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2.
Int J Mol Sci ; 25(13)2024 Jun 21.
Article in English | MEDLINE | ID: mdl-38999931

ABSTRACT

Aging is associated with a decline in the functionality of various cell types, including dermal fibroblasts, which play a crucial role in maintaining skin homeostasis and wound healing. Chronic inflammation and increased reactive oxygen species (ROS) production are hallmark features of aging, contributing to impaired wound healing. MicroRNA-146a (miR-146a) has been implicated as a critical regulator of inflammation and oxidative stress in different cell types, yet its role in aged dermal fibroblasts and its potential relevance to wound healing remains poorly understood. We hypothesize that miR-146a is differentially expressed in aged dermal fibroblasts and that overexpression of miR-146a will decrease aging-induced inflammatory responses and ROS production. Primary dermal fibroblasts were isolated from the skin of 17-week-old (young) and 88-week-old (aged) mice. Overexpression of miR-146a was achieved through miR-146a mimic transfection. ROS were detected using a reliable fluorogenic marker, 2,7-dichlorofluorescin diacetate. Real-time PCR was used to quantify relative gene expression. Our investigation revealed a significant reduction in miR-146a expression in aged dermal fibroblasts compared to their younger counterparts. Moreover, aged dermal fibroblasts exhibited heightened levels of inflammatory responses and increased ROS production. Importantly, the overexpression of miR-146a through miR-146a mimic transfection led to a substantial reduction in inflammatory responses through modulation of the NF-kB pathway in aged dermal fibroblasts. Additionally, the overexpression of miR-146a led to a substantial decrease in ROS production, achieved through the downregulation of NOX4 expression in aged dermal fibroblasts. These findings underscore the pivotal role of miR-146a in mitigating both inflammatory responses and ROS production in aged dermal fibroblasts, highlighting its potential as a therapeutic target for addressing age-related skin wound healing.


Subject(s)
Fibroblasts , Inflammation , MicroRNAs , Reactive Oxygen Species , MicroRNAs/genetics , MicroRNAs/metabolism , Fibroblasts/metabolism , Reactive Oxygen Species/metabolism , Animals , Mice , Inflammation/metabolism , Inflammation/genetics , Inflammation/pathology , NADPH Oxidase 4/metabolism , NADPH Oxidase 4/genetics , Skin/metabolism , Skin/pathology , Skin/cytology , NF-kappa B/metabolism , Cells, Cultured , Aging/metabolism , Aging/genetics , Oxidative Stress
3.
Int J Mol Sci ; 25(13)2024 Jun 26.
Article in English | MEDLINE | ID: mdl-39000078

ABSTRACT

The immunogenicity of allogeneic skin fibroblasts in transplantation has been controversial. Whether this controversy comes from a natural heterogeneity among fibroblast subsets or species-specific differences between human and mouse remains to be addressed. In this study, we sought to investigate whether fibroblasts derived from either adult or neonatal human skin tissues could induce different immune responses toward phagocytosis and T cell activation using in vitro co-culture models. Our results indicate that both phagocytosis and T cell proliferation are reduced in the presence of neonatal skin fibroblasts compared to adult skin fibroblasts. We also show that neonatal skin fibroblasts secrete paracrine factors that are responsible for reduced T cell proliferation. In addition, we show that neonatal skin fibroblasts express less class II human leukocyte antigen (HLA) molecules than adult skin fibroblasts after interferon gamma priming, which might also contribute to reduced T cell proliferation. In conclusion, this study supports the use of allogeneic neonatal skin fibroblasts as a readily available cell source for tissue production and transplantation to treat patients with severe injuries.


