Recombinant T-cell epitope conjugation: A new approach for Dermatophagoides hypoallergen design.

BACKGROUND: Allergen-specific immunotherapy (AIT) is the only clinical approach that can potentially cure some allergic diseases by inducing immunological tolerance. Dermatophagoides pteronyssinus is considered as the most important source of mite allergens worldwide, with high sensitization rates for the major allergens Der p 1, Der p 2 and Der p 23. The aim of this work is to generate a hypoallergenic hybrid molecule containing T-cell epitopes from these three major allergens. METHODS: The hybrid protein termed Der p 2231 containing T-cell epitopes was purified by affinity chromatography. The human IgE reactivity was verified by comparing those with the parental allergens. The hybrid was also characterized immunologically through an in vivo mice model. RESULTS: The hybrid rDer p 2231 stimulated in peripheral blood mononuclear cells (PBMCs) isolated from allergic patients with higher levels of IL- 2, IL-10, IL-15 and IFN-γ, as well as lower levels of IL-4, IL-5, IL-13, TNF-α and GM-CSF. The use of hybrid molecules as a therapeutic model in D. pteronyssinus allergic mice led to the reduction of IgE production and lower eosinophilic peroxidase activity in the airways. We found increased levels of IgG antibodies that blocked the IgE binding to the parental allergens in the serum of allergic patients. Furthermore, the stimulation of splenocytes from mice treated with rDer p 2231 induced higher levels of IL-10 and IFN-γ and decreased the secretion of IL-4 and IL-5, when compared with parental allergens and D. pteronyssinus extract. CONCLUSIONS: rDer p 2231 has the potential to be used in AIT in patients co-sensitized with D. pteronyssinus major allergens, once it was able to reduce IgE production, inducing allergen-specific blocking antibodies, restoring and balancing Th1/Th2 immune responses, and inducing regulatory T-cells.

The hybrid protein BTH2 suppresses allergic airway inflammation in a murine model of HDM-specific immunotherapy.

BACKGROUND: Allergen-specific immunotherapy (AIT) is the only disease-modifying treatment approach to change disease-causing allergens. Hypoallergenic derivatives show promise as potential therapeutics, amongst which BTH2 was designed to induce tolerance against Blomia tropicalis allergy. Our aim was to investigate the hypoallergenicity and immunoregulatory activity of BTH2 in vitro and its therapeutic potential in a mouse model of AIT. METHODS: Recombinant Blo t 5 and Blo t 21 allergens and their hybrid derivatives (BTH1 and BTH2) were expressed and purified. IgE binding capacity was tested by ELISA using sera from Brazilian, Colombian, and Ecuadorian subjects. Secretion of cytokines in supernatants from human cell cultures was measured following stimulation with the four recombinants and controls. The capacity of BTH2 to ameliorate allergic airway inflammation induced by B. tropicalis extract was evaluated in a murine model of AIT. RESULTS: rBlo t 5 and rBlo t 21 were identified as major allergens in Latin American patients, and BTH2 had the lowest IgE binding. In vitro stimulation of human cells induced greater levels of IL-10 and IFN-γ and reduced the secretion of Th2 cytokines. BTH2 ameliorated allergic airway inflammation in B. tropicalis-challenged A/J mice, as evidenced by the histopathological and humoral biomarkers: decreased Th2 cytokines and cellular infiltration (especially eosinophils), lower activity of eosinophil peroxidase, an increase in IgG blocking antibodies and strong reduction of mucus production by goblet cells. CONCLUSIONS: Our study shows that BTH2 represents a promising candidate for the treatment of B. tropicalis allergy with hypoallergenic, immune regulatory and therapeutic properties. Further pre-clinical studies are required in murine models of chronic asthma to further address the efficacy and safety of BTH2 as a vaccine against B. tropicalis-induced allergy.

Genome-wide association study of Helicobacter pylori serological status in Latin American children.

