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Proteomic and Transcriptional Profiles of Human Stem Cell-Derived ? Cells Following Enteroviral Challenge.

ABSTRACT: Enteroviral infections are implicated in islet autoimmunity and type 1 diabetes (T1D) pathogenesis. Significant ?-cell stress and damage occur with viral infection, leading to cells that are dysfunctional and vulnerable to destruction. Human stem cell-derived ? (SC-?) cells are insulin-producing cell clusters that closely resemble native ? cells. To better understand the events precipitated by enteroviral infection of ? cells, we investigated transcriptional and proteomic changes in SC-? cells challenged with coxsackie B virus (CVB). We confirmed infection by demonstrating that viral protein colocalized with insulin-positive SC-? cells by immunostaining. Transcriptome analysis showed a decrease in insulin gene expression following infection, and combined transcriptional and proteomic analysis revealed activation of innate immune pathways, including type I interferon (IFN), IFN-stimulated genes, nuclear factor-kappa B (NF-?B) and downstream inflammatory cytokines, and major histocompatibility complex (MHC) class I. Finally, insulin release by CVB4-infected SC-? cells was impaired. These transcriptional, proteomic, and functional findings are in agreement with responses in primary human islets infected with CVB ex vivo. Human SC-? cells may serve as a surrogate for primary human islets in virus-induced diabetes models. Because human SC-? cells are more genetically tractable and accessible than primary islets, they may provide a preferred platform for investigating T1D pathogenesis and developing new treatments.

SUBMITTER: Nyalwidhe JO

PROVIDER: S-EPMC7074978 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

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Proteomic and Transcriptional Profiles of Human Stem Cell-Derived β Cells Following Enteroviral Challenge.

Nyalwidhe Julius O JO Jurczyk Agata A Satish Basanthi B Redick Sambra S Qaisar Natasha N Trombly Melanie I MI Vangala Pranitha P Racicot Riccardo R Bortell Rita R Harlan David M DM Greiner Dale L DL Brehm Michael A MA Nadler Jerry L JL Wang Jennifer P JP

Microorganisms 20200220 2

Enteroviral infections are implicated in islet autoimmunity and type 1 diabetes (T1D) pathogenesis. Significant β-cell stress and damage occur with viral infection, leading to cells that are dysfunctional and vulnerable to destruction. Human stem cell-derived β (SC-β) cells are insulin-producing cell clusters that closely resemble native β cells. To better understand the events precipitated by enteroviral infection of β cells, we investigated transcriptional and proteomic changes in SC-β cells c ...[more]

PMID: 32093375

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Project description:AimsWe prospectively isolate and characterize first and second heart field- and nodal-like cardiomyocytes using a double reporter line from human embryonic stem cells. Our double reporter line utilizes two important transcription factors in cardiac development, TBX5 and NKX2-5. TBX5 expression marks first heart field progenitors and cardiomyocytes while NKX2-5 is expressed in nearly all myocytes of the developing heart (excluding nodal cells). We address the shortcomings of prior work in the generation of heart field-specific cardiomyocytes from induced pluripotent stem cells and provide a comprehensive early developmental transcriptomic as well as electrophysiological analyses of these three populations.Methods and resultsTranscriptional, immunocytochemical, and functional studies support the cellular identities of isolated populations based on the expression pattern of NKX2-5 and TBX5. Importantly, bulk and single-cell RNA sequencing analyses provide evidence of unique molecular signatures of isolated first and second heart field cardiomyocytes, as well as nodal-like cells. Extensive electrophysiological analyses reveal dominant atrial action potential phenotypes in first and second heart fields in alignment with our findings in single-cell RNA sequencing. Lastly, we identify two novel surface markers, POPDC2 and CORIN, that enable purification of cardiomyocytes and first heart field cardiomyocytes, respectively.ConclusionsWe describe a high-yield approach for isolation and characterization of human embryonic stem cell-derived heart field-specific and nodal-like cardiomyocytes. Obtaining enriched populations of these different cardiomyocyte subtypes increases the resolution of gene expression profiling during early cardiogenesis, arrhythmia modelling, and drug screening. This paves the way for the development of effective stem cell therapy to treat diseases that affect specific regions of the heart- or chamber-specific congenital heart defects.

Project description:Bovine Respiratory Disease (BRD) is a major source of economic loss within the agricultural industry. Vaccination against BRD-associated viruses does not offer complete immune protection and vaccine failure animals present potential routes for disease spread. Serological differentiation of infected from vaccinated animals (DIVA) is possible using antigen-deleted vaccines, but during virus outbreaks DIVA responses are masked by wild-type virus preventing accurate serodiagnosis. Previous work by the authors has established the potential for metabolomic profiling to reveal metabolites associated with systemic immune responses to vaccination. The current study builds on this work by demonstrating for the first time the potential to use plasma metabolite profiling to differentiate between vaccinated and non-vaccinated animals following infection-challenge. Male Holstein Friesian calves were intranasally vaccinated (Pfizer RISPOVAL®PI3+RSV) and subsequently challenged with Bovine Parainfluenza Virus type-3 (BPI3V) via nasal inoculation. Metabolomic plasma profiling revealed that viral challenge led to a shift in acquired plasma metabolite profiles from day 2 to 20 p.i., with 26 metabolites identified whose peak intensities were significantly different following viral challenge depending on vaccination status. Elevated levels of biliverdin and bilirubin and decreased 3-indolepropionic acid in non-vaccinated animals at day 6 p.i. may be associated with increased oxidative stress and reactive oxygen scavenging at periods of peak virus titre. During latter stages of infection, increased levels of N-[(3α,5β,12α)-3,12-dihydroxy-7,24-dioxocholan-24-yl]glycine and lysophosphatidycholine and decreased enterolactone in non-vaccinated animals may reflect suppression of innate immune response mechanisms and progression to adaptive immune responses. Levels of hexahydrohippurate were also shown to be significantly elevated in non-vaccinated animals from days 6 to 20 p.i. These findings demonstrate the potential of metabolomic profiling to identify plasma markers that can be employed in disease diagnostic applications to both differentially identify infected non-vaccinated animals during disease outbreaks and provide greater information on the health status of infected animals.

Dataset Information

Proteomic and Transcriptional Profiles of Human Stem Cell-Derived ? Cells Following Enteroviral Challenge.

Publications

Proteomic and Transcriptional Profiles of Human Stem Cell-Derived β Cells Following Enteroviral Challenge.

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