RESUMO
Skin fibrosis is a hallmark of a wide array of dermatological diseases which can greatly impact the patients' quality of life. Galectin-3 (GAL-3) has emerged as a central regulator of tissue fibrosis, playing an important pro-fibrotic role in numerous organs. Various studies are highlighting its importance as a skin fibrotic diseases biomarker; however, there is a need for further studies that clarify its role. This paper aims to ascertain whether the expression of GAL-3 is increased in relevant in vitro and in vivo models of skin fibrosis. We studied the role of GAL-3 in vitro using normal human dermal fibroblasts (NHDF) and fibrocytes. In addition, we used a skin fibrosis murine model (BALB/c mice) and human biopsies of healthy or keloid tissue. GAL-3 expression was analyzed using real time PCR, Western blot and immunostaining techniques. We report a significantly increased expression of GAL-3 in NHDF and fibrocytes cell cultures following stimulation with transforming growth factor ß1 (TGFß1). In vivo, GAL-3 expression was increased in a murine model of systemic sclerosis and in human keloid biopsies. In sum, this study underlines the involvement of GAL-3 in skin fibrosis using several models of the disease and highlights its role as a relevant target.
Assuntos
Queloide , Escleroderma Sistêmico , Dermatopatias , Humanos , Camundongos , Animais , Galectina 3/genética , Galectina 3/metabolismo , Modelos Animais de Doenças , Qualidade de Vida , Fibrose , Fibroblastos/metabolismo , Dermatopatias/metabolismo , Escleroderma Sistêmico/patologia , Pele/metabolismo , Queloide/metabolismoRESUMO
BACKGROUND: Photothermolysis of unwanted hair depends on the presence of melanin in the hair follicle as the chromophore, but is not effective in patients with non-pigmented, melanin-sparse hair shafts and follicles. This split-scalp, double-blind study was to monitor the efficacy of melanin bound in nanosomes to inject exogenous melanin into the hair follicles thus potentiating successful photothermolysis.
MATERIAL AND METHODS: Twelve patients, phototypes II-III, with white or very fair hair, were treated with a compound containing melanin encapsulated in nanosomes (Melaser®) together with a fluorescent marker. Two equal 6 cm² areas were marked on each side of the occiput of the subjects. The compound was applied to a randomly selected experimental side on each patient (area A), and a saline solution applied in the same manner to the contralateral control side (area B). Penetration of the melanin into the hair follicle was assessed using optical and fluorescence microscopy. Also, condition of hair structure was checked in vivo after standard laser settings used for epilation.
RESULTS: A slight transient erythema was observed in those areas where the compound was applied with some perifollicular edema. No such effects were noticed in those areas where saline solution was applied. No persistent complications such as scarring, hypo- or hyperpigmentation were observed in any of the experimental or control areas. Under fluorescence microscopy, the hair structures in the areas to which the compound had been applied showed a clear melanin deposit confirmed by the immunofluorescence intensity, which was highest at 2 hours after application. By optical microscopy, external melanin was deposited in hair follicles. Tests with standard settings for epilation were efficacious in damaging melanin-marked white hair.
CONCLUSION: This study strongly suggests the safety and efficacy of the application of nanosomes encapsulating melanin for the introduction of melanin into hair follicles. Changes noticed in the hair structure compromising its viability indicated potential application of this external melanin marker for white hair photoepilation.
J Drugs Dermatol. 2016;15(5):615-625.
