RESUMEN

Skin identity is controlled by intrinsic features of the epidermis and dermis and their interactions. Modifying skin identity has clinical potential, such as the conversion of residual limb and stump (nonvolar) skin of amputees to pressure-responsive palmoplantar (volar) skin to enhance prosthesis use and minimize skin breakdown. Greater keratin 9 (KRT9) expression, higher epidermal thickness, keratinocyte cytoplasmic size, collagen length, and elastin are markers of volar skin and likely contribute to volar skin resiliency. Given fibroblasts' capacity to modify keratinocyte differentiation, we hypothesized that volar fibroblasts influence these features. Bioprinted skin constructs confirmed the capacity of volar fibroblasts to induce volar keratinocyte features. A clinical trial of healthy volunteers demonstrated that injecting volar fibroblasts into nonvolar skin increased volar features that lasted up to 5 months, highlighting a potential cellular therapy.

Asunto(s)

Refuerzo Biomédico , Bioimpresión , Dermis , Epidermis , Fibroblastos , Queratinocitos , Adulto , Femenino , Humanos , Masculino , Amputados , Diferenciación Celular , Colágeno/metabolismo , Dermis/citología , Dermis/metabolismo , Elastina/metabolismo , Epidermis/metabolismo , Fibroblastos/citología , Fibroblastos/trasplante , Mano , Queratina-9/metabolismo , Queratinocitos/citología , Queratinocitos/metabolismo , Refuerzo Biomédico/métodos

RESUMEN

BACKGROUND: Burn trauma is one of the major causes of morbidity and mortality worldwide. The standard management of burn wounds consists of early debridement, dressing changes, surgical management, and split-thickness skin autografts (STSGs). However, there are limitations for the standard management that inclines us to find alternative treatment approaches, such as innovative cell-based therapies. We aimed to systematically review the different aspects of cell-based treatment approaches for burn wounds in clinical trials. METHODS: A systematic search through PubMed, Medline, Embase, and Cochrane Library databases was carried out using a combination of keywords, including "Cell transplantation", "Fibroblast", "Keratinocyte", "Melanocyte", or "Stem Cell" with "Burn", "Burn wound", or "Burn injury". Firstly, titles and abstracts of the studies existing in these databases until "February 2024" were screened. Then, the selected studies were read thoroughly, and considering the inclusion and exclusion criteria, final articles were included in this systematic review. Moreover, a manual search was performed through the reference lists of the included studies to minimize the risk of missing reports. RESULTS: Overall, 30 clinical trials with 970 patients were included in our study. Considering the type of cells, six studies used keratinocytes, nine used fibroblasts, eight used combined keratinocytes and fibroblasts, one study used combined keratinocytes and melanocytes, five used combined keratinocytes and fibroblasts and melanocytes, and one study used mesenchymal stem cells (MSCs). Evaluation of the preparation type in these studies showed that cultured method was used in 25 trials, and non-cultured method in 5 trials. Also, the graft type of 17 trials was allogeneic, and of 13 other trials was autologous. CONCLUSIONS: Our study showed that employing cell-based therapies for the treatment of burn wounds have significant results in clinical studies and are promising approaches that can be considered as alternative treatments in many cases. However, choosing appropriate cell-based treatment for each burn wound is essential and depends on the situation of each patient.

Asunto(s)

Quemaduras , Tratamiento Basado en Trasplante de Células y Tejidos , Humanos , Quemaduras/terapia , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Ensayos Clínicos como Asunto , Queratinocitos/citología , Queratinocitos/trasplante , Trasplante de Piel/métodos , Cicatrización de Heridas , Trasplante de Células Madre Mesenquimatosas/métodos

RESUMEN

Epidermal transplantation is a common and widely used surgical technique in clinical medicine. Derivatives of embryonic stem cells have the potential to serve as a source of transplantable cells. However, allograft rejection is one of the main challenges. To investigate the immunogenicity of keratinocytes derived from human embryonic stem cells (ESKCs), we conducted a series of in vivo and in vitro experiments. The results showed that ESKCs have low HLA molecule expression, limited antigen presentation capabilities, and a weak ability to stimulate the proliferation and secretion of inflammatory factors in allogeneic PBMCs in vitro. In humanized immune mouse models, ESKCs elicited weak transplant rejection responses in the host. Overall, we found that ESKCs have low immunogenicity and may have potential applications in the field of regenerative medicine.

