Targeted inhibition of m6A demethylase FTO by FB23 attenuates allergic inflammation in the airway epithelium.

Epithelial cells play a crucial role in asthma, contributing to chronic inflammation and airway hyperresponsiveness. m6A modification, which involves key proteins such as the demethylase fat mass and obesity-associated protein (FTO), is crucial in the regulation of various diseases, including asthma. However, the role of FTO in epithelial cells and the development of asthma remains unclear. In this study, we investigated the demethylase activity of FTO using a small-molecule inhibitor FB23 in epithelial cells and allergic inflammation in vivo and in vitro. We examined the FTO-regulated transcriptome-wide m6A profiling by methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA-seq under FB23 treatment and allergic inflammation conditions. Immunofluorescence staining was performed to assess the tissue-specific expression of FTO in asthmatic bronchial mucosa. We demonstrated that FB23 alleviated allergic inflammation in IL-4/IL-13-treated epithelial cells and house dust mite (HDM)-induced allergic airway inflammation mouse model. The demethylase activity of FTO contributed to the regulation of TNF-α signaling via NF-κB and epithelial-mesenchymal transition-related pathways under allergic inflammation conditions in epithelial cells. FTO was expressed in epithelial, submucosal gland, and smooth muscle cells in human bronchial mucosa. In conclusion, FB23-induced inhibition of FTO alleviates allergic inflammation in epithelial cells and HDM-induced mice, potentially through diverse cellular processes and epithelial-mesenchymal transition signaling pathways, suggesting that FTO is a potential therapeutic target in asthma management.

Regulation of SETD2 maintains immune regulatory function in macrophages to suppress airway allergy.

SET domain-containing 2 (SETD2) is a histone methyltransferase. It regulates the activity of H3K36me3 to enhance gene transcription. Macrophages (Mϕs) are one of the cell types involved in immune response. The purpose of this study is to clarify the role of SETD2 in regulating the immune property of Mϕ. The Mφs were isolated from the bronchoalveolar lavage fluid (BALF) and analysed through flow cytometry and RNA sequencing. A mouse strain carrying Mφs deficient in SETD2 was used. A mouse model of airway allergy was established with the ovalbumin/alum protocol. Less expression of SETD2 was observed in airway Mϕs in patients with allergic asthma. SETD2 of M2 cells was associated with the asthmatic clinical response. Sensitization reduced the expression of SETD2 in mouse respiratory tract M2 cells, which is associated with the allergic reaction. Depletion of SETD2 in Mφs resulted in Th2 pattern inflammation in the lungs. SETD2 maintained the immune regulatory ability in airway M2 cells. SETD2 plays an important role in the maintenance of immune regulatory property of airway Mφs.

Selective production of IL-33-neutralizing autoantibody ameliorates asthma responses and severity.

BACKGROUND: Natural anti-cytokine autoantibodies can regulate homeostasis of infectious and inflammatory diseases. The anti-cytokine autoantibody profile and relevance to the pathogenesis of asthma are unknown. We aim to identify key anti-cytokine autoantibodies in asthma patients, and reveal their immunological function and clinical significance. METHODS: A Luciferase Immunoprecipitation System was used to screen serum autoantibodies against 11 key cytokines in patients with allergic asthma and healthy donors. The antigen-specificity, immunomodulatory functions and clinical significance of anti-cytokine autoantibodies were determined by ELISA, qPCR, neutralization assays and statistical analysis, respectively. Potential conditions for autoantibody induction were revealed by in vitro immunization. RESULTS: Of 11 cytokines tested, only anti-IL-33 autoantibody was significantly increased in asthma, compare to healthy controls, and the proportion positive was higher in patients with mild-to-moderate than severe allergic asthma. In allergic asthma patients, the anti-IL-33 autoantibody level correlated negatively with serum concentration of pathogenic cytokines (e.g., IL-4, IL-13, IL-25 and IL-33), IgE, and blood eosinophil count, but positively with mid-expiratory flow FEF25-75%. The autoantibodies were predominantly IgG isotype, polyclonal and could neutralize IL-33-induced pathogenic responses in vitro and in vivo. The induction of the anti-IL-33 autoantibody in blood B-cells in vitro required peptide IL-33 antigen along with a stimulation cocktail of TLR9 agonist and cytokines IL-2, IL-4 or IL-21. CONCLUSIONS: Serum natural anti-IL-33 autoantibodies are selectively induced in some asthma patients. They ameliorate key asthma inflammatory responses, and may improve lung function of allergic asthma.

