Two pathways for induction of apoptosis by ultraviolet radiation in cultured human keratinocytes.
Loss of attachment may induce apoptosis in epithelial cells, but it is unclear whether substrate adhesion modulates apoptosis triggered by genotoxic agents such as ultraviolet radiation (UV). To investigate this issue, we plated neonatal human keratinocytes on different substrates and irradiated them with UVB. DNA strand breaks were nick-labeled to identify apoptotic nuclei. Keratinocytes grown in monolayers were less susceptible to UV-induced apoptosis than were cells freshly seeded on glass (ED50 2130 +/- 96 J per m2, mean +/- SD, versus 131 +/- 96 J per m2, mean +/- SD, respectively). This phenomenon depended on differences in integrin-mediated adhesion, because blocking of integrin beta1 with a monoclonal antibody increased sensitivity of keratinocyte monolayers to UV and an increase in beta1 integrin receptor occupancy by plating on fibronectin, type IV collagen, or keratinocyte-derived extracellular matrix diminished the UV-dependent apoptosis. Down-regulation of p53 with an anti-sense oligonucleotide did not affect apoptosis in glass-plated keratinocytes but effectively suppressed apoptosis in keratinocytes adhering via beta1 integrin. Thus, in addition to the known p53-dependent pathway, UV was able to induce a p53-independent apoptosis that could be blocked by integrin-mediated cell attachment (the integrin-sensitive pathway). The susceptibility to the p53-dependent apoptosis, but not to the integrin-sensitive process, varied among keratinocytes of different clonogenic potential: transit amplifying cells > stem cells > terminally differentiated cells. The p53-independent integrin-sensitive apoptotic pathway may provide an additional mechanism counteracting UV carcinogenesis in the skin.
Gniadecki
R
R
Department of Dermatology, University of Copenhagen, Bispebjerg Hospital, Denmark.
Hansen
M
M
Wulf
H C
HC
D018829:Adhesins, Bacterial / Q000187:drug effects; D017209:Apoptosis / Q000528:radiation effects*; D015153:Blotting, Western; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D006801:Humans; D019012:Integrin beta1 / Q000494:pharmacology; D015603:Keratinocytes / Q000737:chemistry / Q000166:cytology* / Q000528:radiation effects; D012680:Sensitivity and Specificity; D016159:Tumor Suppressor Protein p53 / Q000032:analysis; D014466:Ultraviolet Rays
D018829:Adhesins, Bacterial; D019012:Integrin beta1; D016159:Tumor Suppressor Protein p53