A20 ameliorates advanced glycation end products-induced melanogenesis by inhibiting NLRP3 inflammasome activation in human dermal fibroblasts.

BACKGROUND: Advanced glycation end products (AGEs) promote melanogenesis through activating NLRP3 inflammasome in fibroblasts. Although A20 has been highlighted to inhibit NLRP3 inflammasome activation, its roles and mechanisms remain elusive in photoaging-associated pigmentation. OBJECTIVES: To determine the significance of fibroblast A20 in AGEs-induced NLRP3 inflammasome activation and pigmentation. METHODS: The correlation between A20 and AGEs or melanin was studied in sun-exposed skin and lesions of melasma and solar lentigo. We then investigated A20 level in AGEs-treated fibroblast and the effect of fibroblast A20 overexpression or knockdown on AGEs-BSA-induced NLRP3 inflammasome activation and pigmentation, respectively. Finally, the severity of NLRP3 inflammasome activation and pigmentation was evaluated after mice were injected intradermally with A20-overexpression adeno-associated virus and AGEs-BSA. RESULTS: Dermal A20 expression was decreased and exhibited negative correlation with either dermal AGEs deposition or epidermal melanin level in sun-exposed skin and pigmentary lesions. Moreover, both AGEs-BSA and AGEs-collagen robustly decreased A20 expression via binding to RAGE in fibroblasts. Further, A20 overexpression or depletion significantly decreased or augmented AGEs-BSA-induced activation of NF-κB pathway and NLRP3 inflammasome and IL-18 production and secretion in fibroblasts, respectively. Importantly, fibroblast A20 potently repressed AGEs-BSA-stimulated melanin content，tyrosinase activity，and expression of microphthalmia-associated transcription factor and tyrosinase in melanocytes. Particularly, fibroblast A20 significantly abrogated AGEs-BSA-promoted melanogenesis in ex vivo skin and mouse models. Additionally, fibroblast A20 inhibited AGEs-BSA-activated MAPKs in melanocytes and the epidermis of ex vivo skin. CONCLUSIONS: Fibroblast A20 suppresses AGEs-stimulate melanogenesis in photoaging-associated hyperpigmentation disorders by inhibiting NLRP3 inflammasome activation.

Advanced Glycation End Products Promote Melanogenesis by Activating NLRP3 Inflammasome in Human Dermal Fibroblasts.

Advanced glycation end product (AGE) accumulation is significantly increased in the dermis of photoaged skin and plays crucial roles in photoaging. Although AGEs have been found to contribute to the yellowish discoloration of photoaged skin, their roles in photoaging-associated hyperpigmentation disorders have not been extensively studied. In this study, we observed that AGEs, NLRP3, and IL-18 were increased in the dermis of sun-exposed skin and lesions of melasma and solar lentigo and that dermal deposition of AGE was positively correlated with epidermal melanin levels. In addition, we found that AGE-BSA potently activated NLRP3 inflammasome and promoted IL-18 production and secretion in cultured fibroblasts, which was mediated by receptor for AGE/NF-κB pathway. Moreover, AGE-BSA significantly promoted melanogenesis by increasing tyrosinase activity and expression of microphthalmia-associated transcription factor and tyrosinase, which was dependent on NLRP3 inflammasome activation and IL-18 secretion in fibroblasts. Notably, AGE-collagen could activate NLRP3 inflammasome in fibroblasts and enhance melanogenesis. Furthermore, we found that IL-18 enhanced melanogenesis by binding to its receptor and activating p38 MAPK and extracellular signal‒regulated kinase 1/2 signaling pathways in melanocytes. Importantly, the promelanogenesis of AGE-BSA was verified in ex vivo cultured skin and mouse models. These findings suggest that dermal AGEs stimulate melanogenesis and contribute to the development of photoaging-associated hyperpigmentation disorders.

Advanced glycation end products alter the m6A-modified RNA profiles in human dermal fibroblasts.

Background: To explore advanced glycation end products (AGEs)-induced m6A modification in fibroblasts and its potential role in photoaging. Methods: We studied m6A modification in AGEs-bovine serum albumin-treated fibroblasts with m6A-mRNA & lncRNA epitranscriptomic microarray and bioinformatics analysis. The m6A modification level was also investigated in skin samples. Results: m6A methylation microarray analysis revealed m6A modification profiles in AGEs-treated fibroblasts. Gene ontology, Kyoto Encyclopedia of Genes and Genomes, protein-protein interaction and competing endogenous RNA network analysis indicated that the genes of differentially methylated mRNAs and lncRNAs were mainly related to inflammation processes. We also found that AGEs-bovine serum albumin dose-dependently increased the m6A level and METTL14 expression in both fibroblasts and sun-exposed skin. Conclusion: Our study provided novel information regarding alterations of m6A modifications in AGEs-induced dermal fibroblasts and potential targets for treatment of photoaging.

UVA-induced photoaging inhibits autophagic degradation by impairing lysosomal function in dermal fibroblasts.

Autophagy has been associated with a variety of diseases especially aging. Human dermal fibroblasts (HDFs) can internalize and then degrade elastin, collagen and advanced glycation end products (AGEs) in lysosomes, which plays prominent roles in extracellular matrix homeostasis and AGEs removal in the dermis. Although autophagy has been reported to be decreased in photoaged fibroblasts, the underlying mechanism and its relevance to photoaging remain elusive. Here, we showed that GFP-LC3 puncta per cell, LC3Ⅰ/Ⅱ conversion and p62 expression were significantly increased, whereas beclin1 expression was not altered in UVA-induced photoaged fibroblasts compared with non-photoaged control. Moreover, autophagic flux was not significantly affected by chloroquine treatment, but was remarkably induced by rapamycin treatment in photoaged fibroblasts, suggesting that UVA-induced photoaging might inhibit autophagy at the degradation stage. Further lysosomal function studies demonstrated that degradation of formed autophagosomes, LC3Ⅱprotein and DQ-Green BSA was all dramatically decreased in photoaged fibroblasts. LysoSensor yellow/blue DND 160 staining and flow cytometry assays demonstrated that photoaging obviously attenuated lysosomal acidification. Also, decreased expression of cathepsin B, L and D was found in photoaged fibroblasts. These data suggest that lowered lysosomal acidity and decreased cathepsins expression might contribute to the inhibition of autophagic degradation, which might be crucial in the development of photoaging through impairing intracellular degradation.