Asunto(s)

Células Madre Embrionarias Humanas , Queratinocitos , Humanos , Queratinocitos/inmunología , Queratinocitos/metabolismo , Queratinocitos/citología , Células Madre Embrionarias Humanas/citología , Células Madre Embrionarias Humanas/inmunología , Células Madre Embrionarias Humanas/metabolismo , Animales , Ratones , Proliferación Celular , Rechazo de Injerto/inmunología , Antígenos HLA/inmunología , Antígenos HLA/metabolismo

RESUMEN

Theranostic sutures are derived from innovative ideas to enhance wound healing results by adding wound diagnostics and therapeutics to typical sutures by functionalizing them with additional materials. Here, we present a new direct electrospinning method for the fast, continuous, inexpensive, and high-throughput production of versatile nanofibrous-coated suture threads, with precise control over various essential microstructural and physical characteristics. The thickness of the coating layer and the alignment of nanofibers with the thread's direction can be adjusted by the user by varying the spooling speed and the displacement between the spinneret needle and thread. To show the flexibility of our method for a range of different materials selected, gelatin, polycaprolactone, silk fibroin, and PEDOT:PSS (poly(3,4-ethylene dioxythiophene):poly(styrene sulfonate)) were the resultant nanofibers characterized by scanning electron microscopy (SEM) imaging and conductivity tests. In a series of in vitro and ex vivo tests (pig skin), sutures were successfully tested for their flexibility and mechanical properties when used as weaving and knotting sutures, and their biocompatibility with a keratinocyte cell line. For temperature-based drug-releasing tests, two fluorescent molecules as drug models with high and low molecular weight, namely fluorescein isothiocyanate-dextran (20 kDa) and rhodamine B (470 Da), were used, and their steady release with incremental increase of temperature to 37 °C over 120 min was seen, which is appropriate for bacterial treatment drugs. Given the advantages of the presented technique, it seems to have promising potential to be used in future medical applications for wound closure and bacterial infection treatment via a temperature-triggered drug release strategy.

Asunto(s)

Nanofibras , Rodaminas , Suturas , Cicatrización de Heridas , Nanofibras/química , Animales , Cicatrización de Heridas/efectos de los fármacos , Humanos , Rodaminas/química , Porcinos , Poliésteres/química , Dextranos/química , Gelatina/química , Nanoporos , Fluoresceína-5-Isotiocianato/química , Materiales Biocompatibles Revestidos/química , Queratinocitos/citología , Queratinocitos/metabolismo , Fibroínas/química , Línea Celular

RESUMEN

The multibiocomponent hybrid alginate hydrogels based on brown and sea algae, containing 100 % ingredients of natural origin were prepared by ionic crosslinking reaction of a polymeric matrix with lipid nanodispersion. To the best of the Authors' knowledge such multicomponent biobased hydrogel of promising medical and cosmetical applications for the first time was obtained in the environment of flower water, received earlier as a waste by-product from various chemical processes. An innovative hybrid alginate hydrogel that is completely biodegradable and eco-friendly was obtained following waterless and upcycling trends that are in line with the principles of sustainable development. The optimal composition of the lipid nanodispersion and the polymeric matrix was selected using the statistical method of design of the experiment. Based on obtained results, multibiocomponent hybrid alginate hydrogels with various ratios of lipid nanodispersion were obtained. Subsequently, the porous structure and elasticity of the hybrid hydrogels were analyzed. Moreover, to confirm the safety of the multibiocomponent alginate hybrid hydrogels the cytotoxic tests were carried out using human fibroblasts and keratinocytes cell lines. As the final product hybrid of hydrolate-swollen alginate hydrogel and lipid nanodispersion containing several active ingredients (silymarin, bakuchiol, spirulina) was obtained.

Asunto(s)

Alginatos , Materiales Biocompatibles , Hidrogeles , Lípidos , Hidrogeles/química , Alginatos/química , Humanos , Lípidos/química , Materiales Biocompatibles/química , Cosméticos/química , Fibroblastos/efectos de los fármacos , Fibroblastos/citología , Queratinocitos/efectos de los fármacos , Queratinocitos/citología , Línea Celular

RESUMEN

The agouti (Dasyprocta prymnolopha) is a medium-sized, wild rodent that is highly rustic and docile. Its size and ease of management make it a viable candidate for an alternative animal model to traditional murine subjects. However, data on the epidermal strata of agoutis are lacking, with significant uncertainties persisting regarding their skin's characterization. This study aimed to describe and quantify the epidermal strata of skin biopsies from male and female agoutis raised in captivity, to further validate the species as a model for dermatological research. Ultrastructural evaluations through atomic force microscopy (AFM) and stereological analyses were conducted, revealing significant differences between the layers of the skin; notably, the dermis exhibited a greater total volume than the epidermis. The findings suggest that the epidermal strata are well-defined, with the volume likely correlating to the size and cellular density of the keratinocytes. Corneodesmosomes and tonofilaments were identified across all epidermal layers, indicating the probable maintenance of anchoring protein activity, even post-cornification of these cells. These results suggest that the agouti may serve as a promising model for dermatological studies, owing to the homogeneity of its cutaneous tissue across different body regions and the distinct volume and morphology of its epithelial stratification, which could enhance the applicability of systematic investigative methods in the future.

