Simon Ayo Y, Sutherland Michael R, Pryzdial Edward L G
Canadian Blood Services, Centre for Innovation, Ottawa, ON, Canada; and Centre for Blood Research/Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, BC, Canada.
Blood. 2015 Jul 16;126(3):378-85. doi: 10.1182/blood-2014-09-598029. Epub 2015 May 5.
Dengue virus (DENV) infection causes ∼200 million cases of severe flulike illness annually, escalating to life-threatening hemorrhagic fever or shock syndrome in ∼500,000. Although thrombocytopenia is typical of both mild and severe diseases, the mechanism triggering platelet reduction is incompletely understood. As a probable initiating event, direct purified DENV-platelet binding was followed in the current study by quantitative reverse transcription-polymerase chain reaction and confirmed antigenically. Approximately 800 viruses specifically bound per platelet at 37°C. Fewer sites were observed at 25°C, the blood bank storage temperature (∼350 sites), or 4°C, known to attenuate virus cell entry (∼200 sites). Dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN) and heparan sulfate proteoglycan were implicated as coreceptors because only the combination of anti-DC-SIGN and low-molecular-weight heparin prevented binding. Interestingly, at 37°C and 25°C, platelets replicated the positive sense single-stranded RNA genome of DENV by up to ∼4-fold over 7 days. Further time course experiments demonstrated production of viral NS1 protein, which is known to be highly antigenic in patient serum. The infectivity of DENV intrinsically decayed in vitro, which was moderated by platelet-mediated generation of viable progeny. This was shown using a transcription inhibitor and confirmed by freeze-denatured platelets being incapable of replicating the DENV genome. For the first time, these data demonstrate that platelets directly bind DENV saturably and produce infectious virus. Thus, expression of antigen encoded by DENV is a novel consideration in the pathogen-induced thrombocytopenia mechanism. These results furthermore draw attention to the possibility that platelets may produce permissive RNA viruses in addition to DENV.
登革病毒(DENV)感染每年导致约2亿例严重流感样疾病,其中约50万例病情会升级为危及生命的出血热或休克综合征。尽管血小板减少在轻症和重症疾病中都很常见,但引发血小板减少的机制尚未完全明确。作为可能的起始事件,本研究通过定量逆转录-聚合酶链反应追踪了直接纯化的DENV与血小板的结合情况,并进行了抗原确认。在37°C时,每个血小板约有800个病毒特异性结合。在血库储存温度25°C(约350个结合位点)或已知会减弱病毒进入细胞能力的4°C(约200个结合位点)时,观察到的结合位点较少。树突状细胞特异性细胞间黏附分子3结合非整合素(DC-SIGN)和硫酸乙酰肝素蛋白聚糖被认为是共受体,因为只有抗DC-SIGN和低分子量肝素的组合才能阻止结合。有趣的是,在37°C和25°C时,血小板在7天内将DENV的正链单链RNA基因组复制了约4倍。进一步的时间进程实验证明了病毒NS1蛋白的产生,已知该蛋白在患者血清中具有高度抗原性。DENV的感染性在体外会自然衰减,而血小板介导的活后代产生可缓解这种衰减。使用转录抑制剂证明了这一点,冻融变性的血小板无法复制DENV基因组也证实了这一点。这些数据首次表明血小板可饱和性地直接结合DENV并产生感染性病毒。因此,DENV编码抗原的表达是病原体诱导血小板减少机制中的一个新考量因素。这些结果还进一步引发了人们对血小板除了DENV之外可能产生允许性RNA病毒的可能性的关注。