Retinoic acid drives hair follicle stem cell activation via Wnt/ß-catenin signalling in androgenetic alopecia.

BACKGROUND: Depletion or permanent quiescence of the hair follicle stem cell (HFSC) pool underlies pathogenesis in androgenetic alopecia (AGA). Reactivation of quiescent HFSCs is considered an efficient treatment strategy for hair loss. The retinoic acid (RA) is critical to ensure stem cell homeostasis and function. However, little is known about whether RA regulates HFSC homeostasis. We aimed to investigate the impact of RA on HFSC homeostasis and the underlying mechanisms, in order to provide new potential targets for medical therapies of AGA. METHODS: Microdissected hair follicles from the occipital and frontal scalp in AGA were obtained for RNA sequencing analysis and test. The C57BL/6 mice model in telogen was established to investigate the effect of exogenous RA. Miniaturized hair follicles from frontal scalp were incubated with or without RA in hair follicle organ culture to test the effects on hair shaft elongation, hair cycling and HFSC activities. A strategy to characterize the effect of RA on HFSC in primary culture was developed to identify novel mechanisms that control HFSC activation. A clinical study was performed to test the efficacy of RA treatment in AGA patients. RESULTS: RA signalling was inhibited in the course of AGA pathogenesis along with HFSC dysfunction. Hair regeneration was retarded in AGA miniaturized hair follicles with RA deficiency, but they tended to recover after treatment with RA. In addition, RA treatment during the telogen phase facilitated HFSC anagen entry and accelerated hair growth. Mechanistically, RA promoted hair growth by stimulating stem cells via Wnt/ß-catenin signalling and accelerating the transition from a dormant to an activated state. Furthermore, a clinical study suggested that RA has obvious advantages in the early intervention of AGA by reactivating HFSCs. CONCLUSIONS: Our study provides insights into the reactivation of HFSCs in AGA and provides potential targets for medical therapies.

Drugs targeting TGF-ß/Notch interaction attenuate hypertrophic scar formation by optic atrophy 1-mediated mitochondrial fusion.

Hypertrophic scar (HS) formation is a cutaneous fibroproliferative disease that occurs after skin injuries and results in severe functional and esthetic disability. To date, few drugs have shown satisfactory outcomes for the treatment of HS formation. Transforming growth factor-beta (TGF-ß)/Notch interaction via small mothers against decapentaplegic 3 (Smad3) could facilitate HS formation; therefore, targeting TGF-ß/ Notch interaction via Smad3 is a potential therapeutic strategy to attenuate HS formation. In addition, optic atrophy 1 (OPA1)-mediated mitochondrial fusion contributes to fibroblast proliferation, and TGF-ß/Smad3 axis and the Notch1 pathway facilitate OPA1-mediated mitochondrial fusion. Thus, the aim of this study was to investigate whether drugs targeting TGF-ß/Notch interaction via Smad3 suppressed fibroblast proliferation to attenuate HS formation through OPA1-mediated mitochondrial fusion. We found that the TGF-ß pathway, Notch pathway, and TGF-ß/Notch interaction via Smad3 were inhibited by pirfenidone, the gamma- secretase inhibitor DAPT, and SIS3 in human keloid fibroblasts (HKF) and an HS rat model, respectively. Protein interaction was detected by co-immunoprecipitation, and mitochondrial morphology was determined by electron microscopy. Our results indicated that pirfenidone, DAPT, and SIS3 suppressed the proliferation of HKFs and attenuated HS formation in the HS rat model by inhibiting TGF-ß/Notch interaction via Smad3. Moreover, pirfenidone, DAPT, and SIS3 hindered OPA1-mediated mitochondrial fusion through inhibiting TGF-ß/Notch interaction, thereby suppressing the proliferation of HS fibroblasts and HS formation. In summary, these findings investigating the effects of drugs targeting TGF-ß/Notch interaction on HS formation might lead to novel drugs for the treatment of HS formation.

Prognostic biomarker CCR6 and its correlation with immune infiltration in cutaneous melanoma.

