16-Kauren-2-beta-18,19-triol inhibits melanosome transport in melanocytes by down-regulation of melanophilin expression

Background: Rab27a, Mlph, and MyoVa form a tripartite complex and relate to melanosome distribution. Melanophilin (Mlph) acts as a linker protein between Rab27a and MyoVa. The biological activity and function of 16-kauren on the expression of Mlph has not yet been studied. Objective: We examined the effect of 16-kauren on melanosome transport and skin pigmentation. Methods: Murine Melan-a melanocytes and SP-1 keratinocytes were used for in vitro analysis. Western blot analysis, quantitative real-time polymerase chain reaction, luciferase assay and immunohistochemical staining in 3D pigmented human skin model were performed. Results: We found that 16-kauren inhibits melanosome transport in Melan-a melanocytes without affecting melanin synthesis. Treatment with 16-kauren reduced melanophilin (Mlph), a key protein in melanosome transport, in Melan-a melanocytes, at both the protein and mRNA levels while it did not affect the expression of Rab27a and MyoVa, the other two key proteins for melanosome transport. Notably, the expression of melanogenic proteins, including tyrosinase, trp1, trp2, and MITF, was not affected by 16-kauren. However, 16-kauren attenuated melanosome distribution in co-culture of Melan-a melanocytes and SP-1 keratinocytes as well as in Melan-a monolayer culture. In further confirmation of the depigmenting effects of 16-kauren on Melanoderm™, a 3D pigmented human skin model, treatment with 16-kauren for 12 days increased the brightness of the tissue as determined by lightness value and reduced the distribution of melanosomes as shown in histological examination. Conclusion: These results demonstrated that 16-kauren is a selective modulator of a melangenic target, Mlph expression, and can be employed as a new depigmenting strategy.