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A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS)

OBJECTIVE: Human precision cut lung slices (PCLS) are widely used as an ex vivo model system for drug discovery and development of new therapies. PCLS reflect the functional heterogeneity of lung tissue and possess relevant lung cell types. We thus determined the use of PCLS in studying non-coding R...

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Autores principales: Niehof, Monika, Reamon-Buettner, Stella Marie, Danov, Olga, Hansen, Tanja, Sewald, Katherina
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8252208/
https://www.ncbi.nlm.nih.gov/pubmed/34215333
http://dx.doi.org/10.1186/s13104-021-05674-w
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author Niehof, Monika
Reamon-Buettner, Stella Marie
Danov, Olga
Hansen, Tanja
Sewald, Katherina
author_facet Niehof, Monika
Reamon-Buettner, Stella Marie
Danov, Olga
Hansen, Tanja
Sewald, Katherina
author_sort Niehof, Monika
collection PubMed
description OBJECTIVE: Human precision cut lung slices (PCLS) are widely used as an ex vivo model system for drug discovery and development of new therapies. PCLS reflect the functional heterogeneity of lung tissue and possess relevant lung cell types. We thus determined the use of PCLS in studying non-coding RNAs notably miRNAs, which are important gene regulatory molecules. Since miRNAs play key role as mediators of respiratory diseases, they can serve as valuable prognostic or diagnostic biomarkers, and in therapeutic interventions, of lung diseases. A technical limitation though is the vast amount of agarose in PCLS which impedes (mi)RNA extraction by standard procedures. Here we modified our recently published protocol for RNA isolation from PCLS to enable miRNA readouts. RESULTS: The modified method relies on the separation of lysis and precipitation steps, and a clean-up procedure with specific magnetic beads. We obtained successfully quality miRNA amenable for downstream applications such as RTqPCR and whole transcriptome miRNA analysis. Comparison of miRNA profiles in PCLS with published data from human lung, identified all important miRNAs regulated in IPF, COPD, asthma or lung cancer. Therefore, this shows suitability of the method for analyzing miRNA targets and biomarkers in the valuable human PCLS model. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13104-021-05674-w.
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spelling pubmed-82522082021-07-06 A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS) Niehof, Monika Reamon-Buettner, Stella Marie Danov, Olga Hansen, Tanja Sewald, Katherina BMC Res Notes Research Note OBJECTIVE: Human precision cut lung slices (PCLS) are widely used as an ex vivo model system for drug discovery and development of new therapies. PCLS reflect the functional heterogeneity of lung tissue and possess relevant lung cell types. We thus determined the use of PCLS in studying non-coding RNAs notably miRNAs, which are important gene regulatory molecules. Since miRNAs play key role as mediators of respiratory diseases, they can serve as valuable prognostic or diagnostic biomarkers, and in therapeutic interventions, of lung diseases. A technical limitation though is the vast amount of agarose in PCLS which impedes (mi)RNA extraction by standard procedures. Here we modified our recently published protocol for RNA isolation from PCLS to enable miRNA readouts. RESULTS: The modified method relies on the separation of lysis and precipitation steps, and a clean-up procedure with specific magnetic beads. We obtained successfully quality miRNA amenable for downstream applications such as RTqPCR and whole transcriptome miRNA analysis. Comparison of miRNA profiles in PCLS with published data from human lung, identified all important miRNAs regulated in IPF, COPD, asthma or lung cancer. Therefore, this shows suitability of the method for analyzing miRNA targets and biomarkers in the valuable human PCLS model. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13104-021-05674-w. BioMed Central 2021-07-02 /pmc/articles/PMC8252208/ /pubmed/34215333 http://dx.doi.org/10.1186/s13104-021-05674-w Text en © The Author(s) 2021 https://creativecommons.org/licenses/by/4.0/Open AccessThis 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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Research Note
Niehof, Monika
Reamon-Buettner, Stella Marie
Danov, Olga
Hansen, Tanja
Sewald, Katherina
A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS)
title A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS)
title_full A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS)
title_fullStr A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS)
title_full_unstemmed A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS)
title_short A modified protocol for successful miRNA profiling in human precision-cut lung slices (PCLS)
title_sort modified protocol for successful mirna profiling in human precision-cut lung slices (pcls)
topic Research Note
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8252208/
https://www.ncbi.nlm.nih.gov/pubmed/34215333
http://dx.doi.org/10.1186/s13104-021-05674-w
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