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Related Experiment Videos

Keratinocyte superoxide generation

C P Turner1, A M Toye, O T Jones

  • 1Department of Biochemistry, University of Bristol, School of Medical Sciences, UK.

Free Radical Biology & Medicine
|January 23, 1998
PubMed
Summary

Keratinocyte cells (H357) produce significant superoxide. Production is modulated by inflammatory cytokines like IL-1beta and IFNgamma, and calcium, but not typical NADPH oxidase activators.

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Area of Science:

  • Biochemistry
  • Cell Biology
  • Dermatology

Background:

  • Superoxide generation is crucial in cellular signaling and inflammation.
  • Keratinocytes, the primary cells of the epidermis, play a role in skin immunity and wound healing.

Purpose of the Study:

  • To investigate superoxide production in the H357 keratinocyte cell line.
  • To identify pathways and stimuli that regulate superoxide generation in these cells.

Main Methods:

  • Quantification of superoxide production using cytochrome c reduction assay.
  • Treatment of H357 cells with various stimuli including cytokines (IL-1beta, IFNgamma), calcium ionophore (Ionomycin), and enzyme inhibitors.
  • Assessment of the effect of cell confluence on superoxide release.

Main Results:

  • H357 keratinocytes generate superoxide at a rate of 8.36 nmol/h/10^6 cells.
  • Superoxide release decreased with increasing cell confluence.
  • Interleukin-1 beta (IL-1beta) and Ionomycin significantly increased superoxide production.
  • Interferon gamma (IFNgamma) reduced superoxide release.
  • Lipoxygenase inhibitors decreased superoxide production, while inhibitors of cyclo-oxygenase, xanthine oxidase, and NADPH oxidase were ineffective.
  • NADH or NADPH addition to cells increased superoxide production.

Conclusions:

  • H357 keratinocytes are a source of significant superoxide production.
  • Superoxide generation in these cells is modulated by inflammatory cytokines and calcium, suggesting a non-phagocytic NADPH oxidase or alternative pathway.
  • Lipoxygenase pathway appears involved in keratinocyte superoxide production.

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