Recent Developments in the Role of Histone Acetylation in Asthma.

BACKGROUND: Epigenetic modifications are known to mediate both beneficial and unfavorable effects of environmental exposures on the development and clinical course of asthma. On the molecular level, epigenetic mechanisms participate in multiple aspects of the emerging and ongoing asthma pathology. SUMMARY: Studies performed in the last several years expand our knowledge on the role of histone acetylation, a classical epigenetic mark, in the regulation of (patho)physiological processes of diverse cells playing a central role in asthma, including those belonging to the immune system (e.g., CD4+ T cells, macrophages) and lung structure (e.g., airway epithelial cells, pulmonary fibroblasts). Those studies demonstrate a number of specific histone acetylation-associated mechanisms and pathways underlying pathological processes characteristic for asthma, as well as report their modification modalities. KEY MESSAGES: Dietary modulation of histone acetylation levels in the immune system might protect against the development of asthma and other allergies. Interfering with the enzymes controlling the histone acetylation status of structural lung and (local) immune cells might provide future therapeutic options for asthmatics. Despite some methodological obstacles, analysis of the histone acetylation levels might improve asthma diagnostics.

Diverse functions of apolipoprotein A-I in lung fibrosis.

Apolipoprotein A-I (apoA-I) mediates reverse cholesterol transport (RCT) out of cells. In addition to its important role in the RTC, apoA-I also possesses anti-inflammatory and antioxidative functions including the ability to activate inflammasome and signal via toll-like receptors. Dysfunctional apoA-I or its low abundance may cause accumulation of cholesterol mass in alveolar macrophages, leading to the formation of foam cells. Increased numbers of foam cells have been noted in the lungs of mice after experimental exposure to cigarette smoke, silica, or bleomycin and in the lungs of patients suffering from different types of lung fibrosis, including idiopathic pulmonary fibrosis (IPF). This suggests that dysregulation of lipid metabolism may be a common event in the pathogenesis of interstitial lung diseases. Recognition of the emerging role of cholesterol in the regulation of lung inflammation and remodeling provides a challenging concept for understanding lung diseases and offers novel and exciting avenues for therapeutic development. Accordingly, a number of preclinical studies demonstrated decreased expression of inflammatory and profibrotic mediators and preserved lung tissue structure following the administration of the apoA-I or its mimetic peptides. This review highlights the role of apoA-I in lung fibrosis and provides evidence for its potential use in the treatment of this pathological condition.

The emerging role of extracellular vesicles as communicators between adipose tissue and pathologic lungs with a special focus on asthma.

Extracellular vesicles (EVs) gain increasing attention due to their (patho-)physiological role in intercellular signaling, specifically in the communication between distant organs. Recent studies highlight a connection between the adipose tissue (AT) and the lung via (immuno-)modulatory EVs in disorders such as obesity-associated asthma and lung cancer-associated cachexia. Although lung cancer-derived EVs induce lipolysis and myotube atrophy in vivo, pathogenic effects were also reported in the opposite direction with the involvement of AT-derived EVs in cancer-promoting responses and potentially in asthma development. In contrast, the majority of studies on AT-derived EVs demonstrate their protective influence on the asthmatic lung. Beneficial effects, such as induction of anti-inflammatory pathways in vitro and in ovalbumin (OVA)-induced asthma mouse models, were particularly conveyed by EVs enriched from AT-derived mesenchymal stem/stromal cells (AT-MSCs), which therefore pose an interesting subject in possible future therapeutic applications. Likewise, AT-MSC-derived EVs exerted beneficial effects in several other pulmonary abnormalities, such as different types of lung injury or pathological changes related to chronic obstructive pulmonary disease. These contradictory findings highlight the need for extensive research to widen the understanding of the role of EVs in the development of diseases and interconnectivity between organs.

Identification of extracellular vesicle microRNA signatures specifically linked to inflammatory and metabolic mechanisms in obesity-associated low type-2 asthma.

