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1.
Carbohydr Polym ; 229: 115437, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31826460

RESUMO

Chitosan nanosystems have been widely explored to deliver therapeutic into cells. The cationic nature of the polymer facilitates its entry into the cell via the negatively charged lipid bilayer. Though the interaction is feasible for successful payload delivery, very little is known about the mechanistic aspects and kinetics of interaction of chitosan nanoparticles (Chnps) with the cellular bilayer membrane. Moreover, the precise mechanism of delivery of therapeutic agents by the Chnps is unknown. The polymerbilayer membrane is anticipated to play a crucial role in deciding its ultimate intracellular fate, while delivering its therapeutic payload. Here, we have made an attempt to understand the interaction of Chnps with the cellular membrane for delivering payload, through experimental analysis and predictive mathematical modeling. We observed that the positively charged, mucoadhesive Chnps lack specificity towards a particular cell type, but are rather successful in the intracellular delivery of nucleic acids.


Assuntos
Quitosana/metabolismo , Portadores de Fármacos/metabolismo , Nanopartículas/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Quitosana/química , Portadores de Fármacos/química , Endocitose/fisiologia , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/química , Humanos , Cinética , Lisossomos/metabolismo , Fluidez de Membrana , Modelos Biológicos , Nanopartículas/química
2.
Biosci Biotechnol Biochem ; 84(4): 800-803, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31842704

RESUMO

Cross-linked albumin films, which are non-cell adhesive, were altered to be cell-adhesive by the combination of varied concentrations of fluorescein isothiocyanate and blue light irradiation. In this system, cell patterning was developed with two different cell lines by sequential seeding.Abbreviations: BSA: bovine serum albumin; EGDE: ethylene glycol diglycidyl ether; PBS: Dulbecco's phosphate buffered saline.


Assuntos
Adesão Celular/efeitos dos fármacos , Adesão Celular/efeitos da radiação , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/química , Luz , Animais , Linhagem Celular , Humanos , Camundongos , Soroalbumina Bovina/química
3.
Chem Commun (Camb) ; 56(2): 213-216, 2019 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-31808495

RESUMO

We developed a peptide-templated gold nanoparticle (AuNP) nanosensor for simultaneous detection of multiple posttranslational modification (PTM) enzymes with a detection limit of 28 pM for histone deacetylase (HDAC) and 0.8 pM for protein tyrosine phosphatase 1B (PTP1B), and it can be further applied for the screening of PTM enzyme inhibitors and the measurement of PTM enzymes in cancer cells.


Assuntos
Histona Desacetilases/análise , Nanopartículas Metálicas/química , Fosfopeptídeos/química , Proteína Tirosina Fosfatase não Receptora Tipo 1/análise , Carbocianinas/química , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/química , Ouro/química , Células HeLa , Inibidores de Histona Desacetilases/química , Histona Desacetilases/química , Humanos , Ácidos Hidroxâmicos/química , Limite de Detecção , Processamento de Proteína Pós-Traducional , Proteína Tirosina Fosfatase não Receptora Tipo 1/antagonistas & inibidores , Proteína Tirosina Fosfatase não Receptora Tipo 1/química , Espectrometria de Fluorescência/métodos , Vanadatos/química
4.
Zhongguo Zhong Yao Za Zhi ; 44(19): 4171-4178, 2019 Oct.
Artigo em Chinês | MEDLINE | ID: mdl-31872695

RESUMO

In order to prepare angiopep-2 modified fluorescein isothiocyanate-labeled neurotoxin nanoparticles( ANG-NPs/FITCNT),emulsion/solvent evaporation method was used with m PEG-PLA and ANG-PEG-PLA( in proper proportions) as carriers and with FITC-NT as drug. With particle size and encapsulation efficiency as comprehensive indexes,the effects of different ultrasound power and ultrasound time combinations on the process were investigated. The in vitro release characteristics of nanoparticles in PBS buffer at p H 7. 4 and p H 6. 5 were investigated by dialysis method. The results indicated that the optimum process for preparing ANG-NPs/FITC-NT was as follows: ultrasonic power 90 W,ultrasonic time 30 s. In such optimal process,ANG-NPs/FITC-NT were well-shaped under the transmission electron microscope,with an average particle size of( 123. 9±0. 5) nm,Zeta potential of(-10. 5±0. 5) m V,encapsulation efficiency of( 68. 1±0. 4) %,and the drug loading of( 0. 82±0. 01) %. The in vitro drug release profiles of the nanoparticles in PBS buffer at p H 7. 4 and p H 6. 5 were both consistent with Ritger-Peppas equation,ln Q = 0. 508 8 lnt-2. 285 0,r = 0. 961 5( p H 7. 4) and ln Q= 0. 449 9 lnt-1. 855 3,r = 0. 970 3( p H 6. 5),respectively. The experiment results proved that the nanoparticles prepared by emulsion/solvent evaporation method had uniform particle size,high encapsulation efficiency and in vitro sustained release characteristic,which might be a potential carrier for NT intracerebral drug delivery.


