therapeutic targets. METHODS: Non-allergic control and ovalbumin (OVA)-allergic wide-type (WT) and Toll-like receptor 2 knockout (Tlr2-/-) mice were exposed to 100 µg of PM (diameter 5.85 µm) or saline by the oropharyngeal instillation. The responses were examined three days after exposure. In the RAW264.7 macrophage cell line, Tlr2 was knocked down by small-interfering RNA or the NF-κB inhibitor JSH-23 was used, and then the cells were stimulated with PMs for 12 h before comparison of the inflammatory responses. RESULTS: PM exposure led to increased inflammatory cell recruitment and airway intensity of PAS + staining in OVA-allergic WT mice, accompanied with an accumulation of inflammatory cells and elevated inflammatory cytokines, such as IL-6 and IL-18, in the bronchoalveolar lavage fluid (BALF). Furthermore, the protein levels of TLR2 and the NLRP3 inflammasome were elevated concomitantly with the airway inflammation post-OVA/PMs challenge. Tlr2 deficiency effectively inhibited the airway inflammation, including pulmonary inflammatory cell recruitment, mucus secretion, serum OVA-specific immunoglobulin E (IgE), and BALF inflammatory cytokine production. Additionally, the P-induced NLRP3 activation in the RAW 264.7 cell line was diminished by the knockdown of Tlr2 or JSH-23 treatment in vitro. CONCLUSION: Our results indicated that PMs exacerbate the allergic airway inflammation mediated by the TLR2/ NF-κB/NLRP3 signaling pathway. Inhibition of NF-κB seems to be a possible treatment.

TNFR2 Signaling Enhances ILC2 Survival, Function, and Induction of Airway Hyperreactivity.

Group 2 innate lymphoid cells (ILC2s) can initiate pathologic inflammation in allergic asthma by secreting copious amounts of type 2 cytokines, promoting lung eosinophilia and airway hyperreactivity (AHR), a cardinal feature of asthma. We discovered that the TNF/TNFR2 axis is a central immune checkpoint in murine and human ILC2s. ILC2s selectively express TNFR2, and blocking the TNF/TNFR2 axis inhibits survival and cytokine production and reduces ILC2-dependent AHR. The mechanism of action of TNFR2 in ILC2s is through the non-canonical NF-κB pathway as an NF-κB-inducing kinase (NIK) inhibitor blocks the costimulatory effect of TNF-α. Similarly, human ILC2s selectively express TNFR2, and using hILC2s, we show that TNFR2 engagement promotes AHR through a NIK-dependent pathway in alymphoid murine recipients. These findings highlight the role of the TNF/TNFR2 axis in pulmonary ILC2s, suggesting that targeting TNFR2 or relevant signaling is a different strategy for treating patients with ILC2-dependent asthma.

Targeted deletion of NFAT-Interacting-Protein-(NIP) 45 resolves experimental asthma by inhibiting Innate Lymphoid Cells group 2 (ILC2).

Here we investigated the role of NFAT-interacting protein (NIP)-45, an Interleukin (IL)-4 inducing Transcription Factor, and its impact on the differentiation of Group 2 Innate -Lymphoid -Cells (ILC2s) in the pathogenesis of asthma. NIP45, a transcription factor regulating NFATc1 activity, mRNA was found to be induced in the Peripheral Blood mononuclear cells (PMBCs) of asthmatic pre-school children with allergies and in the peripheral blood CD4+ T cells from adult asthmatic patients. In PBMCs of asthmatic and control children, NIP45 mRNA directly correlated with NFATc1 but not with T-bet. Targeted deletion of NIP45 in mice resulted in a protective phenotype in experimental asthma with reduced airway mucus production, airway hyperresponsiveness and eosinophils. This phenotype was reversed by intranasal delivery of recombinant r-IL-33. Consistently, ILC2s and not GATA3+ CD4+ T-cells were decreased in the lungs of asthmatic NIP45-/- mice. Reduced cell number spleen ILC2s could be differentiated from NIP45-/- as compared to wild-type mice after in vivo injection of a microcircle-DNA vector expressing IL-25 and decreased cytokines and ILC2 markers in ILC2 differentiated from the bone marrow of NIP45-/- mice. NIP45 thus emerges as a new therapeutic target for the resolution of the airway pathology, down-regulation of ILC2s and mucus production in asthma.

Empagliflozin and Dulaglutide are Effective against Obesity-induced Airway Hyperresponsiveness and Fibrosis in A Murine Model.

Patients with asthma with obesity experience severe symptoms, are unresponsive to conventional asthma treatment, and lack proper pharmacotherapy. Empagliflozin and dulaglutide, developed for diabetes, reduce weight, decrease insulin resistance, and exert additive effects. We evaluated the efficacy of empagliflozin, dulaglutide, and their combination on obesity-induced airway hyperresponsiveness (AHR) and lung fibrosis using a murine model. We assigned C57BL/6J mice to five groups: control, high-fat diet (HFD), and HFD with empagliflozin, dulaglutide, or both. Mice received a 12-week HFD, empagliflozin (5 days/week, oral gavage), and dulaglutide (once weekly, intraperitoneally). Both drugs significantly attenuated HFD-induced weight increase, abnormal glucose metabolism, and abnormal serum levels of leptin and insulin, and co-treatment was more effective. Both drugs significantly alleviated HFD-induced AHR, increased macrophages in bronchoalveolar lavage fluid (BALF), and co-treatment was more effective on AHR. HFD-induced lung fibrosis was decreased by both drugs alone and combined. HFD induced interleukin (IL)-17, transforming growth factor (TGF)-ß1, and IL-1ß mRNA and protein expression, which was significantly reduced by empagliflozin, dulaglutide, and their combination. Tumour necrosis factor (TNF)-α and IL-6 showed similar patterns without significant differences. HFD-enhanced T helper (Th) 1 and Th17 cell differentiation was improved by both drugs. Empagliflozin and dulaglutide could be a promising therapy for obesity-induced asthma and showed additive effects in combination.

DNA methylation and the hygiene hypothesis: connecting respiratory allergy and childhood acute lymphoblastic leukemia.

Aim: The hygiene hypothesis states that a lack of infection in early-life suppresses immune system development, and is linked to respiratory allergy (RA) and childhood acute lymphoblastic leukemia (ALL) risk. Little is known about underlying mechanisms, but DNA methylation is altered in RA and ALL, and in response to infection. We investigated if aberrant methylation may be in common between these diseases and associated with infection. Materials & methods: RA and ALL disease-associated methylation signatures were compared and related to exposure-to-infection signatures. Results: A significant number of genes overlapped between RA and ALL signatures (p = 0.0019). Significant overlaps were observed between exposure-to-infection signatures and disease-associated signatures. Conclusion: DNA methylation may be a mediating mechanism through which the hygiene hypothesis is associated with RA and ALL risk.