Extracellular vesicles from Trypanosoma cruzi-dendritic cell interaction show modulatory properties and confer resistance to lethal infection as a cell-free based therapy strategy

Extracellular vesicles (EVs) include a heterogeneous group of particles. Microvesicles and exosomes are the most characterised vesicles. They can be distinguished by their size, morphology, origin and molecular composition. To date, increasing studies demonstrate that EVs mediates intercellular communication. EVs reach considerable interest in the scientific community due to their role in diverse processes including antigen-presentation, stimulation of anti-tumoral immune responses, tolerogenic or inflammatory effects. In pathogens, EVs shedding is well described in fungus, bacteria, protozoan parasites and helminths. For Trypanosoma cruzi EVs liberation and protein composition was previously described. Dendritic cells (DCs) are key players promoting the immune response against pathogens and also maintaining self-tolerance. In previous reports we have demonstrate that T. cruzi downregulates DCs immunogenicity in vitro and in vivo. Here we analyze EVs from the in vitro interaction between blood circulating tripomastigotes (Tp) and bone-marrow-derived DCs. We found that Tp incremented the number and the size of EVs in cultures with DCs. EVs displayed some exosome markers and intracellular RNA. Protein analysis demonstrated that the parasite changes the protein-EV profile of DCs. We observed that EVs from the interaction of Tp-DCs were easily captured by unstimulated-DCs in comparison with EVs from DCs cultured without the parasite, and also modify the activation status of LPS-stimulated DCs. Noteworthy, we found protection in animal treated with EVs DCs+Tp and challenged with T. cruzi lethal infection. Our goal is to go deep into the molecular characterization of EVs from the DCs-Tp interaction, in order to identify mediators for therapeutic purposes.

细胞外囊泡（Extracellular Vesicles, EVs）是一类异质性颗粒群。微囊泡（Microvesicles）与外泌体（exosomes）是目前研究最为深入的两类囊泡，可通过尺寸、形态、来源及分子组成进行区分。迄今为止，越来越多的研究表明，细胞外囊泡可介导细胞间通讯。因其在抗原呈递、抗肿瘤免疫应答激活、免疫耐受或炎症效应调控等多种生理病理过程中发挥关键作用，细胞外囊泡受到科学界的广泛关注。在病原体领域，真菌、细菌、原生动物寄生虫及蠕虫的细胞外囊泡释放过程已得到充分阐释；针对克氏锥虫（Trypanosoma cruzi）的细胞外囊泡释放及其蛋白组成，此前亦有相关报道。
树突状细胞（Dendritic Cells, DCs）是介导抗病原体免疫应答并维持自身耐受的核心免疫细胞。在既往研究中，我们已证实克氏锥虫可在体内外下调树突状细胞的免疫原性。本研究针对血液循环型锥鞭毛体（tripomastigotes, Tp）与骨髓来源树突状细胞的体外互作所产生的细胞外囊泡展开分析。研究发现，与仅培养树突状细胞的体系相比，加入锥鞭毛体的共培养体系中，细胞外囊泡的数量与尺寸均显著增加；该囊泡携带部分外泌体标志物及胞内RNA。蛋白质组分析结果显示，克氏锥虫可改变树突状细胞所分泌细胞外囊泡的蛋白谱。我们观察到，相较于未接触寄生虫的树突状细胞所分泌的囊泡，Tp-DC互作产生的细胞外囊泡更易被未活化的树突状细胞摄取，同时还可改变脂多糖（Lipopolysaccharide, LPS）活化树突状细胞的活化状态。值得注意的是，经DCs+Tp来源细胞外囊泡处理的实验动物，在受到克氏锥虫致死性感染攻击时可获得保护。本研究旨在深入解析DCs-Tp互作来源细胞外囊泡的分子特征，以筛选可用于治疗的相关介导因子。