Chen Qian, Li Yumeng, Yan Xiaoliang, Sun Zhenjie, Wang Chuan, Liu Shuangquan, Xiao Jian, Lu Chunxue, Wu Yimou
Institution of Pathogenic Biology, Hengyang Medical College, University of South China, Hunan Provincial Key Laboratory for Special Pathogens Prevention and Control, Hunan Province Cooperative Innovation Center for Molecular Target New Drug Study, University of South China, Hengyang, China.
Institute of Clinical Research, The First Affiliated Hospital of University of South China, Hengyang, China.
Front Microbiol. 2020 Dec 3;11:578009. doi: 10.3389/fmicb.2020.578009. eCollection 2020.
The chlamydial plasmid, an essential virulence factor, encodes plasmid proteins that play important roles in chlamydial infection and the corresponding immune response. However, the virulence factors and the molecular mechanisms of are not well understood. In the present study, we investigated the roles and mechanisms of the plasmid-encoded protein CPSIT_P7 of in regulating the inflammatory response in THP-1 cells (human monocytic leukemia cell line). Based on cytokine arrays, CPSIT_P7 induces the expression of interleukin-6 (IL-6), interleukin-8 (IL-8), and monocyte chemoattractant protein-1 (MCP-1) in THP-1 cells. Moreover, the expression levels of IL-6, IL-8, and MCP-1 stimulated by CPSIT_P7 declined after silencing of the Toll-like receptor 4 (TLR4) gene using small interfering RNA and transfection of a dominant negative plasmid encoding TLR4 (pZERO-hTLR4). We further demonstrated that transfection with the dominant negative plasmid encoding MyD88 (pDeNy-hMyD88) and the dominant negative plasmid encoding Mal (pDeNy-hMal) could also abrogate the expression of the corresponding proteins. Western blot and immunofluorescence assay results showed that CPSIT_P7 could activate nuclear factor κB (NF-κB) signaling pathways in THP-1 cells. Altogether, our results indicate that the CPSIT_P7 induces the TLR4/Mal/MyD88/NF-κB signaling axis and therefore contributes to the inflammatory cytokine response.
衣原体质粒是一种重要的毒力因子，它编码的质粒蛋白在衣原体感染及相应免疫反应中发挥重要作用。然而，其毒力因子及分子机制尚未完全明确。在本研究中，我们探究了衣原体编码蛋白CPSIT_P7在调节THP-1细胞（人单核细胞白血病细胞系）炎症反应中的作用及机制。基于细胞因子阵列分析，CPSIT_P7可诱导THP-1细胞中白细胞介素-6（IL-6）、白细胞介素-8（IL-8）和单核细胞趋化蛋白-1（MCP-1）的表达。此外，使用小干扰RNA沉默Toll样受体4（TLR4）基因并转染编码TLR4的显性负体质粒（pZERO-hTLR4）后，CPSIT_P7刺激的IL-6、IL-8和MCP-1表达水平下降。我们进一步证明，转染编码MyD88的显性负体质粒（pDeNy-hMyD88）和编码Mal的显性负体质粒（pDeNy-hMal）也可消除相应蛋白的表达。蛋白质印迹和免疫荧光分析结果表明，CPSIT_P7可激活THP-1细胞中的核因子κB（NF-κB）信号通路。总之，我们的结果表明，CPSIT_P7可诱导TLR4/Mal/MyD88/NF-κB信号轴，从而促进炎性细胞因子反应。
