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1.
Adv Gerontol ; 32(4): 509-515, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31800177

RESUMO

The aim of this work was to examine the content of Yes-associated protein (YAP) in fibroblasts and blood microvessels of human dermis from the development until deep aging (from 20 weeks of pregnancy until 85 years old), and defining of a role of YAP in age-dependent changes in the number of fibroblasts and blood microvessels in the dermis. YAP, proliferating cells nuclear antigen (PCNA), endothelial cells marker CD31 were detected with indirect immunohistochemical technique. Results showed that portion of fibroblasts with positive staining for YAP in the dermis is decreased from 20 weeks of pregnancy to 40 years old. Percent of YAP positive fibroblasts in dermis is increased sufficiently since 41 years old until 60-85 years old group. The content of YAP in blood microvessels in the human dermis is decreased sufficiently from 20 weeks of pregnancy until 40 years old. Age-related changes in the content of YAP in fibroblasts and blood microvessels is not statistically associated with an age-related decrease in total number and percent of PCNA positive fibroblasts, the number of blood vessels in the dermis.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Células Endoteliais , Envelhecimento da Pele , Fatores de Transcrição , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Envelhecimento , Derme/citologia , Derme/metabolismo , Feminino , Fibroblastos/metabolismo , Humanos , Pessoa de Meia-Idade , Gravidez , Envelhecimento da Pele/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
2.
Dermatol Surg ; 45(12): 1580-1584, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31765337

RESUMO

BACKGROUND: Little literature exits on the mechanism of action of implanted polymethylmethacrylate (PMMA) filler. OBJECTIVE: To characterize PMMA-induced dermal extracellular matrix production in the skin. MATERIALS AND METHODS: Single-center, open-label prospective study in healthy volunteers undergoing removal of redundant skin was injected intradermally and subdermally with PMMA dermal filler (Bellafill). Punch biopsies were harvested over a time course and evaluated for the deposition of collagen-3 and procollagen-1, proteoglycans and elastin using immunohistochemistry. Blinded histopathologic readings were performed by a dermatopathologist to characterize the nature of the dermal extracellular matrix findings. RESULTS: Normal inflammatory infiltrate was exhibited at all timepoints after PMMA injection with an influx of fibroblasts and new vasculature. Tissue proteoglycans were noted within the injectate beginning at Week 1 and persisted through the study end point. Increased collagen Type 3 was evident following the first week after injection, peaked at Month 2 and diminished through Months 3 through 6. Procollagen-1 was noted at Month 1 and continued to increase in intensity and organization through the study end point (6 months). Elastin staining was inconclusive. Polymethylmethacrylate microspheres remained within the initial injection area and became encapsulated within new collagen fibers. The growth and pattern of new connective tissue mimicked a normal wound healing response. CONCLUSION: Polymethylmethacrylate-collagen gel filler stimulates collagen-3 and procollagen-1 when injected into human skin. This combination of neocollagenesis followed by microencapsulation of PMMA microspheres in the new tissue provides for long-lasting results.


Assuntos
Colágeno Tipo III/biossíntese , Colágeno Tipo I/biossíntese , Colágeno/administração & dosagem , Preenchedores Dérmicos/administração & dosagem , Derme/efeitos dos fármacos , Polimetil Metacrilato/administração & dosagem , Adulto , Biópsia , Derme/citologia , Derme/metabolismo , Elastina/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Voluntários Saudáveis , Humanos , Injeções Intradérmicas , Microesferas , Pessoa de Meia-Idade , Estudos Prospectivos , Proteoglicanas/metabolismo
3.
Mater Sci Eng C Mater Biol Appl ; 105: 110063, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31546412

RESUMO

Artificial dermal scaffolds, which are made of natural or synthetic materials, can improve new blood vessel formation, cell migration and cell proliferation after being implanted into wounds, and they degrade slowly, playing an important role in dermal reconstruction and scar inhibition, finally achieving the goal of wound healing and functional reconstruction. Although these scaffolds have been widely used in clinical applications, biomaterial-associated infection is a deficiency or even a life-threatening problem that must be addressed, as it greatly affects the survival of the scaffolds. The gallium ion (Ga3+) is a novel metallic antimicrobial whose broad-spectrum antimicrobial properties against most bacteria encountered in burn wound infections have been confirmed, and it has been proposed as a promising candidate to prevent implant-associated infections. In this study, a gallium-loaded antimicrobial artificial dermal scaffold was successfully prepared by gallium ions and a collagen solution. The characterization results showed a porous structure with pore sizes ranging from 50 to 150 µm and a large porosity value of 97.4%. The enzymatic degradation rate in vitro was 19 and 28% after 12 and 24 h, respectively. In vitro antimicrobial testing revealed that the 1 h antibacterial rate against Staphylococcus aureus and Pseudomonas aeruginosa was close to 90%, which indicated its great antimicrobial activity. The results of the cytological evaluation showed slight effect on cell proliferation, with a relative growth rate (RGR) value of 80% and great cytocompatibility with cultured cells according to laser scanning confocal microscopy (LSCM) and scanning electron microscope (SEM). Furthermore, the successful prevention of wound infections in SD rats was confirmed with an in vivo antimicrobial evaluation, and the artificial dermal scaffolds also demonstrated great biocompatibility. This gallium-loaded antimicrobial artificial dermal scaffold exerted excellent antimicrobial activity and great biosafety, warranting further research for future clinical applications.


