Mechanical Stress-Based Cutaneous Autophagy Activation Promotes Hair Regeneration

With the fast pace of modern life,the incidence of pathological hair loss has been increasing.In addition to genetic and drug factors,excessive physical fatigue and psychological stress,as well as irregular work and rest are the most common factors related to hair loss.Hair loss not only affects personal appearance but also causes serious worries and disturbances to patients’ social interactions and psychology,seriously affecting their emotions and quality of life.Pathological hair loss mainly includes seborrheic alopecia,drug-induced alopecia,alopecia areata,and so on.Although the pathogenesis of pathological alopecia is not fully clear,it is crucial to take certain treatments to activate hair follicles into a new growth cycle and promote hair regeneration.It has already been reported that drug-induced autophagy can activate hair follicles to promote hair growth.However,the effective dosage of drug is difficult to control precisely due to the problem of the transdermal and absorptive efficiency.Therefore,a more accessible and controllable stimulation approach to activating autophagy would contribute to the effective treatment of hair loss.This research focuses on physical-mechanical stimulation to activate autophagy for hair regeneration,to investigate safe and effective methods and approaches for treatment of hair loss.Cellular autophagy is an important process of intracellular material turnover and cellular self-renewal,which plays an important role in maintaining the cyclical changes of hair follicles.During pathological hair loss,the hair follicle is in the resting phase for a long-term period,when the level of autophagy is extremely low.Increasing the level of autophagy in the skin during the resting phase can promote hair follicle from the resting phase to the anagen phase and treat hair loss.In traditional Chinese medicine,mechanical stimulation such as skin acupuncture and plum blossom needle has certain clinical therapeutic effects on hair loss,but its mechanism of action is unclear.Inspired by this,we explore the therapeutic effect of mechanical force stimulation on alopecia areata and its related mechanisms based on the changes in the level of skin autophagy.The main research of this paper is described as follows.(1)In the second chapter of the study,we firstly used a model of hair cycle synchronization in mice to explore the effects of different skin stimulation patterns on the effects of resting skin autophagy,hair cycle,and hair growth in the hair follicles.It was found that the autophagy level in the skin was increased in mechanical stimulation groups compared with untreated group,resulting from the upregulation of autophagyrelated protein LC3 and the downregulation of autophagy substrate P62 protein.Among different mechanical stimulation groups,the effect of pointed needle type(plum blossom needle)on autophagy is the best under the same force of stimulation.Moreover,it could stimulate hair follicles to enter the anagen phase earlier than the other groups.Next,we optimized and screened the stimulation parameters of the pointed needle type,including different stimulation force(1.5 m N,15 m N,30 m N),stimulation frequencies(75,150,300,600 times/day),and stimulation days(1,4,7 days).The results showed that the best stimulation parameters were 15 m N,300 times/day,and 4 days of mechanical stimulation(once every other day within a week),which could enhance the level of autophagy in the skin of the resting phase,promote the hair follicle cycle into anagen phase and promote hair regeneration in mice.Finally,to further verify whether mechanical stimulation-induced hair follicle stem cell activation via autophagy,we introduced autophinib,an autophagy inhibitor,to inhibit the autophagy.The experimental results showed that when the autophagy was inhibited by autophinib,the mechanical stimulation could not upregulate the expression of autophagosome-related proteins LC3 or downregulate the expression of P62 protein,indicating that autophinib could inhibit the increase of autophagy level induced by mechanical stimulation in mice.Subsequently,hair growth was also inhibited in the mice.This demonstrates that mechanical stimulation activates hair follicles and promotes hair growth through upregulation of autophagy levels.(2)In the third chapter of the study,we applied the optimized mechanical force stimulation protocol for the treatment of mouse hair loss models,including the alopecia areata model,the corticosterone stress hair loss model,and the high-fat alopecia model.In the alopecia areata model,mechanical force stimulation effectively promoted hair regrowth at the patchy alopecia region,with a significant decrease in the patchy baldness score(p<0.05)and hair growth at the stimulated patchy alopecia region compared with the untreated group,indicating that mechanical force stimulation had a significant treatment effect on patchy hair loss.In the corticosterone stress hair loss model,mechanical stimulation did not show a significant effect on hair growth during continuous application of corticosterone,but the skin in the mechanical force stimulation group entered the anagen phase earlier than the untreated group when corticosterone was withdrawn,indicating that mechanical force stimulation has a role in the recovery of corticosterone stress hair loss.In the high-fat diet-induced alopecia model,normal skin fat cells were mainly distributed in the subcutaneous fat layer and sebaceous glands,while fat particles in the high-fat group were distributed in all layers of the skin and the particles became larger.Especially in the epidermis,fat thickness increased significantly compared with that in normal skin.Mechanical stimulation reduced the percentage of fat particles per unit area in each skin layer,and decreased the thickness of the superficial fat layer.It indicated that mechanical force stimulation regulated fat metabolism in the skin through autophagy and promoted hair follicle activation and hair regeneration.The optimized mechanical force stimulation protocol exhibited better effect on hair regrowth promotion in the baldness model,the corticosterone alopecia model,and the high-fat alopecia model.In conclusion,we found mechanical stimulation could promote hair regulation via autophagy activation,and the optimized mechanical stimulation protocol effectively promoted the hair regeneration of pathological hair loss models.This study not only explored the mechanisms of mechanical stimulation promoting hair regeneration but also provide a safe and effective treatmental approach for hair loss.