Project description:Purpose: this study provided a comprehensive sequence for a systemic view of the transcriptome between mango leaf and fruit, as well as fruit allergens, which will be useful for further genomic research studies and breeding of lower allergenic mango cultivars. Methods:Some allergens have previously been identified in mango (Mangifera indica Linn), including profilins, Bet v 1-like proteins and chitinase. In this paper, 66 potential allergen genes were identified and their relative expressions evaluated in mango fruit and leaf using Illumina RNA-Seq technology. Results:A total of 17.63Gb Clean Data was obtained.The number of %≥Q30 was above 94.58%.RNA-Seq generated 11,751,123 contigs that were assembled into 99,328 unigenes with 16,848 unigenes of >1000 bp. A total of 230,242 unigenes were annotated using public protein databases, with a cut-off E-value above 10−5, of which 27,295, 46,030, 24,227 and 14,023 unigenes were assigned to gene ontology terms, Nr, Swiss-Prot and clusters of orthologous groups, respectively. Allergens mainly belonged to pollen allergen, pathogenesis-related protein Bet v I family and NADPH-dependent FMN reductase.
Project description:Background: Respiratory allergy triggered by pollen allergens is increasing at an alarming rate worldwide. Sunflower pollen is thought to be an important source of inhalant allergens. Present study aims to identify the prevalence of sunflower pollinosis among the Indian allergic population and characterizes the pollen allergens using immuno-proteomic tools. Methodology: Clinico-immunological tests were performed to understand the prevalence of sensitivity towards sunflower pollen among the atopic population. Sera from selected sunflower positive patients were used as probe to detect the IgE-reactive proteins from the one and two dimensionally separated proteome of sunflower pollen. The antigenic nature of the sugar moiety of the glycoprotein allergens was studied by meta-periodate modification of IgE-immunoblot. Finally, these allergens were identified by mass-spectrometry (MALDI TOF/TOF and LC ESI qTOF). MASCOT searching was performed against NCBInr database. However, Helianthus annuus genome is not fully sequenced and partially annotated. So in case of low confidence (p> 0.05) protein identification, searching was performed against EST library of Helianthus annuus. Results: Prevalence of sunflower pollen allergy was observed among 21% of the atopic population and associated with elevated level of specific IgE and histamine in the sera of these patients. Immunoscreening of sunflower pollen proteome with patient serum detected seven IgE-reactive proteins with varying molecular weight and pI. Hierarchical clustering of 2D-immunoblot data highlighted three allergens characterized by a more frequent immuno-reactivity and increased levels of IgE antibodies in the sera of susceptible patients. These allergens were considered as the major allergens of sunflower pollen and were found to have their glycan moiety critical for inducing IgE response. Homology driven search of MS/MS data of these IgE-reactive proteins identified seven previously unreported allergens from sunflower pollen. Three major allergenic proteins were identified as two non-isoformic pectate lyases and a cystein protease. Conclusion: Novelty of the present report is the identification of a panel of seven sunflower pollen allergens for the first time at immuno-biochemical and proteomic level, which substantiated the clinical evidence of sunflower allergy. Further purification and recombinant expression of these allergens will improve component-resolved diagnosis and therapy of pollen allergy.
Project description:Background: The prevalence of individuals allergic to latex, exhibiting cross-hypersensitivity with plant-derived food has been frequently reported as the so-called latex-fruit syndrome. Nonetheless, molecular mechanisms underlying allergy to latex and/or fruit are poorly understood. Objective: The aims of this study were to identify candidate genes that may be associated with the pathogenesis of allergy to latex and /or vegetable food, and to assess if similar molecular pathways are involved in both types of hypersensitivity. Methods: DNA microarray analysis was performed to screen the molecular profiles of peripheral blood mononuclear cells isolated from patients with allergy to latex, to fruit, or with latex-fruit syndrome, and from control healthy subjects. Results: Gene expression profiling identified an overlapping dataset of genes commonly regulated in all the atopic patients enrolled in this study, suggesting that similar molecular mechanisms are involved in the pathogenesis of allergy to the fruit and /or latex. Several regulators of the innate and acquired immunity reported to polarize the immunological response towards a Th2-mediated immune response were overexpressed in patients. Furthermore, evidences suggested that T regulatory cell expression might be defective in allergic patients, as a consequence of a dysregulation of some inflammatory cytokines. Finally, several transcription factors that may be responsible for the Th1/Th2 imbalance were modulated in allergic patients. Conclusion and clinical implications: This study identified relevant genes that may help to elucidate the molecular mechanisms underlying allergic disease. Knowledges of critical targets, along with transcription factors regulating gene activity may facilitate the development of new therapies. Experiment Overall Design: This study aimed at the understanding of the molecular pathways involved in allergy to latex and fruit. As latex and vegetable food allergens cross-react, it could be hypothesized that similar pathways are involved in both types of hypersensitivity. Experiment Overall Design: For this purpose, patients allergic to latex (n=6), allergic to vegetable food (n=5), or suffering from latex-fruit syndrome (n=6) were enrolled from the Unit of Allergy of the Gemelli Hospital of Rome. Moreover, 4 healthy volunteers were added to this study. Five ml of blood were harvested from each individual, and PBMCs were isolated on ficoll gradient. Following sample preparation (as recommended by the manufacturer), each sample was hybridized on Affymetrix human focus array. Data were processed with Affymetrix MAS 5.0 software and averaged intensity of signals from biological replicates were calculated for further analysis. Experiment Overall Design: The CEL files for this study were lost in a computer crash.
