Lawson C M, Bennink J R, Restifo N P, Yewdell J W, Murphy B R
Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Cancer Institute, Bethesda, Maryland 20892.
J Virol. 1994 Jun;68(6):3505-11. doi: 10.1128/JVI.68.6.3505-3511.1994.
The nucleoprotein (NP) of influenza A virus is the dominant antigen recognized by influenza virus-specific cytotoxic T lymphocytes (CTLs), and adoptive transfer of NP-specific CTLs protects mice from influenza A virus infection. BALB/c mouse cells (H-2d) recognize a single Kd-restricted CTL epitope of NP consisting of amino acids 147 to 155. In the present study, mice were immunized with various vaccinia virus recombinant viruses to examine the effect of the induction of primary pulmonary CTLs on resistance to challenge with influenza A/Puerto Rico/8/34 virus. The minigene ESNP(147-155)-VAC construct, composed of a signal sequence from the adenovirus E3/19K glycoprotein (designated ES) and expressing the 9-amino-acid NP natural determinant (amino acids 147 to 155) preceded by an alanine residue, a similar minigene NP(Met 147-155)-VAC lacking ES, and a full-length NP-VAC recombinant of influenza virus were analyzed. The two minigene NP-VAC recombinants induced a greater primary pulmonary CTL response than the full-length NP-VAC recombinant. However, NP-specific CTLs induced by immunization with ESNP(147-155)-VAC did not decrease peak virus titer or accelerate clearance of virus in the lungs of mice challenged intranasally with A/PR/8/34. Furthermore, NP-specific CTLs induced by immunization did not protect mice challenged intranasally with a lethal dose of A/PR/8/34. Sequence analysis of the NP CTL epitope of A/PR/8/34 challenge virus obtained from lungs after 8 days of replication in ESNP(147-155)-VAC-immunized mice showed identity with that of the input virus, demonstrating that an escape mutant had not emerged during replication in vivo. Thus, in contrast to adoptively transferred CTLs, pulmonary NP-specific CTLs induced by recombinant vaccinia virus immunization do not have protective in vivo antiviral activity against influenza virus infection.
甲型流感病毒的核蛋白（NP）是流感病毒特异性细胞毒性T淋巴细胞（CTL）识别的主要抗原，NP特异性CTL的过继转移可保护小鼠免受甲型流感病毒感染。BALB/c小鼠细胞（H-2d）识别由氨基酸147至155组成的NP的单一Kd限制性CTL表位。在本研究中，用各种痘苗病毒重组病毒免疫小鼠，以检查诱导原发性肺CTL对抵抗甲型流感病毒/波多黎各/8/34病毒攻击的影响。分析了由腺病毒E3/19K糖蛋白的信号序列（命名为ES）组成并表达9个氨基酸的NP天然决定簇（氨基酸147至155）且前面有一个丙氨酸残基的小基因ESNP(147-155)-VAC构建体、缺少ES的类似小基因NP(Met 147-155)-VAC以及甲型流感病毒的全长NP-VAC重组体。两种小基因NP-VAC重组体诱导的原发性肺CTL反应比全长NP-VAC重组体更强。然而，用ESNP(147-155)-VAC免疫诱导的NP特异性CTL并没有降低经鼻内接种A/PR/8/34攻击的小鼠肺中的病毒峰值滴度，也没有加速病毒清除。此外，免疫诱导的NP特异性CTL不能保护经鼻内接种致死剂量A/PR/8/34攻击的小鼠。对在ESNP(147-155)-VAC免疫小鼠中复制8天后从肺中获得的A/PR/8/34攻击病毒的NP CTL表位进行序列分析，结果显示与输入病毒相同，表明在体内复制过程中未出现逃逸突变体。因此，与过继转移的CTL不同，重组痘苗病毒免疫诱导的肺NP特异性CTL对流感病毒感染没有体内抗病毒保护活性。