Asunto(s)

Epidermis , Animales , Epidermis/ultraestructura , Epidermis/anatomía & histología , Masculino , Femenino , Dasyproctidae/anatomía & histología , Queratinocitos/ultraestructura , Queratinocitos/citología , Dermatología , Microscopía de Fuerza Atómica

RESUMEN

BACKGROUND/AIMS: Aquaporin-3 (AQP3) is an aquaglyceroporin and peroxiporin that plays a crucial role in skin barrier homeostasis. Dysregulated AQP3 expression has been observed in different inflammatory skin conditions. Hidradenitis Suppurativa (HS) is an autoinflammatory keratinization disease that typically appears between 10 and 21 years of age, characterized by alteration of skin barrier homeostasis. METHODS: To evaluate in vitro the role of AQP3 in the development of HS, we performed real-time PCR and Western blot to analyze gene and protein levels in human keratinocyte cell lines knock-out (KO) for NCSTN and PSENEN genes, simulating genetic-associated HS. Additionally, we investigated the impact of Glyceryl Glucoside (GG) on biological processes by performing MTT, scratch, proliferation assays and proteome studies. RESULTS: We detected a significant decrease of the levels of AQP3 gene and protein in KO cell lines. GG effectively elevated the levels of mRNA and protein, significantly decreased the hyperproliferation rate, and enhanced cell migration in our in vitro model of genetic Hidradenitis Suppurativa. Pathway enrichment analysis further confirmed GG's role in the migration and proliferation pathways of keratinocytes. CONCLUSION: Our results suggest that AQP3 may act as a new novel actor in HS etio-pathogenesis, and GG could be further explored as potential treatment option for managing HS in patients.

Asunto(s)

Acuaporina 3 , Movimiento Celular , Proliferación Celular , Glucósidos , Hidradenitis Supurativa , Queratinocitos , Humanos , Acuaporina 3/metabolismo , Acuaporina 3/genética , Hidradenitis Supurativa/metabolismo , Hidradenitis Supurativa/patología , Hidradenitis Supurativa/tratamiento farmacológico , Hidradenitis Supurativa/genética , Queratinocitos/metabolismo , Queratinocitos/efectos de los fármacos , Queratinocitos/patología , Queratinocitos/citología , Proliferación Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Glucósidos/farmacología , Glucósidos/uso terapéutico , Línea Celular

RESUMEN

Significance: Accurate identification of epidermal cells on reflectance confocal microscopy (RCM) images is important in the study of epidermal architecture and topology of both healthy and diseased skin. However, analysis of these images is currently done manually and therefore time-consuming and subject to human error and inter-expert interpretation. It is also hindered by low image quality due to noise and heterogeneity. Aim: We aimed to design an automated pipeline for the analysis of the epidermal structure from RCM images. Approach: Two attempts have been made at automatically localizing epidermal cells, called keratinocytes, on RCM images: the first is based on a rotationally symmetric error function mask, and the second on cell morphological features. Here, we propose a dual-task network to automatically identify keratinocytes on RCM images. Each task consists of a cycle generative adversarial network. The first task aims to translate real RCM images into binary images, thus learning the noise and texture model of RCM images, whereas the second task maps Gabor-filtered RCM images into binary images, learning the epidermal structure visible on RCM images. The combination of the two tasks allows one task to constrict the solution space of the other, thus improving overall results. We refine our cell identification by applying the pre-trained StarDist algorithm to detect star-convex shapes, thus closing any incomplete membranes and separating neighboring cells. Results: The results are evaluated both on simulated data and manually annotated real RCM data. Accuracy is measured using recall and precision metrics, which is summarized as the F 1 -score. Conclusions: We demonstrate that the proposed fully unsupervised method successfully identifies keratinocytes on RCM images of the epidermis, with an accuracy on par with experts' cell identification, is not constrained by limited available annotated data, and can be extended to images acquired using various imaging techniques without retraining.