Assuntos
Portadores de Fármacos , Nanopartículas , Peptídeos , Fluoresceína-5-Isotiocianato , Tamanho da Partícula , Polietilenoglicóis
5.
Reprod Biol Endocrinol ; 17(1): 109, 2019 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-31856844

RESUMO

BACKGROUND: Capacitation involves physiological changes that spermatozoa must undergo in the female reproductive tract or in vitro to obtain the ability to bind, penetrate and fertilize the egg. Up to date, several methods have been developed to characterize this complex biological process. The goal of the presented study is to mutually compare several fluorescent techniques, check their ability to detect changes in molecular processes during the capacitation progress and determine their ability to predict the percentage of acrosome reacted (AR) sperm after the exposure to solubilized zona pellucida (ZP). The capacitation process was analyzed using four fluorescent techniques: 1. chlortetracycline (CTC) staining, 2. anti-acrosin antibody (ACR.2) assay, 3. anti-phosphotyrosine (pY) antibody assay, 4. fluorescein isothiocyanate-conjugated phalloidin (FITC-phall) assay. All these methods were tested using fluorescent microscopy and flow cytometry. RESULTS: All selected methods are capable to detect the capacitation progress of boar sperm in vitro, but there are significant differences in their outcome when using fluorescent microscopy or flow cytometry experimental arrangements and subsequent statistical analysis (KW-ANOVA). Also, the ability to predict the absolute numbers of sperm which will undergo ZP-induced AR differ significantly (CTC and ACR.2 gave the best predictions). CONCLUSIONS: Our study compared four largely used methods used to characterize capacitation process, highlighted their differences and showed that all are able to detect capacitation progress, CTC and ACR.2 are furthermore able to accurately predict the percentage of AR sperm after ZP-induced AR.


Assuntos
Citometria de Fluxo , Corantes Fluorescentes , Microscopia de Fluorescência , Capacitação Espermática/fisiologia , Sus scrofa/fisiologia , Reação Acrossômica/fisiologia , Animais , Cálcio/análise , Citometria de Fluxo/métodos , Fluoresceína-5-Isotiocianato , Imunofluorescência , Masculino , Microscopia de Fluorescência/métodos , Faloidina , Espermatozoides/fisiologia , Zona Pelúcida/fisiologia
6.
Head Face Med ; 15(1): 27, 2019 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-31711509

RESUMO

BACKGROUND: Controlled release of proteins bound to conventional bone substitutes is still insufficient. Therefore, this study evaluates in-vitro release kinetics of the model protein FITC-BSA (fluorescein conjugated bovine serum albumine) from insoluble bovine collagenous bone matrices (ICBM) with different polymer coatings. Analyzes aim at comparing FITC-BSA release from uncoated versus coated ICBM over time to find bone substitute coatings with consistent release profiles. METHODS: Release kinetics of FITC-BSA from uncoated as well as coated ICBM with five different polymers (RESOMER R 203 H, RG 503 H, RG 504 H, RG 505, L 206 S) were measured over a period of 11 days (d). Measurements were conducted after 6 h (h), 12 h, 24 h, 3 d, 5 d, 7 d, 9 d and 11 d with six samples for each coated ICBM. Two groups were formed (1) with and (2) without medium change at times of measurement. For each group ANOVA with post-hoc Bonferroni testing was used. Scanning electron microscopy assessed morphologic differences between ICBM coating. RESULTS: In group 1 approx. 70% of FITC-BSA release from uncoated ICBM occurred after 6 h compared to approx. 50% in group 2. Only polymers with medium inherent viscosity, i.e. RESOMER RG 503 H, constantly showed significantly more FITC-BSA release throughout 11 d than uncoated ICBM (p = 0.007). The same was found for group 2 (p = 0.005). No significant differences between PLA and PLGA polymers were found. Scanning electron microscopy results indicate a weak adhesion of polymer coatings to ICBM explaining its rather weak retentive effect on overall FITC-BSA release. CONCLUSIONS: Medium molecular size polymers reduce the overall released FITC-BSA from ICBM over time. In clinical practice these polymers may prove ideal for bone substitute materials.