Background: Cutaneous melanoma (CM) is an aggressive type of skin cancer. Even after standard treatment, the recurrence and malignant progression of CM were almost inevitable. The overall survival (OS) of patients with CM varied widely, making it critical for prognostic prediction. Based on the correlation between CCR6 and melanoma incidence, we aimed to investigate the prognostic role of CCR6 and its relationship with immune infiltration in CM. Methods: We obtained RNA sequencing data from The Cancer Genome Atlas (TCGA) to analyze the CM expression. Functional enrichment analyses, immune infiltration analyses, immune checkpoint analyses, and clinicopathology analyses were performed. Univariate and multivariate Cox regression analyses were used to identify independent prognostic factors. A nomogram model had been developed. Kaplan-Meier survival analysis and log-rank test were used to estimate the relationship between OS and CCR6 expression. Results: CCR6 was significantly upregulated in CM. Functional enrichment analyses revealed that CCR6 was correlated with immune response. Most immune cells and immune checkpoints were positively correlated with CCR6 expression. Kaplan-Meier analyses showed that high CCR6 expression was associated with a good outcome in CM and its subtypes. Cox regression showed that CCR6 was an independent prognostic factor in patients with CM (HR = 0.550, 95% CI = 0.332-0.912, p<0.05). Conclusions: CCR6 is considered to be a new prognostic biomarker for patients with CM, and our study provides a potential therapeutic target for CM treatment.

Evaluation of platelet-rich plasma plus basic fibroblast growth factor combined with minoxidil in the treatment of androgenetic alopecia: A randomized controlled trial.

BACKGROUND: Platelet-rich plasma plus basic fibroblast growth factor (PRPF) has been confirmed to be a safe and valuable therapy for androgenetic alopecia (AGA). However, the efficacy of PRPF combined with minoxidil treatment remains unknown. OBJECTIVE: To assess the efficacy of combined PRPF and minoxidil treatment for AGA. METHODS: In this prospective, randomized controlled trial, 75 patients with AGA were randomly divided into three groups and were administered the following treatments: Group 1, direct intradermal PRPF injection; Group 2, topical minoxidil 5% twice daily; and Group 3, PRPF injection combined with minoxidil. The PRPF injection was performed three times, 1 month apart. Hair growth parameters were evaluated using a trichoscope until the sixth month of the study. Patient satisfaction and side effects were recorded during the follow-up. RESULTS: All patients showed improvements (p < 0.05) in hair count, terminal hair, and decrease in telogen hair ratio after treatment. The efficacy of PRPF complex therapy revealed significant improvements (p < 0.05) in hair count, terminal hair and growth rate, compared with monotherapy. LIMITATIONS: Small sample size, short follow-up time and lack of quantification of GFs in PRPF. CONCLUSION: The effect of complex therapy exceed both the effects of PRPF monotherapy and minoxidil treatment, which can be a beneficial AGA treatment strategy.

Platelet lysate promotes hair growth: In vitro and in vivo mechanism and randomized, controlled trial.

BACKGROUND: Platelet lysate (PL), a novel platelet derivative, has been widely used in regenerative medicine and is a potential therapy for improving hair growth. It is necessary to fully clarify the potential mechanism and evaluate preliminary clinical effect of PL on hair growth. METHODS: We used the C57BL/6 model, organ-cultured hair follicles, and RNA-seq analysis to explore the mechanisms of PL regulating hair growth. Then, we performed a randomized, controlled, double-blind study of 107 AGA patients to verify the therapeutic efficacy of PL. RESULTS: The results confirmed that PL improved hair growth and accelerated hair cycling in mice. Organ-cultured hair follicle evaluation confirmed that PL prolonged anagen remarkably and down-regulated IL-6, C-FOS, and p-STAT5a. Clinically, diameter, hair counts, absolute anagen counts and changes from baseline in the PL group showed a significant improvement at 6 months. CONCLUSIONS: We elucidated the specific molecular mechanism of PL action on hair growth and proved equal changes in hair follicle performance after PL vs PRP in AGA patients. This study provided novel knowledge of PL, making it ideal for AGA.

