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1.
Zhonghua Bing Li Xue Za Zhi ; 53(6): 570-577, 2024 Jun 08.
Article in Chinese | MEDLINE | ID: mdl-38825902

ABSTRACT

Objective: To seek the optimal melanin-removal method for hematoxylin and eosin (HE) staining, immunohistochemistry and molecular detection. Methods: Thirty-eight paraffin tissue samples of malignant melanoma diagnosed at the Fujian Cancer Hospital, Fuzhou, China between January 2018 and March 2022 were collected and used to make a tissue microarray. Melanin in these cases was removed using warm hydrogen peroxide, double oxidation depigmentation, modified potassium permanganate-oxalic acid or trichloroisocyanuric acid, followed by HE staining. The cases were divided into two cohorts: one was subject to the one of the above four methods to remove melanin first, followed by immunohistochemistry (SOX-10, Ki-67, HMB45 and Melan A), while the other was subject to immunohistochemical staining first and then a melanin removal. Following that, seventeen melanin-rich paraffin tissue samples were collected and depigmented using the methods described above. DNA extraction was then done, followed by assessments of DNA content and quality. Moreover, the completeness of melanin removal, the effect on HE and immunohistochemical staining, and the quality of DNA were compared between the depigmented methods. Results: Regarding the effectiveness of melanin removal, the modified potassium permanganate-oxalic acid and the warm hydrogen peroxide methods were the most effective, and both showed residual melanin in only 5.26% (2/38) of the cases. The trichloroisocyanuric acid method showed residual melanin in 10.53% (4/38) of the cases. The worst was the double oxidation depigmentation method, which showed pigment residue in 15.79% (6/38) of the cases. For HE staining, the percentage of good staining with the warm hydrogen peroxide method was 92.11%, higher than the other three methods. For immunohistochemical staining, the mean staining scores of immunohistochemistry first followed by melanin removal with modified potassium permanganate-oxalic acid, double oxidation and trichloroisocyanuric acid were 20.84, 26.63 and 35.02, respectively. These immunohistochemical staining scores were higher than those of melanin removal first followed by immunohistochemistry (8.70, 15.41 and 21.22, respectively). The mean staining score of melanin removal by warm hydrogen peroxide method followed by immunohistochemistry was 33.57, superior to that of immunohistochemistry followed by the melanin removal (19.96). Moreover, the staining scores of HMB45, MelanA and Ki-67 with immunohistochemical staining followed by trichloroisocyanuric acid method were 36.45, 33.79, and 36.24, respectively, while the staining score of SOX10 with melanin removal by warm hydrogen peroxide followed by immunohistochemistry was 34.39. The DNA was significantly degraded by modified potassium permanganate-oxalic acid, double oxidation depigmentation and trichloroisocyanuric acid, whereas the mean concentration of DNA extracted after melanin removal by hydrogen peroxide method was 59.59 µg/L, substantially higher than that of DNA extracted without melanin removal (30.3 µg/L, P=0.001). The A260/A280 of DNA extracted after melanin removal by hydrogen peroxide was between 1.8 and 2.0 in all cases, and the A260/A230 was above 2.0 in sixteen cases, suggesting high purity of DNA. However, the DNA extracted without removing the melanin showed poor purity, with A260/A280 below 1.8 in eight cases and A260/A230 below 2.0 in sixteen cases. Conclusions: Warm hydrogen peroxide showed the least melanin residue, superior HE staining and a minimal effect on DNA purity/quality compared to the other three methods. It thus appears most suitable for PCR, NGS and other molecular detection. Melanin removal with trichloroisocyanuric acid after immunohistochemical staining has the least melanin residual, and thus could be the most convenient and efficient. However, it is noted that the efficacy of the same depigmentation method varies with different antibodies. Therefore, the optimal depigmentation method should be selected based on the specific markers of interest.


Subject(s)
Hydrogen Peroxide , Immunohistochemistry , Melanins , Potassium Permanganate , Staining and Labeling , Humans , Melanins/metabolism , Staining and Labeling/methods , Melanoma/metabolism , Melanoma/pathology , Skin Neoplasms/metabolism , Skin Neoplasms/pathology
3.
Exp Dermatol ; 33(5): e15101, 2024 May.
Article in English | MEDLINE | ID: mdl-38770555

ABSTRACT

Skin hyperpigmentation is mainly caused by excessive synthesis of melanin; however, there is still no safe and effective therapy for its removal. Here, we found that the dermal freezer was able to improve UVB-induced hyperpigmentation of guinea pigs without causing obvious epidermal damage. We also mimic freezing stimulation at the cellular level by rapid freezing and observed that freezing treatments <2.5 min could not decrease cell viability or induce cell apoptosis in B16F10 and Melan-A cells. Critically, melanin content and tyrosinase activity in two cells were greatly reduced after freezing treatments. The dramatic decrease in tyrosinase activity was associated with the downregulation of MITF, TYR, TRP-1 and TRP-2 protein expression in response to freezing treatments for two cells. Furthermore, our results first demonstrated that freezing treatments significantly reduced the levels of p-GSK3ß and ß-catenin and the nuclear accumulation of ß-catenin in B16F10 and Melan-A cells. Together, these data suggest that fast freezing treatments can inhibit melanogenesis-related gene expression in melanocytes by regulating the Wnt/ß-catenin signalling pathway. The inhibition of melanin production eventually contributed to the improvement in skin hyperpigmentation induced by UVB. Therefore, fast freezing treatments may be a new alternative of skin whitening in the clinic in the future.