Assuntos
Anti-Infecciosos/química , Derme , Gálio/química , Pseudomonas aeruginosa/crescimento & desenvolvimento , Staphylococcus aureus/crescimento & desenvolvimento , Tecidos Suporte/química , Animais , Linhagem Celular , Derme/química , Derme/metabolismo , Derme/microbiologia , Camundongos , Porosidade
4.
Mater Sci Eng C Mater Biol Appl ; 104: 109841, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31499993

RESUMO

Effective removal of cellular components while retaining extracellular matrix (ECM) proteins is the ultimate goal of decellularization. The aim of this study is to produce a decellularized ECM with highly preserved ECM proteins and to determine the effect of isopropanol as a decellularization solvent on the characteristics of the decellularized porcine skin. Two different protocols were used for porcine skin decellularization. Protocol 1 consisted of Triton-X and sodium dodecyl sulfate (SDS) in water while protocol 2 consisted of Triton-X and SDS in 70% isopropanol. After decellularization, DNA components decreased significantly in protocol 2 with lower amount of lipid content and higher ECM proteins such as collagen (92.91 ±â€¯9.02 µg/mg sample), α-elastin (142.32 ±â€¯6.74 µg/mg sample) and sulfated glycosaminoglycan (sGAG; 7.44 ±â€¯1.30 µg/mg sample) compared with protocol 1 ECM. Higher amount of vascular endothelial growth factor (VEGF; 11.26 ±â€¯0.44 pg/mg sample) content was quantified in protocol 2 compared with protocol 1 while higher trace amount of bone morphogenic protein 2 (BMP-2; 0.28 ±â€¯0.04 pg/mg sample) was also observed in protocol 2 compared with protocol 1. Protocol 2 ECM did not significantly affect the cell viability and exhibited no cytotoxicity when exposed to three different cell lines: L929 fibroblast cells, MC3T3-E1 pre-osteoblast cells, and rat mesenchymal stem cells (BMSC). Subcutaneous implantation after 7 and 21 days revealed higher cell infiltration in protocol 2 ECM and enhanced neovascularization. Isopropanol/surfactants proved to be effective in cell and lipid removal during decellularization while preserving the higher amount of ECM proteins.


Assuntos
Derme/metabolismo , Matriz Extracelular/metabolismo , Engenharia Tecidual/métodos , Animais , Linhagem Celular , Colágeno/metabolismo , DNA/metabolismo , Matriz Extracelular/ultraestrutura , Glicosaminoglicanos/metabolismo , Camundongos , Coelhos , Ratos Sprague-Dawley , Suínos
5.
Biomed Res Int ; 2019: 2626374, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31534956

RESUMO

Wound healing is a complex process regulated by multiple signals and consisting of several phases known as haemostasis, inflammation, proliferation, and remodelling. Keratinocytes, endothelial cells, macrophages, and fibroblasts are the major cell populations involved in wound healing process. Hypoxia plays a critical role in this process since cells sense and respond to hypoxic conditions by changing gene expression. This study assessed the in vitro expression of 77 genes involved in angiogenesis, metabolism, cell growth, proliferation and apoptosis in human keratinocytes (HaCaT), microvascular endothelial cells (HMEC-1), differentiated macrophages (THP-1), and dermal fibroblasts (HDF). Results indicated that the gene expression profiles induced by hypoxia were cell-type specific. In HMEC-1 and differentiated THP-1, most of the genes modulated by hypoxia encode proteins involved in angiogenesis or belonging to cytokines and growth factors. In HaCaT and HDF, hypoxia mainly affected the expression of genes encoding proteins involved in cell metabolism. This work can help to enlarge the current knowledge about the mechanisms through which a hypoxic environment influences wound healing processes at the molecular level.


Assuntos
Apoptose , Proliferação de Células , Derme , Regulação da Expressão Gênica , Neovascularização Fisiológica , Cicatrização , Hipóxia Celular , Derme/irrigação sanguínea , Derme/metabolismo , Células Endoteliais/metabolismo , Fibroblastos/metabolismo , Humanos , Queratinócitos/metabolismo , Macrófagos/metabolismo , Células THP-1
6.
Oxid Med Cell Longev ; 2019: 6146942, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31531185