Assuntos
Substitutos Ósseos , Fluoresceína-5-Isotiocianato/análogos & derivados , Polímeros , Soroalbumina Bovina , Animais , Substitutos Ósseos/farmacocinética , Bovinos , Fluoresceína-5-Isotiocianato/farmacocinética , Cinética , Microscopia Eletrônica de Varredura , Soroalbumina Bovina/farmacocinética
7.
Parasit Vectors ; 12(1): 486, 2019 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-31619276

RESUMO

BACKGROUND: In the animal production sector, enteritis is responsible for serious economic losses, and intestinal parasitism is a major stress factor leading to malnutrition and lowered performance and animal production efficiency. The effect of enteric parasites on the gut function of teleost fish, which represent the most ancient bony vertebrates, is far from being understood. The intestinal myxozoan parasite Enteromyxum leei dwells between gut epithelial cells and causes severe enteritis in gilthead sea bream (Sparus aurata), anorexia, cachexia, growth impairment, reduced marketability and increased mortality. METHODS: This study aimed to outline the gut failure in this fish-parasite model using a multifaceted approach and to find and validate non-lethal serum markers of gut barrier dysfunction. Intestinal integrity was studied in parasitized and non-parasitized fish by immunohistochemistry with specific markers for cellular adhesion (E-cadherin) and tight junctions (Tjp1 and Cldn3) and by functional studies of permeability (oral administration of FITC-dextran) and electrophysiology (Ussing chambers). Serum samples from parasitized and non-parasitized fish were analyzed using non-targeted metabolomics and some significantly altered metabolites were selected to be validated using commercial kits. RESULTS: The immunodetection of Tjp1 and Cldn3 was significantly lower in the intestine of parasitized fish, while no strong differences were found in E-cadherin. Parasitized fish showed a significant increase in paracellular uptake measured by FITC-dextran detection in serum. Electrophysiology showed a decrease in transepithelial resistance in infected animals, which showed a diarrheic profile. Serum metabolomics revealed 3702 ions, from which the differential expression of 20 identified compounds significantly separated control from infected groups in multivariate analyses. Of these compounds, serum inosine (decreased) and creatine (increased) were identified as relevant and validated with commercial kits. CONCLUSIONS: The results demonstrate the disruption of tight junctions and the loss of gut barrier function, a metabolomic profile of absorption dysfunction and anorexia, which further outline the pathophysiological effects of E. leei.


Assuntos
Enterite/veterinária , Doenças dos Peixes/parasitologia , Metabolômica , Myxozoa/patogenicidade , Doenças Parasitárias em Animais/parasitologia , Dourada/parasitologia , Animais , Caderinas/metabolismo , Claudina-3/metabolismo , Creatina/sangue , Dextranos/metabolismo , Modelos Animais de Doenças , Eletrofisiologia , Enterite/parasitologia , Ensaio de Imunoadsorção Enzimática , Fluoresceína-5-Isotiocianato/análogos & derivados , Fluoresceína-5-Isotiocianato/metabolismo , Imuno-Histoquímica , Inosina/sangue , Mucosa Intestinal/metabolismo , Intestinos/parasitologia , Intestinos/patologia , Doenças Parasitárias em Animais/patologia , Permeabilidade , Proteína da Zônula de Oclusão-1/metabolismo
8.
Int J Mol Sci ; 20(19)2019 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-31547100

RESUMO

Naturally existing Chlorogenic acid (CGA) is an antioxidant-rich compound reported to act a chemopreventive agent by scavenging free radicals and suppressing cancer-causing mechanisms. Conversely, the compound's poor thermal and pH (neutral and basic) stability, poor solubility, and low cellular permeability have been a huge hindrance for it to exhibit its efficacy as a nutraceutical compound. Supposedly, encapsulation of CGA in chitosan nanoparticles (CNP), nano-sized colloidal delivery vector, could possibly assist in enhancing its antioxidant properties, in vitro cellular accumulation, and increase chemopreventive efficacy at a lower concentration. Hence, in this study, a stable, monodispersed, non-toxic CNP synthesized via ionic gelation method at an optimum parameter (600 µL of 0.5 mg/mL of chitosan and 200 µL of 0.7 mg/mL of tripolyphosphate), denoted as CNP°, was used to encapsulate CGA. Sequence of physicochemical analyses and morphological studies were performed to discern the successful formation of the CNP°-CGA hybrid. Antioxidant property (studied via DPPH (1,1-diphenyl-2-picrylhydrazyl) assay), in vitro antiproliferative activity of CNP°-CGA, and in vitro accumulation of fluorescently labeled (FITC) CNP°-CGA in cancer cells were evaluated. Findings revealed that successful formation of CNP°-CGA hybrid was reveled through an increase in particle size 134.44 ± 18.29 nm (polydispersity index (PDI) 0.29 ± 0.03) as compared to empty CNP°, 80.89 ± 5.16 nm (PDI 0.26 ± 0.01) with a maximal of 12.04 µM CGA loaded per unit weight of CNP° using 20 µM of CGA. This result correlated with Fourier-Transform Infrared (FTIR) spectroscopic analysis, transmission Electron Microscopy (TEM) and field emission scanning (FESEM) electron microscopy, and ImageJ evaluation. The scavenging activity of CNP°-CGA (IC50 5.2 ± 0.10 µM) were conserved and slightly higher than CNP° (IC50 6.4±0.78 µM). An enhanced cellular accumulation of fluorescently labeled CNP°-CGA in the human renal cancer cells (786-O) as early as 30 min and increased time-dependently were observed through fluorescent microscopic visualization and flow cytometric assessment. A significant concentration-dependent antiproliferation activity of encapsulated CGA was achieved at IC50 of 16.20 µM as compared to CGA itself (unable to determine from the cell proliferative assay), implying that the competent delivery vector, chitosan nanoparticle, is able to enhance the intracellular accumulation, antiproliferative activity, and antioxidant properties of CGA at lower concentration as compared to CGA alone.


