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All‐trans retinoic acid inhibits binding of 1,25‐dihydroxy‐vitamin D 3 to the vitamin D receptor in cultured human keratinocytes
Author(s) -
Fogh K.,
Sølvsten H.,
Jøhnke H.,
Kragballe K.
Publication year - 1996
Publication title -
experimental dermatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.108
H-Index - 96
eISSN - 1600-0625
pISSN - 0906-6705
DOI - 10.1111/j.1600-0625.1996.tb00089.x
Subject(s) - western blot , retinoic acid , vitamin , microbiology and biotechnology , calcitriol receptor , chemistry , monoclonal antibody , receptor , vitamin d binding protein , keratinocyte , calcitriol , blot , antibody , biochemistry , vitamin d and neurology , biology , endocrinology , in vitro , immunology , gene
Psoriasis is characterized by hyperproliferation and impared differentiation of epidermal keratinocytes (KCs). Psoriasis can be treated with derivatives of relinoic acid (RA) and vitamin D 3 . Analogues of vitamin D 3 are able to inhibit proliferation and stimulate differentiation of KCs. In contrast, RA inhibits terminal differentiation of KCs. Interactions are known to occur between RA and vitamin D 3 signalling pathways. The purpose of the present study was to determine the effect of all‐trans RA on the binding of 1,25‐dihydroxy‐vitamin D 3 (1,25 (OH) 2 D 3 ) to the vitamin D 3 , receptor (VDR) of cultured human KCs. Cultured KCs from normal adults were incubated with or without RA (10 −9 –10 −7 M) for 4‐24 h. Cells were then harvested, homogenized and ultrasonicated. The extracted protein was incubated with 3H‐1,25 (OH) 2 D 3 (0.015‐1.0 nM) with or without 250‐fold excess nonradioactive 1,25 (OH) 2 D 3 for 24 h and specific binding was determined by use of the dextran coated charcoal binding assay. Western blot analysis utilizing the monoclonal antibody 9A7γ to VDR was performed on protein extracted from the KCs. The bands resulting from Western blot analysis were visualized by enhanced chemiluminescence. From Scatchard analysis it was found that KCs bind 1,25 (OH) 2 D 3 with high affinity (Kd = 0.175 nM). This binding was dose and time dependently inhibited by RA (60% inhibition at 10 −7 M after 24 h of incubation). By Western blot analysis, RA had no effect on the amount of protein extracted from KCs at any of the RA concentrations tested. In conclusion, these results show that binding of vitamin D 3 to its receptor of human KCs can be inhibited markedly by RA without effecting the amount of protein. These results are in contrast to results with other cell types in which RA upregulates binding of 1,25 (OH) 2 D 3 to the VDR. Because interaction between retinoids and vitamin D 3 may occur at different levels during signal trans‐duction, it is not possible to predict from our results whether RA will inhibit the effects of vitamin D 3 , in vivo.