The in vitro and in vivo skin‐whitening activity of Ectoine through enhanced autophagy in melanocytes and keratinocytes and zebrafish model

Abstract Ectoine, a natural bacterial osmolyte, suppressed UVA irradiated‐α‐melanocyte stimulating hormone (MSH) stimulated melanogenesis through antioxidant Nrf2 pathways in human keratinocytes; however, the underlying skin whitening mechanisms were not elucidated. The depigmenting efficiency of Ectoine (0–400 μM) through antimelanogenesis and melanin degradation by autophagy promotion was investigated in melanoma (B16F10) and melanin‐feeding keratinocyte (HaCaT) cells and in vivo zebrafish model. MTT assay, Western blotting, GFP‐LC3 puncta, AVO formation, melanin assay, immunofluorescence staining, TEM techniques, siLC3 transfection, and zebrafish model were utilized. Ectoine‐induced autophagy in B16F10 and HaCaT cells was shown by enhanced LC3‐II accumulation, autophagosome GFP‐LC3 puncta, autolysosome AVOs formation, ATG4B downregulation, and Beclin‐1/Bcl‐2 dysregulation. The immunoprecipitation data revealed that Ectoine increased the association between LC3‐II and p62 proteins in B16F10 and HaCaT cells. Importantly, antioxidant NAC pretreatment antagonized the Ectoine‐induced ATG4B diminution in B16F10 and HaCaT cells. Ectoine inhibited melanogenesis by suppressing melanosome gp100, tyrosinase, TRP‐1/‐2, and/or melanin formation via autophagy in α‐MSH‐stimulated B16F10 and melanin‐feeding HaCaT cells. TEM findings displayed that Ectoine increased melanosome‐engulfing autophagosomes and autolysosomes in α‐MSH‐stimulated B16F10 and melanin‐feeding HaCaT cells. Ectoine‐inhibited melanogenesis in α‐MSH‐stimulated B16F10 cells and melanin‐feeding HaCaT cells was reversed by pretreatment with the autophagy inhibitor 3‐MA or LC3 silencing. In vivo study demonstrated that Ectoine (5 mM) suppressed endogenous body pigmentation by antimelanogenesis and melanin degradation through autophagy induction in a zebrafish model. The in vitro and in vivo study demonstrated that Ectoine inhibits melanogenesis and enhances melanin degradation by triggering autophagy. Ectoine could be utilized as a whitening ingredient in cosmetic formulations.