Assuntos
Carcinoma de Células Renais , Quitosana , Ácido Clorogênico , Sistemas de Liberação de Medicamentos , Depuradores de Radicais Livres , Neoplasias Renais , Nanopartículas , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Quitosana/química , Quitosana/farmacocinética , Quitosana/farmacologia , Ácido Clorogênico/química , Ácido Clorogênico/farmacocinética , Ácido Clorogênico/farmacologia , Fluoresceína-5-Isotiocianato/química , Fluoresceína-5-Isotiocianato/farmacocinética , Fluoresceína-5-Isotiocianato/farmacologia , Depuradores de Radicais Livres/química , Depuradores de Radicais Livres/farmacocinética , Depuradores de Radicais Livres/farmacologia , Humanos , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/metabolismo , Neoplasias Renais/patologia , Microscopia de Fluorescência , Nanopartículas/química , Nanopartículas/uso terapêutico
9.
Chin J Nat Med ; 17(7): 525-534, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31514984

RESUMO

This study aimed to investigate the mechanisms of Yu-Ping-Feng-San (YPFS) on attenuating allergic inflammation in the initial stage of atopic dermatitis (AD). AD mouse model was established with fluorescein isothiocyanate (FITC) sensitization and elicitation. Epithelial barrier structure was observed with transmission electron microscope. The populations of dendritic cells (DCs) and group 2 innate lymphoid cells (ILC2s) were detected by flow cytometry. Human immortalized keratinocyte (HaCaT) cells were stimulated with Poly(I:C)/TNF-α in vitro to assessthymic stromal lymphopoietin (TSLP), interleukin (IL)-33 and nuclear factor-κB (NF-κB) levels or expressions by immunofluorescence, enzyme linked immunosorbent assay (ELISA) and western blot. In the initial stage of AD, ear swelling and infiltration of inflammatory cells in ear tissues were markedly attenuated with YPFS treatments. The damaged structures of ear epithelium and the increased levels of Th2-cytokines induced by FITC were significantly rescued in YPFS-treated mice. The production of pro-allergic cytokines, TSLP and IL-33, as well as the cell populations of their target cells DCs and ILC2s were decreased in AD model, respectively. Likewise, the levels of TSLP and IL-33 in Poly(I:C)/TNF-α-stimulated HaCaT cells showed the same results. Lower levels of p-NF-κB were detected with YPFS treatment, and the expressions of TSLP and IL-33 could be further decreased with inhibiting of NF-κB. Therefore, YPFS attenuates allergic inflammation in the initial stage of AD probably through regulating NF-κB-TSLP/IL-33 pathway, which may provide a novel effective target for the prevention and treatment of allergic diseases.


Assuntos
Antialérgicos/uso terapêutico , Citocinas/metabolismo , Dermatite Atópica/tratamento farmacológico , Medicamentos de Ervas Chinesas/uso terapêutico , Inflamação/prevenção & controle , Animais , Antialérgicos/farmacologia , Linhagem Celular , Células Dendríticas/patologia , Dermatite Atópica/induzido quimicamente , Dermatite Atópica/metabolismo , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacologia , Células Epiteliais/metabolismo , Fluoresceína-5-Isotiocianato/toxicidade , Inflamação/metabolismo , Inflamação/patologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Linfócitos/metabolismo , Linfócitos/patologia , Camundongos Endogâmicos BALB C , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico
10.
ACS Appl Mater Interfaces ; 11(40): 37313-37321, 2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31517474

