KUVA (khellin plus ultraviolet A) stimulates proliferation and melanogenesis in normal human melanocytes and melanoma cells in vitro

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Abstract

Background

Khellin is a naturally occurring furochromone which, when combined with artificial ultraviolet (UV) A or solar irradiation (KUVA), is reported to repigment vitiligo skin as effectively as PUVA photochemotherapy. The exact mechanism of KUVA-induced repigmentation is unknown.

Objectives

The aim of this study was to test the effect of khellin and KUVA on proliferation and melanogenesis of normal human melanocytes and Mel-1 melanoma cells in vitro.

Methods

Cultured normal human melanocytes, Mel-1 melanoma cells and fibroblasts were treated with khellin, UVA and KUVA and the effect on proliferation determined by cell counting. The effect on melanogenesis was determined by a radiometric melanin formation assay. Changes in gene expression and protein synthesis were determined by Northern blot, reverse transcriptase–polymerase chain reaction (RT–PCR) and Western blot analyses.

Results

Khellin stimulated proliferation of Mel-1 melanoma cells and melanocytes at concentrations between 1 nmol L−1 and 0.5 mmol L−1 with a peak effect at 0.01 mmol L−1 khellin. In contrast, khellin inhibited proliferation of fibroblasts over the entire concentration range tested. At concentrations above 0.5 mmol L−1, khellin was cytotoxic to both melanocytic cells and fibroblasts. Exposure of khellin-treated cells to single doses of UVA between 150 and 280 mJ cm−2 resulted in an enhanced proliferative effect. Khellin and KUVA also stimulated the melanogenic enzyme activity of pigmented cells, with the most effective treatment being 0.01 mmol L−1 khellin with 250 mJ cm−2 UVA. Western blot, Northern blot and RT–PCR analysis revealed that these increases in melanogenic enzyme activity were not due to increases in gene expression or protein synthesis. UVA treatment resulted in an increase in enzyme glycosylation and this correlated with the increase in melanogenesis.

Conclusions

We conclude that khellin activated by UVA stimulates melanocyte proliferation and melanogenesis. Our results point to the possibility that current treatment regimens might be improved if reduced khellin doses are applied and suggest that improved delivery vehicles be tested.

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