RATIONALE AND OBJECTIVE: Plasma extracellular vesicles (EVs) represent a vital source of molecular information about health and disease states. Due to their heterogenous cellular sources, EVs and their cargo may predict specific pathomechanisms behind disease phenotypes. Here we aimed to utilize EV microRNA (miRNA) signatures to gain new insights into underlying molecular mechanisms of obesity-associated low type-2 asthma. METHODS: Obese low type-2 asthma (OA) and non-obese low type-2 asthma (NOA) patients were selected from an asthma cohort conjointly with healthy controls. Plasma EVs were isolated and characterised by nanoparticle tracking analysis. EV-associated small RNAs were extracted, sequenced and bioinformatically analysed. RESULTS: Based on EV miRNA expression profiles, a clear distinction between the three study groups could be established using a principal component analysis. Integrative pathway analysis of potential target genes of the differentially expressed miRNAs revealed inflammatory cytokines (e.g., interleukin-6, transforming growth factor-beta, interferons) and metabolic factors (e.g., insulin, leptin) signalling pathways to be specifically associated with OA. The miR-17-92 and miR-106a-363 clusters were significantly enriched only in OA. These miRNA clusters exhibited discrete bivariate correlations with several key laboratory (e.g., C-reactive protein) and lung function parameters. Plasma EV miRNA signatures mirrored blood-derived CD4+ T-cell transcriptome data, but achieved an even higher sensitivity in identifying specifically affected biological pathways. CONCLUSION: The identified plasma EV miRNA signatures and particularly the miR-17-92 and -106a-363 clusters were capable to disentangle specific mechanisms of the obesity-associated low type-2 asthma phenotype, which may serve as basis for stratified treatment development.

Intranasal administration of Acinetobacter lwoffii in a murine model of asthma induces IL-6-mediated protection associated with cecal microbiota changes.

BACKGROUND: Early-life exposure to certain environmental bacteria including Acinetobacter lwoffii (AL) has been implicated in protection from chronic inflammatory diseases including asthma later in life. However, the underlying mechanisms at the immune-microbe interface remain largely unknown. METHODS: The effects of repeated intranasal AL exposure on local and systemic innate immune responses were investigated in wild-type and Il6-/- , Il10-/- , and Il17-/- mice exposed to ovalbumin-induced allergic airway inflammation. Those investigations were expanded by microbiome analyses. To assess for AL-associated changes in gene expression, the picture arising from animal data was supplemented by in vitro experiments of macrophage and T-cell responses, yielding expression and epigenetic data. RESULTS: The asthma preventive effect of AL was confirmed in the lung. Repeated intranasal AL administration triggered a proinflammatory immune response particularly characterized by elevated levels of IL-6, and consequently, IL-6 induced IL-10 production in CD4+ T-cells. Both IL-6 and IL-10, but not IL-17, were required for asthma protection. AL had a profound impact on the gene regulatory landscape of CD4+ T-cells which could be largely recapitulated by recombinant IL-6. AL administration also induced marked changes in the gastrointestinal microbiome but not in the lung microbiome. By comparing the effects on the microbiota according to mouse genotype and AL-treatment status, we have identified microbial taxa that were associated with either disease protection or activity. CONCLUSION: These experiments provide a novel mechanism of Acinetobacter lwoffii-induced asthma protection operating through IL-6-mediated epigenetic activation of IL-10 production and with associated effects on the intestinal microbiome.

Editorial of Special Issue "Molecular Mechanisms of Allergy and Asthma 2.0".

Similarly to the previous Special Issue entitled "Molecular Mechanisms of Allergy and Asthma" [...].

Epigenetic Mechanisms in Allergy Development and Prevention.

