Galdino Hélio, Saar Gomes Rodrigo, Dos Santos Jessica Cristina, Pessoni Lívia Lara, Maldaner Anetícia Eduarda, Marques Stéfanne Madalena, Gomes Clayson Moura, Dorta Miriam Leandro, de Oliveira Milton Adriano Pelli, Joosten Leo A B, Ribeiro-Dias Fátima
Instituto de Patologia Tropical e Saúde Pública, Universidade Federal de Goiás, Goiânia, Goiás, Brazil.
Instituto de Patologia Tropical e Saúde Pública, Universidade Federal de Goiás, Goiânia, Goiás, Brazil; Department of Internal Medicine, Radboud University Medical Center and Radboud Center of Infectious Diseases (RCI), Nijmegen, The Netherlands.
Cytokine. 2016 Dec;88:184-192. doi: 10.1016/j.cyto.2016.09.009. Epub 2016 Sep 17.
While the role of Toll-like receptors (TLRs) has been investigated in murine models of tegumentary leishmaniasis caused by Leishmania (Viannia) braziliensis, the interaction between TLRs and Leishmania sp. has not been investigated in human cells. The aim of this study was to evaluate the involvement of TLR4 in cytokine production of human peripheral blood mononuclear cells (PBMCs) induced by L. braziliensis, and whether the parasite alters the expression of TLR4 on monocytes/macrophages. Amastigote forms were obtained from mice lesions and PBMCs were isolated from healthy donors. PBMCs were cultured in absence or presence of IFNγ, TLR4 neutralizing antibodies, natural antagonist of TLR4 (Bartonella LPS), TLR4 agonist (E. coli LPS), and amastigote forms. The concentrations of tumor necrosis factor (TNFα) and interleukin 10 (IL-10) were assayed by ELISA and TLR4 expression by flow cytometry. Amastigotes forms of L. braziliensis induced TNFα and IL-10 production only in IFNγ-primed PBMCs. The TNFα and IL-10 production was inhibited by TLR4 neutralization, both with anti-TLR4 antibodies and Bartonella LPS. Interestingly, addition of E. coli LPS further increased TNFα but not IL-10 production induced by L. braziliensis amastigotes. Amastigotes of L. braziliensis strongly reduced membrane TLR4 expression on monocytes/macrophages, apparently by internalization after the infection. The present study reveals that TLR4 drives the production of TNFα and IL-10 induced by L. braziliensis amastigotes and that the parasites decrease TLR4 expression on monocyte surface.
虽然Toll样受体（TLRs）在巴西利什曼原虫（Leishmania (Viannia) braziliensis）引起的皮肤利什曼病小鼠模型中的作用已得到研究，但TLRs与利什曼原虫属之间的相互作用尚未在人类细胞中进行研究。本研究的目的是评估TLR4在巴西利什曼原虫诱导的人外周血单核细胞（PBMCs）细胞因子产生中的作用，以及该寄生虫是否会改变单核细胞/巨噬细胞上TLR4的表达。从小鼠病变中获取无鞭毛体形式，从健康供体中分离PBMCs。将PBMCs在不存在或存在IFNγ、TLR4中和抗体、TLR4天然拮抗剂（巴尔通体LPS）、TLR4激动剂（大肠杆菌LPS）和无鞭毛体形式的情况下进行培养。通过ELISA测定肿瘤坏死因子（TNFα）和白细胞介素10（IL-10）的浓度，并通过流式细胞术测定TLR4的表达。巴西利什曼原虫的无鞭毛体形式仅在IFNγ预处理的PBMCs中诱导TNFα和IL-10的产生。TNFα和IL-10的产生通过抗TLR4抗体和巴尔通体LPS的TLR4中和作用受到抑制。有趣的是，添加大肠杆菌LPS进一步增加了巴西利什曼原虫无鞭毛体诱导的TNFα产生，但未增加IL-10的产生。巴西利什曼原虫的无鞭毛体明显通过感染后的内化作用，强烈降低了单核细胞/巨噬细胞上膜TLR4的表达。本研究表明，TLR4驱动巴西利什曼原虫无鞭毛体诱导的TNFα和IL-10的产生，并且该寄生虫会降低单核细胞表面TLR4的表达。
