[Bimatoprost promotes hair growth of reconstructed hair follicles in mice through activation of the Wnt/ß-catenin signaling pathway].

Objective: To investigate effect of Bimatoprost (BimP) on growth of reconstructed hair follicles in recipient nude mice. Methods: Primary epidermal and dermal cells were isolated from newborn C57BL/6J mice (1-day-old) skins, and the reconstructed hair follicles was implanted in the dorsal skin of Balb/c-nu nude mice using a silicon chamber protocol, then, the 18 nude mice were randomly divided into control group, BimP group and minoxidil group, with 6 mice in each group. After 2 weeks, topical treatment was applied to the grafted area of the nude mice by 2% minoxidil 100 µl, 0.03% BimP 100 µl and saline 100 µl, respectively, once daily for 2 weeks. At day 14 after treatment, the mice were euthanized to measure the length of dorsal hair, and the number and hair cycle of the reconstructed follicles was observed histologically. The total mRNA and proteins expression of Wnt3a, LEF1, ß-catenin and Frizzled7 were determined by qPCR and Western Blotting. The distribution and expression of ß-catenin in the reconstructed follicles was detected by immunofluorescence staining. Results: As compared to the control group, the BimP group had thicker and longer hair [(0.57±0.07) vs (0.36±0.05) cm, P<0.01], no significant difference was seen between the BimP and minoxidil group. The mRNA expression levels of Wnt3a (2.73±0.17 vs 1.00±0.14, P<0.01)、LEF1(1.71±0.12 vs 1.00±0.19, P<0.01)、ß-catenin (2.37±0.21vs 1.00±0.11, P<0.01) and Frizzled7 (2.62±0.15vs 1.00±0.18, P<0.01) were significantly increased in BimP group compared with the control group. Western Blotting showed the same results, the protein expression levels of Wnt3a (1.44±0.21vs 1.00±0.13, P<0.05)、LEF1 (1.36±0.15 vs 1.00±0.09, P<0.05)、ß-catenin (1.60±0.13 vs 1.00±0.16, P<0.01) and Frizzled7 (1.52±0.15 vs 1.00±0.21, P<0.05) in BimP group were higher than those in control group, and the difference was statistically significant. Immunofluorescence staining showed that ß-catenin was strongly expressed in hair bulb cells and sebaceous gland cells of reconstructed hair follicles in BimP group and minoxidil group, whereas barely seen in the control group. Conclusion: BimP directly promotes growth of reconstructed hair follicles in mice by activating canonical Wnt/ß-catenin signaling pathway.

[Decolorization of skin and hair-derived melanin by three ligninolytic enzymes].

Objective: To compare the decolorization efficiency of lignin peroxidase (LiP), manganese peroxidase (MnP) and laccase on eumelanin and pheomelanin, and to investigate the effect of topical administration of LiP solution on hyperpigmented guinea pigs skin induced by 308 nm excimer light. Methods: Pheomelanin-enriched specimens were prepared from human hair and cutaneous melanoma tissue using alkaline lysis method.Synthetic eumelanin was purchased from a commercial supplier.The same amount (0.02%) of melanin was incubated with the equal enzyme activity (0.2 U/ml) of ligninolytic enzymes for 3 h respectively.The absorbance at 475 nm (A(475)) in the enzyme-catalyzed solution was measured using ELISA microplate reader.The experimental hyperpigmentation model was established in the dorsal skin of brownish guinea pigs using 308 nm excimer light radiation.LiP and heat-inactivated LiP solution were topically applied at each site.Meanwhile, 3% hydroquinone and vehicle cream were used as control.The skin color (L value) was recorded using a CR-10 Minolta chromameter.Corneocytes were collected using adhesive taping method.The amount and distribution of melanin in the corneocytes and skin tissues was visualized by Fontana-Masson staining. Results: All three ligninolytic enzymes showed various degree of eumelanin and pheomelanin decolorization activity.The decolorization activity of LiP, MnP and laccase was 40%-70%, 22%-42% and 9%-21%, respectively.The similar lightening was shown in the skin treated with LiP solution and 3% hydroquinone.The amount of melanin granules in the corneocytes was 199±11 by LiP, which was less than that in untreated control (923±12) and heat-inactive control (989±13). The amount of melanin was decreased in the whole epidermis treated with hydroquinone, the epidermis thickness was increased as well. In contrast, melanin of LiP group was decreased only in the superficial epidermis, the epidermis thickness seemed to be normal. Conclusion: LiP exerts a potent decolorization activity for hair- or skin-derived pheomelanin as well as eumelanin.It remains to be further investigated whether LiP serves as a substitute for hydroquinone in skin lightening products.

[Ultrastructural changes of epidermal calcium gradient and lipid lamellar membrane in depigmented skin lesions of vitiligo patients].

OBJECTIVE: To investigate the effects of melanin disappearance in vitiliginous skin on the ultrastructure of epidermal calcium distribution and the stratum corneum (SC) lipid lamellar membranes. METHODS: Five outpatients with stable vitiligo vulgaris and 5 healthy controls were recruited from August 2014 to February 2015 at Department of Dermatology, Renmin Hospital of Wuhan University. The ultrastructural changes of lipid lamellar membranes of the skin were assessed using transmission electron microscopy (TEM) technique in combination with ruthenium tetroxide (RuO4) staining. The concentration and distribution of calcium precipitates in the epidermis were studied using calcium ion-capture cytochemistry combined with TEM. RESULTS: The multilayered lipid lamellae existed within the intercellular space of the normal SC with a characteristic alternating electron-dense and electron-lucent pattern. Expanded intercellular space, fragmentation and lamellar separation were observed in the depigmented skin lesions from vitiligo patients, the bulbous regions of lipid lamellae were filled with electron-dense amorphous materials. Large clumps of calcium precipitates were visualized in the stratum granulosum (SG) of normal skin, fine calcium precipitates and stage Ⅳ melanosomes were noted within the normal stratum basale (SB). In depigmented skin lesions of vitiligo, both the size and number of calcium precipitates in the SG were dramatically decreased. Melanosome was barely seen in the vitiligo SB. CONCLUSION: Disrupted ultrastructure of SC lamellar membranes and disappearance of calcium gradient co-exist in the skin lesion of vitiligo, indicating that melanin in epidermis may play a role in formation of epidermal calcium gradient and maintenance of structural integrity of permeability barrier.