Subject(s)
Freezing , Hyperpigmentation , Melanins , Melanocytes , Monophenol Monooxygenase , Ultraviolet Rays , Wnt Signaling Pathway , beta Catenin , Animals , Melanins/biosynthesis , Melanins/metabolism , Melanocytes/metabolism , Mice , Hyperpigmentation/metabolism , beta Catenin/metabolism , Monophenol Monooxygenase/metabolism , Guinea Pigs , Microphthalmia-Associated Transcription Factor/metabolism , Cell Survival , Intramolecular Oxidoreductases/metabolism , Glycogen Synthase Kinase 3 beta/metabolism , Apoptosis , Oxidoreductases/metabolism , Interferon Type I , Pregnancy Proteins
4.
Toxicol In Vitro ; 98: 105844, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38740103

ABSTRACT

Streptomycin (STR) is an aminoglycoside antibiotic with a broad-spectrum of activity and ototoxic potential. The mechanism of STR-induced inner ear damage has not been fully elucidated. It was previously found that STR binds to melanin, which may result in the accumulation of the drug in melanin-containing tissues. Melanin pigment is present in various parts of the inner ear, including the cochlea and vestibular organ. The present study aimed to assess if streptomycin generates oxidative stress and affects melanogenesis in normal human melanocytes. Moreover the variation of free radical concentration in STR-treated melanocytes was examined by electron paramagnetic resonance spectroscopy (EPR). We found that STR decreases cell metabolic activity and reduces melanin content. The observed changes in the activity of antioxidant enzymes activity in HEMn-DPs treated with streptomycin may suggest that the drug affects redox homeostasis in melanocytes. In this work EPR study expanded knowledge about free radicals in interactions of STR and melanin in melanocytes. The results may help elucidate the mechanisms of STR toxicity on pigment cells, including melanin-producing cells in the inner ear. This is important because understanding the mechanism of STR-induced ototoxicity would be helpful in developing new therapeutic strategies to protect patients' hearing.


Subject(s)
Anti-Bacterial Agents , Melanins , Melanocytes , Oxidative Stress , Streptomycin , Melanins/metabolism , Humans , Electron Spin Resonance Spectroscopy , Oxidative Stress/drug effects , Melanocytes/drug effects , Melanocytes/metabolism , Streptomycin/toxicity , Anti-Bacterial Agents/toxicity , Cells, Cultured , Cell Survival/drug effects , Free Radicals/metabolism , Cell Line
5.
Front Cell Infect Microbiol ; 14: 1369301, 2024.
Article in English | MEDLINE | ID: mdl-38774630

ABSTRACT

Dual-specificity LAMMER kinases are highly evolutionarily conserved in eukaryotes and play pivotal roles in diverse physiological processes, such as growth, differentiation, and stress responses. Although the functions of LAMMER kinase in fungal pathogens in pathogenicity and stress responses have been characterized, its role in Cryptococcus neoformans, a human fungal pathogen and a model yeast of basidiomycetes, remains elusive. In this study, we identified a LKH1 homologous gene and constructed a strain with a deleted LKH1 and a complemented strain. Similar to other fungi, the lkh1Δ mutant showed intrinsic growth defects. We observed that C. neoformans Lkh1 was involved in diverse stress responses, including oxidative stress and cell wall stress. Particularly, Lkh1 regulates DNA damage responses in Rad53-dependent and -independent manners. Furthermore, the absence of LKH1 reduced basidiospore formation. Our observations indicate that Lkh1 becomes hyperphosphorylated upon treatment with rapamycin, a TOR protein inhibitor. Notably, LKH1 deletion led to defects in melanin synthesis and capsule formation. Furthermore, we found that the deletion of LKH1 led to the avirulence of C. neoformans in a systemic cryptococcosis murine model. Taken together, Lkh1 is required for the stress response, sexual differentiation, and virulence of C. neoformans.


Subject(s)
Cryptococcosis , Cryptococcus neoformans , Fungal Proteins , Virulence , Animals , Female , Humans , Mice , Cell Wall/metabolism , Cryptococcosis/microbiology , Cryptococcus neoformans/pathogenicity , Cryptococcus neoformans/genetics , Cryptococcus neoformans/enzymology , Disease Models, Animal , DNA Damage , Fungal Capsules/metabolism , Fungal Capsules/genetics , Fungal Proteins/genetics , Fungal Proteins/metabolism , Gene Deletion , Gene Expression Regulation, Fungal , Melanins/metabolism , Mice, Inbred BALB C , Oxidative Stress , Phosphorylation , Sirolimus/pharmacology , Spores, Fungal/growth & development , Stress, Physiological
6.
Front Biosci (Landmark Ed) ; 29(5): 194, 2024 May 20.
Article in English | MEDLINE | ID: mdl-38812330