Subject(s)
Cell Proliferation , Fibroblasts , Skin , T-Lymphocytes , Humans , Fibroblasts/metabolism , Fibroblasts/immunology , Skin/immunology , Skin/metabolism , Skin/cytology , Infant, Newborn , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Lymphocyte Activation/immunology , Coculture Techniques , Cells, Cultured , Phagocytosis , Adult , Interferon-gamma/metabolism
4.
Cells ; 13(13)2024 Jul 07.
Article in English | MEDLINE | ID: mdl-38995009

ABSTRACT

We developed an automated microregistration method that enables repeated in vivo skin microscopy imaging of the same tissue microlocation and specific cells over a long period of days and weeks with unprecedented precision. Applying this method in conjunction with an in vivo multimodality multiphoton microscope, the behavior of human skin cells such as cell proliferation, melanin upward migration, blood flow dynamics, and epidermal thickness adaptation can be recorded over time, facilitating quantitative cellular dynamics analysis. We demonstrated the usefulness of this method in a skin biology study by successfully monitoring skin cellular responses for a period of two weeks following an acute exposure to ultraviolet light.


Subject(s)
Skin , Humans , Skin/cytology , Skin/diagnostic imaging , Ultraviolet Rays , Cell Tracking/methods , Cell Proliferation , Cell Movement , Microscopy, Fluorescence, Multiphoton/methods , Microscopy/methods
5.
Int J Mol Sci ; 25(14)2024 Jul 12.
Article in English | MEDLINE | ID: mdl-39062918

ABSTRACT

The aging process is linked to numerous cellular changes, among which are modifications in the functionality of dermal fibroblasts. These fibroblasts play a crucial role in sustaining the healing of skin wounds. Reduced cell proliferation is a hallmark feature of aged dermal fibroblasts. Long intergenic non-coding RNA (lincRNAs), such as LincRNA-EPS (Erythroid ProSurvival), has been implicated in various cellular processes. However, its role in aged dermal fibroblasts and its impact on the cell cycle and its regulator, Cyclin D1 (CCND1), remains unclear. Primary dermal fibroblasts were isolated from the skin of 17-week-old (young) and 88-week-old (aged) mice. Overexpression of LincRNA-EPS was achieved through plasmid transfection. Cell proliferation was detected using the MTT assay. Real-time PCR was used to quantify relative gene expressions. Our findings indicate a noteworthy decline in the expression of LincRNA-EPS in aged dermal fibroblasts, accompanied by reduced levels of CCND1 and diminished cell proliferation in these aging cells. Significantly, the overexpression of LincRNA-EPS in aged dermal fibroblasts resulted in an upregulation of CCND1 expression and a substantial increase in cell proliferation. Mechanistically, LincRNA-EPS induces CCND1 expression by sequestering miR-34a, which was dysregulated in aged dermal fibroblasts, and directly targeting CCND1. These outcomes underscore the crucial role of LincRNA-EPS in regulating CCND1 and promoting cell proliferation in aged dermal fibroblasts. Our study provides novel insights into the molecular mechanisms underlying age-related changes in dermal fibroblasts and their implications for skin wound healing. The significant reduction in LincRNA-EPS expression in aged dermal fibroblasts and its ability to induce CCND1 expression and enhance cell proliferation highlight its potential as a therapeutic target for addressing age-related skin wound healing.


Subject(s)
Cell Proliferation , Cyclin D1 , Fibroblasts , RNA, Long Noncoding , Cyclin D1/metabolism , Cyclin D1/genetics , Fibroblasts/metabolism , Fibroblasts/cytology , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , Animals , Mice , Skin/metabolism , Skin/cytology , MicroRNAs/genetics , MicroRNAs/metabolism , Cells, Cultured , Skin Aging/genetics , Dermis/cytology , Dermis/metabolism , Cellular Senescence/genetics , Gene Expression Regulation , Wound Healing/genetics , Aging/genetics
6.
Sci Rep ; 14(1): 12670, 2024 06 03.
Article in English | MEDLINE | ID: mdl-38830883

