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Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics

Dysregulation of cellular metabolism is now a well-recognized hallmark of cancer. Studies investigating the metabolic features of cancer cells have shed new light onto processes in cancer cell biology and have identified many potential novel treatment options. The advancement of mass spectrometry-ba...

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Autores principales: Rushing, Blake R., Schroder, Madison, Sumner, Susan C. J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8879193/
https://www.ncbi.nlm.nih.gov/pubmed/35208242
http://dx.doi.org/10.3390/metabo12020168
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author Rushing, Blake R.
Schroder, Madison
Sumner, Susan C. J.
author_facet Rushing, Blake R.
Schroder, Madison
Sumner, Susan C. J.
author_sort Rushing, Blake R.
collection PubMed
description Dysregulation of cellular metabolism is now a well-recognized hallmark of cancer. Studies investigating the metabolic features of cancer cells have shed new light onto processes in cancer cell biology and have identified many potential novel treatment options. The advancement of mass spectrometry-based metabolomics has improved the ability to monitor multiple metabolic pathways simultaneously in various experimental settings. However, questions still remain as to how certain steps in the metabolite extraction process affect the metabolic profiles of cancer cells. Here, we use ultra-high-performance liquid chromatography–high-resolution mass spectrometry (UHPLC–HRMS) untargeted metabolomics to investigate the effects of different detachment and lysis methods on the types and abundances of metabolites extracted from MDA-MB-231 cells through the use of in-house standards libraries and pathway analysis software. Results indicate that detachment methods (trypsinization vs. scraping) had the greatest effect on metabolic profiles whereas lysis methods (homogenizer beads vs. freeze–thaw cycling) had a lesser, though still significant, effect. No singular method was clearly superior over others, with certain metabolite classes giving higher abundances or lower variation for each detachment–lysis combination. These results indicate the importance of carefully selecting sample preparation methods for cell-based metabolomics to optimize the extraction performance for certain compound classes.
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spelling pubmed-88791932022-02-26 Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics Rushing, Blake R. Schroder, Madison Sumner, Susan C. J. Metabolites Article Dysregulation of cellular metabolism is now a well-recognized hallmark of cancer. Studies investigating the metabolic features of cancer cells have shed new light onto processes in cancer cell biology and have identified many potential novel treatment options. The advancement of mass spectrometry-based metabolomics has improved the ability to monitor multiple metabolic pathways simultaneously in various experimental settings. However, questions still remain as to how certain steps in the metabolite extraction process affect the metabolic profiles of cancer cells. Here, we use ultra-high-performance liquid chromatography–high-resolution mass spectrometry (UHPLC–HRMS) untargeted metabolomics to investigate the effects of different detachment and lysis methods on the types and abundances of metabolites extracted from MDA-MB-231 cells through the use of in-house standards libraries and pathway analysis software. Results indicate that detachment methods (trypsinization vs. scraping) had the greatest effect on metabolic profiles whereas lysis methods (homogenizer beads vs. freeze–thaw cycling) had a lesser, though still significant, effect. No singular method was clearly superior over others, with certain metabolite classes giving higher abundances or lower variation for each detachment–lysis combination. These results indicate the importance of carefully selecting sample preparation methods for cell-based metabolomics to optimize the extraction performance for certain compound classes. MDPI 2022-02-10 /pmc/articles/PMC8879193/ /pubmed/35208242 http://dx.doi.org/10.3390/metabo12020168 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Rushing, Blake R.
Schroder, Madison
Sumner, Susan C. J.
Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics
title Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics
title_full Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics
title_fullStr Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics
title_full_unstemmed Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics
title_short Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics
title_sort comparison of lysis and detachment sample preparation methods for cultured triple-negative breast cancer cells using uhplc–hrms-based metabolomics
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8879193/
https://www.ncbi.nlm.nih.gov/pubmed/35208242
http://dx.doi.org/10.3390/metabo12020168
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