Kent S J, Stallard V, Corey L, Hu S L, Morton W R, Gritz L, Panicali D L, Greenberg P D
Department of Medicine, University of Washington, Seattle 98195.
AIDS Res Hum Retroviruses. 1994 May;10(5):551-60. doi: 10.1089/aid.1994.10.551.
Methods to analyze CD8+ CTL responses to simian immunodeficiency virus (SIV)-encoded proteins are essential to understand lentivirus immunopathogenesis and protective immune responses. Recombinant infectious shuttle vectors are useful for analyzing CTL responses to many viruses, including HIV. Therefore, CTL responses in SIV-infected Macaca fascicularis to SIV env and SIV gag/pol were evaluated using specific antigen stimulation with recombinant vaccinia (rVV) and fowl poxviruses (rFPV) containing SIV genes. Peripheral blood mononuclear cells from SIV-infected animals were stimulated with autologous cells infected with rVV expressing SIV env/gag/pol, and CTLs specific for SIV env and for SIV gag/pol were detected by testing for lytic activity in target cells expressing these genes separately. Lymphocyte subset purifications from the effector population demonstrated that the CTL response was mediated by CD8+ cells, and the use of brefeldin A to selectively block antigen presentation in association with MHC class I products affirmed this cytolytic activity was class I restricted. The use of rVV to analyze responses to SIV genes is potentially problematic in hosts immunized to vaccinia. Fowl poxvirus is an alternative virus that has many of the molecular advantages of vaccinia virus but is genomically distinct. Therefore, the ability of rFPV to expand and detect SIV-specific CTLs was evaluated. Although there was no cytopathic effect following infection with rFPV, macaque cells infected with this vector did express rFPV gene products, and could be used as stimulator and target cells to detect SIV-specific CD8+ CTLs. The results suggest that these recombinant viral vectors can be used to specifically stimulate CD8+, MHC class I-restricted CTLs reactive to SIV proteins, and should facilitate evaluating CTL responses in both SIV-infected animals and animals vaccinated against SIV.
分析CD8 + 细胞毒性T淋巴细胞（CTL）对猴免疫缺陷病毒（SIV）编码蛋白的反应方法对于理解慢病毒免疫发病机制和保护性免疫反应至关重要。重组感染性穿梭载体可用于分析对包括HIV在内的多种病毒的CTL反应。因此，利用含有SIV基因的重组痘苗病毒（rVV）和禽痘病毒（rFPV）进行特异性抗原刺激，评估了SIV感染的食蟹猴中针对SIV env和SIV gag/pol的CTL反应。用表达SIV env/gag/pol的rVV感染的自体细胞刺激来自SIV感染动物的外周血单核细胞，并通过分别检测表达这些基因的靶细胞中的裂解活性来检测对SIV env和SIV gag/pol特异的CTL。从效应细胞群体中纯化淋巴细胞亚群表明CTL反应由CD8 + 细胞介导，使用布雷菲德菌素A选择性阻断与MHC I类产物相关的抗原呈递证实了这种细胞溶解活性受I类限制。在已接种痘苗的宿主中，使用rVV分析对SIV基因的反应可能存在问题。禽痘病毒是一种替代病毒，具有痘苗病毒的许多分子优势，但基因组不同。因此，评估了rFPV扩增和检测SIV特异性CTL的能力。尽管感染rFPV后没有细胞病变效应，但感染该载体的猕猴细胞确实表达rFPV基因产物，并且可以用作刺激细胞和靶细胞来检测SIV特异性CD8 + CTL。结果表明，这些重组病毒载体可用于特异性刺激对SIV蛋白有反应的CD8 + 、MHC I类限制的CTL，并应有助于评估SIV感染动物和接种SIV疫苗动物中的CTL反应。