RESUMO

A simple process is developed for the one-step preparation of dual-compartment alginate microcapsules with controlled size and structure from microfluid-generated water-in-water-in-oil (W/W/O) emulsion droplet. Unlike other methods that rely on transient W/W/O emulsion droplet, we introduce an aqueous two-phase system (ATPS) to form a stable W/W/O emulsion droplet as a template for preparing dual-compartment alginate microcapsules. Two different bioactive molecules are able to be spatially confined encapsulated in the shell and core of alginate microcapsules due to the partitioning effect of ATPS and the high viscosity of alginate solution. Moreover, an enzyme cascade reaction with a spatial confined glucose oxidase and horseradish peroxidase in the shell and core of alginate microcapsules confirms its excellent biocompatibility and high activity. This method provides a green platform for enzyme-catalyzed tandem reactions and controlled sequential release of multiple drugs based on alginate microcapsules.


Assuntos
Alginatos/química , Cápsulas/química , Emulsões/química , Microfluídica , Óleos/química , Água/química , Biocatálise , Fluoresceína-5-Isotiocianato/análogos & derivados , Glucose Oxidase/metabolismo , Peroxidase do Rábano Silvestre/metabolismo , Microfluídica/instrumentação , Tamanho da Partícula , Soroalbumina Bovina
11.
Invest Ophthalmol Vis Sci ; 60(10): 3547-3555, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31415078

RESUMO

Purpose: Current treatments for diabetic retinopathy (DR) have considerable limitations, underpinning the need for new therapeutic options. In this article, the ability of an engineered angiopoietin-1 variant (COMP-Ang1) to ameliorate the injurious effects of hyperglycemia on barrier integrity in a human retinal microvascular endothelial cell (HRMvEC) model is comprehensively investigated. Methods: Confluent HRMvECs were treated (0-72 hours) with d-glucose (5 or 30 mM) in the absence and presence of COMP-Ang1 (10-200 ng/mL). l-glucose (30 mM) was used as osmotic control. Posttreatment, intact cell monolayers were monitored for permeability to FITC-dextran 40 kDa. Cells were also harvested for analysis of interendothelial junction targets by RT-qPCR and Western blotting. The impact of receptor tyrosine kinase Tie2 gene silencing on COMP-Ang1 efficacy was also evaluated. Results: Treatment with 30 mM d-glucose (but not l-glucose) demonstrated a time-dependent elevation in the mean rate of FITC-dextran diffusion across intact HRMvEC monolayers, in parallel with significant reductions in mRNA/protein levels of occludin, claudin-5, ZO-1, and VE-Cadherin. These effects were all attenuated by COMP-Ang1 in a concentration-dependent fashion, with 200 ng/mL recovering barrier function by ∼88%, and recovering reduced interendothelial junction protein levels by more than 50%. Finally, Tie2 knockdown by small interfering RNA silencing blocked the ability of COMP-Ang1 to mitigate against hyperglycemia-induced permeabilization of HRMvECs and depletion of junctional expression levels. Conclusions: In summary, this article presents a reproducible in vitro cell study that quantifies the concentration-dependent efficacy of COMP-Ang1 to mitigate the injurious effects of hyperglycemic challenge on HRMvEC barrier properties via Tie2-mediated signaling.


Assuntos
Barreira Hematorretiniana/fisiologia , Células Endoteliais/efeitos dos fármacos , Hiperglicemia/prevenção & controle , Proteínas Recombinantes de Fusão/farmacologia , Vasos Retinianos/efeitos dos fármacos , Antígenos CD/genética , Western Blotting , Caderinas/genética , Permeabilidade Capilar/efeitos dos fármacos , Células Cultivadas , Claudina-5/genética , Dextranos/metabolismo , Relação Dose-Resposta a Droga , Células Endoteliais/metabolismo , Fluoresceína-5-Isotiocianato/análogos & derivados , Fluoresceína-5-Isotiocianato/metabolismo , Inativação Gênica/fisiologia , Glucose/farmacologia , Humanos , Hiperglicemia/metabolismo , Ocludina/genética , Fenótipo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Receptor TIE-2/genética , Vasos Retinianos/metabolismo
12.
Molecules ; 24(16)2019 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-31409040

