Effects of β-Carotene and Its Cleavage Products in Primary Pneumocyte Type II Cells

Our plain-English summary of Effects of β-Carotene and Its Cleavage Products in Primary Pneumocyte Type II Cells, by C. Haider, F. Ferk, E. Bojaxhi, et al (Antioxidants, 2017).

The study

• Authors: C. Haider, F. Ferk, E. Bojaxhi, G. Martano, H. Stutz, N. Bresgen, S. Knasmüller, A. Alija, P.M. Eckl
• Journal: Antioxidants
• Published: 2017;6(2)
• Cited by: 11 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

β-Carotene has been shown to increase the risk of developing lung cancer in smokers and asbestos workers in two large scale trails, the Beta-Carotene and Retinol Efficacy Trial (CARET) and the Alpha-Tocopherol Beta-carotene Cancer Prevention Trial (ATBC). Based on this observation, it was proposed that genotoxic oxidative breakdown products may cause this effect. In support of this assumption, increased levels of sister chromatid exchanges, micronuclei, and chromosomal aberrations were found in primary hepatocyte cultures treated with a mixture of cleavage products (CPs) and the major product apo-8'carotenal. However, because these findings cannot directly be transferred to the lung due to the exceptional biotransformation capacity of the liver, potential genotoxic and cytotoxic effects of β-carotene under oxidative stress and its CPs were investigated in primary pneumocyte type II cells. The results indicate that increased concentrations of β-carotene in the presence of the redox cycling quinone dimethoxynaphthoquinone (DMNQ) exhibit a cytotoxic potential, as evidenced by an increase of apoptotic cells and loss of cell density at concentrations > 10 µM. On the other hand, the analysis of micronucleated cells gave no clear picture due to the cytotoxicity related reduction of mitotic cells. Last, although CPs induced significant levels of DNA strand breaks even at concentrations ≥ 1 µM and 5 µM, respectively, β-carotene in the presence of DMNQ did not cause DNA damage. Instead, β-carotene appeared to act as an antioxidant. These findings are in contrast with what was demonstrated for primary hepatocytes and may reflect different sensitivities to and different metabolism of β-carotene in the two cell types.

In plain English

This laboratory study of rat lung cells involved no people or asbestos; earlier trials linked β-carotene supplements to more cancer in smokers and asbestos workers. The authors treated type II pneumocytes (air-sac lining cells) from female Fischer 344 rats with a β-carotene breakdown-product mixture, dosed by the β-carotene used to make it. They also tested β-carotene alone or with DMNQ, a chemical that creates oxidative stress. The mixture caused significant DNA strand breaks from 1 µM with serum and 5 µM without serum. It increased apoptotic (self-destructing) cells at 10 µM. Micronucleated cells did not change significantly, but after adjusting for slowed cell division the authors estimated roughly double. β-Carotene alone caused no significant change in cell death or cell division at 50 µM. With DMNQ, 50 µM β-carotene increased cell death and reduced cell division. Micronucleated cells matched control, though the authors warned reduced division could mask a genotoxic effect. DNA damage instead tended to fall (p = 0.073 at 10 µM). The authors concluded pneumocytes were less sensitive than liver cells, though DMNQ may model oxidative stress poorly here.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (cell culture) experiment on primary rat lung cells. The authors' stated rationale was two earlier β-carotene supplementation trials in smokers and asbestos workers. Those trials are background and were not examined in this study.
• Who was studied: Primary pneumocyte type II cells isolated from female Fischer 344 rats weighing about 100 g (number of rats not stated). The work was done at the University of Salzburg and the Medical University of Vienna, Austria.
• Exposure: Cells were treated for three hours with one of the following. (1) A mixture of β-carotene cleavage products (CPs) made by hypochlorite bleaching. Its doses (0.1, 1, 5 and 10 µM) are expressed as µM of β-carotene used for degradation, not as measured amounts of the products. (2) β-carotene at 0.1, 1, 10 and 50 µM, alone or together with the oxidative-stress chemical DMNQ (stated as 40 µM in the methods). No asbestos exposure was studied.
• Main results:
  • Cleavage-product mixture: apoptotic cells increased significantly at 10 µM (µM of β-carotene used for degradation), p less than 0.05.
  • Cleavage-product mixture: DNA damage in the Comet assay increased significantly at 1 µM and above in serum-supplemented cultures. In serum-free cultures it increased significantly at 5 µM and above. These are µM of β-carotene used for degradation.
  • Cleavage-product mixture: no significant changes in micronucleated cells up to 10 µM (data not shown).
  • Cleavage-product mixture: the authors standardized micronucleus levels to a mitotic index of 1% to account for reduced cell division. In controls, 2.45% of cells were micronucleated at a mitotic index of 1.53%, which standardized to 1.6%. At 10 µM, 2.36% were micronucleated at a mitotic index of 0.76%, which standardized to 3.1%. The authors called this indirect evidence of an approximate doubling.
  • β-carotene alone: no statistically significant changes in apoptosis, necrosis or mitotic index, even at 50 µM.
  • β-carotene alone: micronucleated cells were significantly reduced at 50 µM (p less than 0.05).
  • β-carotene plus DMNQ: at 50 µM, apoptotic and necrotic cells increased significantly and mitotic cells decreased significantly. Table 1 gives the values for 50 µM β-carotene plus DMNQ: mitotic cells 0.56 plus or minus 0.12%, apoptotic cells 3.91 plus or minus 2.37%, and necrotic cells 0.66 plus or minus 0.19%.
  • β-carotene plus DMNQ: at 50 µM, micronucleated cells (2.18 plus or minus 1.07) were comparable to the control (2.13 plus or minus 0.48). The authors cautioned that the reduced cell division could mask a genotoxic effect.
  • β-carotene plus DMNQ: increasing β-carotene reduced DNA damage in the Comet assay. The reduction was 'almost significant' (p = 0.073) at 10 µM.
• The authors' conclusion: The authors concluded that, in contrast to hepatocytes (liver cells), pneumocytes were less sensitive to the cleavage products and to β-carotene under oxidative stress. They stated that the results 'could support the assumption' that β-carotene is not causally involved in adverse health effects.
• Limitations the authors state:
  • DMNQ may be an inappropriate way to induce oxidative stress in this cell type.
  • Reduced cell division may have masked a genotoxic effect in the micronucleus analysis.
  • High variation between experiments meant that necrotic cells were not significantly elevated with the cleavage products.
  • Co-culturing pneumocytes with activated macrophages or neutrophils would more realistically reflect the situation of smokers taking β-carotene supplements.

Topics

Medical terms in this study, explained in our Lexicon: Lung Cancer, Micronuclei, Oxidative Stress, Redox Cycling.

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• Primary pleural epithelioid mesothelioma of clear cell type: a case report and review of current literature — C. Gkogkou, K. Samitas, et al. (2011)
• Lobe of origin and histologic type of lung cancer associated with asbestos exposure in the Carotene and Retinol Efficacy Trial (CARET) — C.A. Brodkin, J. McCullough, et al. (1997)
• Role of Beta-Carotene in Lung Cancer Primary Chemoprevention: A Systematic Review with Meta-Analysis and Meta-Regression — J. Kordiak, F. Bielec, et al. (2022)
• Size- and type-specific exposure assessment of an asbestos products factory in China — M.N. Courtice, D.W. Berman, et al. (2016)

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