RESUMO

Background: Nanofat can protect against ultraviolet B- (UVB-) induced damage in nude mice. Fat extract (FE) is a cell-free fraction isolated from nanofat that is enriched with a variety of growth factors. Objective: To determine whether FE can protect against UVB-induced photoaging in cultured dermal fibroblasts and in nude mice. Method: For the in vitro study, human dermal skin fibroblasts were pretreated with FE 24 h prior to UVB irradiation. Generation of reactive oxygen species (ROS) was analyzed immediately following irradiation, while cell cycle analysis was performed 24 h after UVB irradiation. Senescence-associated ß-galactosidase (SA-ß-gal) expression, cell proliferation, and expression of glutathione peroxidase 1 (GPX-1), catalase, superoxide dismutase-1 (SOD-1), SOD-2, and collagen type 1 (COL-1) were investigated 72 h after UVB irradiation. For the in vivo study, the dorsal skin of nude mice was irradiated with UVB and mice were then treated with FE for 8 weeks. The thickness of the dermis, capillary density, and apoptotic cells in skin tissue sections were investigated after treatment. The expression of GPX-1, catalase, SOD-2, SOD-1, and COL-1 in the tissue was also measured. Result: FE significantly increased cell proliferation and protected cells against UVB-induced cell death and cell cycle arrest. FE reduced ROS and the number of aged cells induced by UVB irradiation. FE promoted the expression of COL-1 and GPX-1 in cultured dermal fibroblasts. FE treatment of UVB-irradiated skin increased dermal thickness and capillary density, decreased the number of apoptotic cells, and promoted the expression of COL-1 and GPX-1. Conclusion: FE protects human dermal fibroblasts and the skin of nude mice from UVB-induced photoaging through its antioxidant, antiapoptotic, and proangiogenic activities.


Assuntos
Tecido Adiposo/química , Misturas Complexas/farmacologia , Derme/metabolismo , Fibroblastos/metabolismo , Envelhecimento da Pele , Raios Ultravioleta/efeitos adversos , Animais , Senescência Celular/efeitos dos fármacos , Senescência Celular/efeitos da radiação , Misturas Complexas/química , Derme/patologia , Feminino , Fibroblastos/patologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Oxirredutases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Envelhecimento da Pele/efeitos dos fármacos , Envelhecimento da Pele/efeitos da radiação
7.
ACS Appl Mater Interfaces ; 11(37): 33535-33547, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31369233

RESUMO

Engineering bioscaffolds for improved cutaneous tissue regeneration remains a healthcare challenge because of the increasing number of patients suffering from acute and chronic wounds. To help address this problem, we propose to utilize alfalfa, an ancient medicinal plant that contains antibacterial/oxygenating chlorophylls and bioactive phytoestrogens, as a building block for regenerative wound dressings. Alfalfa carries genistein, which is a major phytoestrogen known to accelerate skin repair. The scaffolds presented herein were built from composite alfalfa and polycaprolactone (PCL) nanofibers with hydrophilic surface and mechanical stiffness that recapitulate the physiological microenvironments of skin. This composite scaffold was engineered to have aligned nanofibrous architecture to accelerate directional cell migration. As a result, alfalfa-based composite nanofibers were found to enhance the cellular proliferation of dermal fibroblasts and epidermal keratinocytes in vitro. Finally, these nanofibers exhibited reproducible regenerative functionality by promoting re-epithelialization and granulation tissue formation in both mouse and human skin, without requiring additional proteins, growth factors, or cells. Overall, these findings demonstrate the potential of alfalfa-based nanofibers as a regenerative platform toward accelerating cutaneous tissue repair.


Assuntos
Derme , Queratinócitos , Medicago sativa/química , Nanocompostos , Nanofibras , Cicatrização/efeitos dos fármacos , Linhagem Celular , Derme/lesões , Derme/metabolismo , Derme/patologia , Humanos , Queratinócitos/metabolismo , Queratinócitos/patologia , Nanocompostos/química , Nanocompostos/uso terapêutico , Nanofibras/química , Nanofibras/uso terapêutico , Poliésteres/química
8.
Int J Mol Sci ; 20(16)2019 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-31412620

RESUMO

Two mesenchymal zinc transporters, ZIP7 and ZIP13, play critical roles in dermal development. ZIP7 and ZIP13 are the closest among the conserved mammalian zinc transporters. However, whether their functions are complementary remains a controversial issue. In the present study, we found that the expression of ZIP13, but not ZIP7, is elevated by transforming growth factor beta (TGF-ß) treatment, indicating that TGF-ß-mediated ZIP13 amplification is crucial for collagen production during dermal development. Genome-wide gene expression analysis revealed that ~26% of genes are dependent on either ZIP7 or ZIP13, which is greater than the ~17% of genes dependent on both of them. ZIP7 depletion induces endoplasmic reticulum (ER) stress in mesenchymal stem cells, resulting in significant inhibition of fibrogenic differentiation. However, ZIP13 depletion does not induce ER stress. Though both ZIP7 and ZIP13 contain traditional ER signal peptides for their intracellular localization, their distributions are distinct. When ZIP7 and ZIP13 are coexpressed, their localizations are distinct; ZIP7 is located on the ER, but ZIP13 is located on both the ER and Golgi, indicating that only ZIP13 is a zinc gatekeeper on the Golgi. Our data illustrate that the different actions of ZIP7 and ZIP13 are crucial for dermal development.


