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Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis

Complex immune dysregulation is a hallmark of sepsis. The occurring phases of immunosuppression and hyperinflammation require rapid detection and close monitoring. Reliable tools to monitor patient’s immune status are yet missing. Currently, microRNAs are being discussed as promising new biomarkers...

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Autores principales: Hirschberger, Simon, Hübner, Max, Strauß, Gabriele, Effinger, David, Bauer, Michael, Weis, Sebastian, Giamarellos-Bourboulis, Evangelos J., Kreth, Simone
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6823537/
https://www.ncbi.nlm.nih.gov/pubmed/31672997
http://dx.doi.org/10.1038/s41598-019-51782-w
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author Hirschberger, Simon
Hübner, Max
Strauß, Gabriele
Effinger, David
Bauer, Michael
Weis, Sebastian
Giamarellos-Bourboulis, Evangelos J.
Kreth, Simone
author_facet Hirschberger, Simon
Hübner, Max
Strauß, Gabriele
Effinger, David
Bauer, Michael
Weis, Sebastian
Giamarellos-Bourboulis, Evangelos J.
Kreth, Simone
author_sort Hirschberger, Simon
collection PubMed
description Complex immune dysregulation is a hallmark of sepsis. The occurring phases of immunosuppression and hyperinflammation require rapid detection and close monitoring. Reliable tools to monitor patient’s immune status are yet missing. Currently, microRNAs are being discussed as promising new biomarkers in sepsis. However, no suitable internal control for normalization of miRNA expression by qPCR has been validated so far, thus hampering their potential benefit. We here present the first evaluation of endogenous controls for miRNA analysis in human sepsis. Novel candidate reference miRNAs were identified via miRNA microArray. TaqMan qPCR assays were performed to evaluate these microRNAs in T-cells and whole blood cells of sepsis patients and healthy controls in two independent cohorts. In T-cells, U48 and miR-320 proved suitable as endogenous controls, while in whole blood cells, U44 and miR-942 provided best stability values for normalization of miRNA quantification. Commonly used snRNA U6 exhibited worst stability in all sample groups. The identified internal controls have been prospectively validated in independent cohorts. The critical importance of housekeeping gene selection is emphasized by exemplary quantification of imuno-miR-150 in sepsis patients. Use of appropriate internal controls could facilitate research on miRNA-based biomarker-use and might even improve treatment strategies in the future.
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spelling pubmed-68235372019-11-12 Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis Hirschberger, Simon Hübner, Max Strauß, Gabriele Effinger, David Bauer, Michael Weis, Sebastian Giamarellos-Bourboulis, Evangelos J. Kreth, Simone Sci Rep Article Complex immune dysregulation is a hallmark of sepsis. The occurring phases of immunosuppression and hyperinflammation require rapid detection and close monitoring. Reliable tools to monitor patient’s immune status are yet missing. Currently, microRNAs are being discussed as promising new biomarkers in sepsis. However, no suitable internal control for normalization of miRNA expression by qPCR has been validated so far, thus hampering their potential benefit. We here present the first evaluation of endogenous controls for miRNA analysis in human sepsis. Novel candidate reference miRNAs were identified via miRNA microArray. TaqMan qPCR assays were performed to evaluate these microRNAs in T-cells and whole blood cells of sepsis patients and healthy controls in two independent cohorts. In T-cells, U48 and miR-320 proved suitable as endogenous controls, while in whole blood cells, U44 and miR-942 provided best stability values for normalization of miRNA quantification. Commonly used snRNA U6 exhibited worst stability in all sample groups. The identified internal controls have been prospectively validated in independent cohorts. The critical importance of housekeeping gene selection is emphasized by exemplary quantification of imuno-miR-150 in sepsis patients. Use of appropriate internal controls could facilitate research on miRNA-based biomarker-use and might even improve treatment strategies in the future. Nature Publishing Group UK 2019-10-31 /pmc/articles/PMC6823537/ /pubmed/31672997 http://dx.doi.org/10.1038/s41598-019-51782-w Text en © The Author(s) 2019 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Hirschberger, Simon
Hübner, Max
Strauß, Gabriele
Effinger, David
Bauer, Michael
Weis, Sebastian
Giamarellos-Bourboulis, Evangelos J.
Kreth, Simone
Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis
title Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis
title_full Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis
title_fullStr Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis
title_full_unstemmed Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis
title_short Identification of suitable controls for miRNA quantification in T-cells and whole blood cells in sepsis
title_sort identification of suitable controls for mirna quantification in t-cells and whole blood cells in sepsis
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6823537/
https://www.ncbi.nlm.nih.gov/pubmed/31672997
http://dx.doi.org/10.1038/s41598-019-51782-w
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