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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...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2021
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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. |
format | Online Article Text |
id | pubmed-8252208 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
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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