Project description:In this study, MLN transcriptome profiling of female BALB/c mice was conducted in order to explore the molecular mechanism(s) of antigen sensitization using three of the most common food allergens; BLG, OVA and PNA. Analysis of cDNA microarray data revealed a complex network of genes that is modulated during BLG, OVA or PNA sensitization. Results from this study implicate the role of differentially expressed genes in the pathogenesis of food allergy. Results from this study will also help better understanding the molecular mechanism of food allergy in animal models and may help in developing new drug target for the therapy of food allergy. This study is likely to add a new dimension in the development of biomarker genes for the determination of potential allergenicity of genetically modified (GM) foods.
Project description:Background: The prevalence of individuals allergic to latex, exhibiting cross-hypersensitivity with plant-derived food has been frequently reported as the so-called latex-fruit syndrome. Nonetheless, molecular mechanisms underlying allergy to latex and/or fruit are poorly understood. Objective: The aims of this study were to identify candidate genes that may be associated with the pathogenesis of allergy to latex and /or vegetable food, and to assess if similar molecular pathways are involved in both types of hypersensitivity. Methods: DNA microarray analysis was performed to screen the molecular profiles of peripheral blood mononuclear cells isolated from patients with allergy to latex, to fruit, or with latex-fruit syndrome, and from control healthy subjects. Results: Gene expression profiling identified an overlapping dataset of genes commonly regulated in all the atopic patients enrolled in this study, suggesting that similar molecular mechanisms are involved in the pathogenesis of allergy to the fruit and /or latex. Several regulators of the innate and acquired immunity reported to polarize the immunological response towards a Th2-mediated immune response were overexpressed in patients. Furthermore, evidences suggested that T regulatory cell expression might be defective in allergic patients, as a consequence of a dysregulation of some inflammatory cytokines. Finally, several transcription factors that may be responsible for the Th1/Th2 imbalance were modulated in allergic patients. Conclusion and clinical implications: This study identified relevant genes that may help to elucidate the molecular mechanisms underlying allergic disease. Knowledges of critical targets, along with transcription factors regulating gene activity may facilitate the development of new therapies.
Project description:CD4+ helper T cells (TH) and regulatory T cells (Treg) that respond to common allergens play an important role in driving and dampening airway inflammation in patients with asthma. Until recently, direct, unbiased molecular analysis of allergen-reactive TH and Treg cells has not been possible. To better understand the diversity of these T cell subsets in allergy and asthma, we analyzed the single-cell transcriptome of ~50,000 house dust mite (HDM) allergen-reactive TH cells and Treg cells from asthmatics with HDM allergy and from three control groups: asthmatics without HDM allergy and non-asthmatics with and without HDM allergy. Our analyses show that HDM allergen-reactive TH and Treg cells are highly heterogeneous, and certain subsets are quantitatively and qualitatively different in subjects with HDM-reactive asthma. The number of interleukin (IL)-9 expressing HDM-reactive TH cells is greater in asthmatics compared with non-asthmatics with HDM allergy, and display enhanced pathogenic properties. More HDM-reactive Th and Treg cells expressing the interferon-response signature (THIFNR and TregIFNR) are present in asthmatics without HDM allergy compared with those with HDM allergy. In cells from these subsets (THIFNR and TregIFNR), expression of TNFSF10 was enriched; its product, TRAIL, dampens activation of TH cells. These findings suggest that the THIFNR and TregIFNR subsets may dampen allergic responses, which may help explain why only some people develop TH2 responses to nearly ubiquitous allergens.
