Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 7.116
Filtrar
1.
Toxicol Lett ; 320: 80-86, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31809884

RESUMO

In vitro cell culture experiments are highly important techniques to accelerate drug discovery, conduct safety testing and reduce the need for animal studies. Therefore, automatization may help to enhance the technical precision, reduce external (including operator's) influence on the data and thus improve reliability. Prior to application in scientific studies, validation of automated systems is absolutely necessary. In this study we present the validation of two combined automated pipetting systems to conduct toxicity studies in HaCaT cells consisting of cell seeding, noxious agent exposure and several assays to assess cell survival, apoptosis and interleukin production. After initial validation of pipetting accuracy, we compared homogeneity after automated seeding to plates seeded by expert laboratory technicians. Moreover, automated dispensing of a potentially unstable noxious agent was analyzed in terms of speed and consistency. We found a 2 % technical imprecision for the cell survival assay and 4.5-6 % for the other assays, bioluminescent and ELISA techniques. Thus, we could demonstrate the excellent technical precision of our assays. In a final step, we found that intraday variations, though acceptable, were much larger than technical variations and had to assume an intraday biological variability between different wells of the same experimental group.


Assuntos
Automação Laboratorial/normas , Substâncias para a Guerra Química/toxicidade , Gás de Mostarda/toxicidade , Técnicas de Cultura de Tecidos/normas , Testes de Toxicidade/normas , Apoptose/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Queratinócitos/patologia , Teste de Materiais , Necrose
2.
Toxicol Lett ; 319: 102-110, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31706006

RESUMO

Crizotinib is a multi-target receptor tyrosine kinase inhibitor which is of great importance for the management of ALK-rearranged non-small cell lung cancer (NSCLC) patients. Serious erythroderma and toxic epidermal necrolysis have been reported associated with crizotinib treatment. The underlying mechanisms have not been examined. In this study, we tested the toxicity of crizotinib on immortal human keratinocytes (HaCaT) and human primary keratinocytes. We found that crizotinib directly cause cytotoxic on these two cells, which could be the explanation of the clinical characteristic of pathology. Apoptosis was observed and Z-VAD-FMK, a pan-caspase inhibitor can almost totally reverse the apoptosis induction effect of crizotinib. However, mitochondrial dysfunction and DNA damage were not involved in crizotinib-induced apoptosis, indicating the intrinsic apoptosis pathway have no connection with this cutaneous toxicity. Further studies showed that crizotinib significantly increased cleaved-caspase-8, a signaling protein of extrinsic apoptosis pathway, in a concentration and time-dependent manner. Moreover, we found the targets of crizotinib were not involved in HaCaT cells apoptosis. Collectively, our findings first report keratinocytes apoptosis is the key cause of crizotinib-induced cutaneous toxicity. We also reveal crizotinib induce apoptosis through the extrinsic apoptosis pathway due to detected up-regulated cleaved-caspase-8. Meanwhile, the apoptosis is independent of mitochondrial dysfunction, DNA damage and related drug targets inhibition.


Assuntos
Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Crizotinibe/toxicidade , Dermatite Esfoliativa/induzido quimicamente , Queratinócitos/efeitos dos fármacos , Clorometilcetonas de Aminoácidos/farmacologia , Caspase 8/metabolismo , Inibidores de Caspase/farmacologia , Linhagem Celular , Dano ao DNA , Dermatite Esfoliativa/patologia , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Cultura Primária de Células , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
3.
J Photochem Photobiol B ; 202: 111704, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31743829

RESUMO

Ultraviolet B (UVB) induces inflammation and causes skin aging. The signs of skin aging, such as wrinkles, discolored spots, loss of skin moisture, and disruption of the skin barrier, are mostly caused by inflammatory signaling among various skin layers. The cells on the outermost surface of the skin are keratinocytes; these cells protect the skin against environmental stress and play an important role in immunomodulation by secreting cytokines in response to environmental stress. In the present study, we found that UVB activates STAT1 to mediate inflammatory signaling, yet STAT1 (S272) and STAT (Y702) shows different responses against UVB exposure. Anhua drak tea is a post-fermented dark tea produced in Anhua and Xinhua country in Hunan province of China. Treatment with 2S,3R-6-methoxycarbonylgallocatechin (MCGE), an epigallocatechin gallate derivative isolated from black tea (Anhua dark tea), effectively suppresses STAT1 activation and inflammatory cytokines, and activates Nrf2 pathway to protect cells from reactive oxygen species production in UVB exposed keratinocyte cells (HaCaT). Interestingly, the effects of MCGE were independent on MAPK signaling pathway. Moreover, MCGE regulates inflammatory cytokines in monocyte-keratinocyte (THP-1, HaCaT) co-culture and macrophage differentiation models. These results suggest that MCGE potentially can be used as a photoprotective agent against UVB-induced inflammatory responses.


