Karine Almeida

Modulation of mucosal/systemic antibody response after sublingual immunotherapy in mite-allergic children

There have been no data on sublingual immunotherapy (SLIT) in Brazilian patients sensitized to house dust mites. This study aimed to evaluate the mucosal/systemic antibody response changes and clinical efficacy after SLIT using Dermatophagoides pteronyssinus (Dpt) allergens with or without bacterial extracts in mite‐allergic Brazilian children.

Immunoproteomics of Brucella abortus reveals differential antibody profiles between S19-vaccinated and naturally infected cattle

Brucella abortus is a Gram‐negative intracellular bacterium that causes infectious abortion in food‐producing animals and chronic infection in humans. This study aimed to characterize a B. abortus S19 antigen preparation obtained by Triton X‐114 (TX‐114) extraction through immunoproteomics to differentiate infected from vaccinated cattle. Three groups of bovine sera were studied: GI, 30 naturally infected cows; GII, 30 S19‐vaccinated heifers; and GIII, 30 nonvaccinated seronegative cows. One‐dimensional (1D) and two‐dimensional electrophoretic profiles of TX‐114 hydrophilic phase antigen revealed a broad spectrum of polypeptides (10–79 kDa). 1D immunoblot showed widespread seroreactivity profile in GI compared with restricted profile in GII. Three antigenic components (10, 12, 17 kDa) were recognized exclusively by GI sera, representing potential markers of infection and excluding vaccinal response. The proteomic characterization revealed 56 protein spots, 27 of which were antigenic spots showing differential seroreactivity profile between GI and GII, especially polypeptides <20 kDa that were recognized exclusively by GI. MS/MS analysis identified five B. abortus S19 proteins (Invasion protein B, Sod, Dps, Ndk, and Bfr), which were related with antigenicity in naturally infected cattle. In conclusion, immunoproteomics of this new antigen preparation enabled the characterization of proteins that could be used as tools to develop sensitive and specific immunoassays for serodiagnosis of bovine brucellosis, with emphasis on differentiation between S19 vaccinated and infected cattle.

Responses of IgE, IgG1, and IgG4 to concanavalin A-binding Blomia tropicalis antigens in allergic patients

Blomia tropicalis (Bt) and Dermatophagoides pteronyssinus (Dp) are the prevalent house dust mites in tropical countries and are associated with allergic diseases. Glycosylated antigens are highly immunogenic and involved in different pathologies. We evaluated the presence of IgE, IgG1, and IgG4 to concanavalin A-binding antigens (Bt-Con-A) isolated from Bt-total extract in sera of allergic and non-allergic subjects. Bt-total and Bt-Con-A extracts were evaluated by SDS-PAGE and ELISA for reacting with IgE, IgG1, and IgG4 in sera of 121 patients with allergic rhinitis and 36 non-allergic individuals. All subjects were skin prick tested with Bt-total extract and inhibition tests were performed for IgE, IgG1, and IgG4 using both extracts (Bt-total and Bt-Con-A). Skin prick test showed that 58% of the patients were sensitized to Bt (Bt+), with 52% reactive to both mites (Bt and Dp) and 6% to Bt only. A broad spectrum of proteins (14-152 kDa) was visualized in Bt-total and components >27 kDa for the Bt-Con-A extract. ELISA showed a similar profile of IgE, IgG1 and IgG4 levels in response to Bt-total and Bt-Con-A extracts in different groups, although Bt+ patients showed a lower IgG4 reactivity to Bt-Con-A extract. Specific IgG1 levels were higher in Bt+ patients than in control subjects, and IgG4 levels showed no significant difference among groups. ELISA inhibition showed a partial IgE and total IgG1 and IgG4 cross-reactivity with Dp extract for Bt-total and Bt-Con-A extracts. We conclude that Con-A-binding components isolated from Bt constitute major allergens and are involved in both allergen sensitization (IgE response) and homeostasis maintenance (IgG1 and IgG4 responses).

Humoral and cellular immune responses to Blomia tropicalis and concanavalin A-binding fractions in atopic patients.

Department of Medicine, Center for Immunity, Inflammation and Regenerative Medicine, University ofVirginia, Charlottesville, VA, USACorrespondence to: E.A. Taketomi, Laboratorio de Alergia e Imunologia Clinica, ICBIM, UFU, Av. Para,1720, Bloco 4C, Campus Umuarama, 38400-902 Uberlândia, MG, BrasilFax: +55-34-3232-8620. E-mail: eat4y@yahoo.com.br

Toxoplasma gondii 70 kDa Heat Shock Protein: Systemic Detection Is Associated with the Death of the Parasites by the Immune Response and Its Increased Expression in the Brain Is Associated with Parasite Replication

