Skin brightening benefit of 4-hexylresorcinol in vivo and in vitro and its underlying mechanism

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.