Successful Treatment of Cryptococcal Meningitis with Amphotericin B in a Patient with Systemic Lupus Erythematosus.

Patients with systemic lupus erythematosus (SLE) have an increased susceptibility to bacterial, viral, fungal and parasitic infections. Cryptococcal infection of the central nervous system (CNS) is a rare but often fatal complication of SLE. Here, we describe a case of cryptococcal meningitis in a female patient with active SLE, who was successfully treated with amphotericin B. This case suggests that the clinical findings of SLE patients with cryptococcal meningitis are non-specific and misleading, and early use of amphotericin B has a good response.

Potent anti-tumour activity of a novel conditionally replicating adenovirus for melanoma via inhibition of migration and invasion.

BACKGROUND: Conditionally replicating adenoviruses (CRAds) represent a novel class of oncological therapeutic agents. One strategy to ensure tumour targeting is to place the essential viral genes under the control of tumour-specific promoters. Ki67 has been selected as a cancer gene therapy target, as it is expressed in most malignant cells but is barely detectable in most normal cells. This study aimed to investigate the effects of a Ki67 promoter-controlled CRAd (Ki67-ZD55-IL-24) on the proliferation and apoptosis of melanoma cells. METHODS: Melanoma cells were independently treated with Ki67-ZD55-IL-24, ZD55-IL-24, Ki67-ZD55, and ZD55-EGFP. The cytotoxic potential of each treatment was assessed using cell viability measurements. Cell migration and invasion were assayed using cell migration and invasion assays. Apoptosis was assayed using the annexin V-FITC assay, western blotting, reverse transcriptase PCR (RT-PCR), haematoxylin and eosin (H&E) staining, and the TUNEL assay. RESULTS: Our results showed that Ki67-ZD55-IL-24 had significantly enhanced anti-tumour activity as it more effectively induced apoptosis in melanoma cells than the other agents. Ki67-ZD55-IL-24 also caused the most significant inhibition of cell migration and invasion of melanoma cells. Furthermore, apoptosis was induced more effectively in melanoma xenografts in nude mice. CONCLUSIONS: This strategy holds promising potential for the further development of an effective approach to treat malignant melanoma.

Photoprotection of bacterial-derived melanin against ultraviolet A-induced cell death and its potential application as an active sunscreen.

BACKGROUND: The increase in the incidence of non-melanoma skin tumours, photoaging, and immunosuppression demand for more effective sunscreen on ultraviolet A (UVA) irradiation. OBJECTIVES: The aim of the study is to evaluate the photoprotective effects of a bacterial-derived melanin against UVA-induced damages in vitro and in vivo. Methods Human fibroblasts were used to assess the role of the bacterial-derived melanin on cell viability against UVA. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and nuclear morphology were employed to evaluate the photoprotection at the cellular level. Fluorometric assays were performed to detect the formation of reactive oxygen species (ROS) in the cells. Evaluations of the bacterial-derived melanin as a sunscreen were measured by transmission test and persistent pigment darkening on human skin. RESULTS: Bacterial-derived melanin efficiently scavenged ROS in the fibroblasts after UVA irradiation. The cell viability of xeroderma pigmentosum (XP) fibroblast treated with varied doses of melanin increased dramatically in comparison with untreated control and the treated XP fibroblasts became more resistant to UVA-induced apoptosis than normal fibroblasts. Although the relative transmission didn't change too much with different concentration of bacterial-derived melanin, this melanin could keep UVA-irradiated skin from pigment darkening and act as an active sunscreen on skin. CONCLUSIONS: The bacterial-derived melanin provided significant protection to fibroblast cell and human skin against the UVA radiation. It has the potential to be developed as an active sunscreen for the patients with photosensitivity skin to sun exposure.

Effects of all-trans retinoic acid on melanogenesis in pigmented skin equivalents and monolayer culture of melanocytes.

The effects of all-trans retinoic acid (RA) on melanogenesis and the mechanism of its action in topical treatment have not been elucidated. The purpose of this study was to determine the effects of RA on melanogenesis in the pigmented skin equivalent as well as in monolayer culture of melanocytes, and to determine whether RA, hydroquinone (HQ), and hydrocortisone (HC) show synergistic depigmenting effects in combined treatments of each other. The suppressing effect of RA on melanogenesis was not observed in pigmented skin equivalents and monolayer culture of murine and human melanocytes, although HQ showed strong inhibition of melanogenesis. The synergistic effects between RA, HQ, and HC were not particularly seen. The results suggested that RA neither has direct inhibitory effects on melanogenesis of melanocytes, nor influences the cell-cell interactions between melanocytes, keratinocytes and fibroblasts, such as paracrine actions with regard to melanin production. The role of RA in bleaching treatments appears to be in other specific actions, such as promotion of keratinocytes proliferation and acceleration of epidermal turnover.