In Utero Ethanol Exposure Causes Mitochondrial Dysfunction, Which Can Result in Apoptotic Cell Death in Fetal Brain: A Potential Role for 4-Hydroxynonenal

Authors

  • Vinitha Ramachandran,

    Corresponding author
    1. University of Texas Health Science Center at San Antonio, Department of Medicine, Division of Gastroenterology and Nutrition, San Antonio, Texas.
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  • Antonio Perez,

    1. University of Texas Health Science Center at San Antonio, Department of Medicine, Division of Gastroenterology and Nutrition, San Antonio, Texas.
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  • Juanjuan Chen,

    1. University of Texas Health Science Center at San Antonio, Department of Medicine, Division of Gastroenterology and Nutrition, San Antonio, Texas.
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  • Duraisamy Senthil,

    1. University of Texas Health Science Center at San Antonio, Department of Medicine, Division of Gastroenterology and Nutrition, San Antonio, Texas.
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  • Steven Schenker,

    1. University of Texas Health Science Center at San Antonio, Department of Medicine, Division of Gastroenterology and Nutrition, San Antonio, Texas.
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  • George I. Henderson

    1. University of Texas Health Science Center at San Antonio, Department of Medicine, Division of Gastroenterology and Nutrition, San Antonio, Texas.
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Reprint requests: Vinitha Ramachandran, PhD, University of Texas Health Science Center at San Antonio, Department of Medicine, Division of Gastroenterology and Nutrition, 7703 Floyd Curl Dr., MSC 7878, San Antonio, TX 78229-3900; Fax: 210-567-1976; E-mail: ramachandrav@uthscsa.edu

Abstract

Background: In utero ethanol exposure causes abnormal fetal brain development that may partly be due to enhanced cell death. The mechanisms underlying this remain to be defined, but ethanol-induced oxidative stress may play a role. The following studies investigated the effects of short-term in utero ethanol exposure on fetal brain mitochondrial events that are known to elicit apoptotic cell death. Evidence is presented suggesting that 4-hydroxynonenal (HNE), a toxic product of lipid oxidation, is a causal factor in the observed mitochondrial damage.

Methods: Mitochondria were isolated from control and ethanol-exposed fetal brains (days 17 and 18 of gestation). Permeability transition was determined spectrophotometrically, and cytochrome c and apoptosis-inducing factor (AIF) release were assessed by Western blotting. Caspase-3 activity and DNA fragmentation were determined both as markers for mitochondrially mediated apoptosis and as consequences of cytochrome c and AIF release.

Results: Maternal ethanol intake caused an increase in mitochondrial permeability transition, and this was accompanied by cytochrome c and AIF release from fetal brain mitochondria that exceeded control values by 62 and 25%, respectively (p < 0.05). In utero ethanol exposure resulted in a 30% increase in caspase-3 activity and a 25% increase in DNA fragmentation (p < 0.05) in the fetal brain. HNE levels were increased by 23% (p < 0.05) in mitochondria by in vivo ethanol exposure. In vitro treatment of fetal brain mitochondria with HNE (25–100 μM) also caused increases in mitochondrial permeability transition, as well as dose-dependent releases of cytochrome c and AIF.

Conclusions: These studies illustrate that in utero ethanol exposure can elicit a cascade of events in the fetal brain that are consistent with mitochondrially mediated apoptotic cell death. Additionally, the increase in mitochondrial content of HNE after ethanol intake and the ability of HNE added to fetal brain mitochondria to mimic these effects of in vivo ethanol exposure support a potential role for HNE in the proapoptotic responses to ethanol.

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