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目的分析HIV-1广谱中和活性感染者基于膜蛋白基因(env)的假病毒对自体血浆和代表性广谱单克隆中和抗体(bm NAbs)的中和表型。方法单基因组扩增(SGA)广谱中和活性感染者第0月血浆的env基因并克隆至pc D-NA~(TM)3.1 Directional TOPO表达载体,将env表达质粒与HIV-1骨架质粒p SG3△env共转染293T/17细胞制备假病毒,用假病毒分别与自体连续时间点血浆和代表性bm NAbs进行中和实验分析中和表型。结果共获得11株功能性假病毒,假病毒对8个连续时间点自体血浆的中和敏感性呈现先升高(第0~15月),后下降(第16~32月),之后再上升(第33~45月)的波动。所有假病毒对10E8、PGT121和VRC01中和敏感(IC_(50)<6μg/m L);各有18.2%(2株)的假病毒对12A21和2G12高度敏感(IC_(50)<1μg/m L);所有病毒均对PGT135呈中和抗性。结论假病毒对当前时间点血浆的中和敏感性低,对之后时间点血浆中和敏感性增强,提示感染者体内病毒与中和抗体的动态进化过程。同一时间点假病毒对特定bm-NAbs的敏感性差异明显,提示感染者准种毒株间中和表型的复杂性。
Objective To analyze the neutralizing phenotype of autologous plasma and representative broad-spectrum monoclonal neutralizing antibody (bm NAbs) based on the envelope protein gene (env) -positive pseudovirus in HIV-1 broad-spectrum neutralizing active infection. Methods The plasma env gene from the first month of single-genome amplification (SGA) neutralizing activity was cloned into pcD-NA ™ (TM) 3.1 Directional TOPO expression vector. The env expression plasmid and HIV-1 backbone plasmid p The pseudoviruses were co-transfected with 293T / 17 cells with SG3Δv env and the neutralization phenotypes were analyzed by neutralization with pseudoviruses and autologous continuous-time plasma and representative bm NAbs, respectively. Results A total of 11 functional pseudoviruses were obtained. The neutralizing sensitivity of pseudoviruses to autologous plasma at 8 consecutive time points increased first (from 0 to 15 months) and then decreased (from 16 to 32 months), and then increased (33 ~ 45 months) fluctuations. All pseudotypes were neutralized to 10E8, PGT121, and VRC01 (IC 50 (6 μg / m L); 18.2% (2 strains) pseudovirions each were highly sensitive to 12A21 and 2G12 (IC50 <1 μg / m L); all viruses showed neutralizing resistance to PGT135. Conclusions Pseudovirus is low sensitive to the neutralization of plasma at the current time point, and its plasma neutralization sensitivity is enhanced at later time point, which indicates the dynamic evolution of virus and neutralizing antibody in infected patients. At the same time, the sensitivity of pseudovirus to specific bm-NAbs was significantly different, suggesting the complexity of neutralizing the phenotype of infected individuals.