Cited by 19Mitochondrial Function and PathologyNitric Oxide and Endothelin EffectsRedox biology and oxidative stress

Modulation of peroxynitrite produced via mitochondrial nitric oxide synthesis during Ca2+ and succinate-induced oxidative stress in cardiac isolated mitochondria

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Gerdes, Harrison J., Yang, Meiying, Heisner, James S., Camara, Amadou K.S., Stowe, David F.·2020·Biochimica et Biophysica Acta (BBA) - Bioenergetics

Abstract

We hypothesized that NO• is generated in isolated cardiac mitochondria as the source for ONOO- production during oxidative stress. We monitored generation of ONOO- from guinea pig isolated cardiac mitochondria subjected to excess Ca 2+ uptake before adding succinate and determined if ONOO- production was dependent on a nitric oxide synthase (NOS) located in cardiac mitochondria (mtNOS). Mitochondria were suspended in experimental buffer at pH 7.15, and treated with CaCl 2 and then the complex II substrate Na-succinate, followed by menadione, a quinone redox cycler, to generate O 2•-. L-tyrosine was added to the mitochondrial suspension where it is oxidized by ONOO- to form dityrosine (diTyr) in proportion to the ONOO- present. We found that exposing mitochondria to excess CaCl 2 before succinate resulted in an increase in diTyr and amplex red fluorescence (H 2 O 2) signals, indicating that mitochondrial oxidant stress, induced by elevated mtCa 2+ and succinate, increased mitochondrial ONOO- production via NO• and O 2•-. Changes in mitochondrial ONOO- production dependent on NOS were evidenced by using NOS inhibitors L-NAME/L-NNA, TEMPOL, a superoxide dismutase (SOD) mimetic, and PTIO, a potent global NO• scavenger. L-NAME and L-NNA decreased succinate and menadione-mediated ONOO- production, PTIO decreased production of ONOO-, and TEMPOL decreased ONOO- levels by converting more O 2•- to H 2 O 2. Electron microscopy showed immuno-gold labeled iNOS and nNOS in mitochondria isolated from cardiomyocytes and heart tissue. Western blots demonstrated iNOS and nNOS bands in total heart tissue, bands for both iNOS and nNOS in β-tubulin-free non-purified (crude) mitochondrial preparations, and a prominent iNOS band, but no nNOS band, in purified (Golgi and ER-free) mitochondria. Prior treatment of guinea pigs with lipopolysacharride (LPS) enhanced expression of iNOS in liver mitochondria but not in heart mitochondria. Our results indicate that release of ONOO- into the buffer is dependent both on O 2•- released from mitochondria and NO• derived from a mtCa 2+-inducible nNOS isoform, possibly attached to mitochondria, and a mtNOS isoform like iNOS that is non-inducible.

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  • Induction of iNOS usually occurs in the presence of an oxidative environment, which produces peroxynitrite when •NO reacts with superoxide, leading to cellular toxicity.

    “Induction of iNOS usually occurs in the presence of an oxidative environment, which produces peroxynitrite when •NO reacts with superoxide [115], leading to cellular toxicity.”

    From The Role of Reactive Species on Innate Immunity (2022) · cites this paper as [115]

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Authors
Gerdes, H. J., Yang, M., Heisner, J. S., Camara, A. K. S., & Stowe, D. F.
Affiliations
  • Medical College of Wisconsin (US)
  • Marquette University (US)
Corresponding author
Stowe, David F.
Venue
Biochimica et Biophysica Acta (BBA) - Bioenergetics
Publisher
Elsevier BV
Published
2020-12
Volume
1861
Issue
12
Pages
148290
ISSN
0005-2728 (ISSN-L)
OpenAlex
W3081476062
Cited by
19
References
73
Citations by year
2026: 2 · 2025: 3 · 2024: 3 · 2023: 6 · 2022: 3 · 2021: 1 · 2020: 1
FWCI
0.89
Type
journal article
Language
en
Open access
Locations
  • Biochimica et Biophysica Acta (BBA) - Bioenergetics (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (acceptedVersion)
Funders
  • U.S. Department of Veterans Affairs
  • National Institutes of Health (5T35 HL072483-34)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
  • Health Sciences › Medicine › Physiology › Nitric Oxide and Endothelin Effects
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Redox biology and oxidative stress
Keywords
peroxynitrite, mitochondrial nitric oxide synthase, inducible nitric oxide synthase, neuronal nitric oxide synthase, heart mitochondria, oxidative stress, calcium overload, succinic acid, superoxide, nitric oxide, dityrosine, menadione, L-NAME, L-NNA, PTIO, tempol, lipopolysaccharide, cardiomyocytes, mitochondrial calcium, superoxide dismutase
MeSH terms
Animals, Calcium, Electron Transport, Guinea Pigs, Hydrogen Peroxide, Isoenzymes, Mitochondria, Heart, Nitric Oxide, Spectrometry, Fluorescence, Stress, Physiological, Superoxide Dismutase, Time Factors, Free Radical Scavengers, Oxidative Stress, Nitric Oxide Synthase, Succinic Acid, Peroxynitrous Acid, Membrane Potential, Mitochondrial

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