Assuntos
Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Derme/embriologia , Derme/metabolismo , Organogênese/genética , Zinco/metabolismo , Animais , Retículo Endoplasmático/metabolismo , Estresse do Retículo Endoplasmático , Genoma , Estudo de Associação Genômica Ampla , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Knockout , Modelos Biológicos
9.
Cell Prolif ; 52(5): e12668, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31379046

RESUMO

OBJECTIVES: Reproducing human hair follicles in vitro is often limited by various reasons such as the lack of a systematic approach to culture distinct hair follicle cell types to reproduce their spatial relationship. Here, we reproduce hair follicle-like constructs resembling the spatial orientation of different cells in vivo, to study the role of keratinocytes in maintaining cellular compartmentalization among hair follicle-related cells. MATERIALS AND METHODS: Dermal papilla (DP) cells, HaCaT keratinocytes and human dermal fibroblast (HDF) cells were seeded sequentially into three-dimensional (3D) microwells fabricated from polyethylene glycol diacrylate hydrogels. Quantitative polymerase chain reaction was used to compare inductive gene expression of 3D and two-dimensional (2D) DP. DP and HaCaT cells were transfected with green fluorescent protein and red fluorescent protein lentivirus, respectively, to enable cell visualization using confocal microscopy. RESULTS: The 3D DP cultures showed significantly enhanced expression of essential DP genes as compared 2D cultures. Core-shell configurations containing keratinocytes forming the outer shell and DP forming the core were observed. Migratory polarization was mediated by cell-cell interaction between the keratinocytes and HDF cells, while preserving the aggregated state of the DP cells. CONCLUSIONS: Keratinocytes may play a role in maintaining compartmentalization between the DP and the surrounding HDF residing in the dermis, and therefore maintains the aggregative state of the DP cells, necessary for hair follicle development and function.


Assuntos
Técnicas de Cultura de Células/métodos , Derme/citologia , Fibroblastos/citologia , Queratinócitos/citologia , Células Cultivadas , Derme/metabolismo , Fibroblastos/metabolismo , Humanos , Hidrogéis/química , Queratinócitos/metabolismo , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Microscopia Confocal
10.
Int J Pharm ; 569: 118547, 2019 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-31377408

RESUMO

The stratum corneum is the main barrier to transdermal drug delivery which has previously resulted in mathematical modelling of solute transport in the skin being primarily directed at this skin layer. However, for topical treatment and skin toxicity studies, the concentration in the epidermis and dermis is important and needs to be modelled mathematically. Hitherto, mathematical models for viable skin layers typically simplified the clearance of solute by blood, either assuming sink condition at the top of the skin capillary loops or assuming a distributed clearance in the dermis. This paper is an attempt to develop a physiologically based mathematical model of drug transport in the viable skin. It incorporates explicit modelling of the capillary loops within the dermis and employs COMSOL Multiphysics® software to model the transport in three dimensions. Previously derived simplified models were compared to the results from this new numerical model. The results of this comparison showed that the simplified model reasonably described the average concentration in the viable skin layers when parameters of the models were chosen appropriately. When the recruitment of the capillary loops in the dermis was full and the top of capillary loops was at a depth of 100µm, the effective depth to place a sink condition in the simpler models was found to be at 150µm. However, when there was only partial recruitment of the capillaries, the effective depth increased to 180µm. The presented modelling is also essential for determining a transdermal flux when the stratum corneum barrier is compromised by such methods as microporation, application of chemical enhancers or microneedles.


Assuntos
Derme/metabolismo , Epiderme/metabolismo , Modelos Biológicos , Transporte Biológico
11.
Int J Pharm ; 570: 118633, 2019 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-31437563

RESUMO

Avobenzone (AVO), oxybenzone (OXY), and octyl methoxycinnamate (OMC), are widely used UV filters. The aim of this study was to investigate the effect of incorporation in mesoporous silica (SBA-15) on their cutaneous deposition and permeation. Stick formulations containing "free" and "incorporated" UV filters (SF1 and SF2, respectively) were prepared and characterized with respect to their physicochemical, thermal, and functional properties. Cutaneous delivery experiments using porcine skin with quantification by UHPLC-MS/MS, demonstrated that skin deposition of AVO and OXY after application of SF2 for 6 and 12 h was significantly lower than that from SF1 at each time-point (Student t-test, p < 0.05): e.g. OXY permeation across the skin was 30-, 12- and 1.5-fold lower after 6, 12 and 24 h, respectively, following application of SF2. Cutaneous biodistribution profiles of AVO and OXY to 800 µm evidenced a significant decrease in the amounts in the viable epidermis and dermis. In contrast, deposition of the more lipophilic OMC was not significantly different (p ˃ 0.05). In vitro photoprotective efficacy results demonstrated that adsorption/entrapment of UV filters enhanced the sun protection factor by 94%. In conclusion, SBA-15, an innovative mesoporous material, increased photoprotection by UV filters while reducing their cutaneous penetration and transdermal permeation.


