Whitteker Jennifer L, Dudani Anil K, Tackaberry Eilleen S
Centre for Biologics Research, Biologics and Genetic Therapies Directorate, Health Canada, Ottawa ON, Canada.
Transplantation. 2008 Jul 15;86(1):155-62. doi: 10.1097/TP.0b013e31817d4823.
Xenotransplantation with pig organs is being considered to alleviate donor organ shortages; however, the risk of introducing porcine viruses into humans is heightened in this setting. The goal of this study was to determine the infectious potential of porcine cytomegalovirus (PCMV), a xenozoonotic virus of interest, in human fibroblasts in vitro.
Confluent human cells were incubated with live PCMV, heat-killed PCMV, or medium alone. Infection was investigated by testing for viral-induced cytopathic effect, assaying viral transcription by nested RT-PCR and subsequent sequencing, and detecting viral protein expression by Western blotting. Plaque neutralization experiments were also performed.
Cells incubated with PCMV demonstrated significant cytopathic effect by 7 days postinfection, and reverse-transcriptase polymerase chain reaction sequencing identified PCMV DNA polymerase in these infected cells. In Western blots, monoclonal antibodies (mAbs) to human CMV glycoprotein B and pig serum presumed to contain anti-PCMV antibodies detected characteristic proteins in experimentally infected human cells and positive controls but not in negative controls. Furthermore, one of these mAbs and the pig serum neutralized PCMV infection in vitro.
These results are a first demonstration that PCMV can infect human fibroblasts in vitro.
猪器官异种移植被认为可缓解供体器官短缺问题;然而,在这种情况下,将猪病毒引入人体的风险增加。本研究的目的是确定猪巨细胞病毒(PCMV)这种感兴趣的异种动物病毒在体外人成纤维细胞中的感染潜力。
将汇合的人细胞与活的PCMV、热灭活的PCMV或仅培养基一起孵育。通过检测病毒诱导的细胞病变效应、用巢式逆转录聚合酶链反应(RT-PCR)和后续测序分析病毒转录以及用蛋白质印迹法检测病毒蛋白表达来研究感染情况。还进行了噬斑中和实验。
与PCMV孵育的细胞在感染后7天显示出明显的细胞病变效应,逆转录聚合酶链反应测序在这些感染细胞中鉴定出PCMV DNA聚合酶。在蛋白质印迹中,针对人巨细胞病毒糖蛋白B的单克隆抗体(mAb)和假定含有抗PCMV抗体的猪血清在实验感染的人细胞和阳性对照中检测到特征性蛋白质,但在阴性对照中未检测到。此外,其中一种mAb和猪血清在体外中和了PCMV感染。
这些结果首次证明PCMV可在体外感染人成纤维细胞。