Review Article
Bioactivation of Nitroglycerin by the Mitochondrial Aldehyde Dehydrogenase

https://doi.org/10.1016/j.tcm.2006.05.001Get rights and content

The mitochondrial aldehyde dehydrogenase (ALDH2, mtALDH) was recently found to catalyze the reduction of nitroglycerin (glyceryl trinitrate [GTN]) to generate nitrite and 1,2-glyceryl dinitrate. The nitrite generated within the mitochondria is metabolized further to generate nitric oxide (NO)-based bioactivity, by reduction to NO and/or by conversion to S-nitrosothiol, as revealed by a series of biochemical, pharmacologic, and genetic studies. These studies also demonstrated that mechanism-based inactivation of mtALDH is involved in the development of GTN tolerance. In mice in which the mtALDH gene was selectively deleted (mtALDH−/−), vascular responsiveness to low but not to high GTN concentrations was eliminated, indicating the existence of an additional mechanism of GTN biotransformation (“high Km” pathway). In addition, bioactivation of isosorbide dinitrate/mononitrate vasodilators is independent of mtALDH. Induction of GTN tolerance in vitro in aortae from normal mice selectively affected responsiveness to low doses of GTN, and the remaining responsiveness to high doses of GTN in mtALDH−/− vasculature did not exhibit tolerance. These findings suggest strongly that the high Km pathway is not involved in the development of GTN tolerance that is mechanism-based. Notably, recent studies indicate that individuals of East Asian origin with the common E487K mutation of mtALDH, which results in decreased mtALDH activity, are significantly less responsive to GTN. These observations in toto provide strong support for the conclusion that mtALDH provides the necessary and sufficient enzymatic mechanism for biotransformation of clinically relevant concentrations of GTN to NO-based vasoactivity and indicate in addition that inactivation of mtALDH plays a significant role in the development of mechanism-based GTN tolerance.

Section snippets

Mitochondrial Aldehyde Dehydrogenase-Catalyzed GTN Biotransformation. How Was It Discovered?

Several candidate enzymes, including glutathionine-S-transferases (GSTs), cytochrome P450 reductase/cytochrome P450, and xanthine oxidoreductase, have been proposed to catalyze GTN biotransformation. However, their possible roles in generating NO bioactivity from GTN remain controversial (Chen et al. 2002). For example, although GSTs can metabolize GTN, the GST inhibitor sulfobromophthalein has no effect on GTN-induced increases in cyclic guanosine monophosphate (cGMP). Moreover, a homozygous

Characterization of mtALDH-Catalyzed GTN Biotransformation

Mitochondrial aldehyde dehydrogenase purified from bovine liver catalyzes predominantly 1,2-GDN formation (1,2-GDN/1,3-GDN, ∼8:1) from low levels of GTN (1 μM). In the presence of NAD+, the 1,2-GDN/1,3-GDN ratio increases to more than 20:1 and the rate of GTN metabolism increases ∼10-fold. The optimum pH of 9.0 is about the same as that of aldehyde dehydrogenase activity (Chen et al. 2002). In a subsequent analysis of overexpressed human mtALDH, Kollau et al. (2005) reported that NAD+ addition

Inhibition of mtALDH-Mediated GTN Reductase Activity by ALDH Inhibitors

Our finding that mtALDH functions as a GTN reductase provides a new explanation for the inhibitory effect of GTN on aldehyde dehydrogenase activity (alcohol-GTN drug interaction) and also indicates that ALDH substrates and inhibitors should suppress GTN reductase activity. We examined the effects of the substrate acetaldehyde and of different classes of ALDH inhibitors on GTN bioactivation.