Assuntos
Derme/metabolismo , Epiderme/metabolismo , Dióxido de Silício/sangue , Absorção Cutânea/efeitos dos fármacos , Raios Ultravioleta/efeitos adversos , Administração Cutânea , Animais , Benzofenonas/química , Cromatografia Líquida de Alta Pressão/métodos , Cinamatos/química , Portadores de Fármacos/química , Composição de Medicamentos/métodos , Estabilidade de Medicamentos , Propiofenonas/química , Dióxido de Silício/química , Fator de Proteção Solar/métodos , Protetores Solares/administração & dosagem , Protetores Solares/química , Suínos , Espectrometria de Massas em Tandem/métodos , Distribuição Tecidual/fisiologia
12.
Int J Nanomedicine ; 14: 5381-5396, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31409994

RESUMO

Background: Tacrolimus (TCR), also known as FK-506, is a biopharmaceutics classification system (BCS) class II drug that is insoluble in water because of its high log P values. After dermal application, TCR remains in the stratum corneum and passes through the skin layers with difficulty. Purpose: The objectives of this study were to develop and evaluate solid lipid nanoparticles (SLNs) with thermosensitive properties to improve penetration and retention. Methods: We prepared TCR-loaded thermosensitive solid lipid nanoparticles (TCR-SLNs) with different types of surfactants on the shell of the particle, which conferred the advantages of enhancing skin permeation and distribution. We also characterized them from a physic point of view and performed in vitro and in vivo evaluations. Results: The TCR contained in the prepared TCR-SLN was in an amorphous state and entrapped in the particles with a high loading efficiency. The assessment of ex vivo skin penetration using excised rat dorsal skin showed that the TCR-SLNs penetrated to a deeper layer than the reference product (0.1% Protopic®). In addition, the in vivo skin penetration test demonstrated that TCR-SLNs delivered more drug into deeper skin layers than the reference product. FT-IR images also confirmed drug distribution of TCR-SLNs into deeper layers of the skin. Conclusion: These results revealed the potential application of thermosensitive SLNs for the delivery of difficult-to-permeate, poorly water-soluble drugs into deep skin layers.


Assuntos
Derme/metabolismo , Lipídeos/química , Nanopartículas/química , Tacrolimo/farmacologia , Temperatura , Administração Cutânea , Animais , Varredura Diferencial de Calorimetria , Derme/efeitos dos fármacos , Liberação Controlada de Fármacos , Irritantes/toxicidade , Nanopartículas/ultraestrutura , Tamanho da Partícula , Coelhos , Ratos Sprague-Dawley , Absorção Cutânea/efeitos dos fármacos , Testes Cutâneos , Espectroscopia de Infravermelho com Transformada de Fourier , Tensoativos/química , Difração de Raios X
13.
PLoS Pathog ; 15(8): e1007993, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31465513

RESUMO

Chikungunya virus (CHIKV) is an arthritogenic alphavirus that acutely causes fever as well as severe joint and muscle pain. Chronic musculoskeletal pain persists in a substantial fraction of patients for months to years after the initial infection, yet we still have a poor understanding of what promotes chronic disease. While replicating virus has not been detected in joint-associated tissues of patients with persistent arthritis nor in various animal models at convalescent time points, viral RNA is detected months after acute infection. To identify the cells that might contribute to pathogenesis during this chronic phase, we developed a recombinant CHIKV that expresses Cre recombinase (CHIKV-3'-Cre). CHIKV-3'-Cre replicated in myoblasts and fibroblasts, and it induced arthritis during the acute phase in mice. Importantly, it also induced chronic disease, including persistent viral RNA and chronic myositis and synovitis similar to wild-type virus. CHIKV-3'-Cre infection of tdTomato reporter mice resulted in a population of tdTomato+ cells that persisted for at least 112 days. Immunofluorescence and flow cytometric profiling revealed that these tdTomato+ cells predominantly were myofibers and dermal and muscle fibroblasts. Treatment with an antibody against Mxra8, a recently defined host receptor for CHIKV, reduced the number of tdTomato+ cells in the chronic phase and diminished the levels of chronic viral RNA, implicating these tdTomato+ cells as the reservoir of chronic viral RNA. Finally, isolation and flow cytometry-based sorting of the tdTomato+ fibroblasts from the skin and ankle and analysis for viral RNA revealed that the tdTomato+ cells harbor most of the persistent CHIKV RNA at chronic time points. Therefore, this CHIKV-3'-Cre and tdTomato reporter mouse system identifies the cells that survive CHIKV infection in vivo and are enriched for persistent CHIKV RNA. This model represents a useful tool for studying CHIKV pathogenesis in the acute and chronic stages of disease.


