Ricardo Riccio Oliveira

Polymyxin B as inhibitor of LPS contamination of Schistosoma mansoni recombinant proteins in human cytokine analysis

BackgroundRecombinant proteins expressed in Escherichia coli vectors are generally contaminated with endotoxin. In this study, we evaluated the ability of Polymyxin B to neutralize the effect of LPS present as contaminant on Schistosoma mansoni recombinant proteins produced in E. coli in inducing TNF-α and IL-10. Peripheral blood mononuclear cells from individuals chronically infected with S. mansoni were stimulated in vitro with recombinant Sm22.6, Sm14 and P24 antigens (10 μg/mL) in the presence of Polymyxin B (10 μg/mL).ResultsThe levels of cytokines were measured using ELISA. There was greater than 90 % reduction (p < 0.05) in the levels of TNF-α and IL-10 when Polymyxin B was added to the cultures stimulated with LPS. In cultures stimulated with S. mansoni recombinant proteins in the presence of Polymyxin B, a reduction in the levels of TNF-α and IL-10 was also observed. However, the percentage of reduction was lower when compared to the cultures stimulated with LPS, probably because these proteins are able to induce the production of these cytokines by themselves.ConclusionThis study showed that Polymyxin B was able to neutralize the effect of endotoxin, as contaminant in S. mansoni recombinant antigens produced in E. coli, in inducing TNF-α and IL-10 production.

Impaired T Helper 2 Response to Aeroallergen in Helminth-Infected Patients with Asthma

Helminthic infections have been shown to inhibit allergy skin-prick tests and to modify the course of asthma. We evaluated Dermatophagoides pteronyssinus-specific immune responses in patients with asthma by measuring levels of T helper 2 (Th2) cytokines in peripheral blood mononuclear cell (PBMC) cultures. PBMCs from Schistosoma mansoni-infected patients with asthma living in an area of polyhelminthic endemicity produced lower levels of interleukin (IL)-5 and IL-4 in response to D. pteronyssinus antigen (Ag) 1 than did PBMCs from helminth-free patients with asthma. In contrast, D. pteronyssinus Ag 1-specific production of IL-10 was higher in helminth-infected patients than in helminth-free patients. The addition of recombinant human IL-10 to D. pteronyssinus Ag 1-stimulated cultures of PBMCs from helminth-free patients led to down-modulation of production of IL-5. After helminth-infected patients with asthma received antihelminthic treatment, there was down-modulation of D. pteronyssinus Ag 1-specific production of IL-10 in vitro. S. mansoni-infected patients with asthma produce lower levels of Th2 cytokines than do helminth-free patients with asthma, and this modulation is likely done by IL-10.

Schistosoma mansoni antigens modulate the allergic response in a murine model of ovalbumin-induced airway inflammation

Schistosoma mansoni infection has been associated with protection against allergies. The mechanisms underlying this association may involve regulatory cells and cytokines. We evaluated the immune response induced by the S. mansoni antigens Sm22·6, PIII and Sm29 in a murine model of ovalbumin (OVA)‐induced airway inflammation. BALB/c mice were sensitized with subcutaneously injected OVA‐alum and challenged with aerolized OVA. Mice were given three doses of the different S. mansoni antigens. Lung histopathology, cellularity of bronchoalveolar lavage (BAL) and eosinophil peroxidase activity in lung were evaluated. Immunoglobulin (Ig)E levels in serum and cytokines in BAL were also measured. Additionally, we evaluated the frequency of CD4+forkhead box P3 (FoxP3)+ T cells in cultures stimulated with OVA and the expression of interleukin (IL)‐10 by these cells. The number of total cells and eosinophils in BAL and the levels of OVA‐specific IgE were reduced in the immunized mice. Also, the levels of IL‐4 and IL‐5 in the BAL of mice immunized with PIII and Sm22·6 were decreased, while the levels of IL‐10 were higher in mice immunized with Sm22·6 compared to the non‐immunized mice. The frequency of CD4+FoxP3+ T cells was higher in the groups of mice who received Sm22·6, Sm29 and PIII, being the expression of IL‐10 by these cells only higher in mice immunized with Sm22·6. We concluded that the S. mansoni antigens used in this study are able to down‐modulate allergic inflammatory mediators in a murine model of airway inflammation and that the CD4+FoxP3+ T cells, even in the absence of IL‐10 expression, might play an important role in this process.