ABSTRACT

BACKGROUNDS: Melanogenesis, regulated by genetic, hormonal, and environmental factors, occurs in melanocytes in the basal layer of the epidermis. Dysregulation of this process can lead to various skin disorders, such as hyperpigmentation and hypopigmentation. Therefore, the present study investigated the effect of ultrasonic-assisted ethanol extract (SHUE) from Sargassum horneri (S. horneri), brown seaweed against melanogenesis in α-melanocyte-stimulating hormone (MSH)-stimulated B16F10 murine melanocytes. METHODS: Firstly, yield and proximate compositional analysis of the samples were conducted. The effect of SHUE on cell viability has been evaluated by using 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. After that, the melanin content and cellular tyrosinase activity in α-MSH-stimulated B16F10 murine melanocytes were examined. Western blot analysis was carried out to investigate the protein expression levels of microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein-1 (TRP1), and tyrosinase-related protein-2 (TRP2). In addition, the effect of extracellular signal-regulated kinase (ERK) on the melanogenesis process was assessed via Western blotting. RESULTS: As per the analysis, SHUE contained the highest average yield on a dry basis at 28.70 ± 3.21%. The findings showed that SHUE reduced the melanin content and cellular tyrosinase activity in α-MSH-stimulated B16F10 murine melanocytes. Additionally, the expression levels of MITF, TRP1, and TRP2 protein were significantly downregulated by SHUE treatment in α-MSH-stimulated B16F10 murine melanocytes. Moreover, SHUE upregulated the phosphorylation of ERK and AKT in α-MSH-stimulated B16F10 murine melanocytes. In addition, experiments conducted using the ERK inhibitor (PD98059) revealed that the activity of SHUE depends on the ERK signaling cascade. CONCLUSION: These results suggest that SHUE has an anti-melanogenic effect and can be used as a material in the formulation of cosmetics related to whitening and lightening.


Subject(s)
Ethanol , Melanins , Melanocytes , Monophenol Monooxygenase , Sargassum , Animals , Sargassum/chemistry , Melanins/biosynthesis , Melanins/metabolism , Monophenol Monooxygenase/metabolism , Monophenol Monooxygenase/antagonists & inhibitors , Melanocytes/drug effects , Melanocytes/metabolism , Mice , Ethanol/chemistry , Microphthalmia-Associated Transcription Factor/metabolism , alpha-MSH/metabolism , Plant Extracts/pharmacology , Plant Extracts/chemistry , Cell Survival/drug effects , Melanoma, Experimental/metabolism , Cell Line, Tumor , Intramolecular Oxidoreductases/metabolism
7.
PLoS One ; 19(5): e0298502, 2024.
Article in English | MEDLINE | ID: mdl-38814922

ABSTRACT

The skin microbiome maintains healthy human skin, and disruption of the microbiome balance leads to inflammatory skin diseases such as folliculitis and atopic dermatitis. Staphylococcus aureus and Cutibacterium acnes are pathogenic bacteria that simultaneously inhabit the skin and cause inflammatory diseases of the skin through the activation of innate immune responses. Silkworms are useful invertebrate animal models for evaluating innate immune responses. In silkworms, phenoloxidase generates melanin as an indicator of innate immune activation upon the recognition of bacterial or fungal components. We hypothesized that S. aureus and C. acnes interact to increase the innate immunity-activating properties of S. aureus. In the present study, we showed that acidification is involved in the activation of silkworm hemolymph melanization by S. aureus. Autoclaved-killed S. aureus (S. aureus [AC]) alone does not greatly activate silkworm hemolymph melanization. On the other hand, applying S. aureus [AC] treated with C. acnes culture supernatant increased the silkworm hemolymph melanization. Adding C. acnes culture supernatant to the medium decreased the pH. S. aureus [AC] treated with propionic acid, acetic acid, or lactic acid induced higher silkworm hemolymph melanization activity than untreated S. aureus [AC]. S. aureus [AC] treated with hydrochloric acid also induced silkworm hemolymph melanization. The silkworm hemolymph melanization activity of S. aureus [AC] treated with hydrochloric acid was inhibited by protease treatment of S. aureus [AC]. These results suggest that acid treatment of S. aureus induces innate immune activation in silkworms and that S. aureus proteins are involved in the induction of innate immunity in silkworms.


Subject(s)
Bombyx , Hemolymph , Melanins , Staphylococcus aureus , Animals , Hemolymph/metabolism , Hemolymph/microbiology , Hemolymph/immunology , Bombyx/microbiology , Bombyx/immunology , Staphylococcus aureus/immunology , Melanins/metabolism , Immunity, Innate , Hydrogen-Ion Concentration , Monophenol Monooxygenase/metabolism
8.
Pigment Cell Melanoma Res ; 37(4): 514-529, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38705722

