1,25-dihydroxyvitamin D(3) protects human keratinocytes against UV-B-induced damage: In vitro analysis of cell viability/proliferation, DNA-damage and -repair.
The skin is the only organ that has the capacity to photo-synthesize the biological active vitamin D metabolite 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] from 7-dehydocholesterol (7-DHC), following exposure to ultraviolet (UV)-B irradiation. The aim of the present work was to investigate the capacity of 1,25(OH)(2)D(3) to protect human keratinocytes (HaCaT) and squamous cell carcinoma cell lines (SCL-1) against the hazardous effects of UV-B irradiation. Human keratinocytes (HaCaT) and squamous cell carcinoma cell lines (SCL-1) were pretreated with 1,25(OH)(2)D(3) over 48 hours and then irradiated once with UVB-radiation. We evaluated the results of several assays (colony-forming-unit-culture assay, WST-1-assay and crystal violet assay), comparing viability/proliferation in 1,25(OH)(2)D(3)-pretreated cells with controls that were pretreated with the carrier substance ethanol alone. Additionally, we analyzed the effects of 1,25(OH)(2)D(3) on UV-induced DNA damage in HaCaT-keratinocytes by detection of cyclobutane pyrimidine dimers (CPDs) via dot blot analysis. We prove that 1,25(OH)(2)D(3), in a concentration of 10(-7) M, protects human keratinocytes (HaCaT) as well as squamous cell carcinoma cell lines (SCL-1) against the hazardous effects of UV-B-radiation (100 J/cm(2)-1,000 J/cm(2)) in vitro. Moreover, we demonstrate that the number of CPDs induced in HaCaT-keratinocytes after irradiation with UV-B (100 J/cm(2)-1,000 J/cm(2)) was decreased after pretreatment with 1,25(OH)(2)D(3), as compared to carrier-treated controls. Analysis of the time course revealed that the elimination of UV-B-induced DNA-damage in HaCaT-keratinocytes occurs quicker when cells are pretreated with 1,25(OH)(2)D(3) (as compared to controls). To put it in a nutshell, our data support the hypothesis that 1,25(OH)(2)D(3) protects cultured human keratinocytes against the hazardous effects of UV-B radiation.
Trémezaygues
Lea
L
Department of Dermatology; The Saarland University Hospital; Homburg/Saar, Germany.
Seifert
Markus
M
Tilgen
Wolfgang
W
Reichrath
Jörg
J