Assuntos
Folículo Piloso/efeitos dos fármacos , Remoção de Cabelo/métodos , Melaninas/administração & dosagem , Nanosferas/administração & dosagem , Fosfolipídeos/administração & dosagem , Transtornos da Pigmentação/tratamento farmacológico , Idoso , Animais , Corantes/administração & dosagem , Corantes/química , Decapodiformes , Portadores de Fármacos/administração & dosagem , Portadores de Fármacos/química , Feminino , Cor de Cabelo/efeitos dos fármacos , Folículo Piloso/patologia , Humanos , Masculino , Melaninas/química , Microscopia de Fluorescência/métodos , Pessoa de Meia-Idade , Nanosferas/química , Agulhas , Fosfolipídeos/química , Transtornos da Pigmentação/patologia , Pigmentos Biológicos/administração & dosagem , Pigmentos Biológicos/química , Estudos Prospectivos , Distribuição AleatóriaRESUMO
BACKGROUND: Glucocorticoid functions are markedly impaired in patients with chronic obstructive pulmonary disease (COPD). The phosphodiesterase 4 inhibitor roflumilast N-oxide (RNO) is the active metabolite of roflumilast approved as a treatment to reduce the risk of exacerbations in patients with severe COPD. OBJECTIVE: We sought to characterize the differential effects of RNO versus corticosteroids and their potential additive/synergistic effect in neutrophils from patients with COPD, thus providing scientific rationale for the combination of roflumilast with corticosteroids in the clinic. METHODS: Peripheral blood neutrophils were isolated from patients with COPD (n = 32), smokers (n = 7), and healthy nonsmokers (n = 25). Levels of IL-8, matrix metallopeptidase 9 (MMP-9), and biomarkers of glucocorticoid resistance were determined by using ELISA and RT-PCR. Neutrophils were incubated with dexamethasone (0.1 nmol/L to 1 µmol/L), RNO (0.1 nmol/L to 1 µmol/L), or the combination of 1 nmol/L RNO plus 10 nmol/L DEX and stimulated with LPS (1 µg/mL) or cigarette smoke extract 5%; levels of IL-8, MMP-9, and other biomarkers were measured at the end of the incubation period. RESULTS: Peripheral neutrophils from patients with COPD showed a primed phenotype with an increased basal release of IL-8 and MMP-9 and expressed a corticosteroid resistance molecular profile characterized by an increase in phosphoinositide 3-kinase δ, macrophage migration inhibitory factor, and glucocorticoid receptor ß expression and a decrease in HDAC activity and mitogen-activated protein kinase phosphatase 1 expression. RNO demonstrated robust anti-inflammatory effects on neutrophils from patients with COPD, reversing their resistance to corticosteroids. The combination of RNO and dexamethasone showed additive/synergistic effects, which were consistent with the reversal of corticosteroid-resistant molecular markers by RNO. CONCLUSION: RNO reverses corticosteroid resistance and shows strong anti-inflammatory effects alone or in combination with corticosteroids on neutrophils from patients with COPD.
Assuntos
Corticosteroides/farmacologia , Aminopiridinas/farmacologia , Benzamidas/farmacologia , Dexametasona/farmacologia , Resistência a Medicamentos/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Inibidores da Fosfodiesterase 4/farmacologia , Idoso , Biomarcadores/metabolismo , Misturas Complexas/isolamento & purificação , Misturas Complexas/farmacologia , Ciclopropanos/farmacologia , Sinergismo Farmacológico , Feminino , Expressão Gênica/efeitos dos fármacos , Histona Desacetilases/metabolismo , Humanos , Interleucina-8/metabolismo , Oxirredutases Intramoleculares/metabolismo , Lipopolissacarídeos/farmacologia , Fatores Inibidores da Migração de Macrófagos/metabolismo , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Pessoa de Meia-Idade , Fosfatases da Proteína Quinase Ativada por Mitógeno/metabolismo , Neutrófilos/metabolismo , Neutrófilos/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Cultura Primária de Células , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologia , Nicotiana/químicaRESUMO
BACKGROUND: Pulmonary hypertension in idiopathic pulmonary fibrosis (IPF) is indicative of a poor prognosis. Recent evidence suggests that tetrahydrobiopterin (BH4), the cofactor of nitric oxide synthase (NOS), is involved in pulmonary hypertension and that pulmonary artery endothelial-to-mesenchymal transition (EnMT) may contribute to pulmonary fibrosis. However, the role of BH4 in pulmonary remodelling secondary to pulmonary fibrosis is unknown. This study examined the BH4 system in plasma and pulmonary arteries from patients with IPF as well as the antiremodelling and antifibrotic effects of the BH4 precursor sepiapterin in rat bleomycin-induced pulmonary fibrosis and in vitro EnMT models. METHODS: BH4 and nitrotyrosine were measured by high-performance liquid chromatography and ELISA, respectively. Expression of sepiapterin reductase (SPR), GTP cyclohydrolase 1 (GCH-1), endothelial NOS (eNOS) and inducible NOS (iNOS) were measured by quantitative PCR and immunohistochemistry. RESULTS: BH4 plasma levels were downregulated in patients with IPF compared with controls while nitrites, nitrates and nitrotyrosine were upregulated. GCH-1 and eNOS were absent in pulmonary arteries of patients with IPF; however, iNOS expression increased while SPR expression was unchanged. In rats, oral sepiapterin (10 mg/kg twice daily) attenuated bleomycin-induced pulmonary fibrosis, mortality, vascular remodelling and pulmonary hypertension by increasing rat plasma BH4, decreasing plasma nitrotyrosine and increasing vascular eNOS and GCH-1 expression. Both transforming growth factor ß1 and endothelin-1 induced EnMT by decreasing BH4 and eNOS expression. In vitro administration of sepiapterin increased endothelial BH4 and inhibited EnMT in human pulmonary artery endothelial cells. CONCLUSIONS: Targeting the BH4 synthesis 'salvage pathway' with sepiapterin may be a new therapeutic strategy to attenuate pulmonary hypertension in IPF.