BACKGROUND: Few genome-wide association studies (GWAS) on Helicobacter pylori infection susceptibility have been conducted for admixed populations from developing countries. Here, we performed a GWAS to identify genetic factors associated with H. pylori serostatus in a cohort of admixed children from a large Latin American urban center. METHODS: A cross-sectional study involving 1161 children from 4 to 11 years old living in poor areas of Salvador, in northeastern Brazil. Logistic regression analysis was performed to detect associations between single-nucleotide variants (SNVs) and H. pylori seropositivity, assuming an additive genetic model. Enrichment analyses were conducted using the MAGMA v1.10 software. RESULTS: We found 22 SNVs to be suggestively associated (p < 10-5 ) with H. pylori seropositivity. The most suggestive SNV was the rs77955022 (p = 4.83e-07) located in an intronic region of EXOC3 at 5p15.33. The second most suggestively associated SNV was rs10914996 (p = 8.97e-07), located in an intergenic region at 1p34.3. Furthermore, we were able to replicate three SNVs (p < 0.05) in the Study of Health in Pomerania (SHIP) cohort: the rs2339212 and rs4795970, both located at 17q12 near TMEM132E, as well as the rs6595814, an intronic variant of FBN2 at 5q23.3. The enrichment analysis indicated the participation of genes and metabolic pathways related to the regulation of the digestive system and gastric acid secretion in the risk of seropositivity for H. pylori. CONCLUSIONS: Additional studies are required to validate these association findings in larger population samples and to get insight into the underlying physiological mechanisms.

Engineering an optimized expression operating unit for improved recombinant protein production in Escherichia coli.

Common strategies to improve recombinant protein production in Escherichia coli often involve the test and optimization of several different variables, when using traditional expression vectors that are commercially available. Now, modern synthetic biology-based strategies allow for extensive modifications of these traditional vectors, or even construction of entirely new modular vectors, so as to permit tunable production of the recombinant proteins of interest. Herein, we describe the engineering of a new expression operating unit (EOU; 938 bp) for producing recombinant proteins in E. coli, through the combinatorial assembly of standardized and well-characterized genetic elements required for transcription and translation (promoter, operator site, RBS, junction RBS-CDS, cloning module, transcriptional terminator). We also constructed a novel T7 promoter variant with increased transcriptional activity (1.7-fold higher), when compared to the canonical wild type T7 promoter sequence. This new EOU yielded an improved production of the reporter protein superfolder GFP (sfGFP) in E. coli BL21(DE3) (relative fluorescence units/RFU = 70.62 ± 1.62 A U.) when compared to a high-producing control expression vector (plasmid BBa_I746909; RFU = 59.68 ± 1.82 A U.). The yields of purified soluble recombinant sfGFP were also higher when using the new EOU (188 mg L-1 culture vs. 108 mg L-1 in the control) and it performed similarly well when inserted into different plasmid backbones (pOPT1.0/AmpR and pOPT2.0/CmR).

Identification of Glycycometus malaysiensis (for the first time in Brazil), Blomia tropicalis and Dermatophagoides pteronyssinus through multiplex PCR.

Blomia tropicalis and Dermatophagoides pteronyssinus play an important role in triggering allergy. Glycycometus malaysiensis causes IgE reaction in sensitive people, but is rarely reported in domestic dust, because it is morphologically similar to B. tropicalis making the identification of these species difficult. The identification of mites is mostly based on morphology, a time-consuming and ambiguous approach. Herein, we describe a multiplex polymerase chain reaction (mPCR) assay based on ribosomal DNA capable to identify mixed cultures of B. tropicalis, D. pteronyssinus and G. malaysiensis, and/or to identify these species from environmental dust. For this, the internal transcribed spacer 2 (ITS2) regions, flanked by partial sequences of the 5.8S and 28S genes, were PCR-amplified, cloned and sequenced. The sequences obtained were aligned with co-specific sequences available in the GenBank database for primer design and phylogenetic studies. Three pairs of primers were chosen to compose the mPCR assay, which was used to verify the frequency of different mites in house dust samples (n = 20) from homes of Salvador, Brazil. Blomia tropicalis was the most frequent, found in 95% of the samples, followed by G. malaysiensis (70%) and D. pteronyssinus (60%). Besides reporting for the first time the occurrence of G. malaysiensis in Brazil, our results confirm the good resolution of the ITS2 region for mite identification. Furthermore, the mPCR assay proved to be a fast and reliable tool for identifying these mites in mixed cultures and could be applied in future epidemiological studies, and for quality control of mite extract production for general use.