RESUMO

Polymer dots (Pdots) represent newly developed semiconductor polymer nanoparticles and exhibit excellent characteristics as fluorescent probes. To improve the sensitivity and biocompatibility of Pdots ratiometric pH biosensors, we synthesized 3 types of water-soluble Pdots: Pdots-PF, Pdots-PP, and Pdots-PPF by different combinations of fluorescent dyes poly(9,9-dioctylfluorenyl-2,7-diyl) (PFO), poly[(9,9-dioctyl-fluorenyl-2,7-diyl)-co-(1,4-benzo-{2,1',3}-thiadazole)] (PFBT), and fluorescein isothiocyanate (FITC). We found that Pdots-PPF exhibits optimal performance on pH sensing. PFO and FITC in Pdots-PPF produce pH-insensitive (λ = 439 nm) and pH-sensitive (λ = 517 nm) fluorescence respectively upon a single excitation at 380 nm wavelength, which enables Pdots-PPF ratiometric pH sensing ability. Förster resonance energy transfer (FRET) together with the use of PFBT amplify the FITC signal, which enables Pdots-PPF robust sensitivity to pH. The emission intensity ratio (I517/I439) of Pdots-PPF changes linearly as a function of pH within the range of pH 3.0 to 8.0. Pdots-PPF also possesses desirable reversibility and stability in pH measurement. More importantly, Pdots-PPF was successfully used for cell imaging in Hela cells, exhibiting effective cellular uptake and low cytotoxicity. Our study suggests the promising potential of Pdots-PPF as an in vivo biomarker.


Assuntos
Técnicas Biossensoriais , Fluorenos/química , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/química , Imagem Óptica/métodos , Polímeros/química , Pontos Quânticos/química , Transferência Ressonante de Energia de Fluorescência , Células HeLa , Humanos , Concentração de Íons de Hidrogênio , Semicondutores , Solubilidade , Água
13.
Talanta ; 205: 120021, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31450404

RESUMO

Herein, we figured out a novel ratiometric fluorescent probe based on Janus silica nanoparticles (SiO2 NPs) where fluorescein isothiocyanate (FITC) and rhodamine B (RB) were asymmetrically functionalized for intracellular pH detection. With this unique strategy, the probe exhibited dual emission bands at 530 and 580 nm upon excitation at a single wavelength of 488 nm. The pH titrations of the probe showed a remarkable increase in ratiometric intensities ratio (I530nm/I580nm) when the pH was switched from 3.0 to 8.0 with a acid dissociation constant (pKa) value of 6.64. The probe was successfully applied to imaging and detection of pH in living cells without noticeable cytotoxicity, indicating that it is a promising candidate for bioanalysis and bioimaging.


Assuntos
Corantes Fluorescentes/química , Concentração de Íons de Hidrogênio , /química , Células A549 , Linhagem Celular Tumoral , Fluoresceína-5-Isotiocianato/química , Transferência Ressonante de Energia de Fluorescência , Humanos , Neoplasias Hepáticas/química , Imagem Molecular , Rodaminas/química , Dióxido de Silício/química , Espectrometria de Fluorescência/métodos
14.
IET Nanobiotechnol ; 13(6): 560-564, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31432786

RESUMO

While cancer is the leading cause of human's deaths worldwide, finding an imaging agent which can detect cancer tumours is needed for cancer diagnosis. In the present study, PEG-citrate dendrimer-G2 was used as a nano-carrier of FITC dye and Iohexol to help passive targeting and uptake of both imaging agents in cancer cells/tumour in vitro and in vivo. Dendrimer was synthesisedand the product characterised using LC-MS, FT-IR, DLS, ELS, AFM, and 1HNMR. After FITC loading into dendrimer, MTT was performed to determine the cytotoxicity of formulation on HEK-293 and MCF-7 cells. In vitro imaging using dendrimer-FITC was done via fluorescent microscope thereafter. Moreover, CT imaging using Iohexol was employed to show the targeting nature and ability of the complex to use as imaging agent in vivo. Data yielded in this study corroborate the notion that the promised dendrimer was synthesised properly and had no toxicity along with FITC on normal cell. Furthermore, CT and fluorescent images showed the targeting nature and imaging ability of Iohexol/FITC loaded dendrimer in vitro and in vivo. Overall, results showed promising characteristics of the novel complexes using dendrimer-G2 both in vitro and in vivo.


Assuntos
Ácido Cítrico/química , Dendrímeros/química , Diagnóstico por Imagem/métodos , Corantes Fluorescentes/química , Polietilenoglicóis/química , Citratos/síntese química , Citratos/química , Dendrímeros/síntese química , Portadores de Fármacos , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/síntese química , Células HEK293 , Humanos , Células MCF-7 , Coloração e Rotulagem/métodos
15.
Int J Pharm ; 568: 118550, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31336152