Assuntos
Artrite Experimental/virologia , Febre de Chikungunya/virologia , Vírus Chikungunya/patogenicidade , Derme/patologia , Fibroblastos/patologia , Músculo Esquelético/patologia , RNA Viral/metabolismo , Animais , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Febre de Chikungunya/metabolismo , Vírus Chikungunya/genética , Derme/metabolismo , Derme/virologia , Modelos Animais de Doenças , Fibroblastos/metabolismo , Fibroblastos/virologia , Camundongos , Camundongos Endogâmicos C57BL , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patologia , Fibras Musculares Esqueléticas/virologia , Músculo Esquelético/metabolismo , Músculo Esquelético/virologia , RNA Viral/genética , Replicação Viral
14.
BMC Genet ; 20(1): 70, 2019 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-31455210

RESUMO

BACKGROUND: Hu sheep, a unique Chinese breed with high reproductive performance, are also well known for their rare white lambskin in China. The quality of lambskin is affected by hair follicles, and dermal papilla cells are an important component of hair follicles that plays a key role in hair follicle growth and development. This study helps elucidate the effect of miR-148a and miR-10a on hair follicle growth and development. RESULTS: Based on the results of gene chip and high-throughput sequencing, bone morphogenetic protein 7 (BMP7) was used as a research object. Bioinformatics analysis and the dual-luciferase reporter system indicated that, along with Western blot and quantitative real-time polymerase chain reaction (qRT-PCR) that miR-148a and miR-10a target relationships with BMP7. BMP7 was the target gene both for miR-148a and miR-10a by the dual-luciferase reporter system and Western blot. Hu sheep dermal papilla cells were successfully isolated and purified, and after transfecting miR-148a/miR-10a mimics and inhibitors into dermal papilla cells, a Cell Counting Kit-8 (CCK-8) was used to determine that miR-148a/miR-10a inhibited the proliferation of Hu sheep dermal papilla cells. In addition, after the overexpression of miR-148a, the expression levels of Smad3 (P < 0.05), Smad6 (P < 0.05), Smad4 (P < 0.01), and Smad5 (P < 0.01) were significantly higher than those of the control groups. After the inhibition of miR-148a, the expression levels of Smad3 (P < 0.05), Smad4 (P < 0.05), and TGF-ß (P < 0.01) were significantly lower than those of the control groups. After the overexpression of miR-10a, the expression levels of Smad1 (P < 0.01), Smad2 (P < 0.05), Smad4 (P < 0.01), Smad5 (P < 0.01), and TGF-ß (P < 0.05) were significantly lower than those of the control groups. After the inhibition of miR-10a, the expression levels of Smad1 (P < 0.01) and Smad2 (P < 0.05) were significantly lower than those of the control groups. CONCLUSIONS: These results revealed the target relationship between miR-148a, miR-10a and BMP7, and the effect of miR-148a and miR-10a on the proliferation of dermal papilla cells. They will provide the basis for a follow-up study on how miR-148a, and miR-10a mediate BMP7 regulation of hair follicle growth and development.


Assuntos
Derme/metabolismo , MicroRNAs/genética , Ovinos/genética , Regiões 3' não Traduzidas , Animais , Células Cultivadas , Perfilação da Expressão Gênica , Genes Reporter , Humanos
15.
Lipids Health Dis ; 18(1): 164, 2019 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-31443723

RESUMO

BACKGROUND: Recent technical advances in the extraction of dermal interstitial fluid (ISF) have stimulated interest in using this rather unexploited biofluid as an alternative to blood for detection and prediction of disease. However, knowledge about the presence of useful biomarkers for health monitoring in ISF is still limited. In this study, we characterized the lipidome of human suction blister fluid (SBF) as a surrogate for pure ISF and compared it to that of plasma. METHODS: Plasma and SBF samples were obtained from 18 healthy human volunteers after an overnight fast. Total lipids were extracted and analyzed by liquid chromatography-tandem mass spectrometry. One hundred ninety-three lipid species covering 10 complex lipid classes were detected and quantified in both plasma and SBF using multiple reaction monitoring. A fraction of the lipid extract was subjected to alkaline transesterification and fatty acid methyl esters were analyzed by gas chromatography-mass spectrometry. RESULTS: The total concentration of lipids in SBF was 17% of the plasma lipid concentration. The molar fraction of lipid species within lipid classes, as well as total fatty acids, showed a generally high correlation between plasma and SBF. However, SBF had larger fractions of lysophospholipids and diglycerides relative to plasma, and consequently less diacylphospholipids and triglycerides. Principal component analysis revealed that the interindividual variation in SBF lipid profiles was considerably larger than the within-subject variation between plasma and SBF. CONCLUSIONS: Plasma and SBF lipid profiles show high correlation and SBF could be used interchangeably with blood for the analysis of major lipids used in health monitoring.