Gene Encoding Duffy Antigen/Receptor for Chemokines Is Associated with Asthma and IgE in Three Populations

RATIONALE Asthma prevalence and severity are high among underserved minorities, including those of African descent. The Duffy antigen/receptor for chemokines is the receptor for Plasmodium vivax on erythrocytes and functions as a chemokine-clearing receptor. Unlike European populations, decreased expression of the receptor on erythrocytes is common among populations of African descent, and results from a functional T-46C polymorphism (rs2814778) in the promoter. This variant provides an evolutionary advantage in malaria-endemic regions, because Duffy antigen/receptor for chemokines-negative erythrocytes are more resistant to infection by P. vivax. OBJECTIVES To determine the role of the rs2814778 polymorphism in asthma and atopy as measured by total serum IgE levels among four populations of African descent (African Caribbean, African American, Brazilian, and Colombian) and a European American population. METHODS Family-based association tests were performed in each of the five populations to test for association between the rs2814778 polymorphism and asthma or total IgE concentration. MEASUREMENTS AND MAIN RESULTS Asthma was significantly associated with the rs2814778 polymorphism in the African Caribbean, Colombian, and Brazilian families (P < 0.05). High total IgE levels were associated with this variant in African Caribbean and Colombian families (P < 0.05). The variant allele was not polymorphic among European Americans. CONCLUSIONS Susceptibility to asthma and atopy among certain populations of African descent is influenced by a functional polymorphism in the gene encoding Duffy antigen/receptor for chemokines. This genetic variant, which confers resistance to malarial parasitic infection, may also partially explain ethnic differences in morbidity of asthma.

Disparate Immunoregulatory Potentials for Double-Negative (CD4− CD8−) αβ and γδ T Cells from Human Patients with Cutaneous Leishmaniasis

ABSTRACT Although most T lymphocytes express the αβ T-cell receptor and either CD4 or CD8 molecules, a small population of cells lacking these coreceptors, CD4− CD8− (double negative [DN]) T cells, has been identified in the peripheral immune system of mice and humans. To better understand the role that this population may have in the human immune response against Leishmania spp., a detailed study defining the activation state, cytokine profile, and the heterogeneity of DN T cells bearing αβ or γδ T-cell receptors was performed with a group of well-defined cutaneous leishmaniasis patients. Strikingly, on average 75% of DN T cells from cutaneous leishmaniasis patients expressed the αβ T-cell receptor, with the remainder expressing the γδ receptor, while healthy donors displayed the opposite distribution with ∼75% of the DN T cells expressing the γδ T-cell receptor. Additionally, αβ DN T cells from cutaneous leishmaniasis patients are compatible with previous antigen exposure and recent activation. Moreover, while αβ DN T cells from Leishmania-infected individuals present a proinflammatory cytokine profile, γδ DN T cells express a regulatory profile exemplified by interleukin-10 production. The balance between these subpopulations could allow for the formation of an effective cellular response while limiting its pathogenic potential.

African Ancestry is a Risk Factor for Asthma and High Total IgE Levels in African Admixed Populations

Characterization of genetic admixture of populations in the Americas and the Caribbean is of interest for anthropological, epidemiological, and historical reasons. Asthma has a higher prevalence and is more severe in populations with a high African component. Association of African ancestry with asthma has been demonstrated. We estimated admixture proportions of samples from six trihybrid populations of African descent and determined the relationship between African ancestry and asthma and total serum IgE levels (tIgE). We genotyped 237 ancestry informative markers in asthmatics and nonasthmatic controls from Barbados (190/277), Jamaica (177/529), Brazil (40/220), Colombia (508/625), African Americans from New York (207/171), and African Americans from Baltimore/Washington, D.C. (625/757). We estimated individual ancestries and evaluated genetic stratification using Structure and principal component analysis. Association of African ancestry and asthma and tIgE was evaluated by regression analysis. Mean ± SD African ancestry ranged from 0.76 ± 0.10 among Barbadians to 0.33 ± 0.13 in Colombians. The European component varied from 0.14 ± 0.05 among Jamaicans and Barbadians to 0.26 ± 0.08 among Colombians. African ancestry was associated with risk for asthma in Colombians (odds ratio (OR) = 4.5, P = 0.001) Brazilians (OR = 136.5, P = 0.003), and African Americans of New York (OR: 4.7; P = 0.040). African ancestry was also associated with higher tIgE levels among Colombians (β = 1.3, P = 0.04), Barbadians (β = 3.8, P = 0.03), and Brazilians (β = 1.6, P = 0.03). Our findings indicate that African ancestry can account for, at least in part, the association between asthma and its associated trait, tIgE levels.

