Qin1,2, J-Z., Chaturvedi1,2, V., Denning1, M.F., Bacon1, P., Panella1, J., Choubey2, D., and Nickoloff1, B.J. 1Department of Pathology, Loyola University Medical Center, Illinois, USA; 2Department of Radiation Oncology, Loyola University Medical Center, Illinois, USA. Oncogene, 21, 2991-3002, (2002).

This study by researchers at Loyola University Medical Center demonstrated that MatTek’s EpiDerm in vitro 3-D human skin tissue equivalent can be used to study the sensitivity and resistance of human epidermal keratinocytes to apoptosis when exposed to UV radiation. The carcinogenic effects of sunlight in human epidermis may be thwarted by either: transient growth arrest and repair of DNA photodamage in keratinocytes (KCs); elimination of keratinocytes with damaged DNA via apoptosis; or by stimulating a senescence switch whereby keratinocytes become irreversibly growth arrested. Using normal human skin organ cultures and living epidermal equivalents, we demonstrate that in the proliferative basal layer, removal of keratinocytes via apoptosis had a rapid onset (beginning within 2 h) following UV-light exposure generating progressively greater numbers of keratinocytes with thymine dimers as the dose of UV-light was increased; involved induction of Apaf-1, activation of caspase-3, and was dependent on p53 activation as addition of a p53 chemical inhibitor blocked the apoptotic response. Suprabasal layer keratinocytes underwent apoptosis at much later time points (48 h). Keratinocytes in the basal layer repaired DNA damage more rapidly than keratinocytes in supra-basal layers. Steady state levels of p53 increased in irradiated cells, and the increase was accompanied by phosphorylation of serine 9 and serine 15, but not serine 6 residues. By contrast, cultured keratinocytes undergoing spontaneous replicative senescence were resistant to UV-induced apoptosis. Senescent keratinocytes constitutively contained low levels of p53, which were neither increased nor phosphorylated or acetylated after UV-exposure and possessed minimal DNA binding activity, indicative of functional inactivation. Furthermore, treatment of senescent keratinocytes with DNA damaging agent adriamycin did not result in activation of latent p53 or apoptosis. When keratinocytes within psoriatic plaques were examined, they resembled senescent keratinocytes in that they expressed p53, which was not phosphorylated or acetylated. Thus, UV-light induces DNA damage in human epidermal keratinocytes triggering p53 activation, and subsequent apoptosis involving distinct cell layers and kinetics. However, the lack of p53 activation as seen in senescent keratinocytes and psoriatic plaques, is associated with a relative resistance of keratinocytes to UV-induced apoptosis. In conclusion, the sensitivity and resistance of keratinocytes to apoptosis depends not only on the location within various layers of epidermis and levels of p53, but may also involve p53 activation via post-translational modifications.


Caspase 3, Caspase 9, EPI-200-3S, EpiDerm, Pro-Caspase 3, Ultraviolet radiation, p53, p53 inhibitor

Materials Tested

Pifithrin-hydrobromide (PFT)

Request a copy of this paper, click here.