Freitas Mateus S, Oliveira Aline F, da Silva Thiago A, Fernandes Fabrício F, Gonçales Relber A, Almeida Fausto, Roque-Barreira Maria C
Departamento de Biologia Celular e Molecular e Bioagentes Patogênicos, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo Ribeirão Preto, Brazil.
Front Microbiol. 2016 Jul 5;7:1003. doi: 10.3389/fmicb.2016.01003. eCollection 2016.
The fungal human pathogen Paracoccidioides brasiliensis contains paracoccin (PCN), a multi-domain protein that has lectin and N-acetyl-glucosaminidase activities, which account for its effects on the growth and morphogenesis of the fungus and on the activation of host macrophages through its interaction with TLR N-glycans. With the purpose of detailing the knowledge on the effects of PCN on macrophages, we used recombinant PCN expressed in Pichia pastoris (p-rPCN) to stimulate isolated murine peritoneal macrophages. The activation of these cells manifested through the release of high levels of inflammatory mediators, such as nitric oxide, TNF-α, IL-12p40, and IL-6. Furthermore, peritoneal macrophages stimulated with p-rPCN increased the relative expression of STAT1, SOCS3, and iNOS2 mRNA (M1 polarization markers). However, the expression of Arginase-1, Ym-1, and FIZZ1 (M2 polarization markers) remained at basal levels. Interestingly, the observed M1 macrophages' polarization triggered by p-rPCN was abolished in cells obtained from knockout Toll-like receptor-4 mice. In this case, the p-rPCN-induced production of pro-inflammatory mediators was blocked too. These results demonstrate that the classical activation of macrophages induced by paracoccin depends on TLR4. Taken together, the results of our study indicate that paracoccin acts as a TLR agonist able to modulate immunity and exerts biological activities that favor its applicability as an immunotherapeutic agent to combat systemic fungal infections.
巴西副球孢子菌是一种人类真菌病原体，它含有副球孢子菌素（PCN），这是一种多结构域蛋白，具有凝集素和N - 乙酰葡糖胺酶活性，这些活性解释了其对真菌生长和形态发生以及通过与Toll样受体（TLR）N - 聚糖相互作用对宿主巨噬细胞激活的影响。为了详细了解PCN对巨噬细胞的影响，我们使用在毕赤酵母中表达的重组PCN（p - rPCN）来刺激分离的小鼠腹腔巨噬细胞。这些细胞的激活表现为释放高水平的炎症介质，如一氧化氮、肿瘤坏死因子 - α、白细胞介素 - 12p40和白细胞介素 - 6。此外，用p - rPCN刺激的腹腔巨噬细胞增加了信号转导和转录激活因子1（STAT1）、细胞因子信号转导抑制因子3（SOCS3）和诱导型一氧化氮合酶2（iNOS2）mRNA（M1极化标志物）的相对表达。然而，精氨酸酶 - 1、Ym - 1和抵抗素样分子α（FIZZ1）（M2极化标志物）的表达保持在基础水平。有趣的是，在从Toll样受体 - 4基因敲除小鼠获得的细胞中，由p - rPCN触发的观察到的M1巨噬细胞极化被消除。在这种情况下，p - rPCN诱导的促炎介质产生也被阻断。这些结果表明，副球孢子菌素诱导的巨噬细胞经典激活依赖于TLR4。综上所述，我们的研究结果表明，副球孢子菌素作为一种能够调节免疫的TLR激动剂，发挥着有利于其作为免疫治疗剂对抗系统性真菌感染的生物活性。