Assuntos
Catequina/farmacologia , Protetores contra Radiação/farmacologia , Transdução de Sinais/efeitos dos fármacos , Chá/química , Raios Ultravioleta , Sítios de Ligação , Catequina/análogos & derivados , Catequina/química , Catequina/isolamento & purificação , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos da radiação , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Técnicas de Cocultura , Citocinas/metabolismo , Humanos , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Queratinócitos/efeitos da radiação , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/efeitos da radiação , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Estrutura Terciária de Proteína , Protetores contra Radiação/química , Protetores contra Radiação/isolamento & purificação , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT1/química , Fator de Transcrição STAT1/metabolismo , Transdução de Sinais/efeitos da radiação , Chá/metabolismo
4.
Life Sci ; 241: 117148, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31830478

RESUMO

Mitochondria are fascinating structures of the cellular compartments that generate energy to run the cells. However, inherent disorders of mitochondria due to diabetes can cause major disruption of metabolism that produces huge amount of reactive oxygen species (ROS). Here we study the elevated level of ROS provoked by high glucose (HG) environment triggered mitochondrial dysfunction, inflammatory response and apoptosis via stress signalling pathway in keratinocytes. Our results demonstrated that elevated glucose level in keratinoctes, increase the accumulations of ROS and decrease in cellular antioxidant capacities. Moreover, excess production of ROS was associated with mitochondrial dysfunction, characterized by loss of mitochondrial membrane potential (ΔΨm), increase in mitochondrial mass, alteration of mitochondrial respiratory complexes, cytochrome c (Cyt c) release, decrease in mitochondrial transcription factor A (TFAM) and increase in mitochondrial DNA (mtDNA) fragmentation. Damaged mtDNA escaped into the cytosol, where it engaged the activation of ERK1/2, PI3K/Akt, tuberin and mTOR via cGAS-STING leading to IRF3 activation. Pre-treatment of pharmacological inhibitors, ERK1/2 or PI3K/Akt suppressed the IRF3 activation. Furthermore, our results demonstrated that activation of IRF3 in HG environment coinciding with increased expression of inflammatory mediators. Excess production of ROS interfered with decreased in cell viability, increased lysosomal content and expression of FoxOs, leading to cell cycle deregulation and apoptosis. Pre-treatment of N-acetyl-l-cysteine (NAC) significantly reduced the HG-induced cell cycle deregulation and apoptosis in keratinocytes. In conclusion, increased oxidative stress underlies the decrease in antioxidant capacities and mitochondrial dysfunction in HG environment correlate with inflammation response and apoptosis via ERK1/2-PI3K/Akt-IRF3 pathway in keratinoctes.


Assuntos
Glucose/farmacologia , Queratinócitos/patologia , Mitocôndrias/patologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Pelados , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Edulcorantes/farmacologia
5.
Chemosphere ; 240: 124746, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31568946

RESUMO

The skin is one of the main organs exposed to airborne particulate matter (PM), which may contain various pollutants linked to a wide range of adverse health endpoints. In the present work, we analyzed the proinflammatory and oxidative effects of some PM components leading to inflammatory responses, cell proliferation or cell death. We investigated four redox-active chemicals, such as Cu (II) metal and quinones generated from polycyclic aromatic hydrocarbons (PAHs), i.e., 9,10 phenanthrenequinone and isomers 1,2 and 1,4 naphthoquinone. We performed in vitro biological tests on human keratinocyte (HaCaT) cells and also acellular assays based on the oxidation of dithiothreitol and ascorbic acid, antioxidants to assess the oxidative potential (OP). We found that treated keratinocytes showed increased activation of the redox-sensitive transcription factor NFκB and increased transcript levels of the NFκB-dependent gene IL8. Moreover, the treatment with Cu(II) and quinones increased the activities and the expression of genes involved in the redox response, SOD1 and GPX, suggesting that PM components induced cellular damage due to redox imbalances. Finally, we found alteration of the mitochondrial ultrastructure and increased apoptosis after 24 h of treatment. The results presented suggest that all of the analyzed pollutant components are able to modulate similar signal transduction pathways, resulting in activation of inflammatory processes in the skin, followed by oxidative damage. Altogether these observations indicate that exposure of skin to air pollutants modifies the redox equilibrium of keratinocytes, which could explain the increased skin damage observed in populations that live in high-pollution cities.


