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Cross‐talks between intracellular zinc increases and reactive oxygen species in hypoxia
Author(s) -
Slepchenko Kira,
Li Yang V
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.779.6
Subject(s) - zinc , hypoxia (environmental) , reactive oxygen species , cytosol , intracellular , mitochondrion , oxygen , chemistry , microbiology and biotechnology , biology , biochemistry , enzyme , organic chemistry
Zinc recently has emerged as a contributing factor in cellular injury and death caused by hypoxia. Reactive oxygen species (ROS) are also known to accumulate during hypoxia, and are damaging to normal cellular function. We hypothesized that there is a connection between these two important signaling events with potential cause and effect relationship between them. To test the hypothesis, we used fluorescent probes to visualize both free cytosolic zinc (FluoZin‐3, AM) and mitochondrial ROS (MitoSOX Red) in live HeLa cells undergoing hypoxia. We observed a sharp increase in free cytosolic zinc within 60 seconds of hypoxia, which was followed by rapid decrease in free zinc. This zinc response resembled a wave because it had the ascending and descending component, we named it phase I (ascending) and phase II (descending). There was a latency period observed, were the zinc levels remained elevated compared to baseline but did not change during this phase. The latency period is a III phase of the hypoxic zinc response. There is a second significant zinc increase after 18–20 minutes of hypoxia and we named it IV phase of zinc response. This is the first detailed description of the initial zinc transients during hypoxia, where we describe four distinct phases. The first mitochondrial ROS increase started after 5 minutes of hypoxic exposure, then leveled off. These data show that zinc increase preceded the initial ROS increase. Removal of intracellular zinc with zinc chelator TPEN during hypoxia caused mitochondrial ROS accumulation to be delayed about 10 minutes, showing that when zinc is removed, the initial mitochondrial ROS accumulation is not initiated, suggesting that zinc plays a major role in initiating mitochondrial ROS increase. In normoxic conditions, the application of exogenous zinc by itself induced mitochondrial ROS accumulation, adding to the evidence that zinc plays a key role in initiating mitochondrial ROS increase. We hypothesized that NADPH oxidase, a known ROS producer, may be a link between zinc and mitochondrial ROS. In cells that were exposed to exogenous zinc and NADPH oxidase was inhibited by apocynin, mitochondrial ROS was not observed, confirming that NADPH oxidase plays an important role in zinc induced ROS increase. Western blot of total extracts of HeLa cells subjected to exogenous zinc showed the higher level of NADPH oxidase when compared to cells without zinc treatment. Thus, these results suggest a new mechanism of cross‐talk between zinc and mitochondrial ROS, where free cytosolic zinc increase precedes and causes initial mitochondrial ROS increase with zinc and NADPH oxidase as key players. The first ROS increase injures mitochondria and they in turn release ROS and zinc, further contributing to cell injury. Hypoxia is an underlying cause of many important diseases like stroke, heart attack and pulmonary embolism and these findings further our understanding of cellular response to hypoxia. Support or Funding Information NIH (NS081629 to Y.V. Li)