Assuntos
Biopterinas/análogos & derivados , Hipertensão Pulmonar/patologia , Artéria Pulmonar/patologia , Fibrose Pulmonar/patologia , Idoso , Oxirredutases do Álcool/metabolismo , Animais , Biopterinas/sangue , Biopterinas/metabolismo , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Ensaio de Imunoadsorção Enzimática , Feminino , GTP Cicloidrolase/metabolismo , Humanos , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/metabolismo , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Artéria Pulmonar/metabolismo , Fibrose Pulmonar/complicações , Fibrose Pulmonar/metabolismo , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
Liposomes have been intensively investigated as carriers for different applications in dermatology and cosmetics. Ascorbic acid has potent antioxidant and anti-inflammatory properties preventing photodamage of keratinocytes; however, due to its instability and low skin penetration, an appropriate carrier is mandatory to obtain desirable efficacy. The present work investigates the ability of a specific ascorbate phosphatidylcholine (PC) liposome to overcome the barrier of the stratum corneum and deliver the active agent into the dermis to prevent photodamage. Abdominal skin from ten patients was used. Penetration of PC liposomes was tested ex vivo in whole skin, epidermis, and dermis by means of fluorescein and sodium ascorbate. Histology and Franz diffusion cells were used to monitor the percutaneous absorption. Ultraviolet (UV)-high performance liquid chromatography was used to analyze diffusion of sodium ascorbate through the different skin layers, while spectrofluorimetry and fluorescent microscopy were used for fluorescein monitoring. UVA/UVB irradiation of whole skin was applied to analyze the antioxidant capacity by Trolox assay and anti-inflammatory effects by tumor necrosis factor alpha and interleukin 1 beta enzyme-linked immunoassay. PC liposomal formulation improved skin penetration of fluorescein and ascorbate. Fluorescein PC liposomes showed better diffusion through epidermis than dermis while ascorbate liposomes showed better diffusion through the dermis than the epidermis. Ascorbate PC liposomes showed preventive antioxidant and anti-inflammatory properties on whole human skin irradiated with UVA/UVB. In summary, ascorbate PC liposomes penetrate through the epidermis and allow nonstable hydrophilic active ingredients reach epidermis and dermis preventing skin photodamage.
RESUMO
Encapsulation of chemicals in liposomes and microneedling are currently used techniques to enhance the penetration of several substances through skin and hair. In this study, we apply a liposomal melanin-fluorescein compound to an ex vivo model of human skin, using a new electrical microneedling device (Nanopore turbo roller). The product was applied by hand massage (A) or with the assistance of the electrical roller for 2 minutes (B). An additional test was performed free of product and with only the E-roller (C). Histological changes and product absorption were evaluated by optical and fluorescent microscopy 60 and 90 minutes after the treatment. Site B showed larger deposits of melanin-fluorescein at superficial and deep levels of hair structures in comparison to site A. Light, epidermal deposits of the melanin-fluorescein complex were also observed. Sites B and C showed a significant widening (47%) of the follicular infundibulum which could explain the increased penetration of the formulation. Microneedling also removed the scales and sebum residues in the neighborhood of the infundibulum. Targeting hair follicles with melanin may be useful to dye poorly pigmented hairs, improving laser hair removal. The procedure accelerates the delivery of melanin into hair structures allowing an even absorption, larger pigment deposits, and deeper penetration of the formulation into the hair.