ABSTRACT

Gelatin-methacryloyl (GelMA) is a highly adaptable biomaterial extensively utilized in skin regeneration applications. However, it is frequently imperative to enhance its physical and biological qualities by including supplementary substances in its composition. The purpose of this study was to fabricate and characterize a bi-layered GelMA-gelatin scaffold using 3D bioprinting. The upper section of the scaffold was encompassed with keratinocytes to simulate the epidermis, while the lower section included fibroblasts and HUVEC cells to mimic the dermis. A further step involved the addition of amniotic membrane extract (AME) to the scaffold in order to promote angiogenesis. The incorporation of gelatin into GelMA was found to enhance its stability and mechanical qualities. While the Alamar blue test demonstrated that a high concentration of GelMA (20%) resulted in a decrease in cell viability, the live/dead cell staining revealed that incorporation of AME increased the quantity of viable HUVECs. Further, gelatin upregulated the expression of KRT10 in keratinocytes and VIM in fibroblasts. Additionally, the histological staining results demonstrated the formation of well-defined skin layers and the creation of extracellular matrix (ECM) in GelMA/gelatin hydrogels during a 14-day culture period. Our study showed that a 3D-bioprinted composite scaffold comprising GelMA, gelatin, and AME can be used to regenerate skin tissues.


Subject(s)
Amnion , Bioprinting , Fibroblasts , Gelatin , Human Umbilical Vein Endothelial Cells , Keratinocytes , Tissue Engineering , Tissue Scaffolds , Keratinocytes/drug effects , Keratinocytes/cytology , Keratinocytes/metabolism , Gelatin/chemistry , Humans , Tissue Engineering/methods , Fibroblasts/drug effects , Fibroblasts/metabolism , Fibroblasts/cytology , Tissue Scaffolds/chemistry , Amnion/cytology , Amnion/metabolism , Amnion/chemistry , Bioprinting/methods , Printing, Three-Dimensional , Skin/metabolism , Skin/cytology , Methacrylates/chemistry , Cell Survival/drug effects , Endothelial Cells/metabolism , Endothelial Cells/drug effects , Endothelial Cells/cytology
7.
Stem Cell Res Ther ; 15(1): 169, 2024 Jun 18.
Article in English | MEDLINE | ID: mdl-38886773

ABSTRACT

With the development of the economy and the increasing prevalence of skin problems, cutaneous medical aesthetics are gaining more and more attention. Skin disorders like poor wound healing, aging, and pigmentation have an impact not only on appearance but also on patients with physical and psychological issues, and even impose a significant financial burden on families and society. However, due to the complexities of its occurrence, present treatment options cannot produce optimal outcomes, indicating a dire need for new and effective treatments. Mesenchymal stem cells (MSCs) and their secretomics treatment is a new regenerative medicine therapy that promotes and regulates endogenous stem cell populations and/or replenishes cell pools to achieve tissue homeostasis and regeneration. It has demonstrated remarkable advantages in several skin-related in vivo and in vitro investigations, aiding in the improvement of skin conditions and the promotion of skin aesthetics. As a result, this review gives a complete description of recent scientific breakthroughs in MSCs for skin aesthetics and the limitations of their clinical applications, aiming to provide new ideas for future research and clinical transformation.


Subject(s)
Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells , Humans , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Mesenchymal Stem Cell Transplantation/methods , Skin/cytology , Regenerative Medicine/methods , Skin Diseases/therapy , Wound Healing , Animals
8.
Int J Mol Sci ; 25(11)2024 May 22.
Article in English | MEDLINE | ID: mdl-38891823

ABSTRACT

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.