Allergen specific immunotherapy regulates macrophage property in the airways.

BACKGROUND: Allergen specific immunotherapy (AIT) has been widely used in allergy clinics. The therapeutic effects of it are to be improved. Macrophages occupy the largest proportion of airway immune cells. The aim of this study is to measure the effects of nasal instillation AIT (nAIT) on airway allergy by regulating macrophage functions. METHODS: An airway allergy mouse model was established with the ovalbumin-alum protocol. nAIT was conducted for mice with airway allergy through nasal instillation. The effects of nAIT were compared with subcutaneous injection AIT (SCIT) and sublingual AIT (SLIT). RESULTS: Mice with airway allergy showed the airway allergic response, including lung inflammation, airway hyper responsiveness, serum specific IgE, increase in the amounts of eosinophil peroxidase, mouse mast cell protease-1, and Th2 cytokines in bronchoalveolar lavage fluid. nAIT had a much better therapeutic effect on the airway allergic response than SCIT and SLIT. Mechanistically, we observed better absorption of allergen in macrophages, better production of IL-10 by macrophages, and better immune suppressive functions in macrophages in mice received nAIT than SCIT and SLIT. CONCLUSIONS: The nAIT has a much better therapeutic effect on suppressing the airway allergic response, in which macrophages play a critical role.

Phospholipid remodeling and its derivatives are associated with COVID-19 severity.

BACKGROUND: Timely medical intervention in severe cases of coronavirus disease 2019 (COVID-19) and better understanding of the disease's pathogenesis are essential for reducing mortality, but early classification of severe cases and its progression is challenging. OBJECTIVE: We investigated the levels of circulating phospholipid metabolites and their relationship with COVID-19 severity, as well as the potential role of phospholipids in disease progression. METHODS: We performed nontargeted lipidomic analysis of plasma samples (n = 150) collected from COVID-19 patients (n = 46) with 3 levels of disease severity, healthy individuals, and subjects with metabolic disease. RESULTS: Phospholipid metabolism was significantly altered in COVID-19 patients. Results of a panel of phosphatidylcholine (PC) and lysophosphatidylcholine (LPC) and of phosphatidylethanolamine and lysophosphatidylethanolamine (LPE) ratios were significantly correlated with COVID-19 severity, in which 16 phospholipid ratios were shown to distinguish between patients with severe disease, mild disease, and healthy controls, 9 of which were at variance with those in subjects with metabolic disease. In particular, relatively lower ratios of circulating (PC16:1/22:6)/LPC 16:1 and (PE18:1/22:6)/LPE 18:1 were the most indicative of severe COVID-19. The elevation of levels of LPC 16:1 and LPE 18:1 contributed to the changes of related lipid ratios. An exploratory functional study of LPC 16:1 and LPE 18:1 demonstrated their ability in causing membrane perturbation, increased intracellular calcium, cytokines, and apoptosis in cellular models. CONCLUSION: Significant Lands cycle remodeling is present in patients with severe COVID-19, suggesting a potential utility of selective phospholipids with functional consequences in evaluating COVID-19's severity and pathogenesis.

Combined BSA-Seq and RNA-Seq Analysis to Identify Candidate Genes Associated with Aluminum Toxicity in Rapeseed (Brassica napus L.).

Exchangeable aluminum (Al) ions released from acidic soils with pH < 5.5 inhibit root elongation of crops, ultimately leading to yield reduced. It is necessary to identify the quantitative trait locus (QTLs) and candidate genes that confer toxicity resistance to understand the mechanism and improve tolerance of rapeseed. In this study, an F2 segregating population was derived from a cross between Al-tolerance inbred line FDH188 (R178) and -sensitive inbred line FDH152 (S169), and the F2:3 were used as materials to map QTLs associated with the relative elongation of taproot (RET) under Al toxicity stress. Based on bulked segregant analysis sequencing (BSA-seq), three QTLs (qAT-A07-1, qAT-A07-2, and qAT-A09-1) were detected as significantly associated with RET, and 656 candidate genes were screened. By combined BSA and RNA-seq analysis, 55 candidate genes showed differentially expressed, including genes encoding ABC transporter G (ABCG), zinc finger protein, NAC, ethylene-responsive transcription factor (ERF), etc. These genes were probably positive factors in coping with Al toxicity stress in rapeseed. This study provides new insight into exploring the QTLs and candidate genes' response to Al toxicity stress by combined BSA-seq and RNA-seq and is helpful to further research on the mechanism of Al resistance in rapeseed.

