Human induced pluripotent stem cell-derived cardiomyocytes as an in vitro model for coxsackievirus B3-induced myocarditis and antiviral drug screening platform.
Human induced pluripotent stem cell-derived cardiomyocytes as an in vitro model for coxsackievirus B3-induced myocarditis and antiviral drug screening platform.
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DOI：
10.1161/circresaha.115.303810
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发表时间：
2014-08-29
影响因子：
20.1
通讯作者：
Wu JC
中科院分区：
文献类型：
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作者：
Sharma A;Marceau C;Hamaguchi R;Burridge PW;Rajarajan K;Churko JM;Wu H;Sallam KI;Matsa E;Sturzu AC;Che Y;Ebert A;Diecke S;Liang P;Red-Horse K;Carette JE;Wu SM;Wu JC
病毒性心肌炎是一种危及生命的疾病，可能导致心力衰竭或心律失常。病毒性心肌炎的主要病原体是柯萨奇病毒的B3株，这是一种正义RNA肠道病毒。然而，人类心脏组织很难获得足够的数量来研究心脏特异性病毒感染的机制。本研究探讨了人类诱导多能干细胞衍生的心肌细胞（hiPSC-CM）是否可用于模拟柯萨奇病毒诱导的病毒性心肌炎的致病过程，并筛选抗病毒治疗的疗效。人iPSC-CM用表达转铁蛋白酶的柯萨奇病毒B3株（CVB3-Luc）感染。利用双视野显微镜、免疫荧光和钙成像来表征病毒感染的hiPSC-CM的细胞形态和钙处理的改变。使用生物发光成像定量hiPSC-CM中的病毒增殖。测试了包括干扰素β 1（IFN β 1）、利巴韦林、吡咯烷二硫代氨基甲酸酯和氟西汀在内的抗病毒化合物在体外消除hiPSC-CM中CVB3-Luc增殖的能力。这些化合物降低hiPSC-CM中CVB3-Luc增殖的能力与先前研究中报告的药物作用一致。通过CVB3-Luc感染的hiPSC-CM的基因表达谱进行的机制分析显示，IFN β 1处理后，病毒RNA和蛋白质清除途径被激活。这项研究表明，hiPSC-CM表达柯萨奇病毒和腺病毒受体，易受柯萨奇病毒感染，并可用于预测抗病毒药物的疗效。我们的研究结果表明，hiPSC-CM/CVB3-Luc检测是一种灵敏的平台，可以以高通量的方式筛选新型抗病毒治疗剂的有效性。
Viral myocarditis is a life-threatening illness that may lead to heart failure or cardiac arrhythmias. A major causative agent for viral myocarditis is the B3 strain of coxsackievirus, a positive-sense RNA enterovirus. However, human cardiac tissues are difficult to procure in sufficient enough quantities for studying the mechanisms of cardiac-specific viral infection. This study examined whether human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) could be used to model the pathogenic processes of coxsackievirus-induced viral myocarditis and to screen antiviral therapeutics for efficacy. Human iPSC-CMs were infected with a luciferase-expressing coxsackievirus B3 strain (CVB3-Luc). Brightfield microscopy, immunofluorescence, and calcium imaging were utilized to characterize virally-infected hiPSC-CMs for alterations in cellular morphology and calcium handling. Viral proliferation in hiPSC-CMs was quantified using bioluminescence imaging. Antiviral compounds including interferon beta 1 (IFNβ1), ribavirin, pyrrolidine dithiocarbamate, and fluoxetine were tested for their capacity to abrogate CVB3-Luc proliferation in hiPSC-CMs in vitro. The ability of these compounds to reduce CVB3-Luc proliferation in hiPSC-CMs was consistent with reported drug effects in previous studies. Mechanistic analyses via gene expression profiling of hiPSC-CMs infected with CVB3-Luc revealed an activation of viral RNA and protein clearance pathways after IFNβ1 treatment. This study demonstrates that hiPSC-CMs express the coxsackievirus and adenovirus receptor, are susceptible to coxsackievirus infection, and can be used to predict antiviral drug efficacy. Our results suggest that the hiPSC-CM/CVB3-Luc assay is a sensitive platform that can screen novel antiviral therapeutics for their effectiveness in a high-throughput fashion.