Subject(s)
Cell Differentiation , Keratinocytes , Stem Cells , Humans , Infant , Stem Cells/cytology , Stem Cells/metabolism , Keratinocytes/metabolism , Keratinocytes/cytology , Child, Preschool , Cell Proliferation , Epidermal Cells/metabolism , Epidermal Cells/cytology , Child , Skin/cytology , Skin/metabolism , Female , Male , Epidermis/metabolism , Cells, Cultured
9.
Toxicol In Vitro ; 99: 105870, 2024 Aug.
Article in English | MEDLINE | ID: mdl-38848825

ABSTRACT

Particulate matter 2.5 (PM2.5) causes skin aging, inflammation, and impaired skin homeostasis. Hyperoside, a flavanol glycoside, has been proposed to reduce the risk of diseases caused by oxidative stress. This study evaluated the cytoprotective potential of hyperoside against PM2.5-induced skin cell damage. Cultured human HaCaT keratinocytes were pretreated with hyperoside and treated with PM2.5. Initially, the cytoprotective and antioxidant ability of hyperoside against PM2.5 was evaluated. Western blotting was further employed to investigate endoplasmic reticulum (ER) stress and cellular senescence and for evaluation of cell cycle regulation-related proteins. Hyperoside inhibited PM2.5-mediated ER stress as well as mitochondrial damage. Colony formation assessment confirmed that PM2.5-impaired cell proliferation was restored by hyperoside. Moreover, hyperoside reduced the activation of PM2.5-induced ER stress-related proteins, such as protein kinase R-like ER kinase, cleaved activating transcription factor 6, and inositol-requiring enzyme 1. Hyperoside promoted cell cycle progression in the G0/G1 phase by upregulating the PM2.5-impaired cell cycle regulatory proteins. Hyperoside significantly reduced the expression of PM2.5-induced senescence-associated ß-galactosidase and matrix metalloproteinases (MMPs), such as MMP-1 and MMP-9. Overall, hyperoside ameliorated PM2.5-impaired cell proliferation, ER stress, and cellular senescence, offering potential therapeutic implications for mitigating the adverse effects of environmental pollutants on skin health.


Subject(s)
Cellular Senescence , Endoplasmic Reticulum Stress , Keratinocytes , Particulate Matter , Quercetin , Humans , Endoplasmic Reticulum Stress/drug effects , Particulate Matter/toxicity , Cellular Senescence/drug effects , Quercetin/pharmacology , Quercetin/analogs & derivatives , Keratinocytes/drug effects , Cell Line , Cell Proliferation/drug effects , Cell Cycle/drug effects , HaCaT Cells , Antioxidants/pharmacology , Skin/drug effects , Skin/metabolism , Skin/cytology
10.
Int J Mol Sci ; 25(12)2024 Jun 07.
Article in English | MEDLINE | ID: mdl-38927998

ABSTRACT

Mesenchymal adipose stromal cells (ASCs) are considered the most promising and accessible material for translational medicine. ASCs can be used independently or within the structure of scaffold-based constructs, as these not only ensure mechanical support, but can also optimize conditions for cell activity, as specific features of the scaffold structure have an impact on the vital activity of the cells. This manuscript presents a study of the secretion and accumulation that occur in a conditioned medium during the cultivation of human ASCs within the structure of such a partial skin-equivalent that is in contact with it. It is demonstrated that the ASCs retain their functional activity during cultivation both within this partial skin-equivalent structure and, separately, on plastic substrates: they proliferate and secrete various proteins that can then accumulate in the conditioned media. Our comparative study of changes in the conditioned media during cultivation of ASCs on plastic and within the partial skin-equivalent structure reveals the different dynamics of the release and accumulation of such secretory factors in the media under a variety of conditions of cell functioning. It is also demonstrated that the optimal markers for assessment of the ASCs' secretory functions in the studied partial skin-equivalent structure are the trophic factors VEGF-A, HGF, MCP, SDF-1α, IL-6 and IL-8. The results will help with the development of an algorithm for preclinical studies of this skin-equivalent in vitro and may be useful in studying various other complex constructs that include ASCs.


