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Articles in PresS, published online ahead of print May 3, 2002
Am J Physiol Lung Cell Mol Physiol, 10.1152/ajplung.00410.2001
Submitted on October 19, 2001
Accepted on April 20, 2002
1 Department of Pediatrics, National Jewish Medical & Research Center, Denver, CO, USA
2 Department of Medicine, National Jewish Medical & Research Center, Denver, CO, USA
* To whom correspondence should be addressed. E-mail: whitec{at}njc.org.
Increased glucose utilization and hexokinase-II expression are adaptive features of lung cells exposed to hypoxia or hyperoxia. Hexokinase-II is the most regulated isoform of hexokinase. Whether its overexpression could be protective against oxidative stress was explored in human lung epithelial-like (A549) cells. Hexokinase-II was overexpressed in A549 cells using a tetracycline-inducible retroviral vector system. Elevated expression of hexokinase-II was confirmed by western blot and activity measurements. Cell death caused by exposure to hyperoxia was decreased in hexokinase-II overexpressing cells. This effect was reversed when hexokinase-II expression was suppressed using doxycycline. A similar protective effect was observed in hexokinase-II overexpressing cells after treatment with 1 mM hydrogen peroxide for 48 hours. At baseline, fluorescence microscopy showed that overexpressed hexokinase-II was localized to mitochondria. Electron microscopic studies showed that hyperoxia-exposed hexokinase-II overexpressors had better preserved and quantitatively smaller mitochondria than those in which the hexokinase-II expression was suppressed or in the nontransduced A549 cells. Mitochondrial membrane potential was increased in hexokinase-II overexpressing cells exposed to hyperoxia when compared to the nontransduced control cells under similar conditions. The present study demonstrates that hexokinase-II protects human lung epithelial-like A549 cells against oxidative insults by protecting the mitochondria.
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