Thioredoxin promotes the regeneration and binding of elastic fibre and basement membrane.

OBJECTIVE: Thioredoxin (TRX), a ubiquitous protein with strong antioxidant activity, decreases in the skin with age. A decrease in TRX is expected to induce cellular senescence, chronic inflammation, and degeneration and loss of extracellular matrix (ECM), such as collagen and elastin within the skin. In this study, we investigated the effects of TRX addition to excised skin or skin models to understand the role of TRX on cells and ECM within the skin. METHODS: To evaluate its effect on skin cells, we cultured a three-dimensional (3D) skin model in a medium containing TRX. The mRNA expression levels of proteins related to elastic and collagen fibres and the basement membrane were determined. Furthermore, 3D imaging and computational analysis were performed to evaluate the effect of TRX on the elastic fibres and extending COL VII structures in excised human skin after coculturing with TRX for 1, 4, 5 and 6 days. RESULTS: Thioredoxin application to a 3D skin model upregulated elastin, COLI and COLVII mRNA expression. Applying TRX to the excised skin increased the number of linear elastic fibres. This effect of TRX demonstrated a daily increment in a dose-dependent manner. Thioredoxin extended the fibrous structure of COL VII into the dermis, expanding its colocalization region with elastic fibres. These structural effects were confirmed using 3D imaging and computational methods. CONCLUSION: Thioredoxin elongates elastic fibres from the dermis to the basement membrane and extends the COL VII structure from the basement membrane to the dermis in excised human skin. These findings suggest the potential of TRX to protect the skin against age-related alterations such as wrinkles and sagging.

OBJECTIF: Thioredoxin (TRX), une protéine ubiquitaire dotée d'une forte activité antioxydante, diminue dans la peau avec l'âge. Une diminution de la TRX est susceptible d'induire la sénescence cellulaire, l'inflammation chronique, et la dégénérescence ainsi que la perte de la matrice extracellulaire (ECM), telle que le collagène et l'élastine de la peau. Dans cette étude, nous avons examiné les effets de l'ajout de TRX à la peau prélevée ou aux modèles de peau afin de comprendre le rôle de TRX sur les cellules et la matrice extracellulaire (ECM) de la peau. MÉTHODES: Pour évaluer son effet sur les cellules cutanées, nous avons cultivé un modèle de peau tridimensionnel (3D) dans un milieu contenant du TRX. Les niveaux d'expression de l'ARNm des protéines liées aux fibres élastiques et de collagène ainsi que de la membrane basale ont été déterminés. De plus, une imagerie 3D et une analyse informatique ont été réalisées pour évaluer l'effet de la TRX sur les fibres élastiques et les structures de COL VII étendues dans la peau humaine prélevée après une coculture avec la TRX pendant 1, 4, 5 et 6 jours. RÉSULTATS: L'application de la Thioredoxin à un modèle de peau en 3D a régulé à la hausse l'expression de l'élastine, du COLI et du COLVII au niveau de l'ARNm. L'application de TRX à la peau excisée a augmenté le nombre de fibres élastiques linéaires. Cet effet du TRX a montré une augmentation quotidienne de manière dose­dépendante. Le Thioredoxin a étendu la structure fibreuse du COL VII dans le derme, élargissant ainsi sa région de colocalisation avec les fibres élastiques. Ces effets structuraux ont été confirmés à l'aide d'imagerie 3D et de méthodes computationnelles. CONCLUSIONS: La Thioredoxin allonge les fibres élastiques du derme à la membrane basale et étend la structure de COL VII de la membrane basale au derme dans la peau humaine excisée. Ces résultats suggèrent le potentiel de la TRX pour protéger la peau contre les altérations liées à l'âge telles que les rides et le relâchement cutané.

Investigation of stratum corneum cell morphology and content using novel machine-learning image analysis.

BACKGROUND: The morphology and content of stratum corneum (SC) cells provide information on the physiological condition of the skin. Although the morphological and biochemical properties of the SC are known, no method is available to fully access and interpret this information. This study aimed to develop a method to comprehensively decode the physiological information of the skin, based on the SC. Therefore, we established a novel image analysis technique based on artificial intelligence (AI) and multivariate analysis to predict skin conditions. MATERIALS AND METHODS: SC samples were collected from participants, imaged, and annotated. Nine biomarkers were measured in the samples using enzyme-linked immunosorbent assay. The data were then used to teach machine-learning models to recognize individual SC cell regions and estimate the levels of the nine biomarkers from the images. Skin physiological indicators (e.g., skin barrier function, facial analysis, and questionnaires) were measured or obtained from the participants. Multivariate analysis, including biomarker levels ​​and structural parameters of the SC as variables, was used to estimate these physiological indicators. RESULTS: We established two machine-learning models. The accuracy of recognition was assessed according to the average intersection over union (0.613), precision (0.953), recall (0.640), and F-value (0.766). The predicted biomarker levels significantly correlated with the measured levels. Skin physiological indicators and questionnaire answers were predicted with strong correlations and correct answer rates. CONCLUSION: Various physiological skin conditions can be predicted from images of the SC using AI models and multivariate analysis. Our method is expected to be useful for dermatological treatment optimization.

Long Hanging Structure of Collagen VII Connects the Elastic Fibers and the Basement Membrane in Young Skin Tissue.