There has been a substantial increase in the incidence and the prevalence of allergic disorders in the recent decades, which seems to be related to rapid environmental and lifestyle changes, such as higher exposure to factors thought to exert pro-allergic effects but less contact with factors known to be associated with protection against the development of allergies. Pollution is the most remarkable example of the former, while less contact with microorganisms, lower proportion of unprocessed natural products in diet, and others resulting from urbanization and westernization of the lifestyle exemplify the latter. It is strongly believed that the effects of environmental factors on allergy susceptibility and development are mediated by epigenetic mechanisms, i.e. biologically relevant biochemical changes of the chromatin carrying transcriptionally-relevant information but not affecting the nucleotide sequence of the genome. Classical epigenetic mechanisms include DNA methylation and histone modifications, for instance acetylation or methylation. In addition, microRNA controls gene expression at the mRNA level. Such epigenetic mechanisms are involved in crucial regulatory processes in cells playing a pivotal role in allergies. Those include centrally managing cells, such as T lymphocytes, as well as specific structural and effector cells in the affected organs, responsible for the local clinical presentation of allergy, e.g. epithelial or airway smooth muscle cells in asthma. Considering that allergic disorders possess multiple clinical (phenotypes) and mechanistic (endotypes) forms, targeted, stratified treatment strategies based on detailed clinical and molecular diagnostics are required. Since conventional diagnostic or therapeutic approaches do not suffice, this gap could possibly be filled out by epigenetic approaches.

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.

Short-Chain Fatty Acids Augment Differentiation and Function of Human Induced Regulatory T Cells.

Regulatory T cells (Tregs) control immune system activity and inhibit inflammation. While, in mice, short-chain fatty acids (SCFAs) are known to be essential regulators of naturally occurring and in vitro induced Tregs (iTregs), data on their contribution to the development of human iTregs are sparse, with no reports of the successful SCFAs-augmented in vitro generation of fully functional human iTregs. Likewise, markers undoubtedly defining human iTregs are missing. Here, we aimed to generate fully functional human iTregs in vitro using protocols involving SCFAs and to characterize the underlying mechanism. Our target was to identify the potential phenotypic markers best characterizing human iTregs. Naïve non-Treg CD4+ cells were isolated from the peripheral blood of 13 healthy adults and cord blood of 12 healthy term newborns. Cells were subjected to differentiation toward iTregs using a transforming growth factor ß (TGF-ß)-based protocol, with or without SCFAs (acetate, butyrate, or propionate). Thereafter, they were subjected to flow cytometric phenotyping or a suppression assay. During differentiation, cells were collected for chromatin-immunoprecipitation (ChIP)-based analysis of histone acetylation. The enrichment of the TGF-ß-based protocol with butyrate or propionate potentiated the in vitro differentiation of human naïve CD4+ non-Tregs towards iTregs and augmented the suppressive capacity of the latter. These seemed to be at least partly underlain by the effects of SCFAs on the histone acetylation levels in differentiating cells. GITR, ICOS, CD39, PD-1, and PD-L1 were proven to be potential markers of human iTregs. Our results might boost the further development of Treg-based therapies against autoimmune, allergic and other chronic inflammatory disorders.

Editorial of Special Issue "Molecular Mechanisms of Allergy and Asthma".

This Special Issue aggregates several high-quality original articles written by renowned researchers [...].

Extracellular Vesicles and Asthma-More Than Just a Co-Existence.

Extracellular vesicles (EVs) are membranous structures, which are secreted by almost every cell type analyzed so far. In addition to their importance for cell-cell communication under physiological conditions, EVs are also released during pathogenesis and mechanistically contribute to this process. Here we summarize their functional relevance in asthma, one of the most common chronic non-communicable diseases. Asthma is a complex persistent inflammatory disorder of the airways characterized by reversible airflow obstruction and, from a long-term perspective, airway remodeling. Overall, mechanistic studies summarized here indicate the importance of different subtypes of EVs and their variable cargoes in the functioning of the pathways underlying asthma, and show some interesting potential for the development of future therapeutic interventions. Association studies in turn demonstrate a good diagnostic potential of EVs in asthma.

Differential Regulation of Interferon Signaling Pathways in CD4+ T Cells of the Low Type-2 Obesity-Associated Asthma Phenotype.