RESUMO
Home parenteral nutrition (PN) is associated with many complications including severe hepatobiliary dysfunction. Commercial ω-6 fatty acid-soybean based-lipid emulsions in PN may mediate long term PN associate liver disease (PNALD) whereas ω-3-fish oil parenteral emulsions have shown to reverse PNALD in children. However, its clinical effectiveness in adults has been scarcely reported. In this work, we study the role of soybean and fish oil lipid commercial emulsions on inflammatory and profibrotic liver markers in adults with long term PNALD and in in vitro cellular models. Inflammatory and profibrotic markers were measured in serum of ten adults with long term PNALD and in culture supernatants of monocytes. Liver epithelial to mesenchymal transition (EMT) was induced by transforming growth factor beta 1 (TGFß1) to evaluate in vitro liver fibrosis. Omegaven®, a 100% fish oil commercial emulsion, was infused during four months in two patients with severe long term PNALD reversing, at the first month, the inflammatory, profibrotic and clinical parameters of PNALD. The effect was maintained during the treatment course but impaired when conventional lipid emulsions were reintroduced. The other patients under chronic soybean oil-based PN showed elevated inflammatory and profibrotic parameters. In vitro human monocytes stimulated with lipopolysaccharide induced a strong inflammatory response that was suppressed by Omegaven®, but increased by soybean emulsions. In other experiments, TGFß1 induced EMT that was suppressed by Omegaven® and enhanced by soybean oil lipid emulsions. Omegaven® improves clinical, anti-inflammatory and anti-fibrotic parameters in adults with long-term home PNALD.
Assuntos
Anti-Inflamatórios/administração & dosagem , Cicatriz/prevenção & controle , Emulsões Gordurosas Intravenosas/administração & dosagem , Óleos de Peixe/administração & dosagem , Hepatopatias/tratamento farmacológico , Nutrição Parenteral no Domicílio/efeitos adversos , Adulto , Idoso , Técnicas de Cultura de Células , Cicatriz/tratamento farmacológico , Transição Epitelial-Mesenquimal , Ácidos Graxos Ômega-6/administração & dosagem , Feminino , Humanos , Hepatopatias/etiologia , Masculino , Pessoa de Meia-Idade , Nutrição Parenteral no Domicílio/métodos , Óleo de Soja/administração & dosagem , TriglicerídeosRESUMO
Eosinophils play a prominent role in the process of allergic inflammation. Non-receptor associated Lyn tyrosine kinases generate key initial signals in eosinophils. Bafetinib, a specific Abl/Lyn tyrosine kinase inhibitor has shown a potent antiproliferative activity in leukemic cells, but its effects on eosinophils have not been reported. Therefore, we studied the effects of bafetinib on functional and mechanistic responses of isolated human eosinophils. Bafetinib was more potent than non-specific tyrosin kinase comparators genistein and tyrphostin inhibiting superoxide anion triggered by N-formyl-Met-Leu-Phe (fMLF; 100 nM) (-log IC50=7.25 ± 0.04 M; 6.1 ± 0.04 M; and 6.55 ± 0.03 M, respectively). Bafetinib, genistein and tyrphostin did not modify the [Ca(2+)]i responses to fMLF. Bafetinib inhibited the release of EPO induced by fMLF with higher potency than genistein and tyrphostin (-log IC50=7.24 ± 0.09 M; 5.36 ± 0.28 M; and 5.37 ± 0.19 M, respectively), and nearly suppressed LTC4, ECP and chemotaxis. Bafetinib, genistein and tyrphostin did not change constitutive apoptosis. However bafetinib inhibited the ability of granulocyte-monocyte colony-stimulating factor to prevent apoptosis. The activation of Lyn tyrosine kinase, p-ERK1/2 and p-38 induced by fMLF was suppressed by bafetinib and attenuated by genistein and tyrphostin. In conclusion, bafetinib inhibits oxidative burst and generation of inflammatory mediators, and reverses the eosinophil survival. Therefore, future anti-allergic therapies based on bafetinib, could help to suppress excessive inflammatory response of eosinophils at inflammatory sites.
Assuntos
Eosinófilos/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/farmacologia , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Movimento Celular/efeitos dos fármacos , Proteína Catiônica de Eosinófilo/biossíntese , Eosinófilos/citologia , Eosinófilos/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Interleucina-5/farmacologia , Leucotrieno C4/biossíntese , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Peroxidase/metabolismo , Superóxidos/metabolismoRESUMO
A simple, sensitive and specific high-performance liquid chromatography method has been developed for the determination of nortriptyline (NT) in plasma samples. The assay involved derivatization with 9H-fluoren-9-ylmethyl chloroformate (Fmoc-Cl) and isocratic reversed-phase (C(18)) chromatography with fluorescence detection. The developed method required only 100 microl of plasma sample, deproteinized and derivatized in one step. Calibration curves were lineal over the concentration range of 5-5000 ng/ml. The derivatization reaction was performed at room temperature in 20 min and the obtained NT derivative was stable for at least 48 h at room temperature. The within-day and between-day relative standard deviation was below 8%. The limit of detection (LOD) was 2 ng/ml, and the lower limit of quantification (LLOQ) was established at 10 ng/ml. The method was applied on plasma collected from rats, at different time intervals, after intravenous administration of 0.5 mg of NT.