Subject(s)
Chemokine CXCL12 , Interleukin-6 , Interleukin-8 , Mesenchymal Stem Cells , Vascular Endothelial Growth Factor A , Humans , Chemokine CXCL12/metabolism , Mesenchymal Stem Cells/metabolism , Mesenchymal Stem Cells/cytology , Culture Media, Conditioned , Vascular Endothelial Growth Factor A/metabolism , Interleukin-6/metabolism , Interleukin-8/metabolism , Hepatocyte Growth Factor/metabolism , Cells, Cultured , Skin/metabolism , Skin/cytology , Cell Proliferation , Chemokine CCL2/metabolism , Adipose Tissue/cytology , Adipose Tissue/metabolism
11.
Methods Mol Biol ; 2849: 1-15, 2024.
Article in English | MEDLINE | ID: mdl-38904915

ABSTRACT

Studying human skin biology can aid in comprehending the pathophysiology of skin diseases and developing novel cell-based therapies, including tissue engineering approaches. This chapter provides a comprehensive guide of methods to determine human skin samples from the perspective of their cellular compositions. We describe as useful technique the histological analysis of tissue sections. We further illustrate the biological characterization of isolated and cultured basal and suprabasal interfollicular keratinocytes by cell sorting, cytospin immunostaining, colony forming efficiency, and long-term dermo-epidermal organotypic cultures.


Subject(s)
Cell Separation , Epidermal Cells , Keratinocytes , Stem Cells , Humans , Keratinocytes/cytology , Keratinocytes/metabolism , Cell Separation/methods , Stem Cells/cytology , Stem Cells/metabolism , Epidermal Cells/cytology , Epidermal Cells/metabolism , Cell Culture Techniques/methods , Cells, Cultured , Skin/cytology , Epidermis/metabolism , Tissue Engineering/methods , Cell Differentiation
12.
Biofabrication ; 16(4)2024 Jul 12.
Article in English | MEDLINE | ID: mdl-38941996

ABSTRACT

Human skin vasculature features a unique anatomy in close proximity to the skin appendages and acts as a gatekeeper for constitutive lymphocyte trafficking to the skin. Approximating such structural complexity and functionality in 3D skin models is an outstanding tissue engineering challenge. In this study, we leverage the capabilities of the digital-light-processing bioprinting to generate an anatomically-relevant and miniaturized 3D skin-on-a-chip (3D-SoC) model in the size of a 6 mm punch biopsy. The 3D-SoC contains a perfusable vascular network resembling the superficial vascular plexus of the skin and closely surrounding bioengineered hair follicles. The perfusion capabilities of the 3D-SoC enables the circulation of immune cells, and high-resolution imaging of the immune cell-endothelial cell interactions, namely tethering, rolling, and extravasation in real-time. Moreover, the vascular pattern in 3D-SoC captures the physiological range of shear rates found in cutaneous blood vessels and allows for studying the effect of shear rate on T cell trafficking. In 3D-SoC, as expected,in vitro-polarized T helper 1 (Th1) cells show a stronger attachment on the vasculature compared to naïve T cells. Both naïve and T cells exhibit higher retention in the low-shear zones in the early stages (<5 min) of T cell attachment. Interestingly, at later stages T cell retention rate becomes independent of the shear rate. The attached Th1 cells further transmigrate from the vessel walls to the extracellular space and migrate toward the bioengineered hair follicles and interfollicular epidermis. When the epidermis is not present, Th1 cell migration toward the epidermis is significantly hindered, underscoring the role of epidermal signals on T cell infiltration. Our data validates the capabilities of 3D-SoC model to study the interactions between immune cells and skin vasculature in the context of epidermal signals. The biopsy-sized 3D-SoC model in this study represents a new level of anatomical and cellular complexity, and brings us a step closer to generating a truly functional human skin with its tissue-specific vasculature and appendages in the presence of circulating immune cells.


Subject(s)
Hair Follicle , Skin , Humans , Skin/blood supply , Skin/cytology , Hair Follicle/cytology , Hair Follicle/blood supply , Cell Movement , Biopsy , Tissue Engineering , Bioprinting
13.
J Photochem Photobiol B ; 257: 112961, 2024 Aug.
Article in English | MEDLINE | ID: mdl-38917719

ABSTRACT

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.