Assuntos
Poluentes Atmosféricos/toxicidade , Queratinócitos/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Material Particulado/toxicidade , Poluentes Atmosféricos/análise , Antioxidantes/metabolismo , Humanos , Metais/análise , Mitocôndrias/metabolismo , Oxirredução , Estresse Oxidativo/efeitos dos fármacos , Material Particulado/análise , Hidrocarbonetos Policíclicos Aromáticos/análise , Quinonas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Pele/efeitos dos fármacos
6.
An Acad Bras Cienc ; 91(4): e20181088, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31800700

RESUMO

Aged skin, featured with dryness and wrinkles, has received mounting attention due to its adverse influences on beauty. ß-Sitosterol and vermicularin are two common active ingredients of Thamnolia vermicularis (Sw.) Ach., a traditional Chinese medicine, of which the anti-aging effect has been discovered. Their protective performance against skin aging was assayed by co-culturing with skin cells in this work. Results showed that ß-sitosterol promoted the biosynthesis of hyaluronic acid by increasing the expression of hyaluronic acid synthases in fibroblasts and enhanced the expression of skin barrier functional proteins including aquaporin 3, loricrin, filaggrin and involucrin in keratinocytes, which conduced to the moisture retention within skin. Moreover, vermicularin might function as an anti-wrinkle agent by preventing the loss of collagen type I. Specifically, vermicularin reduced the amount of reactive oxygen species within hydrogen-peroxide-induced fibroblasts; together with suppressing the activation of mitogen-activated protein kinases, it could inhibit the production of matrix metalloproteinases-1. The present research will contribute to the development of the compounds as anti-aging ingredients for future applications in cosmetic formulations and functional food as well as promote further studies of raw materials containing alike compounds.


Assuntos
Fibroblastos/efeitos dos fármacos , Queratinócitos/efeitos dos fármacos , Extratos Vegetais/farmacologia , Sitosteroides/farmacologia , Envelhecimento da Pele/efeitos dos fármacos , Linhagem Celular , Humanos , Substâncias Protetoras/farmacologia
7.
Anticancer Res ; 39(12): 6673-6684, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31810932

RESUMO

BACKGROUND/AIM: The aim of the study was to evaluate the antitumor potential and combination effects of chemotherapeutic drugs. MATERIALS AND METHODS: The cytotoxicity of 20 drip-type classical and molecular-targeted anticancer drugs was examined against 4 human oral squamous cell carcinoma cell lines and 5 human oral normal mesenchymal and epithelial cells. Cell cycle progression was monitored by a cell sorter. Combination effect was evaluated by combination index. RESULTS: Most of the classical anticancer drugs showed much higher antitumor activity than molecular-targeted drugs, except bortezomib. Among 12 classical anticancer drugs, taxanes and gemsitabine showed the highest tumor-specificity (TS) and potency-selectivity expression (PSE) values, whereas platinum analogs showed the least TS value. Combination of two classical or a classical and a molecular-targeted drug showed mostly additive or antagonistic effect. 5-FU and cisplatin did not produce a subG1 population, but induced G2/M or G1/S arrest, regardless of the addition of cetuximab. Cetuximab, nibolumab and bortezomib showed potent keratinocyte toxicity. CONCLUSION: The present TS monitoring system may provide useful information for building up the treatment regimens of anticancer drugs.


Assuntos
Antineoplásicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma de Células Escamosas/tratamento farmacológico , Células Epiteliais/efeitos dos fármacos , Terapia de Alvo Molecular , Neoplasias Bucais/tratamento farmacológico , Bortezomib/farmacologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Cetuximab/farmacologia , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Fibroblastos/efeitos dos fármacos , Fluoruracila/farmacologia , Hormese , Humanos , Técnicas In Vitro , Queratinócitos/efeitos dos fármacos , Mucosa Bucal/citologia , Nivolumabe/farmacologia , Compostos de Platina/farmacologia , Taxoides/farmacologia
8.
J Photochem Photobiol B ; 201: 111653, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31710929

RESUMO

Autophagy is an important process for maintaining intracellular homeostasis. Our previous study demonstrated that autophagy was down-regulated in ultraviolet B (UVB)-irradiated keratinocytes. Raffinose is a natural oligosaccharide that serves as a novel activator of autophagy and as a balancing agent to regulate the diversity of environmental stress. However, whether raffinose balances ultraviolet stress through the autophagy activation pathway has yet to be established. In this study, we found that raffinose treatment inhibited the LDH release and trypan blue staining in UVB-challenged human keratinocytes cell line HaCaT but did not affect the cleavage of apoptotic markers Caspase-3 and PARP, as well as translocation into nucleus of other cell death markers Endonuclease G and AIF. Moreover, we confirmed that raffinose treatment enhanced autophagy flux in an MTOR-independent manner in HaCaT cells. Importantly, decrease of LC3-II turnover in UVB-irradiated keratinocytes could be rescued by raffinose treatment, indicating that raffinose treatment increased autophagy in UVB-irradiated HaCaT cells. Furthermore, the effect on cell death by raffinose was inhibited when autophagy was suppressed with either a small interfering RNA targeting ATG5 (siATG5) or autophagic inhibitor wortmannin. In conclusion, we demonstrated that raffinose increases MTOR-independent autophagy and reduces cell death in UVB-irradiated keratinocytes. Our study indicated that the natural agent raffinose presents the potential value in opposing photodamage.


Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Rafinose/farmacologia , Raios Ultravioleta , Apoptose/efeitos da radiação , Autofagia/efeitos da radiação , Proteína 5 Relacionada à Autofagia/antagonistas & inibidores , Proteína 5 Relacionada à Autofagia/genética , Proteína 5 Relacionada à Autofagia/metabolismo , Caspase 3/metabolismo , Linhagem Celular , Humanos , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo
9.
Fitoterapia ; 139: 104374, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31629867

RESUMO

Bioassay-guided fractionation of an extract of leaves and twigs of Elaeagnus umbellata led to the isolation of a serotonin derivative, N-[2-(5-hydroxyl-1H-indol-3-yl)ethyl]-butanamide (1), along with six flavonoid glycosides, kaempferol-3-O-ß-d-xylopyranosyl(1 → 2)-ß-d-galactopyranoside-7-O-α-l-rhamnopyranoside (2), kaempferol-3-O-ß-d-galactopyranoside-7-O-α-l-rhamnopyranoside (3), kaempferol-3-O-α-l-rhamnopyranosyl(1 → 6)-ß-d-galactopyranoside-7-O-α-l-rhamnopyranoside (4), kaempferol-3-O-ß-d-xylopyranosyl(1 → 2)-ß-d-galactopyranoside (5), kaempferol-3-O-rutinoside (6), and kaempferol-3-O-ß-d-glucopyranosyl(1 → 2)-ß-d-galactopyranoside-7-O-α-l-rhamnopyranoside (7). Their structures were elucidated using 1D/2D nuclear magnetic resonance spectroscopy and mass spectrometry. Compounds 1-6 were evaluated for their proliferative effects on HaCaT keratinocytes; 1-5 promoted keratinocyte proliferation dose dependently. Compounds 3 and 4 showed potent activities. These results suggest that the leaves and twigs of E. umbellata have wound healing and skin cell regeneration potentials.


Assuntos
Elaeagnaceae/química , Flavonoides/farmacologia , Glicosídeos/farmacologia , Queratinócitos/efeitos dos fármacos , Linhagem Celular , Flavonoides/isolamento & purificação , Glicosídeos/isolamento & purificação , Humanos , Quempferóis/isolamento & purificação , Quempferóis/farmacologia , Estrutura Molecular , Compostos Fitoquímicos/isolamento & purificação , Compostos Fitoquímicos/farmacologia , Folhas de Planta/química , República da Coreia
10.
Int J Nanomedicine ; 14: 7123-7139, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31564869

RESUMO

Background: Poly(amidoamine) (PAMAM) dendrimers are of considerable interest when used as a carrier for topical drugs for the skin, although little is known about their possible side effects. Therefore, our study was about the impact of 2nd and 3rd generation PAMAM dendrimers on human keratinocytes and fibroblasts cells. Methods: The effect of the tested compounds on collagen biosynthesis was determined using 5[3H]-proline incorporation bioassay. Morphological changes accompanying cell growth inhibition were observed using a confocal microscope. To evaluate the percentage of apoptotic/necrotic cells and the cell growth dynamic of apoptotic features, we performed Annexin V/PI double staining assay, assessed caspase activity, and performed cell cycle analysis by flow cytometry. The flow cytometry method was also used to determine the effect of dendrimers on pro-inflammatory cytokines (IL-6, IL-8 IL-1ß). Results: The obtained results showed that as the concentration and the generation of dendrimers increased, collagen biosynthesis decreased. We also observed abnormalities in cell differentiation, which may have caused disturbed secretion of pro-inflammatory cytokines. We found that dendrimers cause chronic inflammation which may cause adverse changes in the skin, ultimately- leading to apoptosis in the case of dendrimers in lower concentrations or necrosis at higher concentrations (especially 3rd generation dendrimers). In addition, the inflammatory path induced by the tested compounds was caused by damage in the mitochondria, which we observed as a significant decrease in the mitochondrial membrane potential. Conclusion: The results of our study showed that PAMAM dendrimers can cause disorders of cell proliferation and differentiation and may be the cause of cell cycle deregulation and chronic adverse inflammation.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Citocinas/metabolismo , Dendrímeros/farmacologia , Fibroblastos/metabolismo , Mediadores da Inflamação/metabolismo , Queratinócitos/metabolismo , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 8/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Colágeno/biossíntese , Dendrímeros/química , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fluorescência , Humanos , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Necrose
11.
Future Microbiol ; 14: 1087-1097, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31512522

RESUMO

Aim: In this work, mastoparan analog peptides from wasp venom were tested against Candida albicans and safety assays were performed using cell culture and model zebrafish. Materials & methods: Minimal inhibitory concentration was determined and toxicity was performed using human skin keratinocyte and embryo zebrafish. Also, permeation of peptides through embryo chorion was performed. Results: The peptides demonstrated anti-C. albicans activity, with low cytotoxicity and nonteratogenicity in Danio rerio. The compounds had different permeation through chorion, suggesting that this occurs due to modifications in their amino acid sequence. Conclusion: The results showed that the studied peptides can be used as structural study models for novel potential antifungal agents.