Project description:CD4+ helper T cells (TH) and regulatory T cells (Treg) that respond to common allergens play an important role in driving and dampening airway inflammation in patients with asthma. Until recently, direct, unbiased molecular analysis of allergen-reactive TH and Treg cells has not been possible. To better understand the diversity of these T cell subsets in allergy and asthma, we analyzed the single-cell transcriptome of ~50,000 house dust mite (HDM) allergen-reactive TH cells and Treg cells from asthmatics with HDM allergy and from three control groups: asthmatics without HDM allergy and non-asthmatics with and without HDM allergy. Our analyses show that HDM allergen-reactive TH and Treg cells are highly heterogeneous, and certain subsets are quantitatively and qualitatively different in subjects with HDM-reactive asthma. The number of interleukin (IL)-9 expressing HDM-reactive TH cells is greater in asthmatics compared with non-asthmatics with HDM allergy, and display enhanced pathogenic properties. More HDM-reactive Th and Treg cells expressing the interferon-response signature (THIFNR and TregIFNR) are present in asthmatics without HDM allergy compared with those with HDM allergy. In cells from these subsets (THIFNR and TregIFNR), expression of TNFSF10 was enriched; its product, TRAIL, dampens activation of TH cells. These findings suggest that the THIFNR and TregIFNR subsets may dampen allergic responses, which may help explain why only some people develop TH2 responses to nearly ubiquitous allergens.
Project description:Purpose: Delonix regia or Gulmohor commonly grows here and there in Indian villages as well as it is used for megacity beautification and environmental management due to its evergreen nature and vibrant flower colour. However, an increasing incidence of seasonal pollinosis was observed among the inhabitants living in close vicinity to this tree suggesting a possible link between the airborne pollen load and the concomitant respiratory hazards. This prompted us to investigate the allergens in the pollen of this dominant avenue tree. Methodology: Allergenicity of D. regia pollen grains was first checked by Skin Prick Test (SPT) and further confirmed by in vitro tests, such as, ELISA, Dot Blot and histamine release assay. The total proteome profiling was done by 2D PAGE and it was confronted with the pooled sera of 10 patients. The IgE reactive proteins were identified by MALDI TOF/TOF in Autoflex speed (Bruker,Germany). The raw spectra were searched against NCBInr database using MASCOT search engine. Result: Delonix regia, pollen grains have been found in considerable amount in the air during its flowering season (May to July). Approximately 31% of atopic individuals were found allergic to D. regia, pollen with elevated level of specific IgE and histamine in the serum. Total 8 IgE reactive proteins have been identified by homology driven proteomics. These proteins are ATP synthase beta subunit (spot no 5), Actin (spot no 6), Hypothetical actin like protein (spot no 7), Recombinant S-adenosylmethionine synthase 2 (spot no 9), UDP-arabinopyranose mutase (spot no 13), Luminal-binding 5 (spot no 2), ELF3-like protein (spot no 8) and hypothetical protein OsJ_04810 (spot no 11). Among these eight identified allergenic proteins, five were previously reported as allergen from different sources whereas the rest three have been reported for the first time as novel allergens. Conclusion: Novelty of this study was to identify 8 allergens from D.regia for the first time using immune-biochemical and proteomic techniques. Further studies will open up new avenues in component resolved diagnosis of pollen allergy.
Project description:Food-specific IgE triggers life-threatening anaphylaxis; however, some people with food-specific IgE are asymptomatic upon allergen consumption. Using murine food allergy models, we discovered an unexpected pathway regulating the ability of food allergens to trigger anaphylaxis. C57BL/6 mice are uniquely resistant to anaphylaxis when challenged orally; we show that their gut barrier was impermeable to food allergens relative to anaphylaxis-susceptible strains, even prior to allergic sensitization. In a forward genetic screen, oral anaphylaxis resistance correlated with Dipeptidase1 (Dpep1) allelic variants. DPEP1 is expressed in intestinal epithelium and catabolizes leukotriene D4. Blocking DPEP1 with cilastatin enhanced allergen absorption in resistant mice. Conversely, pre-treatment of susceptible mice with a leukotriene synthesis inhibitor, zileuton, abrogated allergen absorption and subsequent oral anaphylaxis. Inhibiting leukotrienes may be a novel treatment for food allergy.