ABSTRACT

Epidermal melanin unit integrity is crucial for skin homeostasis and pigmentation. Epidermal growth factor (EGF) receptor (EGFR) is a pivotal player in cell growth, wound healing, and maintaining skin homeostasis. However, its influence on skin pigmentation is relatively unexplored. This study investigates the impact and underlying mechanisms of EGFR inhibitors on skin pigmentation. We evaluated EGF and EGFR expression in various skin cells using quantitative real-time PCR, Western blot, and immunofluorescence. EGF and EGFR were predominantly expressed in epidermal keratinocytes, and treatment with the EGFR tyrosine kinase inhibitors (EGFR-TKIs) gefitinib and PD153035 significantly increased stem cell factor (SCF) and endothelin-1 (ET-1) expression in cultured keratinocytes. Enhanced melanocyte migration and proliferation were observed in co-culture, as evidenced by time-lapse live imaging and single-cell tracking assays. Furthermore, topical application of gefitinib to guinea pig dorsal skin induced increased pigmentation and demonstrated efficacy in mitigating rhododendrol-induced leukoderma. Suppression of EGF signaling indirectly enhanced skin pigmentation by upregulating SCF and ET-1 in epidermal keratinocytes. This novel mechanism highlights the pivotal role of EGF signaling in regulating skin pigmentation, and topical EGFR-TKI therapy at an appropriate dose may be a promising approach for depigmentation disorder management.


Subject(s)
ErbB Receptors , Gefitinib , Hypopigmentation , Keratinocytes , Melanins , Melanocytes , Protein Kinase Inhibitors , ErbB Receptors/metabolism , Animals , Melanins/metabolism , Melanins/biosynthesis , Humans , Protein Kinase Inhibitors/pharmacology , Melanocytes/drug effects , Melanocytes/metabolism , Melanocytes/pathology , Keratinocytes/drug effects , Keratinocytes/metabolism , Keratinocytes/pathology , Hypopigmentation/pathology , Hypopigmentation/drug therapy , Gefitinib/pharmacology , Guinea Pigs , Skin Pigmentation/drug effects , Stem Cell Factor/metabolism , Epidermis/drug effects , Epidermis/pathology , Epidermis/metabolism , Cell Proliferation/drug effects , Cell Movement/drug effects , Endothelin-1/metabolism , Epidermal Growth Factor/metabolism , Epidermal Growth Factor/pharmacology , Quinazolines
9.
Chem Rev ; 124(11): 7165-7213, 2024 Jun 12.
Article in English | MEDLINE | ID: mdl-38758918

ABSTRACT

Melanin, especially integumentary melanin, interacts in numerous ways with electromagnetic radiation, leading to a set of critical functions, including radiation protection, UV-protection, pigmentary and structural color productions, and thermoregulation. By harnessing these functions, melanin and melanin-like materials can be widely applied to diverse applications with extraordinary performance. Here we provide a unified overview of the melanin family (all melanin and melanin-like materials) and their interactions with the complete electromagnetic radiation spectrum (X-ray, Gamma-ray, UV, visible, near-infrared), which until now has been absent from the literature and is needed to establish a solid fundamental base to facilitate their future investigation and development. We begin by discussing the chemistries and morphologies of both natural and artificial melanin, then the fundamentals of melanin-radiation interactions, and finally the exciting new developments in high-performance melanin-based functional materials that exploit these interactions. This Review provides both a comprehensive overview and a discussion of future perspectives for each subfield of melanin that will help direct the future development of melanin from both fundamental and applied perspectives.


Subject(s)
Electromagnetic Radiation , Melanins , Melanins/chemistry , Melanins/metabolism , Humans , Animals
10.
Chemosphere ; 359: 142353, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38761828

ABSTRACT

Microorganisms in the waterline zone can secrete pigments to avoid damage caused by ultraviolet radiation, some of which have corrosive effects. In this work, we found that the secretion of pyomelanin by P3 strain of Pseudoalteromonas lipolytica significantly increases under strong lighting conditions, accelerating the corrosion of the material. Molecular mechanisms indicate that strong light, as a stressful environmental factor, enhances the expression of melanin secretion-related genes to prevent bacteria from being damaged by ultraviolet radiation. Therefore, this work proposes a new corrosion mechanism in the waterline zone, pigment-producing microorganisms are also involved in the waterline corrosion process.


Subject(s)
Alloys , Melanins , Steel , Corrosion , Steel/chemistry , Melanins/metabolism , Alloys/chemistry , Pseudoalteromonas/metabolism , Ultraviolet Rays , Light
11.
J Ethnopharmacol ; 332: 118348, 2024 Oct 05.
Article in English | MEDLINE | ID: mdl-38762211