Assuntos
Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Peptídeos/farmacologia , Venenos de Vespas/farmacologia , Animais , Antifúngicos/administração & dosagem , Antifúngicos/efeitos adversos , Antifúngicos/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/patologia , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/administração & dosagem , Peptídeos e Proteínas de Sinalização Intercelular/efeitos adversos , Peptídeos e Proteínas de Sinalização Intercelular/toxicidade , Queratinócitos/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Peptídeos/administração & dosagem , Peptídeos/efeitos adversos , Peptídeos/toxicidade , Venenos de Vespas/administração & dosagem , Venenos de Vespas/efeitos adversos , Venenos de Vespas/toxicidade , Peixe-Zebra
12.
Int J Med Sci ; 16(8): 1116-1122, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31523174

RESUMO

This study investigated the anti-allergic effect of Poncirus trifoliata (L.) Raf. (PT) on human keratinocytic HaCaT cells in vitro and on 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis-like lesions in vivo. The release of TARC, MCP-1, IL-6 and IL-8 is increased by IFN-γ and TNF-α in HaCaT cells, and PT extract suppressed the increased production of TARC, MCP-1, IL-6, and IL-8. PT extract recovered the expression of filaggrin decreased by IFN-γ and TNF-α. in vivo experiment, PT administration decreased the skin severity score, thickening of the epidermis, movement of inflammatory cells into the dermis, and serum IgE level as compared to DNCB treatment. Moreover, the decrease of filaggrin and loricrin induced by DNCB treatment was recovered by PT administration. The levels of IL-4, IL-5, IL-13 and eotaxin in mouse splenocytes increased after treatment with concanavalin A, and the secretions of IL-4, IL-5, IL-13 and eotaxin were lower in the PT-treated group than in the DNCB group. These findings may indicate that PT is useful in drug development for the treatment of AD.


Assuntos
Dermatite Atópica/tratamento farmacológico , Queratinócitos/efeitos dos fármacos , Queratinócitos/patologia , Extratos Vegetais/farmacologia , Poncirus/química , Animais , Linhagem Celular , Quimiocina CCL11/metabolismo , Citocinas/metabolismo , Dermatite Atópica/induzido quimicamente , Dermatite Atópica/patologia , Dinitroclorobenzeno/toxicidade , Feminino , Humanos , Imunoglobulina E/sangue , Interferon gama/farmacologia , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos , Proteínas S100/metabolismo , Baço/citologia , Baço/metabolismo , Fator de Necrose Tumoral alfa/farmacologia
13.
Molecules ; 24(17)2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-31480216

RESUMO

Atopic dermatitis (AD) is a chronic inflammatory skin disease that can cause skin barrier function damage. Although co-incubation with docosahexaenoic acid (DHA) exerts a positive effect on deficient skin models, no studies have investigated the effects of topical treatment with DHA in an inflammatory reconstructed human epidermis (RHE) model. The effects of DHA on monolayer normal human epidermal keratinocyte (NHEK) cells were evaluated using cell counting kit-8 (CCK-8), real-time quantitative polymerase chain reaction (qPCR), and enzyme-linked immunosorbent assay (ELISA). The skin-related barrier function was assessed using hematoxylin-eosin (HE) staining, Western blot (WB), immunohistofluorescence (IF), and ELISA in normal and inflammatory RHE models. Docosahexaenoic acid upregulated filaggrin and loricrin expression at mRNA levels in addition to suppressing overexpression of tumor necrosis factor-α (TNF-α), interleukin-α (IL-1α), and interleukin-6 (IL-6) stimulated by polyinosinic-polycytidylic acid (poly I:C) plus lipopolysaccharide (LPS) (stimulation cocktail) in cultured NHEK cells. After topical treatment with DHA, cocktail-induced inflammatory characteristics of skin diseases, including barrier morphology, differentiation proteins, and thymic stromal lymphopoietin (TSLP) secretion, were alleviated in RHE models. Supplementation with DHA can improve related barrier function and have anti-inflammation effects in monolayer keratinocytes and RHE models, which indicates that DHA may have potential value for the treatment of inflammation-associated skin diseases.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Ácidos Docosa-Hexaenoicos/farmacologia , Epiderme/patologia , Inflamação/patologia , Queratinócitos/patologia , Modelos Biológicos , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Regulação para Baixo/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Humanos , Inflamação/genética , Mediadores da Inflamação/metabolismo , Queratinócitos/efeitos dos fármacos , Lipopolissacarídeos/farmacologia , Poli I-C/farmacologia
14.
Int J Mol Sci ; 20(17)2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-31480310

