猪δ冠状病毒感染猪树突状细胞及对免疫因子分泌能力的影响
Effects of porcine deltacoronavirus on infection and secretion of immune factors in porcine dendritic cells
胡亚婷 1舒祥力 1赵福杰 1郝晨琳 1曾权 1郑兰兰 2胡慧 2张红垒2
作者信息
- 1. 河南农业大学动物医学院,河南郑州 450002
- 2. 河南农业大学动物医学院,河南郑州 450002;河南省动物性食品安全重点实验室,河南郑州 450002
- 折叠
摘要
以猪树突状细胞(porcine dendritic cells,pDCs)为模型,探究猪 δ 冠状病毒(porcine deltacoronavirus,PDCoV)是否能够对pDCs进行有效感染及对抗病毒细胞因子分泌的影响.采集猪外周抗凝血,分离获得猪外周血单核细胞,经白细胞介素-4(interleukin-4,IL-4)和集落刺激因子(granulocyte-macrophage colony stimulating factor,GM-CSF)诱导培养获得未成熟树突状细胞,利用倒置显微镜和透射电镜观察其生物学形态,通过蛋白质免疫印迹和激光共聚焦试验检测病毒是否可以对pDCs进行感染,qRT-PCR检测抗病毒因子IFN-α和ISG56的表达.结果显示,成功分离培养得到具有典型树突状细胞形态的pDCs,将PDCoV接种至分离培养得到的pDCs中,PDCoV可以有效感染,可以在pDCs中进行复制并产生感染性的子代病毒颗粒,病毒滴度达到4.4 lgTCID50/0.1 mL.同时检测PDCoV感染pDCs时对抗病毒因子表达的影响,结果表明,在病毒感染pDCs后48 h产生了高水平的IFN-α和ISG56.本研究成功诱导培养得到pDCs,首次证实PDCoV可以在体外感染pDCs,并在细胞中进行有效地复制,pDCs在病毒感染后促进IFN-α、ISG56的分泌来发挥抗病毒免疫效应,为后续研究PDCoV的致病机制提供了新的思路.
Abstract
Porcine deltacoronavirus(PDCoV)is a new intestinal coronavirus that causes diarrhea in nursing piglets,which seriously harms the pig industry.This study aims to use porcine dendritic cells(pDCs)as a model to explore whether PDCoV can effectively infect pDCs and its effect on the secretion of anti-viral cytokines.Peripheral anticoagulant cells of porcine were first collected.Por-cine peripheral blood mononuclear cells were then isolated and induced by interleukin-4(IL-4)and colony-stimulating factor(GM-CSF)to obtain immature dendritic cells,and their biological mor-phology was observed by inverted microscopy and transmission electron microscopy.Western blot and laser confocal assays were used to detect whether the virus could infect pDCs,and qRT-PCR were used to detect the expression of antiviral factors IFN-α and ISG56.The results showed that pDCs with typical dendritic cell morphology were successfully isolated and cultured,and PDCoV could effectively infect and replicate in pDCs and produce infectious progeny viral particles,with a viral titer of 4.4 lgTCID50/0.1 mL.At the same time,the effect of PDCoV infection on the expres-sion of antiviral factors in pDCs was detected,and the results showed that high levels of IFN-α and ISG56 were produced 48 h after infection in pDCs.In summary,we successfully obtained pDCs by induced culture,and confirmed for the first time that PDCoV can infect pDCs in vitro and effec-tively replicate in cells,and pDCs promote the secretion of IFN-α and ISG56 after viral infection to exert antiviral immune effects,which provides a novel idea for subsequent research on the patho-genic mechanism of PDCoV.
关键词
猪δ冠状病毒/猪树突状细胞/病毒感染/抗病毒因子Key words
porcine delta coronavirus/porcine dendritic cells/viral infection/antiviral factors引用本文复制引用
基金项目
河南省杰出青年项目(202300410192)
河南省科技攻关计划(212102110363)
河南省高等学校科技创新人才支持计划(21HASTIT039)
出版年
2024
NETL
NSTL
万方数据