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: Tokishakuyakusan (TSS), a traditional Kampo medicine, can effectively alleviate symptoms unique to women, such as menstrual pain and menopausal symptoms, and this effect is believed to be related to its ability to increase the secretion of female hormones. TSS is also believed to be effective against skin pigmentation. However, no studies have examined the effect of TSS on pigmentation. AIM OF THE STUDY: In this study, we conducted basic research to determine the effects of TSS on pigmentation. MATERIALS AND METHODS: Female HRM-2 mice were given free access to a normal diet or a TSS-containing diet for 7 weeks. For 3 weeks starting from the 4th week of treatment, the back of the skin was irradiated with ultraviolet (UV) light, and the melanin level was measured. The expression levels of melanogenesis-related genes and inflammatory markers in the skin were analyzed. RESULTS: The melanin level in the skin of the mice exposed to UV radiation was approximately three times greater than that in the skin of the mice in the non-UV-irradiated group, confirming pigmentation due to UV irradiation. The protein expression levels of tyrosinase (Tyr), tyrosinase-related protein-1 (Tyrp1), and dopachrome tautomerase (Dct), which are important for melanin production, were significantly greater in the UV irradiation group than in the non-UV irradiation group. In contrast, the amount of skin melanin in the mice treated with TSS was significantly lower than that in the UV-irradiated group, and the expression levels of melanogenesis-related enzymes were also lower. Furthermore, TSS significantly decreased the expression of microphthalmia transcription factor (Mitf), a transcription factor for melanogenesis-related enzymes, and the inflammatory cytokines interleukin-1ß and interleukin-6. CONCLUSIONS: TSS inhibits melanin production in melanocytes by suppressing the increase in the expression of melanogenesis-related enzymes caused by UV irradiation. These findings suggested that this effect of TSS is exerted through the combined regulation of MITF expression and anti-inflammatory responses.


Subject(s)
Drugs, Chinese Herbal , Melanins , Monophenol Monooxygenase , Skin Pigmentation , Ultraviolet Rays , Animals , Ultraviolet Rays/adverse effects , Melanins/biosynthesis , Melanins/metabolism , Skin Pigmentation/drug effects , Skin Pigmentation/radiation effects , Female , Mice , Monophenol Monooxygenase/metabolism , Drugs, Chinese Herbal/pharmacology , Skin/drug effects , Skin/radiation effects , Skin/metabolism , Microphthalmia-Associated Transcription Factor/metabolism , Microphthalmia-Associated Transcription Factor/genetics , Medicine, Kampo , Intramolecular Oxidoreductases/metabolism , Intramolecular Oxidoreductases/genetics , Mice, Hairless , Melanogenesis , Membrane Glycoproteins , Oxidoreductases
12.
J Ethnopharmacol ; 332: 118374, 2024 Oct 05.
Article in English | MEDLINE | ID: mdl-38789093

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: Picrasma quassioides (D. Don) Benn is a vascular plant belonging to the genus Picrasma of Simaroubaceae family and grows in Korea, China, India, Taiwan, and Japan. Picrasma quassioides extract has been reported to have anti-inflammatory, anti-bacterial, and anti-cancer properties. Moreover, this plant has been also traditionally used to alleviate symptoms of eczema, atopic dermatitis, psoriasis, scabies, and boils in skin. AIM OF THE STUDY: The Pq-EE has been reported in Chinese pharmacopoeia for its pharmacological effects on skin. However, the detailed mechanism on alleviating skin conditions is not understood. Hence, we investigated the skin improvement potential of Pq-EE against skin damage. MATERIALS AND METHODS: We used the human keratinocyte cell line (HaCaT) and mouse melanoma cell line (B16F10) to study the effects of Pq-EE on the epidermis. Additionally, in vitro antioxidant assays were performed using a solution that included either metal ions or free radicals. RESULTS: In colorimetric antioxidant assays, Pq-EE inhibited free radicals in a dose-dependent manner. The Pq-EE did not affect cell viability and promoted cell survival under UVB exposure conditions in the MTT assay. The Pq-EE downregulated the mRNA levels of apoptotic factors. Moreover, MMP1 and inflammatory cytokine iNOS mRNA levels decreased with Pq-EE treatment. With regard to protein levels, caspases and cleaved caspases were more powerfully inhibited by Pq-EE than UVB-irritated conditions. p53 and Bax also decreased with Pq-EE treatment. The melanin contents and secretion were decreased at nontoxic concentrations of Pq-EE. The pigmentation pathway genes also were inhibited by treatment with Pq-EE. CONCLUSIONS: In summary, we suggest the cell protective potential of Pq-EE against UVB and ROS, indicating its use in UV-protective cosmeceutical materials.


Subject(s)
Anti-Inflammatory Agents , Antioxidants , Apoptosis , Melanins , Picrasma , Plant Extracts , Animals , Anti-Inflammatory Agents/pharmacology , Anti-Inflammatory Agents/isolation & purification , Plant Extracts/pharmacology , Plant Extracts/chemistry , Apoptosis/drug effects , Humans , Mice , Picrasma/chemistry , Antioxidants/pharmacology , Melanins/metabolism , Ethanol/chemistry , HaCaT Cells , Keratinocytes/drug effects , Keratinocytes/radiation effects , Cell Line, Tumor , Melanoma, Experimental/drug therapy , Cell Survival/drug effects , Nitric Oxide Synthase Type II/metabolism , Nitric Oxide Synthase Type II/genetics
13.
Colloids Surf B Biointerfaces ; 239: 113937, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38749166