RESUMO

Particulate matter (PM), a widespread air pollutant, consists of a complex mixture of solid and liquid particles suspended in air. Many diseases have been linked to PM exposure, which induces an imbalance in reactive oxygen species (ROS) generated in cells, and might result in skin diseases (such as aging and atopic dermatitis). New techniques involving nanomedicine and nano-delivery systems are being rapidly developed in the medicinal field. Fullerene, a kind of nanomaterial, acts as a super radical scavenger. Lower water solubility levels limit the bio-applications of fullerene. Hence, to improve the water solubility of fullerene, while retaining its radical scavenger functions, a fullerene derivative, fullerenol C60(OH)36, was synthesized, to examine its biofunctions in PM-exposed human keratinocyte (HaCaT) cells. The PM-induced increase in ROS levels and expression of phosphorylated mitogen-activated protein kinase and Akt could be inhibited via fullerenol pre-treatment. Furthermore, the expression of inflammation-related proteins, cyclooxygenase-2, heme oxygenase-1, and prostaglandin E2 was also suppressed. Fullerenol could preserve the impaired state of skin barrier proteins (filaggrin, involucrin, repetin, and loricrin), which was attributable to PM exposure. These results suggest that fullerenol could act against PM-induced cytotoxicity via ROS scavenging and anti-inflammatory mechanisms, and the maintenance of expression of barrier proteins, and is a potential candidate compound for the treatment of skin diseases.


Assuntos
Poluição do Ar/prevenção & controle , Fulerenos/análise , Material Particulado/toxicidade , Água/química , Apoptose/efeitos dos fármacos , Linhagem Celular , Cidades , Fulerenos/química , Humanos , Inflamação/patologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Queratinócitos/ultraestrutura , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Modelos Biológicos , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Solubilidade
15.
Biol Res ; 52(1): 49, 2019 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-31492195

RESUMO

BACKGROUND: Psoriasis is a common and intractable skin disease affecting the physical and mental health of patients. The accumulation of ROS is involved in the pathogenesis of psoriasis and antioxidants are believed to be therapeutic. This study aimed to investigate the therapeutic efficacy of astilbin on ROS accumulation in psoriasis. RESULTS: The study showed that 50 µg/ml astilbin could inhibit the growth and reduce the accumulation of ROS in HaCaT cells stimulated by IL-17 and TNF-α. Astilbin could elevate the Nrf2 accumulation in the nuclei, eventually leading to the transcriptional activation of various antioxidant proteins and reducing the expression of VEGF. CONCLUSIONS: Our results collectively suggest that astilbin could induce Nrf2 nucleus translocation, which is contribute to reduce the ROS accumulation and VEGF expression, and inhibit the proliferation of HaCaT cells.


Assuntos
Flavonóis/administração & dosagem , Queratinócitos/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/efeitos dos fármacos , Psoríase/tratamento farmacológico , Espécies Reativas de Oxigênio/metabolismo , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Interleucina-17/metabolismo , Queratinócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fator 2 Relacionado a NF-E2/metabolismo , Psoríase/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo
16.
Int J Mol Sci ; 20(16)2019 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-31398894

RESUMO

Claudin-1 (CLDN1) is expressed in the tight junction (TJ) of the skin granular layer and acts as a physiological barrier for the paracellular transport of ions and nonionic molecules. Ultraviolet (UV) and oxidative stress may disrupt the TJ barrier, but the mechanism of and protective agents against this effect have not been clarified. We found that UVB and hydrogen peroxide (H2O2) caused the internalization of CLDN1 and increased the paracellular permeability of lucifer yellow, a fluorescent marker, in human keratinocyte-derived HaCaT cells. Therefore, the mechanism of mislocalization of CLDN1 and the protective effect of an ethanol extract of Brazilian green propolis (EBGP) were investigated. The UVB- and H2O2-induced decreases in CLDN1 localization were rescued by EBGP. H2O2 decreased the phosphorylation level of CLDN1, which was also rescued by EBGP. Wild-type CLDN1 was distributed in the cytosol after treatment with H2O2, whereas T191E, its H2O2-insensitive phosphorylation-mimicking mutant, was localized at the TJ. Both protein kinase C activator and protein phosphatase 2A inhibitor rescued the H2O2-induced decrease in CLDN1 localization. The tight junctional localization of CLDN1 and paracellular permeability showed a negative correlation. Our results indicate that UVB and H2O2 could induce the elevation of paracellular permeability mediated by the dephosphorylation and mislocalization of CLDN1 in HaCaT cells, which was rescued by EBGP. EBGP and its components may be useful in preventing the destruction of the TJ barrier through UV and oxidative stress.


