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.
Recombinant protein production in different Escherichia coli strains is a cornerstone of modern biotechnology. Controlled intracellular processing (CIP) of recombinant passenger proteins that are fused to solubility enhancer proteins has been proposed as an alternative to obtaining satisfactory yields of soluble recombinant proteins without the need for significant post‐processing steps. In this manuscript, we review different genetic systems currently available for controlled intracellular processing of fusion recombinant proteins, regarding system design features, significant advantages, and limitations of the various strategies.
The elucidation of factors that trigger the development of transient wheezing in early childhood may be an important step toward understanding the pathogenesis of asthma and other allergic diseases ...later in life. Transient wheezing has been mainly attributed to viral infections, although sensitisation to aeroallergens and food allergens may occur at an early age. In developing countries, intestinal helminthic infections have also been associated with allergy or atopy-related disorders.
The aim of this study was to explore the association of Trichuris trichiura and Ascaris lumbricoides infections with wheezing and atopy in early childhood.
A cross-sectional study using a Portuguese-language ISAAC phase I questionnaire, adapted for preschool-aged children, nested in a cohort study of childhood diarrhoea, was conducted on 682 children. Two faecal samples per child were examined for the presence of intestinal helminthic infection. IgE antibodies against three allergenic preparations (Dermatophagoides pteronyssinus, Blomia tropicalis and common child food), as well as against A. lumbricoides antigens, were measured in a sub-sample of these children, whose parents allowed the procedure. Atopy was defined by the presence of levels of serum IgE antibodies ≥0.35 kU/L against at least one of the three tested allergenic preparations.
Active T. trichiura infection but not A. lumbricoides infection was positively associated with wheezing in the total studied children population adjusted OR = 2.60; CI = 1.54;4.38 and in the atopic children sub-population adjusted OR = 3.07; CI = 1.00;9.43. The association with atopy was also positive and statistically significant only in the brute analysis OR = 2.13; CI = 1.03;4.40. Anti-A. lumbricoides IgE antibodies, but not current A. lumbricoides infection, were positively associated with wheezing in atopic children adjusted OR = 2.01; CI = 1.00;4.50 and in non-atopic children adjusted OR = 3.07; CI = 1.13;8.35 and it was also associated with atopy adjusted OR = 7.29; CI = 3.90; 13.4. On the other hands, reports of wheezing were not significantly associated with atopy.
These data corroborate previous studies showing that wheezing is predominantly associated with infection in early childhood and shows that anti-A. lumbricoides IgE antibodies, but not active Ascaris infections, are associated with wheezing and atopy. Additionally, the data demonstrate that T. trichiura infection may play a role in the pathogenesis of atopic wheezing in early childhood.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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.
Background: Sanitation affects health, especially that of young children. Residents of Salvador, in Northeast Brazil, have had a high prevalence of intestinal parasites. A citywide sanitation ...intervention started in 1996 aimed to raise the level of sewer coverage from 26% to 80% of households. Objectives: We evaluated the impact of this intervention on the prevalence of Ascaris lumbricoides, Trichuris trichuria, and Giardia duodenalis infections in preschool children. Methods: The evaluation was composed of two cross-sectional studies (1998 and 2003-2004), each of a sample of 681 and 976 children 1—4 years of age, respectively. Children were sampled from 24 sentinel areas chosen to represent the range of environmental conditions in the study site. Data were collected using an individual/household questionnaire, and an environmental survey was conducted in each area before and after the intervention to assess basic household and neighborhood sanitation conditions. Stool samples were examined for the presence of intestinal parasites. The effect of the intervention was estimated by hierarchical modeling, fitting a sequence of multivariate regression models. Findings: The prevalence of A. lumbricoides infection was reduced from 24.4% to 12.0%, T. trichuria from 18.0% to 5.0%, and G. duodenalis from 14.1% to 5.3%. Most of this reduction appeared to be explained by the increased coverage in each neighborhood by the sewage system constructed during the intervention. The key explanatory variable was thus an ecological measure of exposure and not household-based, suggesting that the parasite transmission prevented by the program was mainly in the public (vs. the domestic) domain. Conclusion: This study, using advanced statistical modeling to control for individual and ecological potential confounders, demonstrates the impact on intestinal parasites of sanitation improvements implemented at the scale of a large population.