Asunto(s)

Epidermis , Queratinocitos , Microscopía Confocal , Humanos , Microscopía Confocal/métodos , Epidermis/diagnóstico por imagen , Queratinocitos/citología , Procesamiento de Imagen Asistido por Computador/métodos , Algoritmos , Células Epidérmicas , Redes Neurales de la Computación , Aprendizaje Automático no Supervisado

RESUMEN

Terminal differentiation of epithelial cells is critical for the barrier function of the skin, the growth of skin appendages, such as hair and nails, and the development of the skin of amniotes. Here, we present the hypothesis that the differentiation of cells in the embryonic periderm shares characteristic features with the differentiation of epithelial cells that support the morphogenesis of cornified skin appendages during postnatal life. The periderm prevents aberrant fusion of adjacent epithelial sites during early skin development. It is shed off when keratinocytes of the epidermis form the cornified layer, the stratum corneum. A similar role is played by epithelia that ensheath cornifying skin appendages until they disintegrate to allow the separation of the mature part of the skin appendage from the adjacent tissue. These epithelia, exemplified by the inner root sheath of hair follicles and the epithelia close to the free edge of nails or claws, are referred to as scaffolding epithelia. The periderm and scaffolding epithelia are similar with regard to their transient functions in separating tissues and the conserved expression of trichohyalin and trichohyalin-like genes in mammals and birds. Thus, we propose that parts of the peridermal differentiation program were coopted to a new postnatal function during the evolution of cornified skin appendages in amniotes.

Asunto(s)

Diferenciación Celular , Diferenciación Celular/fisiología , Animales , Piel/embriología , Piel/citología , Piel/metabolismo , Células Epiteliales/citología , Células Epiteliales/metabolismo , Epitelio/embriología , Epitelio/metabolismo , Epidermis/embriología , Epidermis/metabolismo , Queratinocitos/citología , Queratinocitos/metabolismo , Folículo Piloso/embriología , Folículo Piloso/citología , Humanos , Morfogénesis

RESUMEN

Exposure to ultraviolet B (UVB) radiation represents a significant environmental threat to human skin. This study investigates the protective mechanism of Artemisia Capillaris Thunb. (AC) extract against UVB-induced apoptosis and inflammation in HaCaT keratinocytes. AC extract demonstrated a significant protective effect, as evidenced by reduced early apoptosis, late apoptosis, and necrosis, as well as decreased apoptotic cell status upon UVB exposure. Additionally, AC extract effectively inhibited UVB-induced DNA damage, as indicated by diminished γ-H2AX foci formation. Restoration of mitochondrial damage and normalization of mitochondrial membrane potential, along with the reduction of intracellular and mitochondrial reactive oxygen species (ROS) levels, were observed with AC extract pre-treatment. The extract also exhibited anti-inflammatory properties, evidenced by the decreased release of IL-1α, IL-6, and PGE2 from keratinocytes. Additional research on the molecular mechanisms uncovered that the AC extract alters the cGAS/STING pathway, suppressing the mRNA (cGAS, STING, IRF3, IRF7 and TBK1) and protein levels (cGAS, STING, IRF3, IRF7 and NF-κB) linked to this particular pathway. The HPLC analysis identified chlorogenic acid and its derivatives as the major components in AC, constituting up to 16.44% of the total chlorogenic acid content. The cGAS/STING signaling pathway was found to be suppressed by chlorogenic acid and its derivatives, as indicated by molecular docking studies and RT-qPCR analysis. This suppression contributes to the protective effects against cell apoptosis and inflammation induced by UVB. To summarize, AC extract, which is abundant in chlorogenic acid and its derivatives, shows potential in protecting keratinocytes from damage caused by UVB by regulating the cGAS/STING signaling pathway.

Asunto(s)

Apoptosis , Artemisia , Queratinocitos , Proteínas de la Membrana , Nucleotidiltransferasas , Extractos Vegetales , Transducción de Señal , Rayos Ultravioleta , Humanos , Rayos Ultravioleta/efectos adversos , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Extractos Vegetales/farmacología , Extractos Vegetales/química , Artemisia/química , Nucleotidiltransferasas/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/efectos de la radiación , Proteínas de la Membrana/metabolismo , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Queratinocitos/efectos de la radiación , Queratinocitos/citología , Especies Reactivas de Oxígeno/metabolismo , Inflamación/metabolismo , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de la radiación , Daño del ADN/efectos de los fármacos , Daño del ADN/efectos de la radiación , Ácido Clorogénico/farmacología , Ácido Clorogénico/química , Dinoprostona/metabolismo , Células HaCaT , Línea Celular