Assuntos
Claudina-1/metabolismo , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Estresse Oxidativo , Própole/farmacologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Claudina-1/genética , Endocitose/efeitos dos fármacos , Expressão Gênica , Humanos , Peróxido de Hidrogênio/metabolismo , Mutação , Fosforilação , Transporte Proteico , Espécies Reativas de Oxigênio/metabolismo , Junções Íntimas/metabolismo
17.
Int J Nanomedicine ; 14: 5449-5475, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31409998

RESUMO

Purpose: We created and evaluated an enhanced topical delivery system featuring a combination of highly skin-permeable growth factors (GFs), quercetin (QCN), and oxygen; these synergistically accelerated re-epithelialization and granulation tissue formation of/in diabetic wounds by increasing the levels of GFs and antioxidants, and the oxygen partial pressure, at the wound site. Methods: To enhance the therapeutic effects of exogenous administration of GFs for the treatment of diabetic wounds, we prepared highly skin-permeable GF complexes comprised of epidermal growth factor (EGF), insulin-like growth factor-I (IGF-I), platelet-derived growth factor-A (PDGF-A), and basic fibroblast growth factor (bFGF), genetically attached, via the N-termini, to a low-molecular-weight protamine (LMWP) to form LMWP-EGF, LMWP-IGF-I, LMWP-PDGF-A, and LMWP-bFGF, respectively. Furthermore, quercetin (QCN)- and oxygen-carrying 1-bromoperfluorooctane (PFOB)-loaded nanoemulsions (QCN-NE and OXY-PFOB-NE) were developed to improve the topical delivery of QCN and oxygen, respectively. After confirming the enhanced penetration of LMWP-GFs, QCN-NE, and oxygen delivered from OXY-PFOB-NE across human epidermis, we evaluated the effects of combining LMWP-GFs, QCN-NE, and OXY-PFOB-NE on proliferation of keratinocytes and fibroblasts, and the chronic wound closure rate of a diabetic mouse model. Results: The optimal ratios of LMWP-EGF, LMWP-IGF-I, LMWP-PDGF-A, LMWP-bFGF, QCN-NE, and OXY-PFOB-NE were 1, 1, 0.02, 0.02, 0.2, and 60, respectively. Moreover, a Carbopol hydrogel containing LMWP-GFs, QCN-NE, and OXY-PFOB-NE (LMWP-GFs/QCN-NE/OXY-PFOB-NE-GEL) significantly improved scratch-wound recovery of keratinocytes and fibroblasts in vitro compared to that afforded by hydrogels containing each component alone. LMWP-GFs/QCN-NE/OXY-PFOB-NE-GEL significantly accelerated wound-healing in a diabetic mouse model, decreasing wound size by 54 and 35% compared to the vehicle and LMWP-GFs, respectively. Conclusion: LMWP-GFs/QCN-NE/OXY-PFOB-NE-GEL synergistically accelerated the healing of chronic wounds, exerting both rapid and prolonged effects.


Assuntos
Diabetes Mellitus/patologia , Hidrogéis/química , Fator de Crescimento Insulin-Like I/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Oxigênio/metabolismo , Quercetina/farmacologia , Absorção Cutânea , Cicatrização/efeitos dos fármacos , Animais , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Colágeno/biossíntese , Modelos Animais de Doenças , Emulsões/química , Fator de Crescimento Epidérmico/farmacologia , Epiderme/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/farmacologia , Humanos , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Peso Molecular , Nanopartículas/química , Nanopartículas/ultraestrutura , Octanos/química , Fator de Crescimento Derivado de Plaquetas/farmacologia , Protaminas/química , Absorção Cutânea/efeitos dos fármacos
18.
Int J Nanomedicine ; 14: 5033-5050, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31371945

RESUMO

Background: Repairs to deep skin wounds continue to be a difficult issue in clinical practice. A promising approach is to fabricate full-thickness skin substitutes with functions closely similar to those of the natural tissue. For many years, a three-dimensional (3D) collagen hydrogel has been considered to provide a physiological 3D environment for co-cultivation of skin fibroblasts and keratinocytes. This collagen hydrogel is frequently used for fabricating tissue-engineered skin analogues with fibroblasts embedded inside the hydrogel and keratinocytes cultivated on its surface. Despite its unique biological properties, the collagen hydrogel has insufficient stiffness, with a tendency to collapse under the traction forces generated by the embedded cells. Methods: The aim of our study was to develop a two-layer skin construct consisting of a collagen hydrogel reinforced by a nanofibrous poly-L-lactide (PLLA) membrane pre-seeded with fibroblasts. The attractiveness of the membrane for dermal fibroblasts was enhanced by coating it with a thin nanofibrous fibrin mesh. Results: The fibrin mesh promoted the adhesion, proliferation and migration of the fibroblasts upwards into the collagen hydrogel. Moreover, the fibroblasts spontaneously migrating into the collagen hydrogel showed a lower tendency to contract and shrink the hydrogel by their traction forces. The surface of the collagen was seeded with human dermal keratinocytes. The keratinocytes were able to form a basal layer of highly mitotically-active cells, and a suprabasal layer. Conclusion: The two-layer skin construct based on collagen hydrogel with spontaneously immigrated fibroblasts and reinforced by a fibrin-coated nanofibrous membrane seems to be promising for the construction of full-thickness skin substitute.