Assuntos
Diglicerídeos/análise , Líquido Extracelular/química , Ácidos Graxos Ômega-3/análise , Ácidos Graxos Insaturados/análise , Lisofosfolipídeos/análise , Triglicerídeos/análise , Adulto , Cromatografia Líquida , Derme/química , Derme/metabolismo , Líquido Extracelular/metabolismo , Jejum , Feminino , Cromatografia Gasosa-Espectrometria de Massas , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Componente Principal , Espectrometria de Massas em Tandem
16.
Cell Physiol Biochem ; 53(1): 242-257, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31313540

RESUMO

BACKGROUND/AIMS: Excessive exposure to UV radiation negatively affects the human skin, characterized by photo-damage (premature aging & carcinogenesis). UV-B radiation causes about 90% of non-melanoma skin cancers by damaging de-oxy ribonucleic acids (DNA). We have previously reported that UV-B radiation induces skin photodamage through oxidative & Endoplasmic Reticulum (ER) stresses and Glycyrrhizic acid (GA), a natural triterpene, protects skin cells against such stresses. UV-B radiation elicits signalling cascade by activation of proteins involved in sensing, signalling, and repair process of DNA damage. In this study, we explored the effects & mechanisms of Glycyrrhizic acid (GA) against UV-B -induced photodamage using a well established cellular model. METHODS: We used primary human dermal fibroblasts as a cellular model. The cells were cultured in the presence or absence of GA for 3,6, & 24 h. Effect of UV-B was assessed by examining cell viability, cell morphology, oxidative stress, ER stress, DNA damage & cellular autophagy levels through biochemical assays, microscopy & protein expression studies. RESULTS: In this study, we have determined the effect of GA on autophagy mediated DNA damage response system as the main mechanism in preventing photodamage due to UV-B -irradiation to primary human dermal fibroblasts (HDFs). GA treatment to UV-B exposed HDFs, significantly inhibited cell death, oxidative & ER stress responses, prevented Cyclobutane Pyrimidine dimer (CPD) DNA adduct formation, and DNA fragmentation via modulation of UV-B induced autophagic flux. Present results showed that GA treatment quenched reactive oxygen species (ROS), relieved ER stress response, improved autophagy (6 hr's post-UV-B -irradiation) and prevented UV-B induced DNA damage. CONCLUSION: The present study links autophagy induction by GA as the main mechanism in the prevention of DNA damage and provides a mechanistic basis for the photoprotective effect of GA and suggests that GA can be potentially developed as a promising agent against UV-B induced skin photo-damage.


Assuntos
Autofagia , Derme/metabolismo , Fibroblastos/metabolismo , Ácido Glicirrízico/farmacologia , Estresse Oxidativo , Raios Ultravioleta/efeitos adversos , Autofagia/efeitos dos fármacos , Autofagia/efeitos da radiação , Células Cultivadas , Derme/patologia , Fibroblastos/patologia , Humanos , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/efeitos da radiação
17.
PLoS One ; 14(7): e0218536, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31306414

RESUMO

BACKGROUND: Stem cell therapy is the next generation a well-established technique. Cell therapy with mesenchymal stem cells (MSC) has been demonstrated to enhance wound healing in diabetic mice, at least partly due to improved growth factor production. However, it is unclear whether MSC can biomechanically affect wound closure. Utilizing the well-established cell-populated collagen gel contraction model we investigated the interactions between MSC and the extracellular matrix. METHODS: Murine fetal liver-derived Mesenchymal Stem Cells (MSCs) or fetal Dermal Fibroblasts (DFs) were cultured in cell-populated collagen gels (CPCGs). The effect of cell density, conditioned media, growth factors (TGF-B1, FGF, PDGF-BB), cytoskeletal disruptors (colchicine, cytochalasin-D), and relative hypoxia on gel contraction were evaluated. Finally, we also measured the expression of integrin receptors and some growth factors by MSCs within the contracting gels. RESULTS: Our results show that at different densities, MSCs induced a higher gel contraction compared to DFs. Higher cell density resulted in faster and more complete contraction of CPCGs. Cytoskeletal inhibitors either inhibited or prevented MSC-mediated contraction in a dose dependent fashion. Growth factors, conditioned media from both MSC and DF, and hypoxia all influenced CPCG contraction. DISCUSSION: The results suggest that MSCs are capable of directly contributing to wound closure through matrix contraction, and they are more effective than DF. In addition, this study demonstrates the importance of how other factors such as cell concentration, cytokines, and oxygen tension can provide potential modulation of therapies to correct wound healing impairments.