In vivo cleavage of solubility tags as a tool to enhance the levels of soluble recombinant proteins in Escherichia coli.

Recombinant proteins are generally fused with solubility enhancer tags to improve the folding and solubility of the target protein of interest. However, the fusion protein strategy usually requires expensive proteases to perform in vitro proteolysis and additional chromatographic steps to obtain tag-free recombinant proteins. Expression systems based on intracellular processing of solubility tags in Escherichia coli, through co-expression of a site-specific protease, simplify the recombinant protein purification process, and promote the screening of molecules that fail to remain soluble after tag removal. High yields of soluble target proteins have already been achieved using these protease co-expression systems. Herein, we review approaches for controlled intracellular processing systems tailored to produce soluble untagged proteins in E. coli. We discuss the different genetic systems available for intracellular processing of recombinant proteins regarding system design features, advantages, and limitations of the various strategies.

Purification and characterisation of the dimeric group 12 allergen from Blomia tropicalis heterologously expressed by Escherichia coli Top10F´.

Successful research in the wide-ranging field of allergy is usually achieved by definition not only of physicochemical and immunological properties of natural, but also recombinant allergens. Blomia tropicalis mite is a well-known source for various groups of hypersensitivity-causing proteins. The goal of the present work was to produce, purify and characterise by in silico, biochemical and immunological methods the recombinant group-12 allergen of B. tropicalis. The recombinant Blo t 12 aggregation capacity as well as the affinity to antibodies from BALB/c immunised mice and B. tropicalis-sensitised human donors were investigated through in silico analyses, dynamic light scattering, SDS-PAGE, ELISA and Western blot. The presence of Blo t 12 within B. tropicalis extracts was also determined by ELISA and Western blot. High concentrations of dimeric rBlo t 12 were detected through SDS-PAGE next to other aggregates and the results were confirmed by data from DLS and Western blot. The YITVM peptide was predicted to be the most aggregation-prone region. The IgE-reactivity of rBlo t 12 was not completely abolished by aggregate formation but it was significantly decreased compared to rBlo t 5, or B. tropicalis extracts. Natural Blo t 12 may naturally dimerises, but it was detected in non-delipidified B. tropicalis extracts in low amounts. Given that this allergen may be a specific marker for B. tropicalis allergy, the recombinant Blo t 12 herein obtained is characterised as a mid-tier allergen in Brazilian atopic patients and may be useful for the improvement in precision allergy molecular diagnostic applications.

A hybrid of two major Blomia tropicalis allergens as an allergy vaccine candidate.

INTRODUCTION: Allergen-specific immunotherapy (AIT) represents a curative approach for treating allergies. In the tropical and subtropical regions of the world, Blomia tropicalis (Blo t 5 and Blo t 21) is the likely dominant source of indoor allergens. AIM: To generate a hypoallergenic Blo t 5/Blo t 21 hybrid molecule that can treat allergies caused by B tropicalis. METHODS: Using in silico design of B tropicalis hybrid proteins, we chose two hybrid proteins for heterologous expression. Wild-type Blo t 5/Blo t 21 hybrid molecule and a hypoallergenic version, termed BTH1 and BTH2, respectively, were purified by ion exchange and size exclusion chromatography and characterized by physicochemical, as well as in vitro and in vivo immunological, experiments. RESULTS: BTH1, BTH2 and the parental allergens were purified to homogeneity and characterized in detail. BTH2 displayed the lowest IgE reactivity that induced basophil degranulation using sera from allergic rhinitis and asthmatic patients. BTH2 essentially presented the same endolysosomal degradation pattern as the shortened rBlo t 5 and showed a higher resistance towards degradation than the full-length Blo t 5. In vivo immunization of mice with BTH2 led to the production of IgG antibodies that competed with human IgE for allergen binding. Stimulation of splenocytes from BTH2-immunized mice produced higher levels of IL-10 and decreased secretion of IL-4 and IL-5. In addition, BTH2 stimulated T-cell proliferation in PBMCs isolated from allergic patients, with secretion of higher levels of IL-10 and lower levels of IL-5 and IL-13, when compared to parental allergens. CONCLUSIONS AND CLINICAL RELEVANCE: BTH2 is a promising hybrid vaccine candidate for immunotherapy of Blomia allergy. However, further pre-clinical studies addressing its efficacy and safety are needed.