Subject(s)
Apoptosis , Collagen Type I , Elastin , Fibroblasts , Human Growth Hormone , Keratinocytes , Ultraviolet Rays , Humans , Elastin/metabolism , Apoptosis/drug effects , Apoptosis/radiation effects , Cell Line , Fibroblasts/radiation effects , Fibroblasts/metabolism , Fibroblasts/drug effects , Fibroblasts/cytology , Collagen Type I/metabolism , Collagen Type I/genetics , Keratinocytes/radiation effects , Keratinocytes/drug effects , Keratinocytes/metabolism , Keratinocytes/cytology , Human Growth Hormone/metabolism , Human Growth Hormone/pharmacology , Skin/radiation effects , Skin/drug effects , Skin/metabolism , Skin/cytology , Insulin-Like Growth Factor I/metabolism , Mitochondria/metabolism , Mitochondria/radiation effects , Mitochondria/drug effects , RNA, Messenger/metabolism , RNA, Messenger/genetics
14.
J Dermatol Sci ; 115(1): 42-50, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38876908

ABSTRACT

BACKGROUND: Recessive dystrophic epidermolysis bullosa (RDEB) is a severe skin fragility disorder caused by mutations in the COL7A1 gene, which encodes type VII collagen (COL7), the main constituent of anchoring fibrils for attaching the epidermis to the dermis. Persistent skin erosions frequently result in intractable ulcers in RDEB patients. Adipose-derived mesenchymal stromal cells (AD-MSCs) are easily harvested in large quantities and have low immunogenicity. Therefore, they are suitable for clinical use, including applications involving allogeneic cell transplantation. Keratinocyte-like cells transdifferentiated from AD-MSCs (KC-AD-MSCs) express more COL7 than undifferentiated AD-MSCs and facilitate skin wound healing with less contracture. Therefore, these cells can be used for skin ulcer treatment in RDEB patients. OBJECTIVE: We investigated whether KC-AD-MSCs transplantation ameliorated the RDEB phenotype severity in the grafted skin of a RDEB mouse model (col7a1-null) on the back of the immunodeficient mouse. METHODS: KC-AD-MSCs were intradermally injected into the region surrounding the skin grafts, and this procedure was repeated after 7 days. After a further 7-day interval, the skin grafts were harvested. RESULTS: Neodeposition of COL7 and generation of anchoring fibrils at the dermal-epidermal junction were observed, although experiments were based on qualitative. CONCLUSION: KC-AD-MSCs may correct the COL7 insufficiency, repair defective/reduced anchoring fibrils, and improve skin integrity in RDEB patients.


Subject(s)
Collagen Type VII , Disease Models, Animal , Epidermolysis Bullosa Dystrophica , Keratinocytes , Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells , Skin Transplantation , Collagen Type VII/genetics , Collagen Type VII/metabolism , Epidermolysis Bullosa Dystrophica/therapy , Epidermolysis Bullosa Dystrophica/pathology , Epidermolysis Bullosa Dystrophica/genetics , Animals , Humans , Keratinocytes/transplantation , Keratinocytes/metabolism , Mice , Mesenchymal Stem Cell Transplantation/methods , Mesenchymal Stem Cells/metabolism , Skin Transplantation/methods , Skin/pathology , Skin/cytology , Adipose Tissue/cytology , Cell Differentiation , Cells, Cultured , Wound Healing , Mice, Knockout
15.
Biomed Res Int ; 2024: 1041392, 2024.
Article in English | MEDLINE | ID: mdl-38933176

ABSTRACT

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.


