Falkenberg Shollie M, Dassanayake Rohana P, Neill John D, Walz Paul H, Casas Eduardo, Ridpath Julia F, Roth James
Ruminant Disease and Immunology Research Unit, National Animal Disease Center, USDA, Agricultural Research Service, Ames, IA, 50010, United States.
Ruminant Disease and Immunology Research Unit, National Animal Disease Center, USDA, Agricultural Research Service, Ames, IA, 50010, United States.
Vet Immunol Immunopathol. 2020 Mar;221:110024. doi: 10.1016/j.vetimm.2020.110024. Epub 2020 Feb 11.
Current methods for evaluating bovine viral diarrhea virus (BVDV) vaccination response typically rely on measurement of humoral responses as determined by virus neutralizing antibody titers (VNT) against BVDV. While VNT are correlated with increased protection, research has also shown that cell mediated immunity (CMI) is an important component of a protective response against BVDV. For example, improved protection against BVDV by modified-live viral (MLV) vaccines as compared to killed vaccines is thought to be due to better CMI induced by the MLV. The goal of this work was to evaluate the cell mediated response in vaccinated calves using a novel PrimeFlow RNA assay that incorporates cell surface marker staining with intracellular RNA expression of cytokines and viral RNA detection. Results from this study evaluating mRNA for IFN-γ and IL-2 at 24 h post-BVDV stimulation are similar to previous studies in which IFN-γ was detected in the CD4 and CD8 T cell population. However, a novel observation was the detection of IFN-γ mRNA in the NK cell population in vaccinated animals. The NK cell population contributed a significant portion of the IFN-γ produced. This study also demonstrated a decrease in the frequency and amount of BVDV in PBMCs, harvested from vaccinated calves and exposed to BVDV in vitro. Collectively data from this study highlights the association between an increase in IFN-γ and a decreased infection rate of isolated PBMC's, based on the frequency and amount of BVDV positive cells following in vitro exposure. This new method combines not only the ability to evaluate cellular responses, but also the ability to understand potential antiviral properties associated with cellular responses. This is the first assay to describe and simultaneously measure CMI responses and intracellular viral RNA quantity as a method to evaluate protective responses associated with vaccination.
目前评估牛病毒性腹泻病毒（BVDV）疫苗接种反应的方法通常依赖于通过针对BVDV的病毒中和抗体滴度（VNT）来测定体液反应。虽然VNT与增强的保护作用相关，但研究也表明，细胞介导的免疫（CMI）是针对BVDV的保护性反应的重要组成部分。例如，与灭活疫苗相比，改良活病毒（MLV）疫苗对BVDV的保护作用有所改善，这被认为是由于MLV诱导了更好的CMI。这项工作的目的是使用一种新型的PrimeFlow RNA检测方法来评估接种疫苗的小牛的细胞介导反应，该方法将细胞表面标志物染色与细胞因子的细胞内RNA表达和病毒RNA检测相结合。本研究在BVDV刺激后24小时评估IFN-γ和IL-2的mRNA的结果与先前的研究相似，在先前的研究中，在CD4和CD8 T细胞群体中检测到了IFN-γ。然而，一个新的发现是在接种动物的自然杀伤（NK）细胞群体中检测到了IFN-γ mRNA。NK细胞群体产生了很大一部分IFN-γ。这项研究还表明，从接种疫苗的小牛身上采集并在体外暴露于BVDV的外周血单核细胞（PBMC）中，BVDV的频率和数量有所下降。这项研究的总体数据突出了基于体外暴露后BVDV阳性细胞的频率和数量，IFN-γ增加与分离的PBMC感染率降低之间的关联。这种新方法不仅结合了评估细胞反应的能力，还结合了理解与细胞反应相关的潜在抗病毒特性的能力。这是第一种描述并同时测量CMI反应和细胞内病毒RNA数量作为评估与疫苗接种相关的保护性反应的方法。
