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Background: Diabetes impairs wound healing, notably in diabetic foot ulcers (DFU). Stress, marked by the accumulation of lipoylated mitochondrial enzymes and the depletion of Fe-S cluster proteins, triggers cuproptosis-a distinct form of cell death. The involvement of copper in the pathophysiology of DFU has been recognized, and currently, a copper-based therapeutic strategy is emerging as a viable option for enhancing ulcer healing. This study investigates genes linked to copper metabolism in DFU, aiming to uncover potential targets for therapeutic intervention. Methods: Two diabetic wound Gene Expression Omnibus (GEO) datasets were analyzed to study immune cell dysregulation in diabetic wounds. Differentially expressed genes related to copper metabolism were identified and analyzed using machine learning methods. Gene ontology, pathway enrichment, and immune infiltration analyses were performed using DFU samples. The expression of identified genes was validated using qRT-PCR and single-cell RNA sequencing. Results: Ten genes associated with copper metabolism were identified. Among these, SLC31A1 and ADNP were found to be significantly differentially expressed in DFU. Notably, SLC31A1 exhibited higher expression in macrophages, whereas ADNP was found to be highly expressed in fibroblasts and chondrocytes. Conclusion: The study indicates a close link between copper metabolism, the infiltration of immune cells, and DFU. It proposes that copper metabolism could influence the progression of DFU through the activation of immune responses. These observations offer fresh perspectives on the underlying mechanisms of DFU and identify potential targets for therapeutic intervention.
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BACKGROUND AND PURPOSE: Panaxynol (PNN) is a common natural minor component in Umbelliferae plants. Many clinical studies have shown that PNN exhibits nutritional value and anti-inflammatory and other pharmacological activities. However, whether PNN can mediate cardiac ischemia/reperfusion injury (IRI) remains unclear. Here, we aimed to determine the potential effects of PNN on myocardial IRI. METHODS: Myocardial IRI was stimulated in a mouse IRI model, and neonatal rat ventricle myocytes (NRVMs) were exposed to hypoxia/reoxygenation to construct in an vitro model. Myocardial infarction size, myocardial tissue injury, myocardial apoptotic index, hemodynamic monitoring, pyroptosis-related proteins, cardiac enzyme activities and inflammatory responses were examined to assess myocardial injury. RESULTS: It was found that PNN administration markedly reduced myocardial infarct size and apoptosis, suppressed myocardial damage and cell pyroptosis, attenuated pro-inflammatory cytokines and neutrophil infiltration via NLRP3 inhibitor. More importantly, PNN treatment remarkably decreased the expression of TLR4/NF-κB pathway-associated proteins and NLRP3-related pyroptosis proteins by HMGB1 inhibitor. PNN also enhanced cell viability, reduced cardiac enzyme activities, suppressed apoptosis and attenuated inflammation in the isolated NRVMs. Furthermore, vitro studies indicated that MCC950 (a NLRP3 inhibitor) increased the anti-inflammatory and anti-apoptotic effects of PNN on NRVMs via HMGB1/TLR4 pathway. CONCLUSION: To sum up, our results demonstrate that PNN exhibits a cardioprotective effect by modulating heart IRI-induced apoptosis and pyroptosis via HMGB1/TLR4/NF-κB pathway, thereby inhibiting NLRP3 inflammasome stimulation.
Assuntos
Proteína HMGB1 , Infarto do Miocárdio , Traumatismo por Reperfusão Miocárdica , Camundongos , Ratos , Animais , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Piroptose , Proteína HMGB1/metabolismo , Receptor 4 Toll-Like/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Apoptose , Miócitos Cardíacos/metabolismo , Inflamassomos/metabolismo , Infarto do Miocárdio/metabolismo , Modelos Animais de DoençasRESUMO
Baicalin is a traditional Chinese herbal medicine commonly used for hair loss, the precise molecular mechanism of which is unknown. In the present study, the mechanism of baicalin was investigated via the topical application of baicalin to reconstituted hair follicles on mice dorsa and evaluating the effect on canonical Wnt/ßcatenin signaling in the hair follicles and the activity of dermal papillar cells. The results indicate that baicalin stimulates the expression of Wnt3a, Wnt5a, frizzled 7 and disheveled 2 whilst inhibiting the Axin/casein kinase 1α/adenomatous polyposis coli/glycogen synthase kinase 3ß degradation complex, leading to accumulation of ßcatenin and activation of Wnt/ßcatenin signaling. In addition, baicalin was observed to increase the alkaline phosphatase levels in dermal papillar cells, a process which was dependent on Wnt pathway activation. Given its nontoxicity and ease of topical application, baicalin represents a promising treatment for alopecia and other forms of hair loss. Further studies of baicalin using human hair follicle transplants are warranted in preparation for future clinical use.