ABSTRACT

Osteosarcoma conventional chemotherapeutics are known for their side effects, limited options, and induction of drug resistance. This creates the need to develop new therapeutics capable of effectively destroying cancer cells with low toxicity, improving patient survival rate and their life quality. This work reports a novel drug delivery nanoplataform made of Natural Melanin Nanoparticles (MNPs), obtained from Sepia officinalis ink, with 99% incorporation efficiency of doxorubicin (Dox) without the use of non-toxic solvents. A significant photothermal effect was shown by a 36ºC increment after 10 min of laser irradiation, surpassing reported values for synthetic melanin. A sustained drug release of ca. 23% with photothermal stimuli was observed, compared to 15% without stimuli, after 48 h. This nanoplatform is obtained as a food industry side product, which makes it a natural cost-effective biomedical material. Natural MPs were applied in an osteosarcoma cell line (SaOs-2), and internalized by the cells in less than 2 h, showing cytocompatibility up to 1000 µg/mL after 72 h of contact with cells. On the contrary, when natural MNPs loaded with Dox (Dox-MNPs) were placed in contact with the SaOs-2 cells and were simultaneously receiving NIR light it was observed a 93% reduction in cancer cells in 48 h, revealing a synergistic effect between chemotherapy and phototherapy. To our knowledge this is the first time that natural MNPs extracted from Sepia officinalis were tested on an osteosarcoma cell line as chemo-photothermal agent, showing these NPs are an effective, cost-effective, reproducible, non-toxic nanoplatform for osteosarcoma treatment using combined effects.


Subject(s)
Cell Survival , Doxorubicin , Melanins , Nanoparticles , Osteosarcoma , Sepia , Humans , Osteosarcoma/drug therapy , Osteosarcoma/pathology , Doxorubicin/pharmacology , Doxorubicin/chemistry , Melanins/metabolism , Nanoparticles/chemistry , Sepia/chemistry , Cell Survival/drug effects , Cell Line, Tumor , Drug Liberation , Antibiotics, Antineoplastic/pharmacology , Antibiotics, Antineoplastic/chemistry , Bone Neoplasms/drug therapy , Bone Neoplasms/pathology , Particle Size , Cost-Benefit Analysis , Drug Screening Assays, Antitumor
14.
Methods Mol Biol ; 2775: 257-268, 2024.
Article in English | MEDLINE | ID: mdl-38758323

ABSTRACT

Melanin is a complex dark pigment synthetized by the phenoloxidase enzyme laccase in Cryptococcus neoformans. In vitro, this enzyme oxidizes exogenous catecholamines to produce melanin that may be secreted or incorporated into the fungal cell wall. This pigment has multiple roles in C. neoformans virulence during its interaction with different hosts and probably also in protecting fungal cells in the environment against predation and oxidative and radiation stresses, among others. However, it is important to note that laccase also has melanin-independent roles in C. neoformans interactions with host cells. In this chapter, we describe a quantitative laccase assay and a method for evaluating the kinetics of melanin production in C. neoformans colonies.


Subject(s)
Cryptococcus neoformans , Laccase , Melanins , Cryptococcus neoformans/metabolism , Cryptococcus neoformans/enzymology , Laccase/metabolism , Melanins/biosynthesis , Melanins/metabolism , Enzyme Assays/methods
15.
J Photochem Photobiol B ; 255: 112925, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38703451

ABSTRACT

Visible light triggers free radical production in alive and intact Drosophila melanogaster. We exposed fruit flies to red (613-631 nm), green (515-535 nm), and blue (455-475 nm) light while we monitored changes in unpaired electron content with an electron spin resonance spectrometer (ESR/EPR). The immediate response to light is a rapid increase in spin content lasting approximately 10 s followed by a slower, linear increase for approximately 170 s. When the light is turned off, the spin population promptly decays with a similar time course, though never fully returning to baseline. The magnitude and time course of the spin production depends on the wavelength of the light. Initially, we surmised that eumelanin might be responsible for the spin change because of its documented ability for visible light absorption and its highly stable free radical content. To explore this, we utilized different fruit fly strains with varying eumelanin content and clarified the relation of melanin types with the spin response. Our findings revealed that flies with darker cuticle have at least three-fold more unpaired electrons than flies with yellow cuticle. However, to our surprise, the increase in unpaired electron population by light was not drastically different amongst the genotypes. This suggests that light-induced free radical production may not exclusively rely on the presence of black melanin, but may instead be dependent on light effects on quinone-based cuticular polymers.


Subject(s)
Drosophila melanogaster , Light , Melanins , Animals , Free Radicals/chemistry , Drosophila melanogaster/metabolism , Electron Spin Resonance Spectroscopy , Melanins/chemistry , Melanins/metabolism , Melanins/biosynthesis
16.
Exp Dermatol ; 33(5): e15094, 2024 May.
Article in English | MEDLINE | ID: mdl-38742793