RESUMO

Implants offer the opportunity to improve patient adherence and real-world outcomes. However, most polymers used today are hydrophobic and limit drug properties suitable for development. Thermoplastic poly(urethanes) (TPUs) form pores upon hydration and may facilitate the development of implants containing drugs exhibiting broadly different properties. We sought to investigate the effect of drug physicochemical properties on permeability through membranes of varying TPU mixture composition; leverage imaging to visualize microstructural changes to the membrane across the TPU mixture composition range; and quantitatively characterize the membrane microstructure using equivalent pore analysis. We observed a correlation between drug hydrophobicity and its permeability through hydrophobic-rich TPU membranes. Conversely, all compounds diffused through hydrophilic-rich TPU membranes at similar rates, regardless of drug properties. Imaging revealed significant microstructure differences between hydrophobic-rich and hydrophilic-rich TPU membranes, supporting hypotheses proposed in our previous study. The hydrated hydrophilic TPU membrane pore area was determined to be 0.583% and its equivalent pore radius was found to be 128 nm, suggesting that hydrophilic TPU membranes may be used to modify the release of small molecular weight drugs and macromolecules. These findings highlight the benefits of hydrophilic TPUs as rate-controlling membranes to modulate the release rate of drugs with varying physicochemical properties.


Assuntos
Membranas Artificiais , Poliuretanos/química , Dextranos/química , Difusão , Implantes de Medicamento , Emtricitabina/química , Fluoresceína-5-Isotiocianato/análogos & derivados , Fluoresceína-5-Isotiocianato/química , Interações Hidrofóbicas e Hidrofílicas , Ibuprofeno/química , Metoprolol/química , Peso Molecular , Permeabilidade , Porosidade
16.
Analyst ; 144(15): 4477-4482, 2019 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-31268083

RESUMO

An ultra-sensitive sensor was fabricated to measure dopamine through quenching and restoring FITC fluorescence by the competitive binding of dopamine and N-acetylneuraminic acid with mercaptophenylboronic acid anchored on the gold nanoparticles. Assessed by quantifying dopamine in human urine samples, it reached a limit of detection of as low as 50 pM dopamine over a linear range from 10-10 M to 10-7 M. It also features low technical barriers and operation cost, and the strategy proposed could be extendable to other analytes and not restricted to the gold nanoparticles.


Assuntos
Dopamina/análise , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/química , Ouro/química , Nanopartículas Metálicas/química , Ácidos Borônicos/química , Fluorescência , Limite de Detecção , Ácido N-Acetilneuramínico/química , Espectrometria de Fluorescência/métodos , Compostos de Sulfidrila/química
17.
Anal Bioanal Chem ; 411(23): 6021-6029, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31289898

RESUMO

Cytobiological methods for cell nucleus-related studies start with the extraction processes of intranuclear components with many cell lysis buffers, following with the structural characterizations and quantitative analysis of the extracted components. In this study, we tried to evaluate the availability and reliability of these extraction-based analytical methods from their spectral features. We implemented an in situ surface-enhanced Raman scattering spectroscopy (SERS) strategy with the help of the nucleus-targeting nanoprobes to investigate the molecular information of nucleus, in comparison with these ex situ methods. This study provides valuable references for choosing an appropriate detection method according to different detection purposes, and also points out the risks of many developing cell-related analytical methods that combine the traditional cytobiological techniques from exogenous interferences during sample preprocesses. Graphical abstract.


Assuntos
Núcleo Celular/química , DNA/análise , Nanopartículas Metálicas/química , Proteínas Nucleares/análise , Análise Espectral Raman/métodos , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/química , Ouro/química , Células Hep G2 , Humanos , Nanopartículas Metálicas/ultraestrutura , Peptídeos/química , Polietilenoglicóis/química
18.
J Laryngol Otol ; 133(8): 696-699, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31290382

RESUMO

OBJECTIVE: To explore the use of fluorescence lifetime imaging microscopy in thyroid tissues, and to investigate how different thyroid lesions affect fluorescence lifetime. METHOD: Fluorescence lifetime measurements were taken of fresh frozen thyroid surgical specimens stained with fluorescein isothiocyanate tagged anti-thyroglobulin monoclonal antibodies. RESULTS: The mean fluorescence lifetime measurements in 12 patients - 3 with multinodular goitre, 4 with follicular adenoma, 4 with papillary thyroid carcinoma and 1 with follicular carcinoma - were 3.16 ns (range, 2.66-3.52 ns), 3.75 ns (range, 2.99-4.57 ns), 2.97 ns (range, 2.57-3.21 ns) and 3.61 ns, respectively. The fluorescence lifetime of follicular adenoma patients was higher than that of papillary thyroid carcinoma patients by 26 per cent (p = 0.058). The fluorescence lifetime in the follicular carcinoma patient was similar to the follicular adenoma group, but higher than in the papillary thyroid carcinoma group by 22 per cent (p = 0.01). CONCLUSION: Fluorescence lifetime measurements varied in different thyroid pathologies, possibly because of tissue-scale structural influences.


