Effects of calcipotriol (MC 903) on proliferation and differentiation of cultured human epidermal keratinocytes

Background: Calcipotriol (MC 903), a new vitamin D₃ analogue, has been reported to be effective in the treatment of patients with psoriasis. Objective: The purpose of this study is to examine the effects of calcipotriol on proliferation and differentiation of the keratinocytes in monolayer cultures and three-dimensional cultures. Methods: Using monolayer cultures, we examined morphological changes of keratinocytes and performed [³H]thymidine incorporation after calcipotriol was added into the medium. Using three dimensional cultures, we performed two experiments: one with cultures treated with calcipotriol immediately after the keratinocytes had been exposed to the air and another set of cultures treated with calcipotriol after three dimensional morphogenesis of the keratinocytes. We examined morphological changes of keratinocytes and performed a immunohistochemical study for proliferation and differentiation markers. Results: In monolayer cultures, at calcipotriol concentrations of 10^-9 M-10^-6 M, keratinocytes became larger, more irregular, and flattened in a dose-dependent manner. At 10^-9 M-10^-6 M, [³H]thymidine incorporation was decreased dose-dependently as compared to the control culture. In the first experiment using three-dimensional cultures, at 10^-9 M-10^-6 M, total epidermal layers were thinned. This was associated with thinnings of nucleated and horny layers in a dose-dependent manner. In the second experiment using three-dimensional cultures, at 10^-8 M-10^-6 M, nucleated layers were thinned in a dose-dependent manner, but the horny layer was slightly thickened, as compared to the control culture. Immunohistochemical studies showed a reduction of differentiation markers such as keratin 1, involucrin, filaggrin, loricrin consistent with a thinning of nucleated layers in the epidermal architecture in both experiments. In the basal layer, at 10^-9 M-10^-6 M, PCNA-positive cells and BrdU-positive cells were decreased dose-dependently as compared to the control culture. Conclusions: In this study, we demonstrated that at 10^-9 M-10^-6 M calcipotriol inhibited keratinocytes proliferation and stimulated keratinocytes differentiation in a dose-dependent manner.