In the era of personalized medicine, insights into the molecular mechanisms that differentially contribute to disease phenotypes, such as asthma phenotypes including obesity-associated asthma, are urgently needed. Peripheral blood was drawn from 10 obese, non-atopic asthmatic adults with a high body mass index (BMI; 36.67 ± 6.90); 10 non-obese, non-atopic asthmatic adults with normal BMI (23.88 ± 2.73); and 10 healthy controls with normal BMI (23.62 ± 3.74). All asthmatic patients were considered to represent a low type-2 asthma phenotype according to selective clinical parameters. RNA sequencing (RNA-Seq) was conducted on peripheral blood CD4+ T cells. Thousands of differentially expressed genes were identified in both asthma groups compared with heathy controls. The expression of interferon (IFN)-stimulated genes associated with IFN-related signaling pathways was specifically affected in obese asthmatics, while the gap junction and G protein-coupled receptor (GPCR) ligand binding pathways were enriched in both asthma groups. Furthermore, obesity gene markers were also upregulated in CD4+ T cells from obese asthmatics compared with the two other groups. Additionally, the enriched genes of the three abovementioned pathways showed a unique correlation pattern with various laboratory and clinical parameters. The specific activation of IFN-related signaling and viral infection pathways might provide a novel view of the molecular mechanisms associated with the development of the low type-2 obesity-associated asthma phenotype, which is a step ahead in the development of new stratified therapeutic approaches.

Development and characterization of DNAzyme candidates demonstrating significant efficiency against human rhinoviruses.

BACKGROUND: Infections with human rhinoviruses (RVs) are responsible for millions of common cold episodes and the majority of asthma exacerbations, especially in childhood. No drugs specifically targeting RVs are available. OBJECTIVE: We sought to identify specific anti-RV molecules based on DNAzyme technology as candidates to a clinical study. METHODS: A total of 226 candidate DNAzymes were designed against 2 regions of RV RNA genome identified to be sufficiently highly conserved between virus strains (ie, the 5'-untranslated region and cis-acting replication element) by using 3 test strains: RVA1, RVA16, and RVA29. All DNAzymes were screened for their cleavage efficiency against in vitro-expressed viral RNA. Those showing any catalytic activity were subjected to bioinformatic analysis of their reverse complementarity to 322 published RV genomic sequences. Further molecular optimization was conducted for the most promising candidates. Cytotoxic and off-target effects were excluded in HEK293 cell-based systems. Antiviral efficiency was analyzed in infected human bronchial BEAS-2B cells and ex vivo-cultured human sinonasal tissue. RESULTS: Screening phase-generated DNAzymes characterized by either good catalytic activity or by high RV strain coverage but no single molecule represented a satisfactory combination of those 2 features. Modifications in length of the binding domains of 2 lead candidates, Dua-01(-L12R9) and Dua-02(-L10R11), improved their cleavage efficiency to an excellent level, with no loss in eminent strain coverage (about 98%). Both DNAzymes showed highly favorable cytotoxic/off-target profiles. Subsequent testing of Dua-01-L12R9 in BEAS-2B cells and sinonasal tissue demonstrated its significant antiviral efficiency. CONCLUSIONS: Effective and specific management of RV infections with Dua-01-L12R9 might be useful in preventing asthma exacerbations, which should be verified by clinical trials.

Novel Splice Site Mutation in the PROS1 Gene in a Polish Patient with Venous Thromboembolism: c.602-2delA, Splice Acceptor Site of Exon 7.

We identified a novel splice site mutation of the PROS1 gene in a Polish family with protein S (PS) deficiency and explored the molecular pathogenesis of this previously undescribed variant. A novel mutation was detected in a 26-year-old woman with a history of venous thromboembolism (VTE) provoked by oral contraceptives. Her family history of VTE was positive. The sequence analysis of the PROS1 gene was performed in the proband and the proband's family. The proband and their asymptomatic father had lower free PS levels (45% and 50%, respectively) and PS activity (48% and 44%, respectively). Total PS levels were normal (65.6% and 62.4%, respectively). The sequence analysis of the PROS1 gene revealed the presence of heterozygous deletion at the nucleotide position c.602-2 in intron 6, just upstream of exon 7, detected in the proband and her father. This variant alters the splice acceptor site of exon 7, and, according to the in silico prediction, it is highly likely to cause in-frame exon 7 skipping. We also presented follow-up data of two other Polish patients with PS deficiency associated with splice site mutations in PROS1 gene.

