EFFECTS OF 1,25 DIHYDROXYVITAMIN D-3 AND CALCIUM ON GROWTH AND DIFFERENTIATION AND ON C-FOS AND P53 GENE-EXPRESSION IN NORMAL HUMAN KERATINOCYTES

Calcium enhances keratinocyte differentiation, and 1,25 dihydroxyvitamin D-3 (1,25(OH)(2)D-3) is both antiproliferative and prodifferentiative in many cell types, including normal human keratinocytes. In the present study, we examined the combined effects of calcium and 1,25(OH)(2)D-3 on parameters of growth and differentiation and on c-fos and p53 gene expression in normal human keratinocytes. Exposure of normal human keratinocytes to 1,25(OH)(2)D-3 markedly reduced [H-3] thymidine incorporation and cell number at low and high medium Ca++ concentrations. Simultaneously, cells in the G(0)/G(1) phase of the cell cycle increased significantly and those in the S phase fell precipitously. 1,25(OH)(2)D-3 and calcium also induced keratinocyte differentiation independently, as assessed by immunocytochemistry and by induction of involucrin mRNA. Both Ca++ and 1,25(OH)(2)D-3 were shown, by nuclear run-on assays, to increase involucrin gene transcription. A rapid, transient elevation in c-fos protooncogene expression preceded these effects when epidermal growth factor was present alone. When 1,25(OH)(2)D-3 was added to quiescent keratinocytes, there was a marked augmentation of c-fos mRNA accumulation at low and high medium Ca++ concentrations. Varying medium Ca++ concentrations had no effect on c-fos mRNA levels. Increasing medium Ca++ concentrations from 0.15 to 2.0mM produced marked elevations of p53 mRNA accumulation and of the rate of p53 gene transcription, whereas 1,25(OH)(2)D-3 had no effect. These results, therefore, suggest that 1,25(OH)(2)D-3 and calcium act in concert to modulate the expression of two important cell-cycle-associated genes, which may be important components in the initial programming of growth and differentiation of normal human keratinocytes.