Characterization of the single cycle replication of HIV-1 expressing Gaussia luciferase in human PBMCs, macrophages, and in CD4+Tcell-grafted nude mouse. Journal of Virological Methods
Characterization of the single cycle replication of HIV-1 expressing Gaussia luciferase in human PBMCs, macrophages, and in CD4+Tcell-grafted nude mouse. Journal of Virological Methods
复制标题
人类 PBMC、巨噬细胞和 CD4 T 细胞移植裸鼠中表达 HIV-1 的 Gaussia 荧光素酶的单周期复制特征。
DOI：
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发表时间：
2015
影响因子：
3.1
通讯作者：
Zhujun Ao
中科院分区：
文献类型：
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作者：
Zhujun Ao
在这项研究中，我们描述了一种敏感的HIV-1单周期复制病毒，该病毒表达分泌的高斯荧光素酶（Gluc）作为生物标志物。这种单周期复制病毒是由293T细胞与多基因缺失的HIV病毒（RI/E/Gluc）和CMVin-Gag/波兰病毒包膜糖蛋白（Env）质粒共转染产生的。结果表明，该HIV-1病毒在原代CD4+T细胞、巨噬细胞和移植CD4+T细胞的裸鼠体内有效感染并局限于一个复制周期。由于病毒基因组DNA缺乏逆转录酶和整合基因以及env基因的部分缺失，这种反式互补病毒在高度易感的CD4+ C8166 T细胞中多次传代后不能转化为野生型病毒，证明了该系统的安全性。此外，通过检测细胞培养上清或动物腹膜液或血液中的葡萄糖活性，可以很容易地监测该病毒的感染，这比抗p24 ELISA检测更敏感。这种反式互补病毒系统对于在无法获得生物安全3级控制设施的实验室中进行各种HIV单周期感染和病毒表达研究很有价值。
tIn this study, we have described a sensitive HIV-1 single cycle replicating virus that expresses a secretedGaussia luciferase (Gluc) as a biomarker. This single cycle replicating virus was produced by the co-transfection of 293T cells with a multiple gene-deleted HIV provirus (RI/E/Gluc) and CMVin-Gag/Poland viral envelope glycoprotein (Env) plasmids. The results showed that this HIV-1 virus efficientlyinfected and was restricted to one replication cycle in primary CD4+ T cells, macrophages and CD4+T cell-grafted nude mouse. Because the viral genomic DNA lacks reverse transcriptase and integrasegenes and has a partial deletion in the env gene, this trans-complemented virus could not be convertedinto the wild type virus after multiple passages in highly susceptible CD4+ C8166 T cells, demonstratingthe safety of this system. Furthermore, infection with this virus was easily monitored by detecting theGluc activity in the cell culture supernatants or in the animal peritoneal fluid or blood, which was shownto be more sensitive than the anti-p24 ELISA assay. This trans-complemented virus system is valuable forvarious HIV single cycle infection and viral expression studies in the laboratory where a biosafety level-3containment facility is not accessible.