Hepatic gene expression in mouse models of non-alcoholic fatty liver disease after acute exercise.

AIM: Non-alcoholic fatty liver disease (NAFLD) patients benefit from physical exercise. This study aimed to investigate the effect of acute exercise on hepatic gene expression in different mouse models of NAFLD. METHODS: C57BL/6J mice were fed with a control (CD) or a high fat (HFD) diet. AlbCrePtenflox/flox (Pten-KO) and Fxr-/- mice, two genetic models of NAFLD with insulin hypersensitivity and resistance, respectively, were fed with CD. After 4 weeks, mice were randomly assigned to exercise or sedentariness. Mice were killed 15 min or 3 h after the running/sedentary period. Genome-wide hepatic gene expression was evaluated with the Illumina Micro-array platform. Quantitative polymerase chain reaction confirmed changes in gene expression. RESULTS: Acute exercise transiently affected the expression of genes involved in the immune response in C57BL/6 mice fed with CD and this effect normalized in the recovery phase. Acute exercise affected genes involved in gluconeogenesis in the insulin resistant Fxr-/- model. Genes involved in lipid metabolism were affected in C57BL/6 mice fed with CD, but not in mouse models of NAFLD. Genes involved in DNA damage response pathways were deregulated only in C57BL/6 mice fed with CD and not in mouse models of NAFLD. CONCLUSION: The simultaneous analysis of different NAFLD models revealed that an acute exercise bout affects hepatic gene expression differentially according to animal models and that most of the differentially expressed genes are involved in glucose and fatty acid metabolism, immune regulation, and DNA damage response.

Influenza-derived peptides cross-react with allergens and provide asthma protection.

BACKGROUND: The hygiene hypothesis is the leading concept to explain the current asthma epidemic, which is built on the observation that a lack of bacterial contact early in life induces allergic TH2 immune responses. OBJECTIVE: Because little is known about the contribution of respiratory tract viruses in this context, we evaluated the effect of prior influenza infection on the development of allergic asthma. METHODS: Mice were infected with influenza and, once recovered, subjected to an ovalbumin- or house dust mite-induced experimental asthma protocol. Influenza-polarized effector memory T (Tem) cells were transferred adoptively to allergen-sensitized animals before allergen challenge. A comprehensive in silico analysis assessed homologies between virus- and allergen-derived proteins. Influenza-polarized Tem cells were stimulated ex vivo with candidate peptides. Mice were immunized with a pool of virus-derived T-cell epitopes. RESULTS: In 2 murine models we found a long-lasting preventive effect against experimental asthma features. Protection could be attributed about equally to CD4+ and CD8+ Tem cells from influenza-infected mice. An in silico bioinformatic analysis identified 4 influenza- and 3 allergen-derived MHC class I and MHC class II candidate T-cell epitopes with potential antigen-specific cross-reactivity between influenza and allergens. Lymphocytes from influenza-infected mice produced IFN-γ and IL-2 but not IL-5 on stimulation with the aforementioned peptides. Immunization with a mixture of the influenza peptides conferred asthma protection, and peptide-immunized mice transferred protection through CD4+ and CD8+ Tem cells. CONCLUSION: For the first time, our results illustrate heterologous immunity of virus-infected animals toward allergens. This finding extends the original hygiene hypothesis.

Histone Acetylation of Immune Regulatory Genes in Human Placenta in Association with Maternal Intake of Olive Oil and Fish Consumption.