The heat shock protein of Toxoplasma gondii (TgHSP70) is a parasite virulence factor that is expressed during T. gondii stage conversion. To verify the effect of dexamethasone (DXM)-induced infection reactivation in the TgHSP70-specific humoral immune response and the presence of the protein in the mouse brain, we produced recombinant TgHSP70 and anti-TgHSP70 IgY antibodies to detect the protein, the specific antibody and levels of immune complexes (ICs) systemically, as well as the protein in the brain of resistant (BALB/c) and susceptible (C57BL/6) mice. It was observed higher TgHSP70-specific antibody titers in serum samples of BALB/c compared with C57BL/6 mice. However, the susceptible mice presented the highest levels of TgHSP70 systemically and no detection of specific ICs. The DXM treatment induced increased parasitism and lower inflammatory changes in the brain of C57BL/6, but did not interfere with the cerebral parasitism in BALB/c mice. Additionally, DXM treatment decreased the serological TgHSP70 concentration in both mouse lineages. C57BL/6 mice presented high expression of TgHSP70 in the brain with the progression of infection and under DXM treatment. Taken together, these data indicate that the TgHSP70 release into the bloodstream depends on the death of the parasites mediated by the host immune response, whereas the increased TgHSP70 expression in the brain depends on the multiplication rate of the parasite.

Macrophage migration inhibitory factor (MIF) prevents maternal death, but contributes to poor fetal outcome during congenital toxoplasmosis

Migration inhibitory factor (MIF) is a pro-inflammatory cytokine that plays important roles in physiology, pathology, immunology and parasitology, including the control of infection by protozoa parasites such as Toxoplasma gondii. As the MIF function in congenital toxoplasmosis is not fully elucidated yet, the present study brings new insights for T. gondii infection in the absence of MIF based on pregnant C57BL/6MIF-/- mouse models. Pregnant C57BL/6MIF-/- and C57BL/6WT mice were infected with 05 cysts of T. gondii (ME49 strain) on the first day of pregnancy (dop) and were euthanized at 8 dop. Non-pregnant and non-infected females were used as control. Our results demonstrated that MIF-/- mice have more accentuated change in body weight and succumbed to infection first than their WT counterparts. Otherwise, pregnancy outcome was less destructive in MIF-/- mice compared to WT ones, and the former had an increase in the mast cell recruitment and IDO expression and consequently presented less inflammatory cytokine production. Also, MIF receptor (CD74) was upregulated in MIF-/- mice, indicating that a compensatory mechanism may be required in this model of study. The global absence of MIF was associated with attenuation of pathology in congenital toxoplasmosis, but resulted in female death probably because of uncontrolled infection. Altogether, ours results demonstrated that part of the immune response that protects a pregnant female from T. gondii infection, favors fetal damage.

201 Clinical Efficacy and Mucosal/Systemic Antibody Response Changes After Sublingual Immunotherapy in Mite-Allergic Children: A Randomized Double-Blind, Placebo-Controlled Study in Brazil.

Background This study aimed to evaluate the clinical efficacy and mucosal/systemic antibody response changes after sublingual immunotherapy (SLIT) using Dermatophagoides pteronyssinus (Dpt) allergens with or without bacterial extracts in mite-allergic Brazilian children. Methods One-hundred and 2 patients presenting allergic rhinitis with or without asthma were selected for a randomized double-blind, placebo-controlled trial and distributed into 3 groups: DPT (Dpt allergen extract, n = 34), DPT + MRB (Dpt allergen plus mixed respiratory bacterial extracts, n = 36), and Placebo (n = 32). Clinical evaluation and immunological analyses were carried out before and after 12 and 18 months of treatment, including rhinitis/asthma symptom and medication scores, skin prick test (SPT) to Dpt extract, and measurements of Dpt-, Der p 1-, Der p 2-specific IgE, IgG4, and IgG1 in serum and -specific IgA in saliva and nasal lavage fluid. Results Clinical results showed a significant decline in rhinitis/asthma symptom scores in all groups, but medication use decreased only in active DPT group at 12 months. SPT results showed no significant changes and SLIT was generally safe, with no severe systemic reactions. SLIT using Dpt allergen alone induced increased serum IgG4 levels to Dpt, Der p 1 and Der p 2, and increased serum IgG1 and salivary IgA levels to Dpt and Der p 1. SLIT using DPT+MRB was able to decrease IgE levels, particularly to Der p 2, to increase salivary IgA levels to Der p 1, but had no changes on specific IgG4 and IgG1 levels. Conclusions Therefore, SLIT seems to be effective in ameliorating clinical symptoms, but only active SLIT was able to modulate the mucosal and systemic antibody responses. These findings support the role of specific serum IgG4 and IgG1, in addition to salivary IgA, as protective or blocking antibodies as well as biomarkers of tolerance that may be useful for monitoring activation of tolerance-inducing mechanisms during allergen immunotherapy.