Assuntos
Colágeno/metabolismo , Derme/metabolismo , Feto/metabolismo , Fibroblastos/metabolismo , Fígado/metabolismo , Células-Tronco Mesenquimais/metabolismo , Animais , Terapia Baseada em Transplante de Células e Tecidos , Células Cultivadas , Derme/citologia , Feto/citologia , Fibroblastos/citologia , Fígado/citologia , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Transgênicos , Especificidade de Órgãos/fisiologia
18.
Mater Sci Eng C Mater Biol Appl ; 103: 109829, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31349406

RESUMO

The novel biodegradable films of chloroacetated natural rubber/polyvinyl alcohol (CNR/PVA) (55/45 wt%) non-woven nanofiber films encapsulated with kaolin and starch (2.5 and 5 wt%) were produced successfully by green electrospinning technique. The effect of fillers with different content on the physical, chemical, mechanical, biocompatibility and biodegradation properties of CNR/PVA nanofiber films were investigated. The higher crystallinity obtained in CNR/PVA encapsulate with 2.5 wt% kaolin and nanofibers were formed with the maximum diameter distribution and mean value of 40-160 nm and 94.15 ±â€¯54.19 nm respectively. DSC and DMA revealed the kaolin can improve the interfacial adhesion between CNR and PVA and contribute to enhancing the chemical interactions. The mechanical properties improved upon encapsulation of starch and kaolin and more favourable nanofibers with smaller diameter obtained using kaolin rather than starch. The cytotoxicity results revealed the viability of the prepared nanofiber films with human dermal fibroblast cell. Furthermore, the incorporation of starch and kaolin accelerated the degradation rate and the highest enzymatic degradation obtained with 2.5 wt% of starch. The prepared nanofiber films have the potential to be applied for the skin tissue engineering scaffold applications.


Assuntos
Derme/metabolismo , Fibroblastos/metabolismo , Teste de Materiais , Membranas Artificiais , Nanofibras/química , Álcool de Polivinil/química , Borracha/química , Tecidos Suporte/química , Linhagem Celular , Derme/citologia , Fibroblastos/citologia , Humanos , Caulim/química
19.
Photodermatol Photoimmunol Photomed ; 35(5): 360-368, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31166622

RESUMO

BACKGROUND/PURPOSE: Ultraviolet (UV) A (315-400 nm) is the UV light that most frequently reaches the Earth's surface and can penetrate the epidermis through to the dermis, causing various issues, including skin aging and skin cancer. The results of our previous studies have shown that the flavonoid monomer cyanidin-3-o-glucoside (C3G) can effectively inhibit primary human dermal fibroblast (HDF) oxidative damage and apoptosis caused by UVA radiation. Many flavonoids can regulate the level of autophagy. However, whether C3G inhibits UVA-induced oxidative damage to primary HDFs by regulating autophagy levels remains unclear. METHODS AND RESULTS: In this study, we used different doses (0-12 J/cm2 ) of UVA to irradiate cells and showed that the expression levels of autophagy-related gene 5 (Atg5) and microtubule-associated protein 1 light chain 3 (LC3)-II in primary HDFs first increased and then decreased. The expression of Atg5 and LC3-II was significantly decreased under 12 J/cm2 (light-damage model). C3G increased the levels of Atg5 and LC3-II. Primary HDFs were pretreated with C3G, followed by treatment with the autophagy inhibitor 3-methyladenine (3-MA) after 12 J/cm2 UVA irradiation. The inhibitory effects of C3G on morphological changes, oxidative damage, and apoptosis in primary HDFs induced by UVA were significantly decreased. CONCLUSION: C3G can inhibit UVA-induced damage to primary HDFs by inducing autophagy. These results provide a theoretical basis for the application of natural compounds to resist light damage to the skin in the future.


Assuntos
Antocianinas/farmacologia , Autofagia , Derme/metabolismo , Fibroblastos/metabolismo , Glucosídeos/farmacologia , Raios Ultravioleta/efeitos adversos , Regulação para Cima , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Autofagia/efeitos dos fármacos , Autofagia/efeitos da radiação , Células Cultivadas , Derme/patologia , Fibroblastos/patologia , Humanos , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/efeitos da radiação
20.
Immunity ; 50(6): 1482-1497.e7, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31201094

RESUMO

The skin comprises tissue macrophages as the most abundant resident immune cell type. Their diverse tasks including resistance against invading pathogens, attraction of bypassing immune cells from vessels, and tissue repair require dynamic specification. Here, we delineated the postnatal development of dermal macrophages and their differentiation into subsets by adapting single-cell transcriptomics, fate mapping, and imaging. Thereby we identified a phenotypically and transcriptionally distinct subset of prenatally seeded dermal macrophages that self-maintained with very low postnatal exchange by hematopoietic stem cells. These macrophages specifically interacted with sensory nerves and surveilled and trimmed the myelin sheath. Overall, resident dermal macrophages contributed to axon sprouting after mechanical injury. In summary, our data show long-lasting functional specification of macrophages in the dermis that is driven by stepwise adaptation to guiding structures and ensures codevelopment of ontogenetically distinct cells within the same compartment.


Assuntos
Diferenciação Celular/imunologia , Vigilância Imunológica , Macrófagos/imunologia , Regeneração Nervosa , Pele/imunologia , Pele/inervação , Animais , Animais Recém-Nascidos , Biomarcadores , Receptor 1 de Quimiocina CX3C/metabolismo , Derme/citologia , Derme/imunologia , Derme/metabolismo , Imunofenotipagem , Macrófagos/metabolismo , Camundongos , Pele/citologia
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