Comparative Genomics Reveals Insights into the Divergent Evolution of Astigmatic Mites and Household Pest Adaptations.

Highly diversified astigmatic mites comprise many medically important human household pests such as house dust mites causing ∼1-2% of all allergic diseases globally; however, their evolutionary origin and diverse lifestyles including reversible parasitism have not been illustrated at the genomic level, which hampers allergy prevention and our exploration of these household pests. Using six high-quality assembled and annotated genomes, this study not only refuted the monophyly of mites and ticks, but also thoroughly explored the divergence of Acariformes and the diversification of astigmatic mites. In monophyletic Acariformes, Prostigmata known as notorious plant pests first evolved, and then rapidly evolving Astigmata diverged from soil oribatid mites. Within astigmatic mites, a wide range of gene families rapidly expanded via tandem gene duplications, including ionotropic glutamate receptors, triacylglycerol lipases, serine proteases and UDP glucuronosyltransferases. Gene diversification after tandem duplications provides many genetic resources for adaptation to sensing environmental signals, digestion, and detoxification in rapidly changing household environments. Many gene decay events only occurred in the skin-burrowing parasitic mite Sarcoptes scabiei. Throughout the evolution of Acariformes, massive horizontal gene transfer events occurred in gene families such as UDP glucuronosyltransferases and several important fungal cell wall lytic enzymes, which enable detoxification and digestive functions and provide perfect drug targets for pest control. This comparative study sheds light on the divergent evolution and quick adaptation to human household environments of astigmatic mites and provides insights into the genetic adaptations and even control of human household pests.

Characterization of the immune regulatory property of CD22+ CD9+ B cells.

Immunodisruptive homeostasis is recognized in allergic disorders. The mechanism of restoration of immunologic homeostasis in the body is not fully understood. Galectin-9 (Gal9) and CD22 have immune regulatory functions. The goal of this study is to test the role of CD22+ CD9+ B regulatory cells in immune homeostasis the body. A much smaller amount of IL-10 in B10 cells was detected in patients with allergic rhinitis (AR) in contrast to healthy subjects. The IL-10 expression levels in B10 cells were positively correlated with the CD22 expression. CD22 mediated the effects of Gal9 on the enhanced expression of IL-10 in AR B10 cells. Gal9 overcame the refractory induction of IL-10 in B-cells of AR subjects. The immune regulatory ability of AR B10 cells could be restored by Gal9. Combination of Gal9 and SIT induced and activated antigen-specific B10 cells. The B10 cells of Gal9/specific immunotherapy-treated AR mice showed immunosuppressive functions on T-cell activities and induction of type 1 regulatory T cells in an antigen-specific manner. Administration of Gal9 potentiated the effects of specific immunotherapy in mice with AR. In summary, a fraction of regulatory B cells, the CD19+ CD22+ CD9+ B cells, was characterized in the present study. CD22 mediates the effects of Gal9 to promote immunotherapy for allergic diseases by inducing B10 cells. In an antigen-specific manner, the B10 cells suppressed CD4+ T cell activities, and alleviated experimental AR.

Adjuvant role of probiotics in allergen-specific immunotherapy.

Allergy are a global public health problem with serious consequences for human health. Many studies have confirmed that probiotics can play an active role in the treatment of allergies, and early intervention of probiotics can significantly reduce the incidence in infants and young children. As an adjuvant of allergen-specific immunotherapy (AIT), probiotics also show advantages in improving the therapeutic effect and reducing side effects. This paper reviewed the research progress of probiotics in the prevention and alleviation of allergies, focused on the use of probiotics as adjuvants for AIT, and pointed out the possibility of probiotics' extracellular proteins, extracellular vesicles, metabolites, and Toll-like Receptors (TLR) agonist analog as AIT adjuvant alone.

Integration of GWAS and transcriptome analyses to identify SNPs and candidate genes for aluminum tolerance in rapeseed (Brassica napus L.).

BACKGROUND: The exchangeable aluminum (Al), released from the acid soils, is another addition to the environmental stress factors in the form of Al toxicity stress. Al stress affects the normal crop development and reduces the overall yield of rapeseed (Brassica napus L.). The response mechanism of plants to Al toxicity is complicated and difficult to understand with few QTL related studies in rapeseed under Al toxicity stress. RESULT: Using 200,510 SNPs developed by SLAF-seq (specific-locus amplified fragment sequencing) technology, we carried out the genome-wide association analysis (GWAS) in a population of 254 inbred lines of B. napus with large genetic variation and Al-tolerance differences. There were 43 SNPs significantly associated with eight Al-tolerance traits in the seedling stage were detected on 14 chromosomes, and 777 candidate genes were screened at the flanking 100 kb region of these SNPs. Moreover, RNA-seq detected 8291 and 5341 DEGs (the differentially expressed gene) in the Al -tolerant line (ATL) and -sensitive line (ASL), respectively. Based on integration of GWAS and RNA-seq analysis, 64 candidate genes from GWAS analysis differentially expressed at least once in 6 h vs 0 h or 24 h vs 0 h conditions in ATL or ASL. Moreover, four out of sixty-four candidate genes (BnaA03g30320D, BnaA10g11500D, BnaC03g38360D and BnaC06g30030D) were differentially expressed in both 6 h and 24 h compared to 0 h (control) conditions in both lines. The proposed model based on the candidate genes excavated in this study highlighted that Al stress disturb the oxidation-redox balance, causing abnormal synthesis and repair of cell wall and ABA signal transduction, ultimately resulting in inhibition of root elongation. CONCLUSIONS: The integration of GWAS and transcriptome analysis provide an effective strategy to explore the SNPs and candidate genes, which has a potential to develop molecular markers for breeding Al tolerant rapeseed varieties along with theoretical basis of molecular mechanisms for Al toxicity response of Brassica napus plants.

Dual-Channel Heterogeneous Graph Neural Network for Predicting microRNA-Mediated Drug Sensitivity.

Many studies have confirmed that microRNAs (miRNAs) are mediated in the sensitivity of tumor cells to anticancer drugs. MiRNAs are emerging as a type of promising therapeutic targets to overcome drug resistance. However, there is limited attention paid to the computational prediction of the associations between miRNAs and drug sensitivity. In this work, we proposed a heterogeneous network-based representation learning method to predict miRNA-drug sensitivity associations (DGNNMDA). An miRNA-drug heterogeneous network was constructed by integrating miRNA similarity network, drug similarity network, and experimentally validated miRNA-drug sensitivity associations. Next, we developed a dual-channel heterogeneous graph neural network model to perform feature propagation among the homogeneous and heterogeneous nodes so that our method can learn expressive representations for miRNA and drug nodes. On two benchmark datasets, our method outperformed other seven competitive methods. We also verified the effectiveness of the feature propagations on homogeneous and heterogeneous nodes. Moreover, we have conducted two case studies to verify the reliability of our methods and tried to reveal the regulatory mechanism of miRNAs mediated in drug sensitivity. The source code and datasets are freely available at https://github.com/19990915fzy/DGNNMDA.

Stiffness in breast masses with posterior acoustic shadowing: significance of ultrasound real time shear wave elastography.

BACKGROUND: To assess the stiffness of benign breast masses in ultrasound images with posterior acoustic shadowing (PAS) and malignant lesions, and explore the significance of differential diagnosis using ultrasound real time shear wave elastography. MATERIAL AND METHODS: All 117 mammary masses (98 patients) with PAS were assessed by using routine ultrasound examination, and elastic modulus values were obtained with the real time shear wave elastography mode. All breast lesions were confirmed by surgery or biopsy. The significance of differences in ultrasound elastography values between breast benign and malignant masses with posterior acoustic shadowing was assessed, and the ROC curves of elasticity modulus values were analyzed. RESULTS: Among the 117 masses, 72 were benign and 45 were malignant. The two types of breast masses showed significant differences in size, margin, internal echo, calcification, and blood flow characteristics (P < 0.05), although the difference in orientation was not significant (P > 0.05). Emean, Emax and Esd obtained with real time shear wave elastography showed statistically significant differences between benign masses with posterior acoustic shadowing and breast cancer (P < 0.05), while Emin showed no significant difference between them (P = 0.633). Ultrasound real time shear wave elastography showed higher sensitivity and specificity than conventional ultrasound. CONCLUSIONS: Benign and malignant breast masses with PAS show different ultrasound manifestations. Real time shear wave elastography can facilitate the differential diagnosis and treatment planning for these breast masses.

Effects of rAmb a 1-Loaded PLGA-PEG Nanoparticles in a Murine Model of Allergic Conjunctivitis.

Ambrosia artemisiifolia (Amb a) contains many allergens. Allergic conjunctivitis caused by Ambrosia artemisiifolia and its related allergen-specific immunotherapy (AIT) are seldom studied at present. poly(DL-lactide-co-glycolide)-polyethylene glycol (PLGA-PEG) is a very good nano-carrier, which has been applied in the medical field. In this context, we studied the immunotherapy effect and potential mechanism of recombinant Amb a 1 (rAmb a 1)-loaded PLGA-PEG nanoparticles. A mouse allergic conjunctivitis model was established with Ambrosia artemisiifolia crude extract, and the nanoparticles were used for AIT through direct observation of conjunctival tissue, degranulation of mast cells in conjunctival tissue, serum-specific antibodies, cytokines and other assessment models. The treatment of nanoparticles enhanced the secretion of T-helper 1 (Th1) cytokine Interferon-gama (IFN-γ) and the production of immunoglobulin G (IgG)2a (IgG2a), inhibited the secretion of T-helper 2 (Th2) cytokine Interleukin (IL)-13 and IL-4 and the level of IgE. Especially, degranulation of mast cells and expression of mast cell protease-1 (MCP-1) in conjunctival tissue was reduced significantly. In this study, we proved that the nanoparticles prepared by rAmb a 1 and PLGA-PEG have an immunotherapy effect on allergic conjunctivitis in mice.

Meta-Transcriptomic Analysis Reveals the Virome and Viral Genomic Evolution of Medically Important Mites.

Mites are notorious for being vectors transmitting infectious pathogens and source of allergens causing allergic conditions in animals and humans. However, despite their huge impact on public health, the virome of mites remains unknown. Here we characterized the virus diversity and abundance of 14 species of medically important mites based on total RNA sequencing data sets generated in this study as well as those deposited in the Sequence Read Archive (SRA) database. A total of 47 genetically distinct viruses were identified and classified into 17 virus families or virus super-groups, and the viral sequences accounted for as much as 29.67% of total non-rRNA transcriptome in one mite library. The most commonly identified viruses are members of Picornavirales, among which we revealed more than one type of viruses that are evolutionarily related to dicistronic viruses but contain a single open reading frame, thus likely representing a recent example of host (i.e., mite)-related parallel evolution from dicistronic to monocistronic genomic form within the family Dicistroviridae To our best knowledge, this is the first time to perform comprehensive and systematic screening of RNA virome in medically important mites including house dust mites (HDM). Overall, the RNA virome identified here provides not only significant insights into the diversity and evolution of RNA viruses in mites, but also a solid knowledge base for studying their roles in human diseases.IMPORTANCE Mites are important group of arthropods that are associated with a variety of human diseases including scrub typhus and asthma. However, it remains unclear whether or not mites carry viruses that might play a role in human infections or allergic disease. In this study, we used a total transcriptomics approach to characterize and compare the complete RNA virome within mites that are relevant to human health and diseases. Specifically, our data revealed a large diversity, a high abundance, and a flexible genomic evolution for these viruses. Although most of the viruses identified here are unknown to associate with human infectious disease, the abundant presence of viral RNAs may play an immunomodulatory role in the development of allergic reactions such as asthma during environmental exposure to mite allergens, and therefore provide important insights into the mite-induced allergy and preparation of mite allergen vaccines.

Bcl2L12 Contributes to Th2-Biased Inflammation in the Intestinal Mucosa by Regulating CD4+ T Cell Activities.

The Th2-biased inflammation and immune deregulation play a critical role in the pathogenesis of ulcerative colitis (UC). Recent studies indicate that the Bcl2-like protein 12 (Bcl2L12) is associated with immune deregulation of UC. This study aims to investigate the role of Bcl2L12 in the induction of aberrant Th2-biased inflammation. In this study, peripheral blood samples were collected from patients with inflammatory bowel disease. The Th2 cell activities were analyzed by flow cytometry, real-time quantitative RT-PCR, and Western blotting. Mice with Bcl2L12-knockout CD4+ T cells were used in the experiments. The results showed that the expression of Bcl2L12 was detected in peripheral CD4+ T cells, which was significantly higher in UC patients than in healthy subjects. A positive correlation between the expression of Bcl2L12 and Th2 cytokines was detected in CD4+ T cells from UC patients. Naive CD4+ T cells with Bcl2L12 overexpression were prone to differentiate into Th2 cells. Mice with Bcl2L12 deficiency failed to induce the Th2-biased inflammation in the intestine. Bcl2L12 bound GATA3 to form a complex to enhance the binding between GATA3 and the Il4 promoter to enhance the expression of IL-4 in CD4+ T cells. CD4+ T cells with Bcl2L12 overexpression were resistant to apoptosis. In conclusion, the Bcl2L12 is a critical factor in the induction of aberrant Th2 polarization by upregulating Th2 responses and downregulating Th2 cell apoptosis. Bcl2L12 may be a novel therapeutic target in the management of the disorders with Th2-biased inflammation.

T cell epitope of arginine kinase with CpG co-encapsulated nanoparticles attenuates a shrimp allergen-induced Th2-bias food allergy.

T cell peptide-based immunotherapy (PIT) is an appealing therapeutic strategy for modulating allergic responses without IgE cross-linking. We propose a novel PIT that combines a T-cell epitope of the shrimp allergen arginine kinase (AKp) with TLR9 agonist CpG-ODN in nanoparticles (CpG-AKp NPs) to attenuate a shrimp allergen-induced food allergy. Treatment with CpG-AKp NPs demonstrated the attenuation of anaphylaxis responses such as the reduced incidence of diarrhea and hypothermia, lower levels of specific IgE and the induction of IgG2a in serum. Th2 cytokines were suppressed and higher Th1 cytokines were detected in the splenocyte culture supernatants. Treatment of CpG-AKp NPs also enhanced the protein expression of Foxp3 and IL-10 in small intestine but decreased the activation of STAT6 and GATA3 expression, which are related to differentiation of Th2. Our data indicated that CpG-AKp NPs may represent a promising PIT against shrimp allergy.

Vitamin D-deficiency induces eosinophil spontaneous activation.

Eosinophils (Eo) play a critical role in immunity and immune inflammation. The maintenance of Eo homeostasis is not fully understood yet. Vitamin D (VitD) is involved in the regulation of a large number of biochemical reactions. This study tests a hypothesis that VitD receptor (VDR) contributes to the homeostasis of Eos. In this study, EoL-1 cells (an Eo cell line) were cultured in the presence or absence of calcitriol. The Eo-mediators, including major basic protein (MBP), Eo peroxidase (EPX), Eo cationic protein (ECP) and Eo-derived neurotoxin (EDN), were assessed in the culture supernatant and in EoL-1 cells. We observed that, in a VitD deficient environment, EoL-1 cells produced high levels of the Eo-mediators, including MBP, EPX, ECP and EDN, which could be suppressed by the addition of calcitriol to the culture. EoL-1 cells expressed VitD receptor (VDR), which was up regulated by exposure to calcitriol. VDR formed complexes with the transcription factors of the Eo-mediators, which prevented the transcription factors to bind to the promoters of the Eo-mediators, and therefore prevented the Eo-mediated gene transcription. The Eo spontaneous activation was also found in the intestinal mucosa of VDR-deficient mice, in which the intestinal epithelial barrier dysfunction was observed. In conclusion, VDR contributes to the maintenance of the homeostasis of Eos by regulating the gene transcription of the Eo mediators. The VDR-deficiency is one of the causative factors inducing Eo spontaneous activation. This phenomenon may be taken into account in the management of the Eo-related diseases.