title: Why to compare absolute numbers of mitochondria.
abstract: Prompted by pronounced structural differences between rat liver and rat hepatocellular carcinoma mitochondria, we suspected these mitochondrial populations to differ massively in their molecular composition. Aiming to reveal these mitochondrial differences, we came across the issue on how to normalize such comparisons and decided to focus on the absolute number of mitochondria. To this end, fluorescently stained mitochondria were quantified by flow cytometry. For rat liver mitochondria, this approach resulted in mitochondrial protein contents comparable to earlier reports using alternative methods. We determined similar protein contents for rat liver, heart and kidney mitochondria. In contrast, however, lower protein contents were determined for rat brain mitochondria and for mitochondria from the rat hepatocellular carcinoma cell line McA 7777. This result challenges mitochondrial comparisons that rely on equal protein amounts as a typical normalization method. Exemplarily, we therefore compared the activity and susceptibility toward inhibition of complex II of rat liver and hepatocellular carcinoma mitochondria and obtained significant discrepancies by either normalizing to protein amount or to absolute mitochondrial number. Importantly, the latter normalization, in contrast to the former, demonstrated a lower complex II activity and higher susceptibility toward inhibition in hepatocellular carcinoma mitochondria compared to liver mitochondria. These findings demonstrate that solely normalizing to protein amount may obscure essential molecular differences between mitochondrial populations.
authors: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany.
authors: Sabine
authors: S
authors: Schmitt
authors: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany.
authors: Sabine
authors: S
authors: Schulz
authors: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany.
authors: Eva-Maria
authors: EM
authors: Schropp
authors: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany.
authors: Carola
authors: C
authors: Eberhagen
authors: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany.
authors: Alisha
authors: A
authors: Simmons
authors: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany.
authors: Wolfgang
authors: W
authors: Beisker
authors: Research Unit Analytical Pathology-Institute of Pathology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany.
authors: Michaela
authors: M
authors: Aichler
authors: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, D-85764 Neuherberg, Germany. Electronic address: zischka@helmholtz-muenchen.de.
authors: Hans
authors: H
authors: Zischka
mesh_terms: D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D045744:Cell Line, Tumor; D005434:Flow Cytometry; D006801:Humans; D008099:Liver / Q000166:cytology* / Q000378:metabolism; D008113:Liver Neoplasms / Q000378:metabolism*; D008928:Mitochondria / Q000502:physiology*; D024101:Mitochondrial Proteins / Q000378:metabolism; D051381:Rats
chemical_list: D024101:Mitochondrial Proteins
keywords: Metabolic shift; Mitochondria; Mitochondrial number; Mitochondrial protein content; Respiratory complex II