Nicholas J A, Rubino K L, Levely M E, Adams E G, Collins P L
Department of Infectious Diseases, Upjohn Company, Kalamazoo, Michigan 49007.
J Virol. 1990 Sep;64(9):4232-41. doi: 10.1128/JVI.64.9.4232-4241.1990.
Cytolytic T-lymphocyte (CTL) activity specific for respiratory syncytial (RS) virus was investigated after intranasal infection of mice with RS virus, after intraperitoneal infection of mice with a recombinant vaccinia virus expressing the F glycoprotein, and after intramuscular vaccination of mice with Formalin-inactivated RS virus or a chimeric glycoprotein, FG, expressed from a recombinant baculovirus. Spleen cell cultures from mice previously infected with live RS virus or the F-protein recombinant vaccinia virus had significant CTL activity after one cycle of in vitro restimulation with RS virus, and lytic activity was derived from a major histocompatibility complex-restricted, Lyt2.2+ (CD8+) subset. CTL activity was not restimulated in spleen cells from mice that received either the Formalin-inactivated RS virus or the purified glycoprotein, FG. The protein target structures for recognition by murine CD8+ CTL were identified by using target cells infected with recombinant vaccinia viruses that individually express seven structural proteins of RS virus. Quantitation of cytolytic activity against cells expressing each target structure suggested that 22K was the major target protein for CD8+ CTL, equivalent to recognition of cells infected with RS virus, followed by intermediate recognition of F or N, slight recognition of P, and no recognition of G, SH, or M. Repeated stimulation of murine CTL with RS virus resulted in outgrowth of CD4+ CTL which, over time, became the exclusive subset in culture. Murine CD4+ CTL were highly cytolytic for RS virus-infected cells, but they did not recognize target cells infected with any of the recombinant vaccinia viruses expressing the seven RS virus structural proteins. Finally, the CTL response in peripheral blood mononuclear cells of adult human volunteers was investigated. The detection of significant levels of RS virus-specific cytolytic activity in these cells was dependent on at least two restimulations with RS virus in vitro, and cytolytic activity was derived primarily from the CD4+ subset.
在用呼吸道合胞（RS）病毒经鼻感染小鼠后、用表达F糖蛋白的重组痘苗病毒经腹腔感染小鼠后、以及用福尔马林灭活的RS病毒或由重组杆状病毒表达的嵌合糖蛋白FG经肌肉接种小鼠后，研究了针对RS病毒的溶细胞性T淋巴细胞（CTL）活性。先前感染过活RS病毒或F蛋白重组痘苗病毒的小鼠的脾细胞培养物，在用RS病毒进行一轮体外再刺激后具有显著的CTL活性，并且裂解活性源自主要组织相容性复合体限制的Lyt2.2 +（CD8 +）亚群。在用福尔马林灭活的RS病毒或纯化的糖蛋白FG处理的小鼠的脾细胞中，CTL活性未被再刺激。通过使用感染了分别表达RS病毒七种结构蛋白的重组痘苗病毒的靶细胞，确定了小鼠CD8 + CTL识别的蛋白质靶标结构。对表达每种靶标结构的细胞的溶细胞活性定量分析表明，22K是CD8 + CTL的主要靶蛋白，等同于对感染RS病毒的细胞的识别，其次是对F或N的中等识别、对P的轻微识别，以及对G、SH或M无识别。用RS病毒反复刺激小鼠CTL导致CD4 + CTL的增殖，随着时间的推移，CD4 + CTL成为培养物中的唯一亚群。小鼠CD4 + CTL对RS病毒感染的细胞具有高度溶细胞性，但它们不识别感染了表达七种RS病毒结构蛋白中任何一种的重组痘苗病毒的靶细胞。最后，研究了成年人类志愿者外周血单个核细胞中的CTL反应。在这些细胞中检测到显著水平的RS病毒特异性溶细胞活性依赖于在体外至少用RS病毒进行两次再刺激，并且溶细胞活性主要源自CD4 +亚群。
