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Aphidicolin induces 6-thioguanine resistant mutants in human diploid fibroblasts

Ada Kolman1, Natalia Kotova, Jan Grawé

  • 1Department of Molecular Biology and Functional Genomics, Stockholm University, SE-106 91, Stockholm, Sweden. ada.kolman@molbio.su.se

Mutation Research
|February 6, 2002
PubMed

Insights

Aphidicolin (APC), a DNA polymerase inhibitor, increases mutations and cell death in human fibroblasts. Longer exposure to APC significantly enhances these cytotoxic and mutagenic effects.

Area of Science:

  • Cell Biology
  • Toxicology
  • Genetics

Background:

  • Aphidicolin (APC) is a known inhibitor of DNA polymerases alpha and delta.
  • Understanding the cytotoxic and mutagenic potential of APC is crucial for assessing its biological impact.

Purpose of the Study:

  • To investigate the cytotoxic and mutagenic effects of aphidicolin (APC) in human diploid VH-10 fibroblasts.
  • To determine the influence of treatment duration and concentration on APC's effects.
  • To analyze the impact of APC on cell cycle progression and the mode of cell death.

Main Methods:

  • Human diploid VH-10 fibroblasts were treated with varying concentrations (10-40 microM) and durations (2 or 4h) of APC.
  • Mutagenicity was assessed by measuring the frequency of 6-thioguanine (6-TG) resistant mutants at the HPRT locus.
  • Cell cycle distribution was analyzed using flow cytometry, and cell death was evaluated via fluorescein-diacetate (FDA) staining and TUNEL assay.

Main Results:

  • APC treatment significantly reduced cell survival, with a more pronounced effect after 4h compared to 2h.
  • A 5-fold and 10-fold increase in 6-TG resistant mutant frequencies was observed after 2h and 4h of APC treatment, respectively.
  • APC induced a G2 cell cycle block and cell death primarily through necrosis, not apoptosis.

Conclusions:

  • Aphidicolin exhibits significant cytotoxic and mutagenic effects on human fibroblasts.
  • Treatment duration is a critical factor influencing the severity of APC's impact.
  • APC-induced cell death occurs mainly via necrosis, with cells arrested in the G2 phase of the cell cycle.

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