Wan Yuepeng,Hu Genhua,Wu Zhiqing

Guangzhou Uniasia Cosmetics Technology Co.,Ltd.,China

Lin Xianting,Yang Jianzhong

Beauty Hi-tech Innovation Co.,Ltd.,China

Abstract Hair loss and gray hair are the ultimate manifestations of scalp and hair aging.Improving metabolic activities in cells is a fundamental for anti-aging while autophagy activity of cell is a key measure.Hinokitiol,one of the more than 30 quasi-drug actives approved in Japan in the field of hair tonic products has two known mechanisms for its effectivities,i.e.promoting cell activity and anti-microbial.Through this study,we identified a new mechanism of hinokitiol for hair loss prevention,i.e.promotion of autophagy and mitophagy activities of dermal papilla cells.

Key words Hinokitiol; hair loss prevention; anti-aging; autophagy; mitophagy

1 Hinokitiol as an active ingredient for hair loss prevention

Hair loss has been a common problem in our society now,especially among the stressed people.In addition to medication after hair loss,more and more people choose to use the medicated cosmetics to prevent hair loss in daily life.In Japan,cosmetics are legally classified into two groups:Cosmetics and Quasi-drugs.The safety and efficacy of quasidrugs is tightly regulated and a permission is required prior to selling the product.The ingredients used for quasi-drugs include actives and additives.The actives are more strictly regulated in terms of its claimable efficacy,dosage,and application filed.Table1 shows the main active ingredients of quasi-drugs for hair growth and loss prevention.Hinokitiol is among the actives having two mechanisms for its effectivities,i.e.promoting cell activity and anti-microbial.[1]Herein,we would like to describe a discovery of a new mechanism for hinokitiol as a Japan approved active ingredient for hair growth and hair loss prevention,i.e.promoting autophagy activity of dermal papilla cells.

Table 1.Active ingredients for hair loss prevention quasi-drugs in Japan

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2 Known effects of hinokitiol

Hinokitiol (β-thujaplicin),properties shown in Figure 1,is one of more than 30 quasi-drug actives approved in Japan in the field of hair tonic products.Hinokitiol is a natural monoterpenoid found in the wood of Cupressaceae trees,which was originally isolated from the essential oil component of Taiwan cypress in 1936 by Tetsuo Nozoe.[2]

Figure 1.Properties of hinokitiol

It has much been reported that hinokitiol exhibits a broad spectrum of antibacterial activity.For example,application of hinokitiol resulted in a reduction in the number of staphylococcus aureus on the skin surface,and an improvement in skin condition after treatment.[3]Hinokitiol also has antibacterial activity to Malassezia.[4]An in-vitro study assessed the anti-inflammatory effect and mechanism of action of hinokitiol in human corneal epithelial cells and reported that it may be clinically used as an external medicine.[5,6]Furthermore,hinokitiol may cure hair loss by suppressing factors that promote follicular apoptosis,such as TNF-alpha,in addition to stimulating new hair growth.[7]It has also been reported that the hair growing activities were enhanced by using the encapsulated hinokitiol nanospheres via in-vitro and in-vivo studies,which transformed the hair growth cycle from the resting phase to the growing phase.[8]

3 Anti-aging via autophagy regulation

Anti-aging is definitely an attractive benefit to many people around the world.In the cosmetic industry,antiaging of skin,scalp and hair has been a key focus and in recent years.Hair loss prevention can be classified as a part of anti-aging for scalp and hair.Improving the metabolic activities in cells is a fundamental to anti-aging and the autophagy activity is a key to determine the metabolic cycle in cells.[9]Autophagy includes a variety of complex physiological and pathophysiological functions,such as adaptation to malnutrition,removal of damaged intracellular proteins and organelles,cell development,anti-aging,elimination of microorganisms,cell death,cancer inhibition and antigen presentation.[10]Mitophagy is a mechanism of selective removal of defective mitochondria due to oxidative stress or DNA damage.The defective mitochondria are degraded by autophagosome and digested with lysosome.[11]The 2016 Nobel Prize in Physiology or Medicine was awarded to the Japanese professor,Yoshinori Ohsumi for his contributions in the discovery of mechanisms for autophagy.This brings a new trend in applying autophagy theory into development of new anti-aging technologies in the field of cosmetics.For example,a study by POLA Chemical Industries Inc.indicates that autophagy helps decompose the old substances accumulated with age into basic substances such as amino acids for recycle,to improve reduced nicotinamide adenine dinucleotide in fibroblasts for skin gloss.[11]

Hair loss and gray hair are the ultimate manifestation of scalp and hair aging.Both are closely related to hair growth cycle.The hair growth cycle includes three stages:anagen,catagen,telogen.The hair follicle functions activity and the matrix cells divide rapidly to form hair in anagen.Furthermore,the melanosomes,tyrosinase and the related proteins are formed to promote the melanocytes synthesizing melanin and make hair coloring in anagen.In contrast,the mitotic activity of matrix cells going down till to stopped and the melanocytes also stop to synthesize melanin when the hair move to catagen.

The research about matrix cells reported that the autophagy activity of hair follicles in telogen is lower than anagen,and the hair follicles in anagen will rapidly move to telogen.In addition,apoptosis will be reduced due to the activation and prolongation of autophagy.[12]Therefore,improving the autophagy activity of hair follicles to accelerate the metabolism of hair follicles is an important means for anti-aging of scalp and hair.Herein,we evaluated the autophagy activity promoting effect of hinokitiol by using human hair follicle dermal papilla cells.

4 New mechanism of hinokitiol for hair loss prevention

Hair dermal papilla cells exist in the dermal papilla located at the bottom of hair follicles,playing crucial roles in hair formation,growth,and cycling.Autophagy and mitophagy evaluations using human dermal papilla cells were performed as follows:①Autophagy:culturing the human dermal papilla cells; adding testing sample to culture;measuring by fluorescent plate reader after stained with Hoechst 33342 nucleic acid stain and cyto ID green detection reagent.② Mitophagy:culturing the human dermal papilla cells; adding testing sample to culture; collecting cells with trypsin EDTA solution;measuring the percentage of cells stained with mitophagy dye detection kit via Tali image cytometer.

Figure 2 and 3 show that hinokitiol can effectively promote both autophagy and mitophagy activities of dermal papilla cells.

Figure 2.Evaluation of autophagy promotion effect

Figure 3.Evaluation of mitophagy promotion effect

These experimental results indicate that the effect of hinokitiol on hair loss prevention may have a new mechanism,i.e.promotion of autophagy activity of dermal papilla cells.

5 Conclusion

Hinokitiol is widely used in personal care products as an active ingredient for hair loss prevention.In this study,we discovered a new mechanism of hinokitiol,i.e.promotion of autophagy and mitophagy activities of dermal papilla cells.