Vissers Marloes, Habets Marrit N, Ahout Inge M L, Jans Jop, de Jonge Marien I, Diavatopoulos Dimitri A, Ferwerda Gerben
Laboratory of Pediatric Infectious Diseases, Department of Pediatrics, Radboud university medical center.
J Vis Exp. 2013 Dec 10(82):e50766. doi: 10.3791/50766.
Human respiratory syncytial virus (HRSV) infections present a broad spectrum of disease severity, ranging from mild infections to life-threatening bronchiolitis. An important part of the pathogenesis of severe disease is an enhanced immune response leading to immunopathology. Here, we describe a protocol used to investigate the immune response of human immune cells to an HRSV infection. First, we describe methods used for culturing, purification and quantification of HRSV. Subsequently, we describe a human in vitro model in which peripheral blood mononuclear cells (PBMCs) are stimulated with live HRSV. This model system can be used to study multiple parameters that may contribute to disease severity, including the innate and adaptive immune response. These responses can be measured at the transcriptional and translational level. Moreover, viral infection of cells can easily be measured using flow cytometry. Taken together, stimulation of PBMC with live HRSV provides a fast and reproducible model system to examine mechanisms involved in HRSV-induced disease.
人呼吸道合胞病毒（HRSV）感染呈现出广泛的疾病严重程度，从轻度感染到危及生命的细支气管炎。严重疾病发病机制的一个重要部分是增强的免疫反应导致免疫病理。在此，我们描述一种用于研究人类免疫细胞对HRSV感染的免疫反应的方案。首先，我们描述用于培养、纯化和定量HRSV的方法。随后，我们描述一种体外模型，其中外周血单个核细胞（PBMC）用活的HRSV刺激。该模型系统可用于研究可能导致疾病严重程度的多个参数，包括先天性和适应性免疫反应。这些反应可以在转录和翻译水平上进行测量。此外，使用流式细胞术可以轻松测量细胞的病毒感染情况。总之，用活的HRSV刺激PBMC提供了一个快速且可重复的模型系统，用于研究HRSV诱导疾病所涉及的机制。