1,2-GDN formation by purified bovine liver mtALDH was inhibited by the classic substrate analog ALDH

Mitochondrial Nitrite Reductase Activity

The results discussed above suggest that nitrite generated from GTN by mtALDH within the mitochondria (Eq. (1)) is further reduced or converted to NO bioactivity and that this NO bioactivity is exported from mitochondria to the cytosol, where it activates sGC (sGC is not detected in purified mitochondria by immunoblotting; Chen and Stamler, unpublished observation). To demonstrate directly the generation and export of NO bioactivity by mitochondria, we used a reporter assay to test whether

Biochemistry

As expected, GTN biotransformation to 1,2-GDN is essentially eliminated in the mitochondria of mtALDH−/− mice (>90% decrease compared to wild-type mitochondria), whereas 1,3-GDN production is not affected. Accordingly, mtALDH−/− mitochondria almost completely lose the ability to generate cGMP from 1 μM GTN in the RFL-6 cell reporter assay (Chen et al. 2005).

In intact aorta, biotransformation of GTN to 1,2-GDN was eliminated at low GTN concentrations (<1 μM) and significantly attenuated at 1–10

Clinical Implications

Nitrovasodilator therapy has not provided survival benefits. It is possible that mitochondrial damage and impaired respiration (Needleman and Hunter 1966) that are consequent upon the localization of GTN biotransformation to mitochondria may provide at least a partial explanation for the failure of long-term GTN therapy to improve mortality. Glyceryl trinitrate-related adverse atherogenic effects and increased morbidity also merit further investigation (Nakamura et al., 1999, Gori and Parker,

Summary and Perspective

Recent biochemical, pharmacologic, and genetics studies have demonstrated clearly that mtALDH is the essential enzyme in the biotransformation of clinical levels of GTN, and that it plays a significant role in the development of GTN tolerance. These studies, and in particular analyses in mtALDH knockout mice, also indicate that there is an additional mechanism for GTN biotransformation (high Km pathway), but that this pathway probably plays no role in the development of mechanism-based GTN

Acknowledgments

We thank Dr. Douglas T. Hess for valuable discussions and critical reading and editing of the manuscript.

References (49)

  • KH Moon et al.

    Inhibition of mitochondrial aldehyde dehydrogenase by nitric oxide-mediated S-nitrosylation

    FEBS Lett

    (2005)
  • N Mukerjee et al.

    Inactivation of human aldehyde dehydrogenase by isosorbide dinitrate

    J Biol Chem

    (1994)
  • Y Nakamura et al.

    Long-term nitrate use may be deleterious in ischemic heart disease: A study using the databases from two large-scale postinfarction studies

    Am Heart J

    (1999)
  • ML Shen et al.

    Role of disulfiram in the in vitro inhibition of rat liver mitochondrial aldehyde dehydrogenase

    Biochem Pharmacol

    (2000)
  • GR Thatcher et al.

    Nitrates and NO release: Contemporary aspects in biological and medicinal chemistry

    Free Radic Biol Med

    (2004)
  • K Baxter et al.

    Heart preservation with Celsior solution improved by the addition of nitroglycerine

    Transplantation

    (2001)
  • GW Brudvig et al.

    Reactions of nitric oxide with cytochrome c oxidase

    Biochemistry

    (1980)
  • Z Chen et al.

    Identification of the enzymatic mechanism of nitroglycerin bioactivation

    Proc Natl Acad Sci U S A

    (2002)
  • Z Chen et al.

    An essential role for mitochondrial aldehyde dehydrogenase in nitroglycerin bioactivation

    Proc Natl Acad Sci U S A

    (2005)
  • A Daiber et al.

    Oxidative stress and mitochondrial aldehyde dehydrogenase activity: A comparison of pentaerythritol tetranitrate with other organic nitrates

    Mol Pharmacol

    (2004)
  • A Daiber et al.

    Heterozygous deficiency of manganese superoxide dismutase in mice (Mn-SOD+/−): A novel approach to assess the role of oxidative stress for the development of nitrate tolerance

    Mol Pharmacol

    (2005)
  • J DiFabio et al.

    Role of mitochondrial aldehyde dehydrogenase in nitrate tolerance

    Mol Pharmacol

    (2003)
  • HL Fung

    Biochemical mechanism of nitroglycerin action and tolerance: Is this old mystery solved?

    Annu Rev Pharmacol Toxicol

    (2004)
  • T Gori et al.

    The puzzle of nitrate tolerance: Pieces smaller than we thought?

    Circulation

    (2002)
  • Cited by (0)

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