Toxoplasma gondii protects from IgE sensitization and induces Th1/Th2 immune profile.

Toxoplasma gondii (T gondii) infection has been associated with protection against allergy and autoimmune diseases. We investigated the effects of T gondii infection on cytokine and antibody responses in atopic and nonatopic Brazilian subjects. We have measured in whole-blood cultures, Th1 (IFN-γ and IL-12), Th2 (IL-5) and regulatory cytokine IL-10 in blood cells unstimulated and stimulated with pokeweed mitogen or T gondii soluble tachyzoites antigen (STAg) or Dermatophagoides pteronyssinus antigen. A significant negative association was found between high levels of anti-dust mite IgE and T gondii seropositivity (OR = 0.46; 95%CI = 0.25-0.85). STAg stimulation induced a mixed profile of Th1 and Th2 cytokines (IFN-γ, IL-12 and IL-5) in Tg-positive atopic individuals compared with Tg-negative atopic individuals (P < .0001, P = .033 and P = .003, respectively). In contrast, IL-10 production was not different between these groups. No association was found between T gondii infection and asthma. We hypothesized that the protective effect on atopy might be related to the strong Th1 immune response to T gondii found on the seropositive subjects. From our knowledge, this is the first study to investigate the association between atopy and T gondii infection in Brazilian subjects, analysing the cellular immune responses.

Genetic variants in RORA are associated with asthma and allergy markers in an admixed population.

Asthma and allergy affect hundreds of millions of people from childhood to old age. In most of them, the inflammatory process of respiratory allergies involves the participation of type 2 cytokines, derived from T helper-2 (Th2)-cell, and Group 2 Innate Lymphoid (ILC2) Cells. An efficient memory Th2 cell response is dependent on IL-13 produced by ILC2s, causing allergic lung inflammation and elevated serum levels of immunoglobulin E. ILC2 cells are derived from common lymphoid progenitors and their growing depends on the transcription factor RORA. The aim of this work was to identify genetic variants in RORA associated with asthma phenotypes and allergy markers. Genomic DNA samples of 1246 individuals participating from Social Changes Asthma and Allergy in Latin America Program (SCAALA) have been genotyped using Illumina Human 2.5 Omni Beadchip. Logistics regressions have been performed to analyze the association among RORA variants and asthma, skin prick tests (SPT), specific IgE and type 2 cytokine production. Twelve single nucleotide variants (SNVs) were significantly associated with atopy (P < 0.01), in which four of them, rs10162630, rs17191519, rs17270243, and rs55796775 and their haplotypes were strongly and positively associated (P < 0.001). Furthermore, these variants increased the RORA gene expression in silico analysis. Other SNVs in RORA were associated with allergy markers, atopic and non-atopic asthma. Therefore, it is believed that variants in RORA gene may influence immunologic features of asthma and allergies and could be possible targets for future treatment of allergic diseases.

Proteomic Analysis Reveals Allergen Variability among Breeds of the Dust Mite Blomia tropicalis.

BACKGROUND: The dawn of the "omics" technologies has changed allergy research, increasing the knowledge and identification of new allergens. However, these studies have been almost restricted to Dermatophagoides spp. Although Blomia tropicalis has long been established as a clinically important source of allergens, a thorough proteomic characterization is still lacking for this dust mite. OBJECTIVE: To increase knowledge of B. tropicalis allergens through proteomic analysis. METHODS: Eleven in-bred lineages of B. tropicalis were obtained from 11 unique different pregnant females. Their somatic extracts were analyzed and compared with a commercially available extract by liquid chromatography tandem mass spectrometry (LC-MS/MS). RESULTS: Considerable differences in the protein expression profiles were found among the breeds, and most of them displayed higher expression levels of major allergens than the commercially available extract. Blo t 2 was the most prominent allergenic protein in the analyzed extracts. Six identified allergens and 14 isoforms have not yet been recognized by IUIS. Conversely, 3 previously recognized B. tropicalis allergens were not found. CONCLUSIONS: The clear impact of inbreeding on allergen content shown by our study leads us to conclude that the quantification and/or identification of allergens from in-bred lines should be routinely considered for mite cultivation in order to select breeds with higher amounts of major allergens. In this sense, LC-MS/MS may be a useful method to achieve this quality control for research and commercial purposes.