Assuntos
Adenoma/diagnóstico por imagem , Bócio Nodular/diagnóstico por imagem , Glândula Tireoide/diagnóstico por imagem , Neoplasias da Glândula Tireoide/diagnóstico por imagem , Adenoma/metabolismo , Feminino , Fluoresceína-5-Isotiocianato/farmacologia , Técnica Direta de Fluorescência para Anticorpo , Bócio Nodular/metabolismo , Humanos , Masculino , Microscopia de Fluorescência , Tireoglobulina/antagonistas & inibidores , Glândula Tireoide/patologia
19.
Int J Pharm ; 568: 118469, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31265884

RESUMO

Resveratrol is a small molecule produced by various plants with a remarkable range of beneficial functions in animals. One of these is stimulating signaling pathways in adipose tissue that protect against obesity. Unfortunately, resveratrol suffers from poor bioavailability that inhibits its accumulation in target tissues, including fat, thus hindering the realization of its therapeutic potential. To address this, we are developing biodegradable microparticles as drug depots for controlled release of resveratrol within fat. In this study, resveratrol was encapsulated into poly(lactide-co-glycolide) microparticles using an oil-in-water emulsion/solvent evaporation technique. The oil phase consisted of resveratrol and poly(lactide-co-glycolide) dissolved in a mixture of dichloromethane and ethanol; meanwhile, the aqueous phase contained poly(vinyl alcohol) as the emulsifier. Increasing ethanol's volume ratio increased resveratrol's solubility in the oil phase and particle drug loading. The maximal loading achieved was 65 µg/mg (6.5%) and occurred when the ethanol to dichloromethane ratio was 1:3. Under these conditions, particles exhibited ruffled surfaces, which resulted in variable drug release over the first three days of a six-week release assay. By decreasing resveratrol and ethanol in the oil phase and increasing poly(vinyl alcohol) in the aqueous phase, smooth particles were achieved, but they suffered a 15-25-fold decrease in drug loading depending on size. Small particles exhibited higher drug loading and burst drug release compared to larger particles because of their higher specific surface area. Utilizing mild chemistry, we functionalized poly(vinyl alcohol) with fluorescein isothiocyanate and demonstrated that encapsulation of resveratrol in the particle decreases the amount of fluorescent polymer on the particle surface, suggesting resveratrol displaces the emulsifier during particle formation. Taken together, resveratrol can be encapsulated into poly(lactide-co-glycolide) microparticles, but it accumulates at the particle surface impacting drug loading, surface roughness, and drug release.


Assuntos
Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Álcool de Polivinil/química , Resveratrol/química , Células 3T3-L1 , Tecido Adiposo , Animais , Preparações de Ação Retardada/química , Liberação Controlada de Fármacos , Fluoresceína-5-Isotiocianato/química , Camundongos , Tamanho da Partícula
20.
Talanta ; 204: 875-881, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31357376

RESUMO

It is of paramount importance to be able to accurately quantify surface coverage of antibodies on gold nanoparticles (AuNP) so as to optimise the sensitivity of AuNP-based immunosensors. Herein, we developed a fluorescence-based method to directly quantify rabbit immunoglobulin G (IgG) used as antibody model bound to AuNP. Rabbit IgG was first labelled with fluorescein-5-isothiocyanate (FITC) prior to conjugation to AuNP via either physisorption or chemisorption. IgG-conjugated AuNP were treated with NaCN to dissolve the AuNP and restore the fluorescence emission that was quenched in the presence of the metallic colloids, followed by quantification of fluorescein by spectrofluorimetry. This direct assay gave about 4 IgG bound to each 15-nm diameter AuNP for both immobilization strategies. This surface coverage value was in good agreement with that determined from the theoretical value calculated from the Localized Surface Plasmon Resonance (LSPR) band shift. For comparison, we also applied two indirect methods based on the quantitation of excess IgG remaining in the supernatant using fluorescence assay or enzyme-linked immunosorbent assay (ELISA). The indirect assays, either fluorescence or ELISA, commonly used to assess the antibody coverage on AuNP, overestimated the IgG surface coverage to a large extent, since up to 3 to 4 times higher coverages were measured. Therefore, the direct fluorescence method reported in this paper appears as a valuable method for quantification of surface coverage of antibody on AuNP.


Assuntos
Anticorpos Imobilizados/química , Fluoresceína-5-Isotiocianato/química , Corantes Fluorescentes/química , Ouro/química , Imunoglobulina G/química , Nanopartículas Metálicas/química , Animais , Anticorpos Imobilizados/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Cabras , Imunoglobulina G/imunologia , Coelhos , Espectrometria de Fluorescência/métodos , Propriedades de Superfície
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