Modeling human gray hair by irradiation as a valuable tool to study aspects of tissue aging.

As one of the earliest and most visible phenomenon of aging, gray hair makes it a unique model system for investigating the mechanism of aging. Ionizing radiation successfully induces gray hair in mice, and also provides a venue to establish an organ-cultured human gray hair model. To establish a suitable organ-cultured human gray HF model by IR, which imitates gray hair in the elderly, and to explore the mechanisms behind the model. By detecting growth parameters, melanotic and senescence markers of the model, we found that the model of 5 Gy accords best with features of elderly gray hair. Then, we investigated the formation mechanisms of the model by RNA-sequencing. We demonstrated that the model of organ-cultured gray HFs after 5 Gy irradiation is closest to the older gray HFs. Moreover, the 5 Gy inhibited the expression of TRP-1, Tyr, Pmel17, and MITF in hair bulbs/ORS of HFs. The 5 Gy also significantly induced ectopically pigmented melanocytes and increased the expression of DNA damage and senescence in HFs. Finally, RNA-seq analysis of the model suggested that IR resulted in cell DNA damage, and the accumulation of oxidative stress in the keratinocytes. Oxidative stress and DNA damage caused cell dysfunction and decreased melanin synthesis in the gray HFs. We found that HFs irradiated at 5 Gy successfully constructed an appropriate aging HF model. This may provide a useful model for cost-effective and predictable treatment strategies to human hair graying and the process of aging.

Relieving postoperative pain using tumescent solution with ropivacaine in follicular unit excision.

BACKGROUND: Local tumescent anesthesia relieves postoperative pain. OBJECTIVE: The objective of the study was to compare the effect of injecting a tumescent solution with/without ropivacaine on postoperative pain. METHODS: A randomized, double-blind control study was conducted in 314 patients who underwent first follicular unit excision after obtaining informed consent and ethics committee approval. The patients were randomly divided into three groups: intra-groups (group 1, injected with tumescent solution with ropivacaine; group 2, without ropivacaine) and inter-group (group 3, right-head/left-head side with/without ropivacaine). Postoperative pain was recorded using the 5-point Wong-Baker Faces Pain Scale. No preoperative analgesic was administered to any patient. The survival rate of hair follicles was measured using dermoscopy during follow-up. Data were statistically analyzed. RESULTS: Of the 314 patients included in the study, 166 were men and 148 were women with a mean age of 32.15 ± 4.58 (range, 25-45) years. Postoperative pain with ropivacaine was significantly more relieved compared with that without ropivacaine in both groups (p < 0.05). There was no significant difference between sex and survival rate of hair follicles in the intra- or inter-group. CONCLUSION: A tumescent solution with ropivacaine has proven to relieve postoperative pain and is a safe and valuable form of local anesthesia in follicular unit excision.

Emerging Role of Dermal White Adipose Tissue in Modulating Hair Follicle Development During Aging.

Hair follicle stem cells are extensively reprogrammed by the aging process, manifesting as diminished self-renewal and delayed responsiveness to activating cues, orchestrated by both intrinsic microenvironmental and extrinsic macroenvironmental regulators. Dermal white adipose tissue (dWAT) is one of the peripheral tissues directly adjacent to hair follicles (HFs) and acts as a critical macroenvironmental niche of HF. dWAT directly contributes to HF aging by paracrine signal secretion. However, the altered interrelationship between dWAT and HF with aging has not been thoroughly understood. Here, through microdissection, we separated dWAT from the skin of aged mice (18 months) and young mice (2 months) in telogen and depilation-induced anagen for transcriptome comparing. Notably, compared with young dWAT, aberrant inflammatory regulators were recapitulated in aging dWAT in telogen, including substantial overexpressed inflammatory cytokines, matrix metalloproteinases, and prostaglandin members. Nonetheless, with anagen initiation, inflammation programs were mostly abolished in aging dWAT, and instead of which, impaired collagen biosynthesis, angiogenesis, and melanin synthesis were identified. Furthermore, we confirmed the inhibitory effect on hair growth of CXCL1, one of the most significantly upregulated inflammation cytokines in aging dWAT. Besides this, we also identified the under-expressed genes related to Wnt signaling fibroblast growth factor family members and increased BMP signaling in aging dWAT, further unraveling the emerging role of dWAT in aging HFs malfunction. Finally, we proved that relieving inflammation of aging dWAT by injecting high-level veratric acid stimulated HF regenerative behavior in aged mice. Concomitantly, significantly decreased TNF-a, CCL2, IL-5, CSF2, and increased IL10 in dWAT was identified. Overall, the results elaborated on the complex physiological cycling changes of dWAT during aging, providing a basis for the potential regulatory effect of dWAT on aging HFs.

