Age-related differences in cigarette smoke extract-induced H 2O 2 production by lung endothelial cells

Research output: Contribution to journalArticle

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Abstract

Cigarette smoke causes oxidative stress in the lung resulting in injury and disease. The purpose of this study was to determine if there were age-related differences in cigarette smoke extract (CSE)-induced production of reactive species in single and co-cultures of alveolar epithelial type I (AT I) cells and microvascular endothelial cells harvested from the lungs (MVECLs) of neonatal, young and old male Fischer 344 rats. Cultures of AT I cells and MVECLs grown separately (single culture) and together (co-culture) were exposed to CSE (1, 10, 50, 100%). Cultures were assayed for the production of intracellular reactive oxygen species (ROS), hydroxyl radical (OH), peroxynitrite (ONOO -), nitric oxide (NO) and extracellular hydrogen peroxide (H 2O 2). Single and co-cultures of AT I cells and MVECLs from all three ages produced minimal intracellular ROS in response to CSE. All ages of MVECLs produced H 2O 2 in response to CSE, but young MVECLs produced significantly less H 2O 2 compared to neonatal and old MVECLs. Interestingly, when grown as a co-culture with age-matched AT I cells, neonatal and old MVECLs demonstrated ~50% reduction in H 2O 2 production in response to CSE. However, H 2O 2 production in young MVECLs grown as a co-culture with young AT I cells did not change with CSE exposure. To begin investigating for a potential mechanism to explain the reduction in H 2O 2 production in the co-cultures, we evaluated single and co-cultures for extracellular total antioxidant capacity. We also performed gene expression profiling specific to oxidant and anti-oxidant pathways. The total antioxidant capacity of the AT I cell supernatant was ~5 times greater than that of the MVECLs, and when grown as a co-culture and exposed to CSE (≥10%), the total antioxidant capacity of the supernatant was reduced by ~50%. There were no age-related differences in total antioxidant capacity of the cell supernatants. Gene expression profiling found eight genes to be significantly up-regulated or down-regulated. This is the first study to describe age-related differences in MVECLs exposed to CSE.

Original languageEnglish (US)
Pages (from-to)311-317
Number of pages7
JournalMicrovascular Research
Volume82
Issue number3
DOIs
StatePublished - Nov 2011

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Endothelial cells
Smoke
Tobacco Products
Endothelial Cells
Coculture Techniques
Alveolar Epithelial Cells
Lung
Cell culture
Antioxidants
Gene Expression Profiling
Oxidants
Gene expression
Reactive Oxygen Species
Peroxynitrous Acid
Oxidative stress
Inbred F344 Rats
Hydroxyl Radical
Hydrogen Peroxide
Rats
Nitric Oxide

ASJC Scopus subject areas

  • Biochemistry
  • Cardiology and Cardiovascular Medicine
  • Cell Biology

Cite this

Age-related differences in cigarette smoke extract-induced H 2O 2 production by lung endothelial cells. / Downs, Charles A.; Montgomery, David W; Merkle, Carrie J.

In: Microvascular Research, Vol. 82, No. 3, 11.2011, p. 311-317.

Research output: Contribution to journalArticle

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abstract = "Cigarette smoke causes oxidative stress in the lung resulting in injury and disease. The purpose of this study was to determine if there were age-related differences in cigarette smoke extract (CSE)-induced production of reactive species in single and co-cultures of alveolar epithelial type I (AT I) cells and microvascular endothelial cells harvested from the lungs (MVECLs) of neonatal, young and old male Fischer 344 rats. Cultures of AT I cells and MVECLs grown separately (single culture) and together (co-culture) were exposed to CSE (1, 10, 50, 100{\%}). Cultures were assayed for the production of intracellular reactive oxygen species (ROS), hydroxyl radical (OH), peroxynitrite (ONOO -), nitric oxide (NO) and extracellular hydrogen peroxide (H 2O 2). Single and co-cultures of AT I cells and MVECLs from all three ages produced minimal intracellular ROS in response to CSE. All ages of MVECLs produced H 2O 2 in response to CSE, but young MVECLs produced significantly less H 2O 2 compared to neonatal and old MVECLs. Interestingly, when grown as a co-culture with age-matched AT I cells, neonatal and old MVECLs demonstrated ~50{\%} reduction in H 2O 2 production in response to CSE. However, H 2O 2 production in young MVECLs grown as a co-culture with young AT I cells did not change with CSE exposure. To begin investigating for a potential mechanism to explain the reduction in H 2O 2 production in the co-cultures, we evaluated single and co-cultures for extracellular total antioxidant capacity. We also performed gene expression profiling specific to oxidant and anti-oxidant pathways. The total antioxidant capacity of the AT I cell supernatant was ~5 times greater than that of the MVECLs, and when grown as a co-culture and exposed to CSE (≥10{\%}), the total antioxidant capacity of the supernatant was reduced by ~50{\%}. There were no age-related differences in total antioxidant capacity of the cell supernatants. Gene expression profiling found eight genes to be significantly up-regulated or down-regulated. This is the first study to describe age-related differences in MVECLs exposed to CSE.",
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