Aging leads to substantial structural changes in the skin. Elastic fibers maintain skin structure, but their degeneration and loss of function with age result in wrinkle formation and loss of skin elasticity. Oxytalan fiber, a type of elastic fiber, extends close to the dermal-epidermal junction (DEJ) from the back of the dermis. Oxytalan fibers are abundant in the papillary layer and contribute to skin elasticity and texture. However, to accurately understand the mechanisms of skin elasticity, the interaction between elastic fibers and DEJ should be elucidated. Here, we investigated elastic fibers and DEJ and their structural alterations with aging. Several basement membrane proteins [collagen (COL) IV, COLVII, and laminin 332], fibrous tropoelastin, and fibrillin-1 in excised human skin tissue were observed using three-dimensional imaging. Age-related alterations in COLVII, elastic fibers, and fibrillin-1 were evaluated. We found that COLVII forms long hanging structures and is co-localized with fibrous tropoelastin in young skin but not aged skin. Fibrillin-1-rich regions were observed at the tips of elastin fibers in young skin tissue, but rarely in aged skin. This co-localization of elastic fiber and COLVII may maintain skin structure, thereby preventing wrinkling and sagging. COLVII is a potential therapeutic target for skin wrinkling.

Elastin microfibril interface-located protein 1 and its catabolic enzyme, cathepsin K, regulate the age-related structure of elastic fibers in the skin.

INTRODUCTION: The elastic fiber structure becomes shorter, thicker, and curved with age. Nonetheless, the proteins and catabolic enzymes influencing the maintenance of and change in the three-dimensional (3D) structure of elastic fibers remain unknown. This study aimed to identify the proteins involved in the maintenance and degeneration of elastic fiber structures. METHODS: We performed a combined 3D structural analysis using tissue decolorization technology and mRNA abundance and comprehensive protein expression of tissue-derived cells. The relationship between the proteins was evaluated. RESULTS: Elastin microfibril interface-located protein 1 (EMILIN-1) and cathepsin K (CTSK) were implicated in structural changes in elastic fibers with aging. EMILIN-1 and CTSK levels were highly correlated and changed with age. CTSK was identified as the degrading enzyme of EMILIN-1. CTSK fragmented the otherwise linearly existing dermal elastic fiber structure, with more evident changes in oxytalan fibers. EMILIN-1 expression in fibroblasts was increased by co-culturing with keratinocytes. Furthermore, CTSK expression was increased by UV stress in keratinocytes, resulting in decreased EMILIN-1 expression. CONCLUSION: Using our new assessment strategy, we observed that EMILIN-1 and CTSK are highly linked to changes in the elastic fiber structure with aging. These results indicate that suppressing CTSK expression and increasing EMILIN-1 expression might be an effective approach to prevent elastic fiber morphological changes that lead to wrinkles and sagging. Furthermore, EMILIN-1 in the dermis increases due to interaction with the epidermis, which could provide a new target for the therapeutic care of elastic fibers (including preservation of oxytalan fibers) in epidermis-dermis interaction.

The protective role of DJ-1 in ultraviolet-induced damage of human skin: DJ-1 levels in the stratum corneum as an indicator of antioxidative defense.

DJ-1 is a multifunctional protein associated with Parkinson's disease and plays a significant role in protecting nerve cells from oxidative stress. DJ-1 is expressed in the skin, although its function there is unknown. In this study, we investigated DJ-1 function in keratinocytes. DJ-1 was induced by H2O2 exposure and UV irradiation in keratinocytes. DJ-1 knockdown with small interfering RNA (siRNA) increased reactive oxygen species (ROS) and lactate dehydrogenase (LDH) release after UVB irradiation, suggesting that DJ-1 reduces ROS and might protect skin cells from UV damage in vitro. To investigate the in vivo role of DJ-1 in the skin, we determined DJ-1 levels in human stratum corneum samples obtained by the tape-stripping method. DJ-1 levels in the stratum corneum (scDJ-1) correlated with total antioxidant capacity. We also examined the effect of scDJ-1 on changes in skin after UVB irradiation. DJ-1 was elevated in SC from the upper arm 1 to 2 weeks after UVB irradiation. One day after UVB irradiation, L* (brightness) and a* (redness) values, indicators of skin color, were altered regardless of scDJ-1 expression. However, these values recovered more quickly in subjects with high scDJ-1 expression than in those with low scDJ-1 expression. These data suggest that DJ-1 in skin plays a significant role in protection against UV radiation and oxidative stress, and that DJ-1 levels in the SC might be an indicator of antioxidative defense against UV-induced damage.

Macrophage migration inhibitory factor (MIF) in the stratum corneum: a marker of the local severity of atopic dermatitis.

Different biomarkers are used to evaluate the severity of atopic dermatitis (AD); however, it remains difficult to determine the severity of localized skin lesions. MIF plays an essential role in the pathophysiology of skin inflammation. To establish whether the MIF level in the stratum corneum (SC) serves as a marker of the severity of AD lesions, we examined the SC MIF (scMIF) levels in AD patients. The SC of the cheek, neck and upper arm skin was collected using tape stripping, and the scMIF levels were measured. Consequently, the scMIF levels were found to be significantly higher in the involved skin lesions than the uninvolved areas within the same patient. Moreover, the scMIF levels were significantly correlated with the severity of local skin lesions. These findings suggest that the scMIF level can be used as an effective marker for evaluating the local severity of AD.