High-Throughput Microscopy Analysis of Mitochondrial Membrane Potential in 2D and 3D Models

Recent proteomic, metabolomic, and transcriptomic studies have highlighted a connection between changes in mitochondria physiology and cellular pathophysiological mechanisms. Secondary assays to assess the function of these organelles appear fundamental to validate these -omics findings. Although mi...

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Bibliographic Details
Published in:Cells (Basel, Switzerland) Vol. 12; no. 7; p. 1089
Main Authors: Vianello, Caterina, Dal Bello, Federica, Shin, Sang Hun, Schiavon, Sara, Bean, Camilla, Magalhães Rebelo, Ana Paula, Knedlík, Tomáš, Esfahani, Emad Norouzi, Costiniti, Veronica, Lacruz, Rodrigo S., Covello, Giuseppina, Munari, Fabio, Scolaro, Tommaso, Viola, Antonella, Rampazzo, Elena, Persano, Luca, Zumerle, Sara, Scorrano, Luca, Gianelle, Alessio, Giacomello, Marta
Format: Journal Article
Language:English
Published: Switzerland MDPI AG 01.04.2023
MDPI
Subjects:
Amino acids
Analysis
Blood & organ donations
Care and treatment
Cell culture
co-culture
Diagnosis
Dosage and administration
Dyes
Fibroblasts
Fibroblasts - metabolism
Health aspects
High-throughput screening (Biochemical assaying)
Homeostasis
Humans
Image processing
Kinetics
Macrophages
Melanoma
Membrane potential
Membrane Potential, Mitochondrial
Metabolomics
Microscopy
Mitochondria
Mitochondria - metabolism
Mitochondrial diseases
mitochondrial membrane potential
Neural stem cells
NSCs
Organelles
Proteomics
Respiration
single muscle fibers
Spheroids
Tetracycline
Tetracyclines
TMRM
Transcriptomics
Italy
single muscle fibers
NSCs
TMRM
mitochondrial membrane potential
machine learning
spheroids
co-culture
ISSN:2073-4409, 2073-4409
Online Access:Get full text
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Summary:Recent proteomic, metabolomic, and transcriptomic studies have highlighted a connection between changes in mitochondria physiology and cellular pathophysiological mechanisms. Secondary assays to assess the function of these organelles appear fundamental to validate these -omics findings. Although mitochondrial membrane potential is widely recognized as an indicator of mitochondrial activity, high-content imaging-based approaches coupled to multiparametric to measure it have not been established yet. In this paper, we describe a methodology for the unbiased high-throughput quantification of mitochondrial membrane potential in vitro, which is suitable for 2D to 3D models. We successfully used our method to analyze mitochondrial membrane potential in monolayers of human fibroblasts, neural stem cells, spheroids, and isolated muscle fibers. Moreover, by combining automated image analysis and machine learning, we were able to discriminate melanoma cells from macrophages in co-culture and to analyze the subpopulations separately. Our data demonstrated that our method is a widely applicable strategy for large-scale profiling of mitochondrial activity.
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Current address: Department of Biochemistry and Molecular Pharmacology, New York University Medical Center, New York, NY 10016, USA.
These authors contributed equally to this work.
ISSN:2073-4409
2073-4409
DOI:10.3390/cells12071089