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Updated: Jun 30, 2025
Assessment of Glutamine as a Fuel Source for Alveolar Macrophages Exposed to Chronic Ethanol Using an Extracellular Flux Bioanalyzer
Published on: November 15, 2024
Pioglitazone reverses alcohol-induced alterations in alveolar macrophage mitochondrial phenotype
Kathryn M Crotty1,2, Shayaan A Kabir1,2, Sarah S Chang1,2
1Division of Pulmonary, Allergy, Critical Care and Sleep Medicine, Department of Medicine, Emory University, Atlanta, Georgia, USA.
Background:
People with alcohol use disorder (AUD) have an increased risk of developing pneumonia and pulmonary diseases. Alveolar macrophages (AMs) are immune cells of the lower respiratory tract that are necessary for clearance of pathogens. However, alcohol causes AM oxidative stress, mitochondrial damage and dysfunction, and diminished phagocytic capacity, leading to lung injury and immune suppression.
Methods:
AMs were isolated by bronchoalveolar lavage from people with AUD and male and female C57BL/6J mice given chronic ethanol (20% w/v, 12 weeks) in drinking water. The peroxisome proliferator-activated receptor γ ligand, pioglitazone, was used to treat human AMs ex vivo (10 μM, 24 h) and mice in vivo by oral gavage (10 mg/kg/day). Levels of AM mitochondrial superoxide and hypoxia-inducible factor-1 alpha (HIF-1α) mRNA, a marker of oxidative stress, were measured by fluorescence microscopy and RT-qPCR, respectively. Mouse AM phagocytic ability was determined by internalized Staphylococcus aureus, and mitochondrial capacity, dependency, and flexibility for glucose, long-chain fatty acid, and glutamine oxidation were measured using an extracellular flux analyzer. In vitro studies used a murine AM cell line, MH-S (±0.08% ethanol, 72 h) to investigate mitochondrial fuel oxidation and ATP-linked respiration.
Results:
Pioglitazone treatment decreased mitochondrial superoxide in AMs from people with AUD and ethanol-fed mice and HIF-1α mRNA in ethanol-fed mouse lungs. Pioglitazone also reversed mouse AM glutamine oxidation and glucose or long-chain fatty acid flexibility to meet basal oxidation needs. In vitro, ethanol decreased the rate of AM mitochondrial and total ATP production, and pioglitazone improved changes in glucose and glutamine oxidation.
Conclusions:
Pioglitazone reversed chronic alcohol-induced oxidative stress in human AM and mitochondrial substrate oxidation flexibility and superoxide levels in mouse AM. Decreased ethanol-induced AM HIF-1α mRNA with pioglitazone suggests that this pathway may be a focus for metabolic-targeted therapeutics to improve morbidity and mortality in people with AUD.
Insights
Pioglitazone treatment reduced oxidative stress and improved mitochondrial function in alveolar macrophages (AMs) affected by alcohol use disorder (AUD). This suggests pioglitazone as a potential therapeutic for alcohol-related lung conditions.
Area of Science:
- Immunology
- Cell Biology
- Pharmacology
Background:
- Alcohol use disorder (AUD) elevates pneumonia risk due to impaired alveolar macrophage (AM) function.
- Alcohol induces oxidative stress and mitochondrial dysfunction in AMs, compromising lung immunity.
Purpose of the Study:
- To investigate the therapeutic potential of pioglitazone in mitigating alcohol-induced damage to AMs.
- To assess pioglitazone's effects on oxidative stress and mitochondrial metabolism in AMs from AUD patients and animal models.
Main Methods:
- Alveolar macrophages (AMs) were isolated from human AUD patients and ethanol-fed mice.
- Pioglitazone was administered ex vivo and in vivo to assess its impact on AM mitochondrial superoxide and HIF-1α mRNA.
- Mitochondrial respiration and phagocytic capacity were evaluated using extracellular flux analysis and Staphylococcus aureus uptake assays.
Main Results:
- Pioglitazone significantly reduced mitochondrial superoxide and HIF-1α mRNA in AMs from AUD subjects and ethanol-fed mice.
- Treatment with pioglitazone restored mitochondrial substrate oxidation flexibility and improved ATP production in ethanol-exposed AMs.
- In vitro studies confirmed that pioglitazone counteracted ethanol-induced impairments in glucose and glutamine oxidation.
Conclusions:
- Pioglitazone effectively reversed alcohol-induced oxidative stress and normalized mitochondrial function in AMs.
- The findings highlight the potential of targeting the HIF-1α pathway with metabolic therapeutics like pioglitazone to improve outcomes for individuals with AUD.
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