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Medicamentos de Ervas Chinesas/farmacologia , Flavonoides/farmacologia , Folículo Piloso/efeitos dos fármacos , Via de Sinalização Wnt/efeitos dos fármacos , Fosfatase Alcalina/metabolismo , Alopecia/tratamento farmacológico , Alopecia/metabolismo , Animais , Células Cultivadas , Feminino , Folículo Piloso/citologia , Folículo Piloso/metabolismo , Folículo Piloso/ultraestrutura , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Wnt/metabolismo , beta Catenina/metabolismoRESUMO
Melanosomes are membrane-bound intracellular organelles that are uniquely generated by melanocytes (MCs) in the basal layer of human epidermis. Highly pigmented mature melanosomes are transferred from MCs to keratinocytes (KCs), and then positioned in the supra-nuclear region to ensure protection against ultraviolet radiation (UVR). However, the molecular mechanism underlying melanosome (or melanin pigment) transfer remains enigmatic. Emerging evidence shows that exo-/endo-cytosis of the melanosome core (termed melanocore) has been considered as the main transfer manner between MCs and KCs. As KCs in the skin migrate up from the basal layer and undergo terminal differentiation, the melanocores they have taken up from MCs are subjected to degradation. In this study, we isolated individual melanocores from human MCs in culture and then induced their destruction/disruption using a physical approach. The results demonstrate that the ultrastructural integrity of melanocores is essential for their antioxidant and photoprotective properties. In addition, we also show that cathepsin V (CTSV), a lysosomal acid protease, is involved in melanocore degradation in calcium-induced differentiated KCs and is also suppressed in KCs following exposure to UVA or UVB radiation. Thus, our study demonstrates that change in the proportion of melanocores in the intact/undegraded state by CTSV-related degradation in KCs affects photoprotection of the skin.
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Catepsinas/metabolismo , Cisteína Endopeptidases/metabolismo , Fibroblastos/efeitos da radiação , Queratinócitos/efeitos da radiação , Melanócitos/efeitos da radiação , Melanossomas/efeitos da radiação , Antioxidantes/metabolismo , Transporte Biológico , Catepsinas/genética , Diferenciação Celular , Fracionamento Celular , Cisteína Endopeptidases/genética , Fibroblastos/metabolismo , Fibroblastos/ultraestrutura , Prepúcio do Pênis/citologia , Prepúcio do Pênis/metabolismo , Expressão Gênica , Humanos , Queratinócitos/metabolismo , Queratinócitos/ultraestrutura , Masculino , Melaninas/química , Melaninas/metabolismo , Melanócitos/metabolismo , Melanócitos/ultraestrutura , Melanossomas/química , Melanossomas/metabolismo , Cultura Primária de Células , Proteólise , Raios UltravioletaRESUMO
OBJECTIVES: The transfer of melanosomes from melanocytes to neighbouring keratinocytes is critical to protect the skin from the deleterious effects of ultraviolet A (UVA) and ultraviolet B (UVB) irradiation; however, the initial factor(s) that stimulates melanosome transfer remains unclear. In this study, we investigated the induction of retinal-dependent calcium (Ca2+ ) influx in melanocytes (MCs) by UVA or UVB irradiation and the effect of transient receptor potential cation channel subfamily M member 1 (TRPM1) (melastatin1)-related Ca2+ influx on melanosome transfer. MATERIALS AND METHODS: Primary human epidermal MCs were exposed to physiological doses of UVB or UVA light and loaded with a calcium indicator Fluo-4 dye. The change of intracellular calcium of MCs was monitored using a two-photon confocal fluorescence microscopy. MCs were co-cultured with human epidermal keratinocytes (KCs) in the absence or presence of voriconazole (a TRPM1 blocker) or calcium chelators. MCs were also transfected with TRPM1 siRNA for silencing the expression of TRPM1 gene. The melanosome transfer in the co-cultured cells was quantitatively analysed using flow cytometry and was further confirmed by immunofluorescent double-staining. The protein levels and distributions of TRPM1, OPN3 and OPN5 in MCs were measured by Western blotting or immunofluorescent staining. RESULTS: The retinal-dependent Ca2+ influx of UVA-exposed melanocytes differed greatly from that of UVB-exposed melanocytes in the timing-phase. The protein expression of TRPM1 in mono- and co-cultured MCs was dose-dependently up-regulated by UVA and UVB. TRPM1 siRNA-mediated knockdown and the blockage of TRPM1 channel using a putative antagonist (voriconazole) significantly inhibited melanosome transfer in co-cultures following UVA or UVB exposure. CONCLUSIONS: The distinct time-phases of Ca2+ influx in MCs induced by UVA or UVB contribute to the consecutive stimulation of melanosome transfer, thereby providing a potent photoprotection against harmful UV radiation.