Laderoute Marian P, Larocque Louise J, Giulivi Antonio, Diaz-Mitoma Francisco
Bloodborne Pathogens Division, Blood Zoonotics Unit, Public Health Agency of Canada, Ottawa, Ontario Canada; Department of Pathology and Laboratory Medicine, The University of Ottawa, Ottawa, Ontario Canada.
Bloodborne Pathogens Division, Blood Zoonotics Unit, Public Health Agency of Canada, Ottawa, Ontario Canada.
Open AIDS J. 2015 Dec 7;9:112-22. doi: 10.2174/1874613601509010112. eCollection 2015.
The goals of the research were to determine if a foamy effect on macrophages was due to human endogenous retrovirus K102 (HERV-K102) replication, and to further address its potential significance in HIV-1 infection.
An RT-PCR HERV-K HML-2 pol method was used to screen the unknown HERV, and isolated bands were sent for sequencing. Confirmation of RNA expression was performed by a real time quantitative PCR (qPCR) pol ddCt method. Rabbit antibodies to Env peptides were used to assess expression by immunohistology and processing of Env by western blots. A qPCR pol ddCt method to ascertain genomic copy number was performed on genomic DNA isolated from plasma comparing HIV-1 exposed seronegative (HESN) commercial sex workers (CSW) to normal controls and contrasted with HIV-1 patients.
HERV-K102 expression, particle production and replication were associated with foamy macrophage generation in the cultures of cord blood mononuclear cells under permissive conditions. A five-fold increased HERV-K102 pol genomic copy number was found in the HESN cohort over normal which was not found in HIV-1 positive patients (p=0.0005).
This work extends the evidence that HERV-K102 has foamy virus attributes, is replication competent, and is capable of high replication rate in vivo and in vitro. This may be the first characterization of a replication-competent, foamy-like virus of humans. High particle production inferred by increased integration in the HESN cohort over HIV-1 patients raises the issue of the clinical importance of HERV-K102 particle production as an early protective innate immune response against HIV-1 replication.
本研究的目的是确定巨噬细胞上的泡沫样效应是否归因于人类内源性逆转录病毒K102（HERV-K102）的复制，并进一步探讨其在HIV-1感染中的潜在意义。
采用逆转录聚合酶链反应（RT-PCR）的HERV-K HML-2 pol方法筛选未知的HERV，并将分离出的条带送去测序。通过实时定量PCR（qPCR）的pol ddCt方法确认RNA表达。使用针对Env肽的兔抗体通过免疫组织化学和蛋白质印迹法检测Env的表达。对从血浆中分离的基因组DNA进行qPCR的pol ddCt方法以确定基因组拷贝数，将暴露于HIV-1的血清阴性（HESN）商业性工作者（CSW）与正常对照进行比较，并与HIV-1患者进行对比。
在允许条件下，脐带血单个核细胞培养物中HERV-K102的表达、颗粒产生和复制与泡沫样巨噬细胞的生成相关。在HESN队列中发现HERV-K102 pol基因组拷贝数比正常情况增加了五倍，而在HIV-1阳性患者中未发现这种情况（p = 0.0005）。
这项工作进一步证明了HERV-K102具有泡沫病毒属性，具有复制能力，并且能够在体内和体外以高复制率进行复制。这可能是人类中具有复制能力的泡沫样病毒的首次特征描述。与HIV-1患者相比，HESN队列中整合增加所推断的高颗粒产生提出了HERV-K102颗粒产生作为针对HIV-1复制的早期保护性固有免疫反应的临床重要性问题。
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