Facial Defect Repair Using a Flap Based on the Superficial Temporal Artery.

BACKGROUND: Although a local flap repair is optimal for facial defects, an extra flap or split-thickness skin graft may be needed if a sufficient local flap area is not available. In this study, we developed a distant axial pedicle flap procedure based on the inner transverse perforator of the ascending frontal branch of the superficial temporal artery to repair facial defects while meeting patients' requirements for a like-for-like tissue reconstructive outcome. METHODS: For defect repair after facial tumor removal, we designed upper frontal facial pedicle flaps based on the inner transverse perforator of the ascending frontal branch of the superficial temporal artery. Facelift procedures were applied concomitantly for donor site repair. RESULTS: We applied the procedure to 12 patients who underwent curative lesion resection. Notably, all flaps survived. Venous congestion developed in only 1 case, in which the wound was covered with heparin sodium gauze to inhibit wound coagulation until the congestion gradually resolved. In all cases, the frontal donor site scars were adjacent to the hairline and were concealed very well by hair growth. During postoperative follow-ups of 8-43 months, the patients experienced only minor complications. CONCLUSIONS: The flap based on the inner transverse perforator of the ascending frontal branch of the superficial temporal artery is a useful alternative for facial defect repair surgery. The low incidence of complications and easy concealment of the donor site underscore the safe and aesthetically acceptable nature of the procedure.

Establishment of an Efficient Primary Culture System for Human Hair Follicle Stem Cells Using the Rho-Associated Protein Kinase Inhibitor Y-27632.

BACKGROUND: Hair follicle tissue engineering is a promising strategy for treating hair loss. Human hair follicle stem cells (hHFSCs), which play a key role in the hair cycle, have potential applications in regenerative medicine. However, previous studies did not achieve efficient hHFSC expansion in vitro using feeder cells. Therefore, there is a need to develop an efficient primary culture system for the expansion and maintenance of hHFSCs. METHODS: The hHFSCs were obtained by two-step proteolytic digestion combined with microscopy. The cell culture dishes were coated with human fibronectin and inoculated with hHFSCs. The hHFSCs were harvested using a differential enrichment procedure. The effect of Rho-associated protein kinase (ROCK) inhibitor Y-27632, supplemented in keratinocyte serum-free medium (K-SFM), on adhesion, proliferation, and stemness of hHFSCs and the underlying molecular mechanisms were evaluated. RESULTS: The hHFSCs cultured in K-SFM, supplemented with Y-27632, exhibited enhanced adhesion and proliferation. Additionally, Y-27632 treatment maintained the stemness of hHFSCs and promoted the ability of hHFSCs to regenerate hair follicles in vivo. However, Y-27632-induced proliferation and stemness in hHFSCs were conditional and reversible. Furthermore, Y-27632 maintained propagation and stemness of hHFSCs through the ERK/MAPK pathway. CONCLUSION: An efficient short-term culture system for primary hHFSCs was successfully established using human fibronectin and the ROCK inhibitor Y-27632, which promoted the proliferation, maintained the stemness of hHFSCs and promoted the ability to regenerate hair follicles in vivo. The xenofree culturing method used in this study provided a large number of high-quality seed cells, which have applications in hair follicle tissue engineering and stem cell therapy.