Tropical Diseases Department, Botucatu Medical School - UNESP, Botucatu, Brazil.
Bioprocess and Biotechnology Department, Agronomic Sciences School - UNESP, Botucatu, Brazil.
PLoS Negl Trop Dis. 2020 Jan 21;14(1):e0007949. doi: 10.1371/journal.pntd.0007949. eCollection 2020 Jan.
Leishmaniasis is caused by intracellular parasites transmitted to vertebrates by sandfly bites. Clinical manifestations include cutaneous, mucosal or visceral involvement depending upon the host immune response and the parasite species. To assure their survival inside macrophages, these parasites developed a plethora of highly successful strategies to manipulate various immune system pathways. Considering that inflammasome activation is critical for the establishment of a protective immune response in many parasite infections, in this study we determined the transcriptome of THP-1 cells after infection with L. infantum, with a particular focus on the inflammasome components. To this end, the human cell line THP-1, previously differentiated into macrophages by PMA treatment, was infected with L. infantum promastigotes. Differentiated THP-1 cells were also stimulated with LPS to be used as a comparative parameter. The gene expression signature was determined 8 hours after by RNA-seq technique. Infected or uninfected THP-1 cells were stimulated with nigericin (NIG) to measure active caspase-1 and TNF-α, IL-6 and IL-1β levels in culture supernatants after 8, 24 and 48 hours. L. infantum triggered a gene expression pattern more similar to non-infected THP-1 cells and very distinct from LPS-stimulated cells. Some of the most up-regulated genes in L. infantum-infected cells were CDC20, CSF1, RPS6KA1, CD36, DUSP2, DUSP5, DUSP7 and TNFAIP3. Some up-regulated GO terms in infected cells included cell coagulation, regulation of MAPK cascade, response to peptide hormone stimulus, negative regulation of transcription from RNA polymerase II promoter and nerve growth factor receptor signaling pathway. Infection was not able to induce the expression of genes associated with the inflammasome signaling pathway. This finding was confirmed by the absence of caspase-1 activation and IL-1β production after 8, 24 and 48 hours of infection. Our results indicate that L. infantum was unable to activate the inflammasomes during the initial interaction with THP-1 cells.
利什曼病是由沙蝇叮咬传播给脊椎动物的细胞内寄生虫引起的。临床表现包括皮肤、黏膜或内脏受累,这取决于宿主的免疫反应和寄生虫种类。为了确保它们在巨噬细胞内生存,这些寄生虫发展了大量非常成功的策略来操纵各种免疫系统途径。考虑到炎性体激活对于许多寄生虫感染中建立保护性免疫反应至关重要,在这项研究中,我们确定了感染 L. infantum 后 THP-1 细胞的转录组,特别关注炎性体成分。为此,我们使用 PMA 处理将先前分化为巨噬细胞的人细胞系 THP-1 感染 L. infantum 前体。还使用 LPS 刺激分化的 THP-1 细胞作为比较参数。使用 RNA-seq 技术在 8 小时后确定基因表达特征。用 Nigericin(NIG)刺激感染或未感染的 THP-1 细胞,在 8、24 和 48 小时后测量培养上清液中活性 caspase-1 和 TNF-α、IL-6 和 IL-1β 的水平。L. infantum 触发的基因表达模式与未感染的 THP-1 细胞更相似,与 LPS 刺激的细胞非常不同。在 L. infantum 感染的细胞中,一些上调幅度最大的基因是 CDC20、CSF1、RPS6KA1、CD36、DUSP2、DUSP5、DUSP7 和 TNFAIP3。感染细胞中一些上调的 GO 术语包括细胞凝结、MAPK 级联的调节、对肽激素刺激的反应、RNA 聚合酶 II 启动子转录的负调节和神经生长因子受体信号通路。感染不能诱导与炎性体信号通路相关的基因表达。这一发现通过在感染后 8、24 和 48 小时缺乏 caspase-1 激活和 IL-1β 产生得到了证实。我们的结果表明,在与 THP-1 细胞的初始相互作用过程中,L. infantum 无法激活炎性体。