Persistent p53 mutations in single cells from normal human skin - PubMed (original) (raw)

Persistent p53 mutations in single cells from normal human skin

G Ling et al. Am J Pathol. 2001 Oct.

Abstract

Epidermal clones of p53-mutated keratinocytes are abundant in chronically sun-exposed skin and may play an important role in early development of skin cancer. Advanced laser capture microdissection enables genetic analysis of targeted cells from tissue sections without contamination from neighboring cells. In this study p53 gene mutations were characterized in single cells from normal, chronically sun-exposed skin. Biopsies were obtained from skin subjected to daily summer sun and skin totally protected from the sun by blue denim fabric. Using laser capture microdissection, 172 single-cell samples were retrieved from four biopsies and analyzed using single-cell polymerase chain reaction and direct DNA sequencing. A total of 14 different mutations were identified in 26 of 99 keratinocytes from which the p53 gene could be amplified. Mutations displayed a typical UV signature and were detected in both scattered keratinocytes and in a small cluster of p53-immunoreactive keratinocytes. This minute epidermal p53 clone had a diameter of 10 to 15 basal cells. Two missense mutations were found in all layers of epidermis within the p53 clone. The presented data show that p53 mutations are common in normal skin and that a clone of keratinocytes with a mutated p53 gene prevailed despite 2 months of total protection from ultraviolet light.

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Figures

Figure 1.

Figure 1.

Immunohistochemically stained epidermis from skin protected from UV radiation during 2 summer months. Note rare p53-immunoreactive keratinocyte in epidermis before microdissection (A) and after isolation using laser-assisted microdissection (B). Original magnifications, ×400.

Figure 2.

Figure 2.

Immunohistochemically stained (no counter stain) epidermis from B, revealing a small cluster of p53-immunoreactive keratinocytes found in normal skin after 2 months of total protection from UV radiation. Original low magnification, ×100 (A) and original high magnification ×400 (B). The epidermal p53 clone measures 10 to 15 cells in diameter and is enclosed within an area of 0.05 mm2.

Figure 3.

Figure 3.

Topography of the detected p53 mutations identified in the small cluster of p53-immunoreactive keratinocytes from Figure 2B ▶ is illustrated. Two frequent missense mutations (codon 241 and codon 218) were present in all three layers of epidermis. Both p53-immunoreactive (red) and nonimmunoreactive (yellow) keratinocytes show mutations.

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