ANALYSIS OF IMMUNOREGULATORY VACCINIA GENES

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 天花病毒和牛痘病毒含有大量的免疫调节基因产物，它们被认为通过调节宿主的免疫反应来促进病毒的复制。干扰素-γ的分泌(IFNGamma)是体外细胞免疫反应的常用指标。我们推测，痘苗病毒B8R蛋白，一种分泌的IFNG受体类似物，可能干扰ELISPOT法检测IFNGamma。ELISPOT法检测外周血单个核细胞(PBMC)在痘苗病毒或EB病毒(EBV)转化的自体B细胞刺激下分泌的干扰素-γ(IFN-γ)。痘苗病毒感染可抑制CD8+T细胞对EBV转化的自体B细胞株的干扰素-γELISPOT反应达70%。相比之下，EBV特异性的干扰素-g ELISPOT反应不会被感染改良的安卡拉牛痘(MVA)所抑制，MVA是一种不含B8R的减毒牛痘菌株。用缺失B8R基因的痘苗病毒重组体刺激PBMC，与野生型毒株相比，检测到的IFNGamma分泌增加了75%。用MVA刺激痘苗接种者的PBMC，与野生型相比，干扰素-γ的分泌增加了12倍。痘苗病毒免疫调节基因产物干扰细胞免疫反应的体外检测，包括痘苗特异性和EBV特异性反应。痘苗病毒可溶性干扰素-γ受体(B8R)的缺失导致了这种作用的不完全消除，这表明MVA中缺乏的其他基因也有助于逃避CD8+T细胞介导的免疫反应。这些结果表明，使用痘苗病毒作为刺激来评估IFNG的分泌可能低估了宿主的免疫反应。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Variola virus and vaccinia virus contain a large number of immunoregulatory gene products which are thought to enhance viral reproduction by modulating host immune responses. Secretion of interferon-gamma (IFNgamma) is a commonly used measure of cellular immune responses in vitro. We hypothesized that the vaccinia virus B8R protein, a secreted IFNg-receptor analog, might interfere with detection of IFNgamma by the ELISPOT assay. IFNgamma secretion by peripheral blood mononuclear cells (PBMCs) in response to stimulation with vaccinia virus or autologous B cells transformed by Epstein-Barr virus (EBV) was quantitated by ELISPOT. Vaccinia infection inhibited CD8+ T cell IFN-gamma ELISPOT responses to autologous EBV-transformed B cell lines by 70%. In contrast, the EBV-specific IFN-g ELISPOT response was not inhibited by infection with modified vaccinia Ankara (MVA), an attenuated vaccinia strain that does not contain B8R. Stimulation of PBMCs with a vaccinia virus recombinant that has had the B8R gene deleted resulted in a 75 percent increase in detection of IFNgamma secretion compared with the wild-type strain. Stimulation of PBMCs from vaccinia vaccinees with MVA resulted in a 12-fold increase in IFNgamma secretion compared with the wild-type strain. Vaccinia virus immunoregulatory gene products interfere with in vitro detection of cellular immune responses, including vaccinia-specific and EBV-specific responses. Deletion of the soluble IFN-gamma receptor (B8R) from vaccinia virus resulted in an incomplete abrogation of this effect, suggesting that other genes absent in MVA also contribute to evasion of CD8+ T cell mediated immune responses. These results suggest that use of vaccinia virus as a stimulus to assess IFNg secretion may underestimate the host immune response.