Parasites and allergy: Observations from Brazil.

Brazil is a middle-income country undergoing the epidemiological transition. Effects of changes in daily life habits and access to clean water, sanitation and urban services on a growing urban population have contributed to a double burden of both infectious and noncommunicable chronic diseases. Studies have indicated that parasite infections may modulate the human immune system and influence the development of allergic conditions such as asthma. However, there is no consensus in the published literature on the effects of parasitic infections on allergy, perhaps as a consequence of factors determining the epidemiology of these infections that vary between populations such as age of first infection, duration and chronicity of infections, parasite burden and species, and host genetic susceptibility. In this review, we discuss the observations from Brazil concerning the relationship between parasite infections and allergy.

Relationship between African Biogeographical Ancestry and Helicobacter pylori infection in children of a large Latin American urban center.

BACKGROUND AND AIM: The relationship between race/ethnicity and H pylori infection has been extensively reported, with a higher prevalence of infection observed in black individuals. Whether such differences are due to genetic factors underlying African ancestry remains to be clarified. In the present study, we evaluated the association between the proportion of individual African ancestry and H pylori infection in a sample of 1046 children living in a large Latin American urban center. MATERIALS AND METHODS: Estimation of individual biogeographical ancestry was based on 370,539 SNPs and performed using the ADMIXTURE software. Multivariate logistic regression models and mediation analysis considering the influence of previously recognized socioenvironmental risk factors to H pylori infection were performed. All analyses were conducted using the statistical package STATA v.14.0. RESULTS: Each 10% increase in the proportion of individual African ancestry was positively and independently associated with H pylori infection in our population (adjusted OR = 1.22, 95% CI = 1.10-1.36, P < .001). Mediation analysis demonstrated that only 9.23% of the effect of the individual African ancestry on H pylori infection was explained by factors such as household income, the absence of street paving and crowding. CONCLUSIONS: The results suggest that genetic variants that covariate with African ancestry may explain an important part of the racial differences observed for the prevalence of H pylori infection.

Sambucus australis Modulates Inflammatory Response via Inhibition of Nuclear Factor Kappa B (NF-kB) in vitro.

Medicinal plants have long been used as an alternative to traditional drugs for the treatment of inflammatory conditions due to the classical side effects and restricted access of various commercially available drugs, such as steroids (GCs) and nonsteroidal anti-inflammatory drugs (NSAIDs). Sambucus australis is a Brazilian herb that is commonly used to treat inflammatory diseases; however, few studies have examined the use of this species in the treatment of inflammatory conditions. The present study aims to evaluate the potential anti-inflammatory activity of S. australis in vitro. We established spleen cell cultures stimulated with pokeweed mitogen (PWM) to evaluate the production of proinflammatory cytokines, such as IL-4, IL-5, IFN-y, and IL-10 (by ELISA), and the expression of the transcription factor NF-kB (by RT-PCR). In addition, we evaluated the levels of nitric oxide in macrophage cultures and the membrane-stabilizing activity of S. australis methanolic extract (EMSA). Treatment with EMSA at concentrations of 100, 50, 25 and 12.5 µg/ml significantly decreased IL-4 (p<0.001) and IL-5 (p<0.001) levels. Treatment with 100 µg/ml EMSA reduced IFN-у (p<0.001) levels. Moreover, at 100 mg/ml, EMSA also increased IL-10 production and reduced NF-kB expression (p<0.01). In macrophage cultures stimulated with LPS, EMSA decreased nitric oxide levels (p<0.001) at all concentrations tested (100, 50, 25 and 12.5 µg/ml). Additionally, EMSA had a protective effect in the erythrocyte membrane stabilization assay. Taken together, these results suggest that S. australis has anti-inflammatory potential in vitro, characterized by the reduction of both inflammatory cytokines and the expression of NF-kB along with the up-regulation of IL-10.

Multiobjective grammar-based genetic programming applied to the study of asthma and allergy epidemiology.