Objective
Sufficient vitamin D (25‐hydroxyvitamin D 25(OH)D) serum levels are associated with decreased asthma symptoms. Our aim was to investigate associations between vitamin D and atopy, asthma, ...asthma severity, and asthma phenotypes in Brazilian teenagers.
Methods
This cross‐sectional study involved 942 individuals (11–19 years old) engaged in an asthma cohort. The ISAAC questionnaire was employed to diagnosis asthma and asthma severity. Serum allergen‐specific immunoglobulin E (sIgE) was measured by ImmunoCap and serum 25(OH)D was measured by ELISA. We calculated the correlation between sIgE and 25(OH)D. We used multivariate logistic regression analysis to assess associations of interest.
Results
We found that 25(OH)D deficiency was positively associated with atopy (OR 1.45, confidence interval CI 1.05–2.00) and high levels of this vitamin negatively correlated with sIgE to Dermatophagoides pteronyssinus (r = −0.11, p = 0.019). The average 25(OH)D serum level was 27.0 ± 9.5 ng/ml; 366 individuals (38.8%) had a sufficient level. There was no association between 25(OH)D and asthma, asthma severity or asthma phenotypes in the population. However, sex was a possible effect modifier of the association between vitamin D and asthma: insufficiency in asthmatic women (86%) was higher than in asthmatic men (42%), and there was an association between insufficient vitamin D levels and greater asthma risk only in women (OR = 3.06, 95% CI 1.16–8.07).
Conclusion
We have shown that vitamin D deficiency was associated with greater risk of atopy in both sexes and vitamin D insufficiency was associated with asthma only in women. There was no association between vitamin D levels and asthma phenotypes or asthma severity.
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).
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.
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 .
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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.
•Genetic variants in ADIPOQ and LEP are associated with asthma.•Increasing in the number of risk allele is associated with a higher risk of asthma.•BMI changes the association of genetic variants ...with asthma and atopy.
Asthma and atopy are considered condition associated with obesity, being affected by genetic and environmental factors. The LEP and ADIPOQ genes, responsible for the expression and secretion of leptin and adiponectin, respectively, and polymorphisms in such genes have been linked to both diseases, independently, and also with the obesity-associated asthma phenotype in populations with high European ancestry and high-income countries. However, in mixed populations, there are few studies evaluating the impact of these variants in genes associated with the phenotype of asthma and obesity. Thus, the aim of this study was to investigate variants in LEP and ADIPOQ associated with asthma and atopy, and whether overweight modifies that effect.
The study involved 203 asthmatics children and 813 control subjects (between 5 and 11 years old), with or without overweight, from the SCAALA (Asthma and Allergy Social Changes in Latin America) program. Among them, 831 had data for allergy markers, being 258 atopic and 573 non-atopic. Genotyping was performed using a commercial panel Omnium Illumina 2.5. Logistic regression was performed to identify associations expected by using PLINK 1.09 and three genetic models: additive, dominant and recessive adjusted for sex, age, helminth infection, BMI and Principal Components (PC) 1 and 2, for ancestry, in order to control the confounding factor by population structure.
For asthma, G allele of rs822396, in ADIPOQ, was positively associated in additive model (OR 1.4, 95% CI 1.08–1.83) and T allele of rs1063537 in dominant model (OR 1.52, 95% CI 1.01–2.30). In LEP, rs11763517 (C allele) and rs11760956 (A allele) were both negatively associated with asthma in the additive model (OR 0.70, 95% CI 0.54–0.91; OR 0.66, 95% CI 0.50–0.89) respectively, and the A allele of rs2167270 in dominant model (OR 0.71, 95% CI 0.51–0.98). The G allele of rs12706832 showed a positive association with asthma in the recessive model (OR 1.66, 95% CI 1.06–2.61). When the population was stratified by the BMI / Age Z-Score, the protection observed for asthma between the variants rs11760956, rs11763517 and rs2167270 was lost overweight individuals; The protection observed for atopy was lost in all variants (rs16861205, rs2167270 and rs17151919) in the overweight group.
These results suggest that SNPs on the LEP and ADIPOQ genes may have an impact on atopy and asthma. Furthermore, we also show that the asthma and atopy protection attributed to variants on LEP and ADIPOQ genes is lost in individuals exposed to overweight.
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.
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.
To increase knowledge of B. tropicalis allergens through proteomic analysis.
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).
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.
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.
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