Characterisation of apoptotic responses in human keratinocytes expressnig alpha-6 integrin and CD44
Abstract
a6 integrin has previously been proposed as a human keratinocyte stem cell
markers. CD44-positive cells in epithelial tumours have been identified as
potential cancer stem cells (CSCs) and display increased survival following
chemotherapy.
Keratinocytes expressing CD44 and a high level of a6 integrin (a6
integrinhigh+/CD44+ cells) displayed stem cell-like growth qualities in vitro
including the formation of holoclone-like colonies and a high proliferative
capacity. Irradiation with ultraviolet-B (UVB) initiated apoptosis in the total
keratinocyte population. However, the a6 integrinhigh+/CD44+ cells were
significantly resistant to UVB-induced apoptosis. This fraction was also
resistant to apoptosis following treatment with the genotoxic agents etoposide,
camptothecin and bleomycin. In contrast, the population was highly apoptotic
following cisplatin treatment. Addition of the PI3 Kinase inhibitors Wortmannin
and LY294002 prior to inducement increased the apoptotic sensitivity of the a6
integrinhigh+/CD44+ cells to UVB. However, pAkt was not directly involved due to
the addition of a specific Akt-inhibitor not affecting the resistance of the cells to
UVB.
Cisplatin can induce p63-mediated apoptosis via c-abl activation. Preincubation
with the c-abl kinase inhibitor imatinib significantly reduced the level
of cisplatin-induced apoptosis in the a6 integrinhigh+/CD44+ population.
Interestingly, the a6 integrinhigh+/CD44+ cells displayed a significantly higher
level of TAp63 mRNA compared to the rest of the cell population. The nuclear
expression of p63 was reduced, not increased, following cisplatin treatment and
imatinib inhibited this loss.
This thesis has identified a pattern of apoptotic behaviour not previously
characterised in human keratinocytes. a6 integrinhigh+/CD44+ cells display a
consistent apoptotic resistance to UVB, etoposide and camptothecin and a
3
sensitivity to cisplatin in vitro. Increased levels of TAp63 and evidence of PI3K
and c-abl signalling have provided clues as to the pathways involved in this
behaviour.
Authors
Wray, HelenCollections
- Theses [3706]