BACKGROUND: Asthma and allergies prevalence increased in recent decades, being a serious global health problem. They are complex diseases with strong contextual influence, so that the use of advanced machine learning tools such as genetic programming could be important for the understanding the causal mechanisms explaining those conditions. Here, we applied a multiobjective grammar-based genetic programming (MGGP) to a dataset composed by 1047 subjects. The dataset contains information on the environmental, psychosocial, socioeconomics, nutritional and infectious factors collected from participating children. The objective of this work is to generate models that explain the occurrence of asthma, and two markers of allergy: presence of IgE antibody against common allergens, and skin prick test positivity for common allergens (SPT). RESULTS: The average of the accuracies of the models for asthma higher in MGGP than C4.5. IgE were higher in MGGP than in both, logistic regression and C4.5. MGGP had levels of accuracy similar to RF, but unlike RF, MGGP was able to generate models that were easy to interpret. CONCLUSIONS: MGGP has shown that infections, psychosocial, nutritional, hygiene, and socioeconomic factors may be related in such an intricate way, that could be hardly detected using traditional regression based epidemiological techniques. The algorithm MGGP was implemented in c ++ and is available on repository: http://bitbucket.org/ciml-ufjf/ciml-lib .

Global issues in allergy and immunology: Parasitic infections and allergy.

Allergic diseases are on the increase globally in parallel with a decrease in parasitic infection. The inverse association between parasitic infections and allergy at an ecological level suggests a causal association. Studies in human subjects have generated a large knowledge base on the complexity of the interrelationship between parasitic infection and allergy. There is evidence for causal links, but the data from animal models are the most compelling: despite the strong type 2 immune responses they induce, helminth infections can suppress allergy through regulatory pathways. Conversely, many helminths can cause allergic-type inflammation, including symptoms of "classical" allergic disease. From an evolutionary perspective, subjects with an effective immune response against helminths can be more susceptible to allergy. This narrative review aims to inform readers of the most relevant up-to-date evidence on the relationship between parasites and allergy. Experiments in animal models have demonstrated the potential benefits of helminth infection or administration of helminth-derived molecules on chronic inflammatory diseases, but thus far, clinical trials in human subjects have not demonstrated unequivocal clinical benefits. Nevertheless, there is sufficiently strong evidence to support continued investigation of the potential benefits of helminth-derived therapies for the prevention or treatment of allergic and other inflammatory diseases.

Allergens of Blomia tropicalis: An Overview of Recombinant Molecules.

Allergic diseases are considered a major problem for healthcare systems in both developed and developing countries. House dust mites are well-known triggers of allergic manifestations. While the Dermatophagoides genus is widely distributed globally, Blomia tropicalis is the most prominent mite species in the tropical and subtropical regions of the world. Over the last decades, an increase in sensitization rates to B. tropicalis has been reported, leading to increased research efforts on Blomia allergens. In fact, 8 new allergens have been identified and characterized to different degrees. Here, we provide an overview of recent developments concerning the identification and production of recombinant Blomia allergens, as well as their structural and immunological characterization. Although considerable progress has been achieved, detailed molecule-based studies are still needed to better define the clinical relevance of Blomia allergens. Thus, the establishment of a well-standardized and fully characterized panel of allergens remains a challenge for the development of better diagnosis and therapy of allergic diseases induced by B. tropicalis.

Effect of dietary consumption as a modifier on the association between FTO gene variants and excess body weight in children from an admixed population in Brazil: the Social Changes, Asthma and Allergy in Latin America (SCAALA) cohort study.

Previous studies have shown associations of variants of the FTO gene with body weight, but none of these have involved Latin American populations with a high level of miscegenation, as is seen in the north-eastern Brazilian population. This study evaluated the association between SNP in the FTO gene and excess weight in Salvador, Bahia, Brazil. In addition, the effect of diet as a modifier on this association was also investigated. This cross-sectional study included 1191 participants aged 4-11 years, who were genotyped for 400 variants of the FTO gene. Direct anthropometric measures were made and dietary data were obtained by 24-h food recall. Multivariate logistic regression analyses were used to assess the associations of interest. Overall, 11·2 % of the individuals included in the study were overweight/obese. Interactions were identified between the percentage energy intake from proteins and obesity risk linked to the rs62048379 SNP (P interaction=0·01) and also between fat intake (PUFA:SFA ratio) and obesity risk linked to the rs62048379 SNP (P interaction=0·01). The T allele for the variant rs62048379 was positively associated with overweight/obesity in individuals whose percentage energy intake from protein was above the median (OR 2·00; 95 % CI 1·05, 3·82). The rs62048379 SNP was also associated with overweight/obesity in individuals whose PUFA:SFA ratio was below the median (OR 1·63; 95 % CI 1·05, 2·55). The association between FTO gene variants and excess body weight can be modulated by dietary characteristics, particularly by fatty acid distribution and dietary protein intake in children.