ABSTRACT

Melasma is a common condition of hyperpigmented facial skin. Picosecond lasers are reported to be effective for the treatment of melasma. We aimed to identify the most effective therapeutic mode and elucidate the potential molecular mechanisms of picosecond lasers for the treatment of melasma. Female Kunming mice with melasma-like conditions were treated using four different picosecond laser modes. Concurrently, in vitro experiments were conducted to assess changes in melanin and autophagy in mouse melanoma B16-F10 cells treated with these laser modes. Changes in melanin in mouse skin were detected via Fontana-Masson staining, and melanin particles were evaluated in B16-F10 cells. Real-time polymerase chain reaction and western blotting were used to analyse the expression levels of melanosome and autophagy-related messenger ribonucleic acid (mRNA) and proteins. A combination of large-spot low-fluence 1064-nm and fractional 1064-nm picosecond lasers resulted insignificant decreases in melanin as well as in mRNA and protein expression of melanin-synthesizing enzymes (TYR, TRP-1 and MITF). This combination also led to increased expression of the autophagy-related proteins, Beclin1 and ATG5, with a marked decrease in p62 expression. Intervention with the PI3K activator, 740 Y-P, increased TYR, TRP-1, MITF, p-PI3K, p-AKT, p-mTOR and p62 expression but decreased the expression of LC3, ATG5 and Beclin1. A combination of large-spot low-fluence 1064-nm and fractional 1064-nm picosecond lasers proved more effective and safer. It inhibits melanin production, downregulates the PI3K/AKT/mTOR pathway, enhances melanocyte autophagy and accelerates melanin metabolism, thereby reducing melanin content.


Subject(s)
Autophagy , Melanins , Melanosis , Melanosomes , Phosphatidylinositol 3-Kinases , Proto-Oncogene Proteins c-akt , Signal Transduction , TOR Serine-Threonine Kinases , Animals , Melanosis/metabolism , TOR Serine-Threonine Kinases/metabolism , Female , Mice , Proto-Oncogene Proteins c-akt/metabolism , Melanins/metabolism , Melanosomes/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Low-Level Light Therapy , Autophagy-Related Protein 5/metabolism , Autophagy-Related Protein 5/genetics , Melanoma, Experimental/metabolism , Melanoma, Experimental/radiotherapy
17.
Exp Dermatol ; 33(5): e15093, 2024 May.
Article in English | MEDLINE | ID: mdl-38742821

ABSTRACT

Senile skin hyperpigmentation displays remarkable histopathological features of dermal aging. The crosstalk between melanocytes and dermal fibroblasts plays crucial roles in aging-related pigmentation. While senescent fibroblasts can upregulate pro-melanogenic factors, the role of anti-melanogenic factors, such as dickkopf1 (DKK1), and the upstream regulatory mechanism during aging remain obscure. This study investigated the roles of yes-associated protein (YAP) and DKK1 in the regulation of dermal fibroblast senescence and melanogenesis. Our findings demonstrated decreased YAP activity and DKK1 levels in intrinsic and extrinsic senescent fibroblasts. YAP depletion induced fibroblast senescence and downregulated the expression and secretion of DKK1, whereas YAP overexpression partially reversed the effect. The transcriptional regulation of DKK1 by YAP was supported by dual-luciferase reporter and chromatin immunoprecipitation assays. Moreover, YAP depletion in fibroblasts upregulated Wnt/ß-catenin in melanocytes and stimulated melanogenesis, which was partially rescued by the re-supplementation of DKK1. Conversely, overexpression of YAP in senescent fibroblasts decreased Wnt/ß-catenin levels in melanocytes and inhibited melanogenesis. Additionally, reduced levels of YAP and DKK1 were verified in the dermis of solar lentigines. These findings suggest that, during skin aging, epidermal pigmentation may be influenced by YAP in the dermal microenvironment via the paracrine effect of DKK1.


Subject(s)
Adaptor Proteins, Signal Transducing , Cellular Senescence , Fibroblasts , Intercellular Signaling Peptides and Proteins , Melanins , Melanocytes , Paracrine Communication , Skin Aging , Transcription Factors , YAP-Signaling Proteins , Fibroblasts/metabolism , Intercellular Signaling Peptides and Proteins/metabolism , Humans , Melanocytes/metabolism , YAP-Signaling Proteins/metabolism , Transcription Factors/metabolism , Transcription Factors/genetics , Adaptor Proteins, Signal Transducing/metabolism , Melanins/metabolism , Melanins/biosynthesis , Wnt Signaling Pathway , Dermis/cytology , Cells, Cultured , Melanogenesis
18.
Lipids Health Dis ; 23(1): 138, 2024 May 11.
Article in English | MEDLINE | ID: mdl-38734619

ABSTRACT

BACKGROUND: Skin barrier alterations play a crucial function in melasma development. Past researches have demonstrated variations in lipid content between the epidermis of melasma lesions and normal tissues, along with the varied expression of lipid-related genes in melasma. This study aimed to analyze the lipidome profiles of skin surface lipids (SSL) in patients with melasma before and after treatment to understand associated abnormalities. METHODS: Melasma was treated with tranexamic acid orally and hydroquinone cream topically. Disease was assessed using the Melasma Area and Severity Index (MASI), and the impact to life was evaluated with Melasma Quality of Life (MELASQoL) score. Epidermal melanin particles were observed using reflection confocal microscopy (RCM), whereas epidermal pigment and blood vessel morphology were observed using dermoscopy, and SSL samples were collected. Specific information regarding alterations in lipid composition was obtained through multivariate analysis of the liquid chromatography-mass spectrometry data. RESULTS: After treatment, patients with melasma exhibited decreased MASI and MELASQoL scores (P < 0.001); RCM revealed reduced melanin content in the lesions, and dermoscopy revealed fewer blood vessels. Fifteen lipid subclasses and 382 lipid molecules were identified using lipidomic assays. The expression levels of total lipids, phosphatidylcholine, and phosphatidylethanolamine in the melasma lesions decreased after treatment (P < 0.05). CONCLUSION: This study revealed alterations in the SSL composition after effective melasma treatment, suggesting a compensatory role for lipids in melasma barrier function. The mechanism involving SSL and the lipid barrier, which influences melasma's occurrence, needs further elucidation.


