Laboratory of Immunoparasitology, Oswaldo Cruz Institute, FIOCRUZ, Rio de Janeiro, Rio de Janeiro, Brazil.
Flow Cytometry Core Facility, Oswaldo Cruz Institute, FIOCRUZ, Rio de Janeiro, Rio de Janeiro, Brazil.
PLoS One. 2020 Mar 23;15(3):e0229400. doi: 10.1371/journal.pone.0229400. eCollection 2020.
The pathogenesis of cutaneous leishmaniasis (CL) caused by Leishmania (Viannia) braziliensis is dictated mainly by the immune-mediated-tissue inflammation developed. The understanding of the immunological mechanisms that generate tissue damage or resolution of lesions is the key to the development of effective vaccine protocols and proper therapeutic schemes. It is clear that the specific immune response mediated by T cells is responsible for the beneficial outcome of the disease, however, the roles of CD4+ T, CD8+ T, NK and NKT cell subpopulations in immunopathogenesis of CL need to be elucidated. Peripheral blood cells from patients before, during and after the antimonial therapy, as well as healthy individuals (HI) were cultured with (LbAgS) or without (NS) L. braziliensis antigens (LbAg). Afterwards, the frequencies of LbAg-specific-cytotoxic CD8+ T, CD4+ T, NK and CD3+CD56+ NKT cells, as well as their activation and exhaustion profiles, were defined by flow cytometry. We observed higher frequencies of CD8+ T, NK and CD3+CD56+ NKT cells and lower frequencies of CD4+ T lymphocytes in LbAgS cell cultures from patients before treatment. The specific response to LbAg resulted in an expansion of cytotoxic-activated CD4+ T, CD8+ T, and NK cells, before and during treatment, indicating specificity in the response by these cells against L. braziliensis. Furthermore, comparing the differences of frequencies of cytotoxic-activated CD4+T, CD8+T, and NK cells, among before and during treatment patients and HI groups, we conclude that these cell populations are in charge of immune response elicited by antimonial therapy. Interestingly, we also observed that NK cells were induced by LbAg to an exhaustion profile during all clinical stages of the disease. The increased antigen-specific activation and cytotoxic activity are in line with the strong inflammatory response described in this disease, a likely cause of tissue damage. These findings reinforce the involvement of these distinct cytotoxic-activated cell populations in the immunopathogenesis of CL, showing a character of specificity in this immune response.
巴西利什曼原虫引起的皮肤利什曼病（CL）的发病机制主要由免疫介导的组织炎症发展决定。了解产生组织损伤或病变消退的免疫机制是开发有效疫苗方案和适当治疗方案的关键。显然，T 细胞介导的特异性免疫反应是疾病有益结局的原因，然而，CD4+T、CD8+T、NK 和 NKT 细胞亚群在 CL 免疫发病机制中的作用仍需阐明。在锑剂治疗前、治疗中和治疗后，以及健康个体（HI）中，从患者的外周血细胞中培养了（LbAgS）或没有（NS）巴西利什曼原虫抗原（LbAg）。随后，通过流式细胞术定义了 LbAg 特异性细胞毒性 CD8+T、CD4+T、NK 和 CD3+CD56+NKT 细胞的频率以及它们的激活和耗竭特征。我们观察到，在治疗前，与 NS 细胞培养相比，LbAgS 细胞培养中的 CD8+T、NK 和 CD3+CD56+NKT 细胞频率更高，CD4+T 淋巴细胞频率更低。在治疗前和治疗期间，对 LbAg 的特异性反应导致细胞毒性激活的 CD4+T、CD8+T 和 NK 细胞的扩增，表明这些细胞对 L. braziliensis 的反应具有特异性。此外，通过比较治疗前和治疗期间患者和 HI 组之间细胞毒性激活的 CD4+T、CD8+T 和 NK 细胞频率的差异，我们得出结论，这些细胞群负责锑剂治疗引起的免疫反应。有趣的是，我们还观察到 NK 细胞在疾病的所有临床阶段均被 LbAg 诱导至衰竭表型。增加的抗原特异性激活和细胞毒性活性与该疾病中描述的强烈炎症反应一致，这可能是组织损伤的原因。这些发现加强了这些不同的细胞毒性激活细胞群在 CL 免疫发病机制中的参与，并显示出这种免疫反应的特异性特征。