Effect of polymorphisms on TGFB1 on allergic asthma and helminth infection in an African admixed population.

BACKGROUND: Allergic asthma is a complex disorder that results from a combination of genetic and environmental factors. Studies suggest that helminth infections can activate a regulatory network characterized by the production of regulatory cytokines, such as interleukin 10 and transforming growth factor ß1 (TGF-ß1) and subsequently protect against immune-mediated diseases, such as asthma. On the other hand, TGF-ß1 is increased in the lungs of individuals with asthma and may modulate airway inflammation. The role of TGF- ß 1 single-nucleotide polymorphisms (SNPs) in allergic disease remains inconclusive. OBJECTIVE: To evaluate the effects of genetic variations in the TGF-ß1 on allergy and helminths infections in children. METHODS: We tested for association among 4 TGF-ß1 SNPs and allergic asthma, specific IgE, skin prick test result, and IL-10 production in 1,335 Brazilians. In addition, we analyzed the association with markers of helminth infection (parasite burden, anti-Ascaris IgE, and worm specific IgG4). The polymorphisms were genotyped using Taq Man probes. RESULTS: We found an association between rs1800470 (C allele) and atopic wheezing (odds ratio [OR], 0.60; 95% confidence interval [CI], 0.37-0.95) and markers of allergy (OR, 0.41; 95% CI, 0.22-0.79). In contrast, a positive association was observed between the haplotype ACCA and Trichuris trichiura infection (OR, 1.85; P = .003) and Ascaris lumbricoides infection (OR, 2.01; P < .001). This haplotype was also associated with increased IL-10 production (ß = 50.7; P < .001). CONCLUSION: Individuals with TGF-ß1 polymorphisms have an increased susceptibility to helminth infections and a lower risk of developing allergy. These studies suggest that immune modulation of allergic disease results not only from environmental factors but also from genetic susceptibility and IL-10 production.

Applied immuno-epidemiological research: an approach for integrating existing knowledge into the statistical analysis of multiple immune markers.

BACKGROUND: Immunologists often measure several correlated immunological markers, such as concentrations of different cytokines produced by different immune cells and/or measured under different conditions, to draw insights from complex immunological mechanisms. Although there have been recent methodological efforts to improve the statistical analysis of immunological data, a framework is still needed for the simultaneous analysis of multiple, often correlated, immune markers. This framework would allow the immunologists' hypotheses about the underlying biological mechanisms to be integrated. RESULTS: We present an analytical approach for statistical analysis of correlated immune markers, such as those commonly collected in modern immuno-epidemiological studies. We demonstrate i) how to deal with interdependencies among multiple measurements of the same immune marker, ii) how to analyse association patterns among different markers, iii) how to aggregate different measures and/or markers to immunological summary scores, iv) how to model the inter-relationships among these scores, and v) how to use these scores in epidemiological association analyses. We illustrate the application of our approach to multiple cytokine measurements from 818 children enrolled in a large immuno-epidemiological study (SCAALA Salvador), which aimed to quantify the major immunological mechanisms underlying atopic diseases or asthma. We demonstrate how to aggregate systematically the information captured in multiple cytokine measurements to immunological summary scores aimed at reflecting the presumed underlying immunological mechanisms (Th1/Th2 balance and immune regulatory network). We show how these aggregated immune scores can be used as predictors in regression models with outcomes of immunological studies (e.g. specific IgE) and compare the results to those obtained by a traditional multivariate regression approach. CONCLUSION: The proposed analytical approach may be especially useful to quantify complex immune responses in immuno-epidemiological studies, where investigators examine the relationship among epidemiological patterns, immune response, and disease outcomes.