Subject(s)
Hydroquinones , Lipidomics , Melanosis , Quality of Life , Humans , Melanosis/drug therapy , Female , Adult , Hydroquinones/therapeutic use , Hydroquinones/administration & dosage , Tranexamic Acid/therapeutic use , Middle Aged , Melanins/metabolism , Male , Lipids/blood , Lipids/analysis , Epidermis/metabolism , Epidermis/drug effects , Epidermis/pathology , Phosphatidylethanolamines/metabolism , Phosphatidylcholines/metabolism , Skin/pathology , Skin/drug effects , Skin/metabolism , Lipid Metabolism/drug effects
19.
Sci Rep ; 14(1): 10621, 2024 05 09.
Article in English | MEDLINE | ID: mdl-38729969

ABSTRACT

Asymptomatic Leucine-Rich Repeat Kinase 2 Gene (LRRK2) carriers are at risk for developing Parkinson's disease (PD). We studied presymptomatic substantia nigra pars compacta (SNc) regional neurodegeneration in asymptomatic LRRK2 carriers compared to idiopathic PD patients using neuromelanin-sensitive MRI technique (NM-MRI). Fifteen asymptomatic LRRK2 carriers, 22 idiopathic PD patients, and 30 healthy controls (HCs) were scanned using NM-MRI. We computed volume and contrast-to-noise ratio (CNR) derived from the whole SNc and the sensorimotor, associative, and limbic SNc regions. An analysis of covariance was performed to explore the differences of whole and regional NM-MRI values among the groups while controlling the effect of age and sex. In whole SNc, LRRK2 had significantly lower CNR than HCs but non-significantly higher volume and CNR than PD patients, and PD patients significantly lower volume and CNR compared to HCs. Inside SNc regions, there were significant group effects for CNR in all regions and for volumes in the associative region, with a trend in the sensorimotor region but no significant changes in the limbic region. PD had reduced volume and CNR in all regions compared to HCs. Asymptomatic LRRK2 carriers showed globally decreased SNc volume and CNR suggesting early nigral neurodegeneration in these subjects at risk of developing PD.


Subject(s)
Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 , Magnetic Resonance Imaging , Melanins , Parkinson Disease , Substantia Nigra , Humans , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Male , Female , Middle Aged , Melanins/metabolism , Magnetic Resonance Imaging/methods , Parkinson Disease/genetics , Parkinson Disease/diagnostic imaging , Parkinson Disease/pathology , Parkinson Disease/metabolism , Substantia Nigra/diagnostic imaging , Substantia Nigra/pathology , Substantia Nigra/metabolism , Aged , Heterozygote , Adult , Case-Control Studies
20.
BMC Genomics ; 25(1): 505, 2024 May 22.
Article in English | MEDLINE | ID: mdl-38778258

ABSTRACT

BACKGROUND: In day-old Hungarian white goose goslings, there is a noticeable difference in dorsal down coloration between males and females, with females having darker dorsal plumage and males having lighter plumage. The ability to autosex day-old goslings based on their dorsal down coloration is important for managing them efficiently and planning their nutrition in the poultry industry. The aim of this study was to determine the biological and genetic factors underlying this difference in dorsal down colorationthrough histological analysis, biochemical assays, transcriptomic profiling, and q‒PCR analysis. RESULTS: Tissue analysis and biochemical assays revealed that compared with males, 17-day-old embryos and day-old goslings of female geese exhibited a greater density of melanin-containing feather follicles and a greater melanin concentration in these follicles during development. Both female and male goslings had lower melanin concentrations in their dorsal skin compared to 17-day-old embryos. Transcriptome analysis identified a set of differentially expressed genes (DEGs) (MC1R, TYR, TYRP1, DCT and MITF) associated with melanogenesis pathways that were downregulated or silenced specifically in the dorsal skin of day-old goslings compared to 17-day-old embryos, affecting melanin synthesis in feather follicles. Additionally, two key genes (MC1R and MITF) associated with feather coloration showed differences between males and females, with females having higher expression levels correlated with increased melanin synthesis and darker plumage. CONCLUSION: The expression of multiple melanogenesis genes determines melanin synthesis in goose feather follicles. The dorsal down coloration of day-old Hungarian white goose goslings shows sexual dimorphism, likely due to differences in the expression of the MC1R and MITF genes between males and females. These results could help us better understand why male and female goslings exhibit different plumage patterns.


Subject(s)
Geese , Gene Expression Profiling , Melanins , Pigmentation , Sex Characteristics , Animals , Female , Male , Geese/genetics , Geese/metabolism , Melanins/metabolism , Pigmentation/genetics , Feathers/metabolism , Feathers/growth & development , Transcriptome
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