Subject(s)
Coculture Techniques , Keratinocytes , Coculture Techniques/methods , Humans , Keratinocytes/cytology , Keratinocytes/metabolism , Keratinocytes/drug effects , Epidermis/metabolism , Fibroblasts/cytology , Fibroblasts/metabolism , Fibroblasts/drug effects , Cell Survival/drug effects , Skin/cytology , Skin/metabolism , Models, Biological
16.
J Mech Behav Biomed Mater ; 156: 106597, 2024 Aug.
Article in English | MEDLINE | ID: mdl-38810542

ABSTRACT

The skin, the outermost organ of the human body, is vital for sensing and responding to stimuli through mechanotransduction. It is constantly exposed to mechanical stress. Consequently, various mechanical therapies, including compression, massage, and microneedling, have become routine practices for skin healing and regeneration. However, these traditional methods require direct skin contact, restricting their applicability. To address this constraint, we developed shear wave stimulation (SWS), a contactless mechanical stimulation technique. The effectiveness of SWS was compared with that of a commercial compression bioreactor used on reconstructed skin at various stages of maturity. Despite the distinct stimulus conditions applied by the two methods, SWS yielded remarkable outcomes, similar to the effects of the compression bioreactor. It significantly increased the shear modulus of tissue-engineered skin, heightened the density of collagen and elastin fibers, and resulted in an augmentation of fibroblasts in terms of their number and length. Notably, SWS exhibited diverse effects in the low- and high-frequency modes, highlighting the importance of fine-tuning the stimulus intensity. These results unequivocally demonstrated the capability of SWS to enhance the mechanical functions of the skin in vitro, making it a promising option for addressing wound healing and stretch mark recovery.


Subject(s)
Skin , Skin/cytology , Humans , Stress, Mechanical , Tissue Engineering , Mechanical Phenomena , Biomechanical Phenomena , Fibroblasts/cytology , Animals , Collagen , Shear Strength , Elastin/metabolism
17.
Clin Transl Med ; 14(5): e1720, 2024 May.
Article in English | MEDLINE | ID: mdl-38778457

ABSTRACT

Melanocyte stem cells (MSCs), melanocyte lineage-specific skin stem cells derived from the neural crest, are observed in the mammalian hair follicle, the epidermis or the sweat gland. MSCs differentiate into mature melanin-producing melanocytes, which confer skin and hair pigmentation and uphold vital skin functions. In controlling and coordinating the homeostasis, repair and regeneration of skin tissue, MSCs play a vital role. Decreased numbers or impaired functions of MSCs are closely associated with the development and therapy of many skin conditions, such as hair graying, vitiligo, wound healing and melanoma. With the advancement of stem cell technology, the relevant features of MSCs have been further elaborated. In this review, we provide an exhaustive overview of cutaneous MSCs and highlight the latest advances in MSC research. A better understanding of the biological characteristics and micro-environmental regulatory mechanisms of MSCs will help to improve clinical applications in regenerative medicine, skin pigmentation disorders and cancer therapy. KEY POINTS: This review provides a concise summary of the origin, biological characteristics, homeostatic maintenance and therapeutic potential of cutaneous MSCs. The role and potential application value of MSCs in skin pigmentation disorders are discussed. The significance of single-cell RNA sequencing, CRISPR-Cas9 technology and practical models in MSCs research is highlighted.


Subject(s)
Melanocytes , Humans , Melanocytes/cytology , Homeostasis , Skin/cytology , Stem Cells/cytology , Stem Cells/metabolism , Animals , Cell Differentiation
18.
Molecules ; 29(9)2024 Apr 30.
Article in English | MEDLINE | ID: mdl-38731569

ABSTRACT

Skin wounds, leading to infections and death, have a huge negative impact on healthcare systems around the world. Antibacterial therapy and the suppression of excessive inflammation help wounds heal. To date, the application of wound dressings, biologics and biomaterials (hydrogels, epidermal growth factor, stem cells, etc.) is limited due to their difficult and expensive preparation process. Cinnamomum burmannii (Nees & T. Nees) Blume is an herb in traditional medicine, and its essential oil is rich in D-borneol, with antibacterial and anti-inflammatory effects. However, it is not clear whether Cinnamomum burmannii essential oil has the function of promoting wound healing. This study analyzed 32 main components and their relative contents of essential oil using GC-MS. Then, network pharmacology was used to predict the possible targets of this essential oil in wound healing. We first proved this essential oil's effects in vitro and in vivo. Cinnamomum burmannii essential oil could not only promote the proliferation and migration of skin stromal cells, but also promote M2-type polarization of macrophages while inhibiting the expression of pro-inflammatory cytokines. This study explored the possible mechanism by which Cinnamomum burmannii essential oil promotes wound healing, providing a cheap and effective strategy for promoting wound healing.


