Ovariectomized OVA-sensitized mice display increased frequency of CD4+Foxp3+ T regulatory cells in the periphery

It is well established that female sex hormones have a pivotal role in inflammation. For instance, our group has previously reported that estradiol has proinflammatory actions during allergic lung response in animal models. Based on these findings, we have decided to further investigate whether T regulatory cells are affected by female sex hormones absence after ovariectomy. We evaluated by flow cytometry the frequencies of CD4+Foxp3+ T regulatory cells (Tregs) in central and peripheral lymphoid organs, such as the thymus, spleen and lymph nodes. Moreover, we have also used the murine model of allergic lung inflammation a to evaluate how female sex hormones would affect the immune response in vivo. To address that, ovariectomized or sham operated female Balb/c mice were sensitized or not with ovalbumin 7 and 14 days later and subsequently challenged twice by aerosolized ovalbumin on day 21. Besides the frequency of CD4+Foxp3+ T regulatory cells, we also measured the cytokines IL-4, IL-5, IL-10, IL-13 and IL-17 in the bronchoalveolar lavage from lungs of ovalbumine challenged groups. Our results demonstrate that the absence of female sex hormones after ovariectomy is able to increase the frequency of Tregs in the periphery. As we did not observe differences in the thymus-derived natural occurring Tregs, our data may indicate expansion or conversion of peripheral adaptive Tregs. In accordance with Treg suppressive activity, ovariectomized and ovalbumine-sensitized and challenged animals had significantly reduced lung inflammation. This was observed after cytokine analysis of lung explants showing significant reduction of pro-inflammatory cytokines, such as IL-4, IL-5, IL-13 and IL-17, associated to increased amount of IL-10. In summary, our data clearly demonstrates that OVA sensitization 7 days after ovariectomy culminates in reduced lung inflammation, which may be directly correlated with the expansion of Tregs in the periphery and further higher IL-10 secretion in the lungs.

Effects of Aedes aegypti salivary components on dendritic cell and lymphocyte biology

BACKGROUND: Saliva is a key element of interaction between hematophagous mosquitoes and their vertebrate hosts. In addition to allowing a successful blood meal by neutralizing or delaying hemostatic responses, the salivary cocktail is also able to modulate the effector mechanisms of host immune responses facilitating, in turn, the transmission of several types of microorganisms. Understanding how the mosquito uses its salivary components to circumvent host immunity might help to clarify the mechanisms of transmission of such pathogens and disease establishment. METHODS: Flow cytometry was used to evaluate if increasing concentrations of A. aegypti salivary gland extract (SGE) affects bone marrow-derived DC differentiation and maturation. Lymphocyte proliferation in the presence of SGE was estimated by a colorimetric assay. Western blot and Annexin V staining assays were used to assess apoptosis in these cells. Naïve and memory cells from mosquito-bite exposed mice or OVA-immunized mice and their respective controls were analyzed by flow cytometry. RESULTS: Concentration-response curves were employed to evaluate A. aegypti SGE effects on DC and lymphocyte biology. DCs differentiation from bone marrow precursors, their maturation and function were not directly affected by A. aegypti SGE (concentrations ranging from 2.5 to 40 μg/mL). On the other hand, lymphocytes were very sensitive to the salivary components and died in the presence of A. aegypti SGE, even at concentrations as low as 0.1 μg/mL. In addition, A. aegypti SGE was shown to induce apoptosis in all lymphocyte populations evaluated (CD4+ and CD8+ T cells, and B cells) through a mechanism involving caspase-3 and caspase-8, but not Bim. By using different approaches to generate memory cells, we were able to verify that these cells are resistant to SGE effects. CONCLUSION: Our results show that lymphocytes, and not DCs, are the primary target of A. aegypti salivary components. In the presence of A. aegypti SGE, naïve lymphocyte populations die by apoptosis in a caspase-3- and caspase-8-dependent pathway, while memory cells are selectively more resistant to its effects. The present work contributes to elucidate the activities of A. aegypti salivary molecules on the antigen presenting cell-lymphocyte axis and in the biology of these cells.