Maternal diet modifies epigenetic programming in offspring, a potentially critical factor in the immune dysregulation of modern societies. We previously found that prenatal fish oil supplementation affects neonatal T-cell histone acetylation of genes implicated in adaptive immunity including PRKCZ, IL13, and TBX21. In this study, we measured H3 and H4 histone acetylation levels by chromatin immunoprecipitation in 173 term placentas collected in the prospective birth cohort, ALADDIN, in which information on lifestyle and diet is thoroughly recorded. In anthroposophic families, regular olive oil usage during pregnancy was associated with increased H3 acetylation at FOXP3 (p = 0.004), IL10RA (p = 0.008), and IL7R (p = 0.007) promoters, which remained significant after adjustment by offspring gender. Furthermore, maternal fish consumption was associated with increased H4 acetylation at the CD14 gene in placentas of female offspring (p = 0.009). In conclusion, prenatal olive oil intake can affect placental histone acetylation in immune regulatory genes, confirming previously observed pro-acetylation effects of olive oil polyphenols. The association with fish consumption may implicate ω-3 polyunsaturated fatty acids present in fish oil. Altered histone acetylation in placentas from mothers who regularly include fish or olive oil in their diets could influence immune priming in the newborn.

Increased activity of lipoprotein-associated phospholipase A2 in non-severe asthma.

BACKGROUND: Given increased risk of cardiovascular events in asthma we hypothesized that lipoprotein-associated phospholipase A2 (Lp-PLA2), an enzyme involved in atherosclerosis, is associated with proinflammatory and prothrombotic blood alterations in this disease. METHODS: In 164 adult asthmatics (63 with severe asthma) we measured plasma Lp-PLA2 activity using the PLAC test. We determined its relations to inflammation and prothrombotic blood alterations. RESULTS: In asthma, Lp-PLA2 was inversely related to the age (ß = -0.1 [-0.18 to -0.02]) and was lower in women (n = 122 [74%], 205 [182-242] vs. 243 [203-262] nmol/min/ml, p = 0.001). Interestingly, Lp-PLA2 correlated negatively with the asthma severity score (ß = -0.15 [-0.23 to -0.07]), being 10.3% higher in those with non-severe (mild or moderate) asthma (n = 101, 62%) as compared to the severe disease subtype (224 [191-261] vs. 203 [181-229], p = 0.006 after adjustment for potential confounders). Lp-PLA2 activity was positively related to the levels of low-density lipoprotein (ß = 0.1 [0.02-0.18]), triglycerides (ß = 0.11 [0.03-0.19]) and glucose (ß = 0.1 [0.02-0.18]) and inversely to the tumor necrosis factor α (ß = -0.27 [-0.35 to -0.2]), high sensitivity C-reactive protein (ß = -0.1 [-0.19 to -0.02]) and fibrinogen (ß = -0.12 [-0.21 to -0.03]), as well as prothrombin (ß = -0.16 [-0.24 to -0.08]), and parameters describing thrombin generation potential, such as endogenous thrombin potential (ß = -0.14 [-0.21 to -0.06]) and peak thrombin generated (ß = -0.2 [-0.28 to -0.12]). CONCLUSIONS: Elevated Lp-PLA2 activity in non-severe asthmatics suggests increased atherosclerotic risk in this group. Lower Lp-PLA2 activity accompanied by its inverse relationship to inflammatory or prothrombotic blood biomarkers observed in turn in severe asthmatics might be related to the pathogenesis of more severe asthma phenotype.

Antisense molecules: A new class of drugs.

An improved understanding of disease pathogenesis leads to identification of novel therapeutic targets. From a pharmacologic point of view, these can be addressed by small chemical compounds, so-called biologicals (eg, mAbs and recombinant proteins), or by a rather new class of molecule based on the antisense concept. Recently, a new wave of clinical studies exploring antisense strategies is evolving. In addition to cancer, they include predominantly trials on infectious and noninfectious diseases, such as chronic inflammatory and metabolic conditions. This article, based on a systematic PubMed literature search, highlights recent developments in this emerging field.