Cortisol synthesis by primary human keratinocytes
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Cortisol analogues have been used to treat skin disorders, such as psoriasis and atopic
dermatitis, for over 50 years but the ability of normal human keratinocytes to synthesise
cortisol has not been reported. Keratinocytes are capable of de novo cholesterol
synthesis, they express P450 enzymes that are required for steroidogenesis and can
metabolise androgens and estrogens. In addition, steroidogenic acute regulatory protein
(StAR) that controls the rate determining step of acute steroidogenesis has been
identified in the epidermis. The aim of this thesis was to identify de novo cortisol
steroidogenesis by keratinocytes and investigate the function of cortisol in keratinocytes
in vitro.
Normal epidermis was shown to express three cholesterol transporters that are
associated with promoting steroid synthesis; StAR was identified in the basal layer,
metastatic lymph node 64 (MLN64) in the suprabasal layers and translocator protein
(TSPO) was detected throughout the epidermal layers. In addition, the nuclear receptor
DAX1, a negative regulator of StAR, was identified in the cytoplasm of cells that form
normal epidermis. Comparatively, the expression of these proteins was altered in
psoriasis and atopic dermatitis, where DAX1 was localised to the nucleus of most
diseased tissue and StAR was not detected. This suggests that acute steroid synthesis is
ablated in these hyperproliferative skin conditions.
The ability of normal primary human keratinocytes to synthesise cortisol was
investigated. Radioimmunoassay demonstrated keratinocytes were capable of de novo
pregnenolone synthesis, which was promoted with the cortisol analogue dexamethasone
(dex). Interestingly, 25-hydroxycholesterol, which bypasses StAR, did not further
enhance steroid synthesis. This suggests that there is an alternative rate determining
step of steroid synthesis in cultured primary keratinocytes. Thin layer chromatography
demonstrated keratinocytes could metabolise pregnenolone to progesterone and
progesterone to cortisol. Progesterone metabolism to cortisol was also confirmed with
liquid chromatography/mass spectroscopy.
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Dex was shown to maintain keratinocyte viability and was implicated in promoting
cellular redox potential. Since redox potential is a critical regulator of steroidogenesis,
this observation could provide a mechanism for dex-induced pregnenolone synthesis in
cultured keratinocytes. These observations led to a hypothesis that local cortisol
synthesis functions to regulate cellular redox potential to prevent cell death as part of a
positive feedback system. Therefore, this thesis has identified the cortisol steroidogenic
pathway in primary human keratinocytes and a potential functional mechanism for the
pathway.
Authors
Hannen, Rosalind FrancescaCollections
- Theses [3367]