Skin test reactivity and Der p-induced interleukin 10 production in patients with asthma or rhinitis infected with Ascaris

BACKGROUND Helminth infections have been associated with reduced reactivity to aeroallergens, which could be related to interleukin 10 (IL-10) production, as reported in schistosomiasis. OBJECTIVE To compare skin responses to aeroallergens with Der p specific IL-10 production in patients with asthma or rhinitis according to Ascaris infection status. METHODS Cross-sectional study of 113 patients with asthma or rhinitis from a region endemic for geohelminths. Stool examinations and skin prick tests to aeroallergens were performed in all the patients. Der p specific IL-10 production was measured in the supernatant of peripheral blood mononuclear cell (PBMC) cultures of a subsample of 53 patients. RESULTS Forty-seven patients had Ascaris in their stool samples. Skin test results were positive in 77% of Ascaris-infected patients and in 71% of uninfected individuals. Median levels of Der p specific IL- 10 in PBMC cultures of infected and uninfected patients were similar (7.8 and 8.4 pg/mL, respectively). The lack of association remained when parasite load was taken into account and when patients with evidence of previous infection were removed from the uninfected group. CONCLUSIONS In patients with asthma or rhinitis living in an urban area endemic for geohelminths, we found no association between Ascaris infection and skin reactivity to aeroallergens. Furthermore, there was no difference in Der p specific IL-10 production by PBMCs. These negative findings indicate that different from what is observed in Schistosoma infection, Ascaris lumbricoides infection in individuals living in an urban area does not induce strong regulatory responses.

A continuum of admixture in the Western Hemisphere revealed by the African Diaspora genome

The African Diaspora in the Western Hemisphere represents one of the largest forced migrations in history and had a profound impact on genetic diversity in modern populations. To date, the fine-scale population structure of descendants of the African Diaspora remains largely uncharacterized. Here we present genetic variation from deeply sequenced genomes of 642 individuals from North and South American, Caribbean and West African populations, substantially increasing the lexicon of human genomic variation and suggesting much variation remains to be discovered in African-admixed populations in the Americas. We summarize genetic variation in these populations, quantifying the postcolonial sex-biased European gene flow across multiple regions. Moreover, we refine estimates on the burden of deleterious variants carried across populations and how this varies with African ancestry. Our data are an important resource for empowering disease mapping studies in African-admixed individuals and will facilitate gene discovery for diseases disproportionately affecting individuals of African ancestry.

High Heritability but Uncertain Mode of Inheritance for Total Serum IgE Level and Schistosoma mansoni Infection Intensity in a Schistosomiasis-Endemic Brazilian Population

BACKGROUND Evidence of genetic control for total serum IgE (tIgE) level has been reported in multiple populations, although populations with substantial exposure to helminths have yielded lower estimates of heritability, despite evidence suggesting that genes also control a significant portion of the variation in the number of Schistosoma mansoni eggs per gram of fecal matter. METHODS By use of a whole-population ascertainment scheme, 822 individuals were enrolled from a schistosomiasis-endemic area in Conde, Bahia, in Brazil. Heritability was estimated by using an additive polygenic model, and segregation analysis was performed for 2 quantitative traits, tIgE level and egg count. RESULTS After adjusting for nongenetic covariates, the heritability of log-transformed tIgE level and log-transformed egg count was estimated at 60% and 31%, respectively. No evidence for a single major gene controlling tIgE level or egg count was observed in segregation analysis for 781 individuals and 403 individuals, respectively, in 318 families, however, which suggests complex biological control. CONCLUSIONS The high heritability of tIgE level indicates that genetic factors are likely to control tIgE level even in the presence of helminthic infection. Substantial heritability for the burden of S. mansoni infection was confirmed in these Brazilian families. Further genetic studies will be needed to dissect the specific genetic factors that underlie these traits.

Schistosoma mansoni antigen-driven interleukin-10 production in infected asthmatic individuals

UNLABELLED Asthmatics infected with Schistosoma mansoni have a less severe course of asthma and an inhibition of the Th2 inflammatory response that seems to be mediated by interleukin (IL-10). The objective of this study was to evaluate the capacity of some S. mansoni antigens to stimulate IL-10 production in vitro by cells of asthmatic infected individuals. Peripheral bloods mononuclear cells were stimulated with the S. mansoni recombinant antigens Sm22.6, Sm14, P24, and PIII antigen. IL-10 was measured in the supernatants of cultures. As the recombinant antigens were cloned in Escherichia coli, we blocked contaminant endotoxin with polymyxin B added to the cultures. We demonstrated that all antigens used drove high production of IL-10 in S. mansoni infected individuals (n = 13, 408 +/- 514 and 401 +/- 383 pg/ml, 484 +/- 245 pg/ml, 579 +/- 468 pg/ml, respectively). In asthmatics infected with S. mansoni (n = 21) rP24 induced higher levels of IL-10 (565 +/- 377 pg/ml) when compared to PIII, rSm14 and rSm22.6 (184 +/- 209 pg/ml; 292 +/- 243 pg/ml; 156 +/- 247 pg/ml, respectively). CONCLUSION the S. mansoni antigens evaluated in this study stimulated IL-10 production by cells from infected individuals and therefore they have the potential to be used as a modulator of the inflammatory response in asthma.