非洲猪瘟病毒C717R蛋白诱导小鼠炎症应答

Inflammatory response induced by African swine fever virus C717R protein in mice

摘要:

【目的】通过炎症应答系统筛选，发现非洲猪瘟病毒(African swine fever virus, ASFV) C717R蛋白可诱导炎症反应，本研究旨在首次鉴定C717R蛋白功能，通过构建C717R重组慢病毒并感染BALB/c小鼠，探究其对炎症应答产生的影响。【方法】通过炎症小体表达系统筛选出诱导炎症应答的C717R蛋白，并构建C717R重组慢病毒。利用C717R重组慢病毒感染小鼠，使C717R在小鼠组织中表达。经实时荧光定量、蛋白质免疫印迹等方法检测C717R慢病毒包装、蛋白表达以及促炎细胞因子TNF-α、IL-1β、IL-6、IFN-β的变化。【结果】C717R蛋白在小鼠组织中正常表达。酶联免疫吸附测定(enzyme linked immunosorbent assay, ELISA)检测发现，表达C717R蛋白的小鼠，血清中促炎细胞因子TNF-α、IL-1β、IL-6、IFN-β及IFN-γ分泌水平显著升高。实时荧光定量检测表达C717R蛋白的小鼠组织证实，C717R、TNF-α、IL-1β、IL-6的mRNA转录水平显著上调；蛋白质免疫印迹证实，C717R可诱导小鼠不同组织caspase-1和IL-1β的成熟。组织病理切片结果显示，表达C717R蛋白的BALB/c小鼠和对照组和相比，肝脏、心脏、肺脏等器官炎性细胞浸润程度较对照组更严重。【结论】ASFV的C717R蛋白表达诱导BALB/c小鼠产生炎症应答，为鉴定和阐明C717R蛋白介导的促炎新机制提供了重要依据。

Abstract:

[Objective] By screening the inflammatory response system, we discovered that the African swine fever virus (ASFV) protein C717R induced an inflammatory response. This study aims to reveal the function of C717R protein by constructing recombinant C717R-lentivirus and observing the recombinant-induced inflammatory response in BALB/c mice. [Methods] C717R was screened out by the inflammasome expression system and used to construct the recombinant C717R-lentivirus. Mice were infected with the recombinant C717R-lentivirus to enable the expression of C717R in mouse tissue. Quantitative real-time PCR, western blotting, and enzyme-linked immunosorbent assay were employed to examine the package of C717R-lentivirus, protein expression, and the serum levels of pro-inflammatory cytokines including tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), IL-6, and interferon-beta (IFN-β), respectively. [Results] C717R protein was normally expressed in mouse tissue. The serum levels of TNF-α, IL-1β, IL-6, IFN-γ, and IFN-β were elevated in the expression of C717R protein in mice. The mRNA levels of C717R, TNF-α, IL-1β, and IL-6 were significantly up-regulated in the expression of C717R protein in mice. Western blotting demonstrated that expression of C717R protein induced the caspase-1 and IL-1β in different tissues. The lesions in the liver, heart, lungs, and other organs of BALB/c mice expressing C717R protein were severer than those in the control mice. [Conclusion] The expression of C717R protein induced an inflammatory response in BALB/c mice. The findings provide a basis for deciphering the mechanism of C717R protein-mediated inflammation.