RESUMEN

Plakophilin 4 (PKP4) is a component of cell-cell junctions that regulates intercellular adhesion and Rho-signaling during cytokinesis with an unknown function during epidermal differentiation. Here we show that keratinocytes lacking PKP4 fail to develop a cortical actin ring, preventing adherens junction maturation and generation of tissue tension. Instead, PKP4-depleted cells display increased stress fibers. PKP4-dependent RhoA localization at AJs was required to activate a RhoA-ROCK2-MLCK-MLC2 axis and organize actin into a cortical ring. AJ-associated PKP4 provided a scaffold for the Rho activator ARHGEF2 and the RhoA effectors MLCK and MLC2, facilitating the spatio-temporal activation of RhoA signaling at cell junctions to allow cortical ring formation and actomyosin contraction. In contrast, association of PKP4 with the Rho suppressor ARHGAP23 reduced ARHGAP23 binding to RhoA which prevented RhoA activation in the cytoplasm and stress fiber formation. These data identify PKP4 as an AJ component that transduces mechanical signals into cytoskeletal organization.

Asunto(s)

Actinas , Uniones Adherentes , Placofilinas , Proteína de Unión al GTP rhoA , Placofilinas/metabolismo , Placofilinas/genética , Proteína de Unión al GTP rhoA/metabolismo , Uniones Adherentes/metabolismo , Humanos , Actinas/metabolismo , Queratinocitos/metabolismo , Queratinocitos/citología , Proteínas Activadoras de GTPasa/metabolismo , Proteínas Activadoras de GTPasa/genética , Quinasas Asociadas a rho/metabolismo , Quinasas Asociadas a rho/genética , Transducción de Señal , Fibras de Estrés/metabolismo , Células Cultivadas , Animales

RESUMEN

This study investigated the three-dimensional (3D) cellular interactions and tunneling nanotubes (TNTs) during fetal mouse skin regeneration on embryonic days 13 (E13) and 15 (E15). We aimed to understand spatial relationships among cell types involved in skin regeneration and assess the potential role of TNTs. Full-thickness skin incisions were performed in E13 and E15 embryos. Wound sites were collected, embedded in epoxy resin, processed for 3D reconstruction (1 µm thickness sections), and subjected to whole-mount immunostaining. We conducted in vitro co-culture experiments with fetal macrophages and fibroblasts to observe TNT formation. To assess the effect of TNTs on skin regeneration, an inhibiting agent (cytochalasin B) was administered to amniotic fluid. Results revealed that E13 epidermal keratinocytes interacted with dermal fibroblasts and macrophages, facilitating skin regrowth. TNT structures were observed at the E13-cell wound sites, among macrophages, and between macrophages and fibroblasts, confirmed through in vitro co-culture experiments. In vitro and utero cytochalasin B administration hindered those formation and inefficient skin texture regeneration at E13 wound sites. This emphasizes the necessity of 3D cellular interactions between epidermal and dermal cells during skin regeneration in mouse embryos at E13. The prevalence of TNT structures indicated their involvement in achieving complete skin texture restoration.

Asunto(s)

Técnicas de Cocultivo , Fibroblastos , Nanotubos , Regeneración , Piel , Animales , Ratones , Regeneración/fisiología , Piel/metabolismo , Nanotubos/química , Queratinocitos/citología , Queratinocitos/fisiología , Macrófagos/metabolismo , Feto , Femenino , Cicatrización de Heridas/efectos de los fármacos , Cicatrización de Heridas/fisiología , Comunicación Celular , Citocalasina B/farmacología

RESUMEN

Epithelia consist of proliferating and differentiating cells that often display patterned arrangements. However, the mechanism regulating these spatial arrangements remains unclear. Here, we show that cell-cell adhesion dictates multicellular patterning in stratified epithelia. When cultured keratinocytes, a type of epithelial cell in the skin, are subjected to starvation, they spontaneously develop a pattern characterized by areas of high and low cell density. Pharmacological and knockout experiments show that adherens junctions are essential for patterning, whereas the mathematical model that only considers local cell-cell adhesion as a source of attractive interactions can form regions with high/low cell density. This phenomenon, called cell-cell adhesion-induced patterning (CAIP), influences cell differentiation and proliferation through Yes-associated protein modulation. Starvation, which induces CAIP, enhances the stratification of the epithelia. These findings highlight the intrinsic self-organizing property of epithelial cells.

Asunto(s)

Uniones Adherentes , Adhesión Celular , Diferenciación Celular , Proliferación Celular , Células Epiteliales , Queratinocitos , Adhesión Celular/fisiología , Queratinocitos/metabolismo , Queratinocitos/citología , Diferenciación Celular/genética , Humanos , Células Epiteliales/metabolismo , Células Epiteliales/citología , Uniones Adherentes/metabolismo , Animales , Epitelio/metabolismo , Ratones , Células Cultivadas

RESUMEN

Mesenchymal stem/stromal cells (MSCs) and their extracellular vesicles (MSC-EVs) have been described to have important roles in tissue regeneration, including tissue repair, control of inflammation, enhancing angiogenesis, and regulating extracellular matrix remodeling. MSC-EVs have many advantages for use in regeneration therapies such as facility for dosage, histocompatibility, and low immunogenicity, thus possessing a lower possibility of rejection. In this work, we address the potential activity of MSC-EVs isolated from adipose-derived MSCs (ADMSC-EVs) cultured on cross-linked dextran microcarriers, applied to test the scalability and reproducibility of EV production. Isolated ADMSC-EVs were added into cultured human dermal fibroblasts (NHDF-1), keratinocytes (HaCat), endothelial cells (HUVEC), and THP-1 cell-derived macrophages to evaluate cellular responses (i.e., cell proliferation, cell migration, angiogenesis induction, and macrophage phenotype-switching). ADMSC viability and phenotype were assessed during cell culture and isolated ADMSC-EVs were monitored by nanotracking particle analysis, electron microscopy, and immunophenotyping. We observed an enhancement of HaCat proliferation; NHDF-1 and HaCat migration; endothelial tube formation on HUVEC; and the expression of inflammatory cytokines in THP-1-derived macrophages. The increased expression of TGF-ß and IL-1ß was observed in M1 macrophages treated with higher doses of ADMSC-EVs. Hence, EVs from microcarrier-cultivated ADMSCs are shown to modulate cell behavior, being able to induce skin tissue related cells to migrate and proliferate as well as stimulate angiogenesis and cause balance between pro- and anti-inflammatory responses in macrophages. Based on these findings, we suggest that the isolation of EVs from ADMSC suspension cultures makes it possible to induce in vitro cellular responses of interest and obtain sufficient particle numbers for the development of in vivo concept tests for tissue regeneration studies.

Asunto(s)

Proliferación Celular , Vesículas Extracelulares , Macrófagos , Células Madre Mesenquimatosas , Humanos , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Vesículas Extracelulares/metabolismo , Macrófagos/metabolismo , Macrófagos/citología , Movimiento Celular , Células THP-1 , Fibroblastos/metabolismo , Fibroblastos/citología , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Técnicas de Cultivo de Célula/métodos , Células Cultivadas , Queratinocitos/metabolismo , Queratinocitos/citología , Citocinas/metabolismo

RESUMEN

Intracellular organelles support cellular physiology in diverse conditions. In the skin, epidermal keratinocytes undergo differentiation with gradual changes in cellular physiology, accompanying remodeling of lysosomes and the Golgi apparatus. However, it was not known whether changes in Golgi and lysosome morphology and their redistribution were linked. Here, we show that disassembled Golgi is distributed in close physical apposition to lysosomes in differentiated keratinocytes. This atypical localization requires the Golgi tethering protein GRASP65, which is associated with both the Golgi and lysosome membranes. Depletion of GRASP65 results in the loss of Golgi-lysosome apposition and the malformation of lysosomes, defined by their aberrant morphology, size, and function. Surprisingly, a trans-Golgi enzyme and secretory Golgi cargoes are extensively localized to the lysosome lumen and secreted to the cell surface, contributing to total protein secretion of differentiated keratinocytes but not in proliferative precursors, indicating that lysosomes acquire specialization during differentiation. We further demonstrate that the secretory function of the Golgi apparatus is critical to maintain keratinocyte lysosomes. Our study uncovers a novel form of Golgi-lysosome cross-talk and its role in maintaining specialized secretory lysosomes in differentiated keratinocytes.

Asunto(s)

Diferenciación Celular , Aparato de Golgi , Proteínas de la Matriz de Golgi , Queratinocitos , Lisosomas , Lisosomas/metabolismo , Queratinocitos/metabolismo , Queratinocitos/citología , Aparato de Golgi/metabolismo , Humanos , Proteínas de la Matriz de Golgi/metabolismo , Proteínas de la Matriz de Golgi/genética , Proteínas de la Membrana/metabolismo

RESUMEN

Autophagy participates in the regulation of ferroptosis. Among numerous autophagy-related genes (ATGs), ATG5 plays a pivotal role in ferroptosis. However, how ATG5-mediated ferroptosis functions in UVB-induced skin inflammation is still unclear. In this study, we unveil that the core ferroptosis inhibitor GPX4 is significantly decreased in human skin tissue exposed to sunlight. We report that ATG5 deletion in mouse keratinocytes strongly protects against UVB-induced keratinocyte ferroptosis and skin inflammation. Mechanistically, ATG5 promotes the autophagy-dependent degradation of GPX4 in UVB-exposed keratinocytes, which leads to UVB-induced keratinocyte ferroptosis. Furthermore, we find that IFN-γ secreted by ferroptotic keratinocytes facilitates the M1 polarization of macrophages, which results in the exacerbation of UVB-induced skin inflammation. Together, our data indicate that ATG5 exacerbates UVB-induced keratinocyte ferroptosis in the epidermis, which subsequently gives rise to the secretion of IFN-γ and M1 polarization. Our study provides novel evidence that targeting ATG5 may serve as a potential therapeutic strategy for the amelioration of UVB-caused skin damage.

Asunto(s)

Proteína 5 Relacionada con la Autofagia , Ferroptosis , Interferón gamma , Queratinocitos , Macrófagos , Rayos Ultravioleta , Queratinocitos/metabolismo , Queratinocitos/efectos de la radiación , Queratinocitos/citología , Proteína 5 Relacionada con la Autofagia/metabolismo , Proteína 5 Relacionada con la Autofagia/genética , Animales , Ratones , Interferón gamma/metabolismo , Macrófagos/metabolismo , Macrófagos/efectos de la radiación , Macrófagos/citología , Humanos , Piel/efectos de la radiación , Piel/metabolismo , Piel/patología , Fosfolípido Hidroperóxido Glutatión Peroxidasa/metabolismo , Autofagia/efectos de la radiación , Inflamación/metabolismo , Inflamación/patología

RESUMEN

BACKGROUND: Ultraviolet-B (UVB) radiation is the leading environmental cause of skin damage and photoaging. The epidermis and dermis layers of the skin mainly absorb UVB. UVB stimulates apoptosis, cell cycle arrest, generation of reactive oxygen species, and degradation of collagen and elastin fibers. OBJECTIVE: This study investigated the potential of human growth hormone (hGH) in protecting the skin fibroblasts and keratinocytes (HFFF-2 and HaCaT cell lines) from UVB-induced damage. METHODS: The MTT assay was performed to evaluate UVB-induced mitochondrial damage via assessing the mitochondrial dehydrogenase activity, and flow cytometry was carried out to investigate the effects of UVB and hGH on the cell cycle and apoptosis of UVB-irradiated cells. In addition, the fold change mRNA expression levels of Type I collagen and elastin in HFFF-2 cells were evaluated using the qRT-PCR method following UVB exposure. RESULTS: We observed that treatment of cells with hGH before UVB exposure inhibited UVB-induced loss of mitochondrial dehydrogenase activity, apoptosis, and sub-G1 population formation in both cell lines. We also found that hGH-treated HFFF-2 cells showed up-regulated mRNA expression of Type I collagen, elastin, and IGF-1 in response to UVB irradiation. CONCLUSION: These findings suggest hGH as a potential anti-UVB compound that can protect skin cells from UVB-induced damage. Our findings merit further investigation and can be used to better understand the role of hGH in skin photoaging.

Asunto(s)

Apoptosis , Colágeno Tipo I , Elastina , Fibroblastos , Hormona de Crecimiento Humana , Queratinocitos , Rayos Ultravioleta , Humanos , Elastina/metabolismo , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Línea Celular , Fibroblastos/efectos de la radiación , Fibroblastos/metabolismo , Fibroblastos/efectos de los fármacos , Fibroblastos/citología , Colágeno Tipo I/metabolismo , Colágeno Tipo I/genética , Queratinocitos/efectos de la radiación , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Queratinocitos/citología , Hormona de Crecimiento Humana/metabolismo , Hormona de Crecimiento Humana/farmacología , Piel/efectos de la radiación , Piel/efectos de los fármacos , Piel/metabolismo , Piel/citología , Factor I del Crecimiento Similar a la Insulina/metabolismo , Mitocondrias/metabolismo , Mitocondrias/efectos de la radiación , Mitocondrias/efectos de los fármacos , ARN Mensajero/metabolismo , ARN Mensajero/genética

RESUMEN

In the present study, pyroligneous acid, also known as wood vinegar, has been employed as reducing and stabilizing agent in the synthesis of silver nanoparticles (AgNPs) anchored on nanocellulose (NC). The idea is to confer the latter bactericidal properties for its typical uses such as in cosmetics and food-packing. It has been demonstrated that AgNPs can be directly produced onto NC in one-pot fashion while dramatically enhancing the kinetics of AgNPs synthesis (2 h for reaction completion) in comparison to the NC-less counterpart (10 days for reaction completion). Furthermore, NC allowed for a narrower size distribution of AgNPs. NC-supported and non-supported AgNPs had sizes of 5.1 ± 1.6 nm and 16.7 ± 4.62 nm, respectively. Immortalized human keratinocytes (HaCat) cells were then employed as model to evaluate the cytotoxicity of the AgNPs-NC compound. The latter was found not to impact cell proliferation at any formulation, while decreasing the viability by only 6.8% after 72 h. This study contributes to the development of more environmentally benign routes to produce nanomaterials and to the understanding of their impact on cells.

Asunto(s)

Supervivencia Celular , Celulosa , Células HaCaT , Nanopartículas del Metal , Plata , Humanos , Plata/química , Nanopartículas del Metal/química , Celulosa/química , Celulosa/farmacología , Supervivencia Celular/efectos de los fármacos , Queratinocitos/efectos de los fármacos , Queratinocitos/citología , Tamaño de la Partícula , Proliferación Celular/efectos de los fármacos , Ácido Acético/química , Ácido Acético/farmacología

RESUMEN

Two-dimensional (2D) cell culture is an important tool in the discovery of skin-active agents. Fibroblasts and keratinocytes, more rarely fibroblast-keratinocyte cocultures, are usually used for that purpose, where test compounds are added by mixing with the overlaying growth medium. However, such an approach is suboptimal because it lacks the stratum corneum component. The stratum corneum acts as a selective gatekeeper and opposes the intradermal permeation of many compounds that are bioactive when placed in direct contact with cells. One solution is to use reconstituted epidermis, but this approach is costly and time consuming. Here, a model is proposed, where the simplicity and convenience of the 2D cell culture is combined with the advantage of a hydrophobic barrier reminiscent of the skin horny layer. This model was tested with skin-relevant solvents, as well as with "naked" hydrophilic and encapsulated compounds. Cell viability and collagen stimulation were used as readouts. The results showed that the incorporation of a stratum corneum-substitute barrier on top of a 2D cell culture reduced the cytotoxicity of a common cosmetic solvent, dimethyl isosorbide (DMI), in cell culture and modified the bioactivity of the added actives (magnesium ascorbyl phosphate [MAP] and oligomeric proanthocyanidins [OPCs]/levan biopolymer), which became dependent on their ability to penetrate through a lipidic layer. Taken together, these results indicate a better physiological relevance of this cell culture model in workflows aimed at the discovery and analysis of skin-active compounds than conventional 2D systems.

Asunto(s)

Técnicas de Cocultivo , Queratinocitos , Técnicas de Cocultivo/métodos , Humanos , Queratinocitos/citología , Queratinocitos/metabolismo , Queratinocitos/efectos de los fármacos , Epidermis/metabolismo , Fibroblastos/citología , Fibroblastos/metabolismo , Fibroblastos/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Piel/citología , Piel/metabolismo , Modelos Biológicos

RESUMEN

In the interfollicular epidermis, keratinocyte stem cells (KSC) generate a short-lived population of transit amplifying (TA) cells that undergo terminal differentiation after several cell divisions. Recently, we isolated and characterized a highly proliferative keratinocyte cell population, named "early" TA (ETA) cell, representing the first KSC progenitor with exclusive features. This work aims to evaluate epidermis, with a focus on KSC and ETA cells, during transition from infancy to childhood. Reconstructed human epidermis (RHE) generated from infant keratinocytes is more damaged by UV irradiation, as compared to RHE from young children. Moreover, the expression of several differentiation and barrier genes increases with age, while the expression of genes related to stemness is reduced from infancy to childhood. The proliferation rate of KSC and ETA cells is higher in cells derived from infants' skin samples than of those derived from young children, as well as the capacity of forming colonies is more pronounced in KSC derived from infants than from young children's skin samples. Finally, infants-KSC show the greatest regenerative capacity in skin equivalents, while young children ETA cells express higher levels of differentiation markers, as compared to infants-ETA. KSC and ETA cells undergo substantial changes during transition from infancy to childhood. The study presents a novel insight into pediatric skin, and sheds light on the correlation between age and structural maturation of the skin.

Asunto(s)

Diferenciación Celular , Queratinocitos , Células Madre , Humanos , Lactante , Células Madre/citología , Células Madre/metabolismo , Queratinocitos/metabolismo , Queratinocitos/citología , Preescolar , Proliferación Celular , Células Epidérmicas/metabolismo , Células Epidérmicas/citología , Niño , Piel/citología , Piel/metabolismo , Femenino , Masculino , Epidermis/metabolismo , Células Cultivadas