Abstract:

4-Hexylresorcinol （4-HR）, a potent tyrosinase inhibitor, has been used as an even-tone active ingredient for skin care application since 2007. While the skin brightening efficacy of 4-HR in Chinese population has not been thoroughly investigated and its significance in keratinocytes has not been fully raveled. This study aims to evaluate the skin brightening potential of 4-HR in vivo and in vitro and explore its new mechanism of action through transcriptome approach. The skin brightening effect of 0.4% 4-HR in a facial serum was assessed in an 8-week, double-blinded, placebo-controlled, and randomized clinical study in 67 Chinese participants. ITA°, melanin index （MI） and visual grading were measured at baseline and 2, 4 and 8 weeks after use. A pigmented living skin equivalent （pLSE） model constructed from Asian skin cells was utilized to assess the brightening efficacy of 0.4% 4-HR by measuring the model’s brightness （L^* value） and melanin content. Then, transcriptomic analysis of 4-HR treated human epidermal keratinocytes was conducted, and the two in vitro models were adopted for hypothesis validation afterwards. In the clinical study, the result shows both 0.4% 4-HR serum and placebo chassis can significantly improve all measures as compared to baseline at the 2, 4, and 8 weeks. Furthermore, 0.4% HR serum demonstrates a better performance in increasing ITA° as early as 2 weeks of application and decreasing MI value than the placebo group at Week 2. In the pLSE model, 0.4% 4-HR with topical application evidently increases L^* value by 15.88% and decreases melanin content by 47.61% compared to UVB group. RNA-sequencing analysis implies that 4-HR can regulate multiple biological processes including skin development, keratinocyte differentiation, oxidant activity and autophagy function. In the blue-light challenged human keratinocytes model, 4-HR shows a significant ROS suppression capacity. In the keratinocytes-melanocytes co-culture model, 4-HR prompts autophagy activity and decreases melanin content. Most importantly, the melanin inhibitory activity of 4-HR is compromised after co-treating with Chloroquine, an autophagy inhibitor, suggesting autophagy regulation property of 4-HR may partially contribute to its skin brightening efficacy. Taken together, these data demonstrate skin brightening efficacy of 0.4% 4-HR in vivo and in vitro, in addition to acting as a tyrosinase inhibitor, 4-HR can contribute to skin brightening benefit via enhancing cellular antioxidant capacity and autophagy activation.

Key words: skin brightening, 4-hexylresorcinol, autophagy, oxidation, transcriptomics