Subject(s)
Cinnamomum , Oils, Volatile , Skin , Wound Healing , Cinnamomum/chemistry , Oils, Volatile/chemistry , Oils, Volatile/pharmacology , Wound Healing/drug effects , Gas Chromatography-Mass Spectrometry , Cell Proliferation/drug effects , Cell Movement/drug effects , Skin/cytology , Skin/drug effects , Skin/injuries , Skin/microbiology , Stromal Cells/drug effects , Escherichia coli/drug effects , Staphylococcus aureus/drug effects , Animals , Mice , Humans , RAW 264.7 Cells , HaCaT Cells
19.
PLoS One ; 19(5): e0304602, 2024.
Article in English | MEDLINE | ID: mdl-38809935

ABSTRACT

This study aims to investigate if high-concentration HOCl fogging disinfection causes cytotoxicity and genotoxicity to cultured primary human skin fibroblasts. The cells were exposed to a dry fog of HOCl produced from solutions with a concentration of 300 ppm (5.72 mM) or 500 ppm (9.53 mM). After four times when fibroblasts were exposed to aerosolized HOCl at a concentration of 500 ppm for 9 minutes, significant cytotoxicity and genotoxicity effects were observed. Significant changes in the morphology of fibroblasts and cell death due to membrane disruption were observed, independent of the number of exposures. Flow cytometry analyses performed under these experimental conditions indicated a decrease in the number of cells with an intact cell membrane in the exposed samples compared to the sham samples, dropping to 49.1% of the total cells. Additionally, under the same conditions, the neutral comet assay results demonstrated significant DNA damage in the exposed cells. However, no analogous damages were found when the cells were exposed to aerosolized HOCl generated from a 300-ppm solution for 3 minutes, whether once or four times. Therefore, we have concluded that aerosolized HOCl in dry fog, with a concentration exceeding 300 ppm, can cause cytotoxic and genotoxic effects on human skin fibroblasts.


Subject(s)
DNA Damage , Fibroblasts , Hypochlorous Acid , Humans , Fibroblasts/drug effects , Hypochlorous Acid/toxicity , DNA Damage/drug effects , Cells, Cultured , Comet Assay , Skin/drug effects , Skin/cytology , Aerosols , Cell Survival/drug effects
20.
J Invest Dermatol ; 144(5): 936-949, 2024 May.
Article in English | MEDLINE | ID: mdl-38643988

ABSTRACT

The epidermis is the body's first line of protection against dehydration and pathogens, continually regenerating the outermost protective skin layers throughout life. During both embryonic development and wound healing, epidermal stem and progenitor cells must respond to external stimuli and insults to build, maintain, and repair the cutaneous barrier. Recent advances in CRISPR-based methods for cell lineage tracing have remarkably expanded the potential for experiments that track stem and progenitor cell proliferation and differentiation over the course of tissue and even organismal development. Additional tools for DNA-based recording of cellular signaling cues promise to deepen our understanding of the mechanisms driving normal skin morphogenesis and response to stressors as well as the dysregulation of cell proliferation and differentiation in skin diseases and cancer. In this review, we highlight cutting-edge methods for cell lineage tracing, including in organoids and model organisms, and explore how cutaneous biology researchers might leverage these techniques to elucidate the developmental programs that support the regenerative capacity and plasticity of the skin.


Subject(s)
Cell Differentiation , Cell Lineage , Humans , Animals , Skin/cytology , Stem Cells/cytology , Cell Proliferation , Regeneration/physiology
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