Assuntos
Colágeno/farmacologia , Fibrina/farmacologia , Hidrogéis/farmacologia , Membranas Artificiais , Nanofibras/química , Poliésteres/farmacologia , Pele Artificial , Animais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Derme/citologia , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Humanos , Recém-Nascido , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Ratos
19.
Molecules ; 24(17)2019 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-31454971

RESUMO

Quercus mongolica Fisch. ex Ledeb. (QM) has been used as an oriental traditional medicine to relieve hemorrhoids, fever, and enteritis. We screened the inhibitory activities of the extracts and compounds (1-6) isolated from QM on the production of inflammatory cytokines and chemokines to evaluate their anti-inflammatory activities. Further, we evaluated the expression levels of cytokines, chemokines, and immune factors on pedunculagin (PC, 1), which was selected from isolated compounds (1-6) because of its potential anti-inflammation effect. Additionally, we evaluated whether the inflammation mitigation effects of PC (1) following UVB exposure in keratinocytes occurred because of nuclear factor (NF)-κB and signal transducer and activator of transcription (STAT)/Janus kinase (JAK) activation. PC (1) remarkably suppressed interleukin (IL)-6, IL-10, IL-13, and monocyte chemoattractant protein-1 (MCP-1) mRNA expression and reduced the mRNA expression level of Cyclooxygenase-2 (COX-2) and also reduced the phosphorylation of p38 mitogen-activated protein kinases (p38), c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) in a concentration-dependent manner.


Assuntos
Anti-Inflamatórios/farmacologia , Queratinócitos/citologia , Fenóis/farmacologia , Quercus/química , Taninos/farmacologia , Anti-Inflamatórios/química , Linhagem Celular , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Avaliação Pré-Clínica de Medicamentos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/efeitos da radiação , Lipopolissacarídeos/efeitos adversos , NF-kappa B , Fenóis/química , Fosforilação , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Transdução de Sinais/efeitos dos fármacos , Taninos/química , Raios Ultravioleta/efeitos adversos
20.
Int J Mol Sci ; 20(16)2019 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-31426336

RESUMO

Reactive oxygen species (ROS) are generated from diverse cellular processes or external sources such as chemicals, pollutants, or ultraviolet (UV) irradiation. Accumulation of radicals causes cell damage that can result in degenerative diseases. Antioxidants remove radicals by eliminating unpaired electrons from other molecules. In skin health, antioxidants are essential to protect cells from the environment and prevent skin aging. (-)-Epigallocatechin-3-(3″-O-methyl) gallate (3″Me-EGCG) has been found in limited oolong teas or green teas with distinctive methylated form, but its precise activities have not been fully elucidated. In this study, we examined the antioxidant roles of 3″Me-EGCG in keratinocytes (HaCaT cells). 3″Me-EGCG showed scavenging effects in cell and cell-free systems. Under H2O2 exposure, 3″Me-EGCG recovered cell viability and increased the expression of heme oxygenase 1 (HO-1). Under ultraviolet B (UVB) and sodium nitroprusside (SNP) exposure, 3″Me-EGCG protected keratinocytes and regulated the survival protein AKT1. By regulating the AKT1/NF-κB pathway, 3″Me-EGCG augmented cell survival and proliferation in HaCaT cells. These results indicate that 3″Me-EGCG exhibits antioxidant properties, resulting in cytoprotection against various external stimuli. In conclusion, our findings suggest that 3″Me-EGCG can be used as an ingredient of cosmetic products or health supplements.


Assuntos
Antioxidantes/farmacologia , Catequina/análogos & derivados , Citoproteção/efeitos dos fármacos , Ácido Gálico/análogos & derivados , Queratinócitos/efeitos dos fármacos , Antioxidantes/química , Catequina/química , Catequina/farmacologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Citoproteção/efeitos da radiação , Ácido Gálico/química , Ácido Gálico/farmacologia , Humanos , Queratinócitos/citologia , Queratinócitos/metabolismo , Queratinócitos/efeitos da radiação , Protetores contra Radiação/química , Protetores contra Radiação/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Raios Ultravioleta/efeitos adversos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA