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Related Experiment Videos

Human keratinocytes catabolize thymidine.

P M Schwartz1, L C Kugelman, Y Coifman

  • 1Department of Dermatology, Veterans Administration Medical Center, West Haven, Connecticut 06516.

The Journal of Investigative Dermatology
|January 1, 1988
PubMed
Summary

Human keratinocytes rapidly break down thymidine, affecting DNA incorporation studies. Continuous thymidine addition is needed for accurate in vitro proliferation measurements.

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Area of Science:

  • Dermatology
  • Cell Biology
  • Biochemistry

Background:

  • Human neonatal foreskin keratinocytes proliferate in vitro and incorporate exogenous thymidine into DNA.
  • Keratinocytes' ability to catabolize thymidine has not been previously reported.
  • Thymidine catabolism varies significantly between different tissues and species.

Purpose of the Study:

  • To investigate the catabolism of thymidine by human keratinocytes.
  • To determine the implications of thymidine catabolism for in vitro proliferation studies.
  • To explore the potential impact on thymidine analog therapies for skin diseases.

Main Methods:

  • Cultured human neonatal foreskin keratinocytes were used to study thymidine incorporation and catabolism.
  • Stratified keratinocyte cultures were assessed for their ability to reduce thymidine levels in the medium.

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  • Soluble extracts from various human and animal skin tissues, fibroblasts, and melanocytes were analyzed for thymidine catabolic activity.

    • Main Results:

    • Human keratinocytes, including those from neonatal foreskin and adult skin, actively catabolize thymidine.
    • Stratified keratinocyte cultures reduced medium thymidine by over 90% within 2-4 hours.
    • Linear incorporation of thymidine into DNA was observed for only 2 hours without continuous replenishment, necessitating ongoing addition of radioactive thymidine for extended studies.
    • Skin extracts from mice, rabbits, and guinea pigs showed no thymidine catabolism, unlike human skin.
    • Cultivated human fibroblasts and melanocytes exhibited minimal to no thymidine catabolic activity.

    Conclusions:

    • Keratinocytes possess a significant capacity to catabolize thymidine, impacting [3H]thymidine incorporation assays.
    • Accurate assessment of keratinocyte proliferation in vitro requires strategies to overcome rapid thymidine degradation, such as continuous thymidine addition.
    • Understanding thymidine catabolism in keratinocytes is crucial for the development and application of thymidine analog therapies for cutaneous conditions.