title: Mitochondrial respiratory function induces endogenous hypoxia.
abstract: Hypoxia influences many key biological functions. In cancer, it is generally believed that hypoxic condition is generated deep inside the tumor because of the lack of oxygen supply. However, consumption of oxygen by cancer should be one of the key means of regulating oxygen concentration to induce hypoxia but has not been well studied. Here, we provide direct evidence of the mitochondrial role in the induction of intracellular hypoxia. We used Acetylacetonatobis [2-(2'-benzothienyl) pyridinato-kN, kC3'] iridium (III) (BTP), a novel oxygen sensor, to detect intracellular hypoxia in living cells via microscopy. The well-differentiated cancer cell lines, LNCaP and MCF-7, showed intracellular hypoxia without exogenous hypoxia in an open environment. This may be caused by high oxygen consumption, low oxygen diffusion in water, and low oxygen incorporation to the cells. In contrast, the poorly-differentiated cancer cell lines: PC-3 and MDAMB231 exhibited intracellular normoxia by low oxygen consumption. The specific complex I inhibitor, rotenone, and the reduction of mitochondrial DNA (mtDNA) content reduced intracellular hypoxia, indicating that intracellular oxygen concentration is regulated by the consumption of oxygen by mitochondria. HIF-1α was activated in endogenously hypoxic LNCaP and the activation was dependent on mitochondrial respiratory function. Intracellular hypoxic status is regulated by glucose by parabolic dose response. The low concentration of glucose (0.045 mg/ml) induced strongest intracellular hypoxia possibly because of the Crabtree effect. Addition of FCS to the media induced intracellular hypoxia in LNCaP, and this effect was partially mimicked by an androgen analog, R1881, and inhibited by the anti-androgen, flutamide. These results indicate that mitochondrial respiratory function determines intracellular hypoxic status and may regulate oxygen-dependent biological functions.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.
authors.forename: Sara
authors.initials: S
authors.lastname: Prior
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.
authors.forename: Ara
authors.initials: A
authors.lastname: Kim
authors.affiliation: Department of Chemistry and Chemical Biology, Graduate School of Engineering, University of Gunma, Kiryu, Gunma, Japan.
authors.forename: Toshitada
authors.initials: T
authors.lastname: Yoshihara
authors.affiliation: Department of Chemistry and Chemical Biology, Graduate School of Engineering, University of Gunma, Kiryu, Gunma, Japan.
authors.forename: Seiji
authors.initials: S
authors.lastname: Tobita
authors.affiliation: Department of Molecular Medicine, Institute for Molecular and Cellular Regulation, University of Gunma, Maebashi, Gunma, Japan.
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Takeuchi
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Higuchi
mesh_terms: D015374:Biosensing Techniques / Q000379:methods; D015153:Blotting, Western; D015687:Cell Hypoxia / Q000502:physiology*; D045744:Cell Line, Tumor; D019069:Cell Respiration / Q000502:physiology*; D056831:Coordination Complexes; D005947:Glucose / Q000378:metabolism; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000502:physiology*; D009369:Neoplasms / Q000503:physiopathology*; D010100:Oxygen / Q000032:analysis*; D012402:Rotenone
chemical_list: D056831:Coordination Complexes; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; C000594111:acetylacetonatobis(2-(2'-benzothienyl)pyridinato-kN,kC3')iridium(III); D012402:Rotenone; D005947:Glucose; D010100:Oxygen