Unknown,Transcriptomics,Genomics,Proteomics

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Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes as an In Vitro Model for Coxsackievirus B3-Induced Myocarditis and Antiviral Drug Screening Platform

ABSTRACT: ABSTRACT Background: Viral myocarditis is a life-threatening illness that may lead to heart failure or cardiac arrhythmias. 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. Methods and Results: Human iPSC-CMs were infected with a luciferase-expressing mutant of the coxsackievirus B3 strain (CVB3-Luc). Brightfield microscopy, immunofluorescence, and calcium imaging were used to characterize virally infected hiPSC-CMs. Viral proliferation on hiPSC-CMs was subsequently quantified using bioluminescence imaging. For drug screening, select antiviral compounds including interferon beta 1 (IFNβ1), ribavirin, pyrrolidine dithiocarbamate (PDTC), and fluoxetine were tested for their capacity to abrogate CVB3-Luc proliferation in hiPSC-CMs in vitro. The ability of some of these compounds to reduce CVB3-Luc proliferation in hiPSC-CMs was consistent with the reported drug effects in previous studies. Finally, mechanistic analyses via gene expression profiling of hiPSC-CMs infected with CVB3-Luc revealed an activation of viral RNA and protein clearance pathways within these hiPSC-CMs after IFNβ1 treatment. Conclusions: This study demonstrates that hiPSC-CMs express the coxsackievirus and adenovirus receptor, are susceptible to coxsackievirus infection, and can be used to confirm antiviral drug efficacy. Our results suggest that the hiPSC-CM/CVB3-Luc assay is a sensitive platform that could be used to screen novel antiviral therapeutics for their effectiveness in a high-throughput fashion. For this experiment, human induced pluripotent stem cell derived cardiomyocytes were infected with coxsackievirus at multiplicity of infection (MOI) of 5 for 8 hours. Cells were treated with and without interferon beta 1 in order to determine if treatment activates antiviral response genes and/or viral clearance pathways. 4 total samples (2 for each condition) were analyzed

ORGANISM(S): Homo sapiens

SUBMITTER: Jared Churko

PROVIDER: E-GEOD-57781 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:ABSTRACT Background: Viral myocarditis is a life-threatening illness that may lead to heart failure or cardiac arrhythmias. 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. Methods and Results: Human iPSC-CMs were infected with a luciferase-expressing mutant of the coxsackievirus B3 strain (CVB3-Luc). Brightfield microscopy, immunofluorescence, and calcium imaging were used to characterize virally infected hiPSC-CMs. Viral proliferation on hiPSC-CMs was subsequently quantified using bioluminescence imaging. For drug screening, select antiviral compounds including interferon beta 1 (IFNβ1), ribavirin, pyrrolidine dithiocarbamate (PDTC), and fluoxetine were tested for their capacity to abrogate CVB3-Luc proliferation in hiPSC-CMs in vitro. The ability of some of these compounds to reduce CVB3-Luc proliferation in hiPSC-CMs was consistent with the reported drug effects in previous studies. Finally, mechanistic analyses via gene expression profiling of hiPSC-CMs infected with CVB3-Luc revealed an activation of viral RNA and protein clearance pathways within these hiPSC-CMs after IFNβ1 treatment. Conclusions: This study demonstrates that hiPSC-CMs express the coxsackievirus and adenovirus receptor, are susceptible to coxsackievirus infection, and can be used to confirm antiviral drug efficacy. Our results suggest that the hiPSC-CM/CVB3-Luc assay is a sensitive platform that could be used to screen novel antiviral therapeutics for their effectiveness in a high-throughput fashion. For this experiment, human induced pluripotent stem cell derived cardiomyocytes were infected with coxsackievirus at multiplicity of infection (MOI) of 5 for 8 hours. Cells were treated with and without interferon beta 1 in order to determine if treatment activates antiviral response genes and/or viral clearance pathways.

Project description:ABSTRACT: Viral aseptic meningitis is a neuroinflammatory condition that occurs when viruses gain access to the central nervous system (CNS) and induce inflammation. The blood-brain barrier (BBB) is comprised of brain endothelial cells (BECs) that stringently regulate the passage of molecules, toxins, and pathogens from the circulation into the CNS. Through their unique properties, such as complex tight junctions, reduced rates of endocytosis, expression of efflux transporters, and restricted expression of leukocyte adhesion molecules, the BBB is often able to limit pathogen entry into the brain; however, certain neurotropic pathogens, such as coxsackievirus B3 (CVB3) are able to infect the CNS. We have previously demonstrated that CVB3 can infect and disrupt induced pluripotent stem cell-derived brain-like endothelial cells (iBECs), but the host response to this infection remains unknown. Here, we investigate global host transcriptional changes during CVB3 infection of iBECs using RNA sequencing. We validated our data set by exploring pathways altered by CVB3 using quantitative real-time PCR (qPCR) and enzyme-linked immunosorbent assay of upregulated cytokines and interferon signaling molecules. IMPORTANCE: Coxsackievirus B3 (CVB3) is a leading cause of viral aseptic meningitis that can cause severe disease in susceptible individuals. To gain access to the central nervous system, CVB3 must cross central nervous system barriers, such as the blood-brain barrier. Previously, we have shown that CVB3 infects a human stem cell-derived brain-like endothelial cell model. Here, we report the global transcriptome of stem cell-derived brain-like endothelial cells to CVB3 infection and provide proof-of-concept validation of the dataset using molecular biology techniques. These data could inform novel mechanisms of CVB3-mediated blood-brain barrier dysfunction. KEYWORDS: Coxsackievirus B3, blood-brain barrier, brain endothelial cells, RNA sequencing, induced pluripotent stem cells

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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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