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1.
Chemosphere ; 238: 124594, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31445334

RESUMO

Enhancing the biodegradation efficiency of atrazine, a kind of commonly applied herbicide, has been attracted much more concern. Here, Zn2+ which has long been considered essential in adjusting cell physiological status was selected to investigate its role on the biodegradation of atrazine by Arthrobacter sp. DNS10 as well as the transmembrane transport of atrazine during the biodegradation period. The results of gas chromatography showed that the atrazine removal percentages (initial concentration was 100 mg L-1) in 0.05 mM Zn2+ and 1.0 mM Zn2+ treatments were 94.42% and 86.02% respectively at 48 h, while there was also 66.43% of atrazine left in the treatment without exogenous Zn2+ existence. The expression of atrazine chlorohydrolase gene trzN in the strain DNS10 cultured with 0.05 mM and 1.0 mM Zn2+ was 7.30- and 4.67- times respectively compared with that of the non-zinc treatment. In addition, the flow cytometry test suggests that 0.05 mM of Zn2+ could better adjust the membrane permeability of strain DNS10, meanwhile, the amount of atrazine accumulation in the strain DNS10 co-cultured with this level Zn2+ was 2.21 times of that of the strain without Zn2+. This study may facilitate a better understanding of the mechanisms that exogenous Zn2+ enhances the biodegradation of atrazine by Arthrobacter sp. DNS10.


Assuntos
Arthrobacter/metabolismo , Atrazina/análise , Herbicidas/análise , Hidrolases/biossíntese , Zinco/farmacologia , Biodegradação Ambiental/efeitos dos fármacos , Transporte Biológico/fisiologia , Hidrolases/genética , Permeabilidade/efeitos dos fármacos , Microbiologia do Solo
2.
J Photochem Photobiol B ; 200: 111645, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31671371

RESUMO

Antimicrobial peptide W3R6 was derived from chensinin-1b and showed potential as a novel antibiotics. However, W3R6 was susceptible to protease cleavage, which limited its therapeutic application. To improve the proteolytic resistance of W3R6, D-amino acids were incorporated into its sequence by specific amino acid substitution or whole sequence substitution according to the specificity of the cleavage site. In this study, partially substituted analog D-Arg-W3R6 and completely substituted D-enantiomer D-W3R6 were synthesized. The resistance of D-Arg-W3R6 and D-W3R6 to cleavage by the tested protease increased, particularly of D-W3R6. The antimicrobial activity of D-Arg-W3R6 was almost the same as that of the parent peptide W3R6, but the antimicrobial activity of D-W3R6 was slightly decreased. The hemolytic activity of both D-Arg-W3R6 and D-W3R6 was negligible. The CD spectrum of D-W3R6 exhibited symmetry with that of W3R6 in a membrane-mimetic environment. The membrane interaction between the D-amino acid substituted analogs and a real/mimic bacterial cell membrane was examined. The outer membrane depolarization, inner membrane permeability and dye leakage in three types of liposomes treated with D-Arg-W3R6 and D-W3R6 were not obviously different from W3R6, which could be due to the similar physical and chemical properties. In addition, these three peptides showed the binding ability with LPS micelles detected by ITC, and their ability to disrupt the LPS micelles was examined by DLS experiment and even neutralize the surface negative charge of E. coli cells. These results suggest that D-Arg-W3R6 is a promising antibiotic molecule.


Assuntos
Aminoácidos/química , Anti-Infecciosos/química , Peptídeos Catiônicos Antimicrobianos/química , Anti-Infecciosos/síntese química , Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/síntese química , Peptídeos Catiônicos Antimicrobianos/farmacologia , Parede Celular/química , Parede Celular/efeitos dos fármacos , Parede Celular/metabolismo , Eritrócitos/citologia , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Hemólise/efeitos dos fármacos , Humanos , Lipossomos/química , Lipossomos/metabolismo , Peptídeos/química , Permeabilidade/efeitos dos fármacos , Estabilidade Proteica
3.
Cell Physiol Biochem ; 53(5): 865-886, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31724838

RESUMO

BACKGROUND/AIMS: Heart failure is characterized by chronic low-grade vascular inflammation, which in itself can lead to endothelial dysfunction. Clinical trials showed reductions in heart failure-related hospitalizations of type 2 diabetic patients using sodium glucose co-transporter 2 inhibitors (SGLT2i's). Whether and how SGLT2i's directly affect the endothelium under inflammatory conditions is not completely understood. The aim of the study was to investigate whether the SGLT2i Empagliflozin (EMPA) and Dapagliflozin (DAPA) reduce tumor necrosis factor α (TNFα) induced endothelial inflammation in vitro. METHODS: Human coronary arterial endothelial cells (HCAECs) and human umbilical vein endothelial cells (HUVECs) were (pre-)incubated with 1 µM EMPA or DAPA and subsequently exposed to 10 ng/ml TNFα. ROS and NO were measured using live cell imaging. Target proteins were either determined by infrared western blotting or fluorescence activated cell sorting (FACS). The connection between Cav-1 and eNOS was determined by co-immunoprecipitation. RESULTS: Nitric oxide (NO) bioavailability was reduced by TNFα and both EMPA and DAPA restored NO levels in TNFα-stimulated HCAECs. Intracellular ROS was increased by TNFα, and this increase was completely abolished by EMPA and DAPA in HCAECs by means of live cell imaging. eNOS signaling was significantly disturbed after 24 h when cells were exposed to TNFα for 24h, yet the presence of both SGLT2is did not prevent this disruption. TNFα-induced enhanced permeability at t=24h was unaffected in HUVECs by EMPA. Similarly, adhesion molecule expression (VCAM-1 and ICAM-1) was elevated after 4h TNFα (1.5-5.5 fold increase of VCAM-1 and 4-12 fold increase of ICAM-1) but were unaffected by EMPA and DAPA in both cell types. Although we detected expression of SGLT2 protein levels, the fact that we could not silence this expression by means of siRNA and the mRNA levels of SGLT2 were not detectable in HCAECs, suggests aspecificity or our SGLT2 antibody and absence of SGLT2 in our cells. CONCLUSION: These data suggest that EMPA and DAPA rather restore NO bioavailability by inhibiting ROS generation than by affecting eNOS expression or signaling, barrier function and adhesion molecules expression in TNFα-induced endothelial cells. Furthermore, the observed effects cannot be ascribed to the inhibition of SGLT2 in endothelial cells.


Assuntos
Compostos Benzidrílicos/farmacologia , Regulação para Baixo/efeitos dos fármacos , Glucosídeos/farmacologia , Óxido Nítrico/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Vasos Coronários/citologia , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Permeabilidade/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transportador 2 de Glucose-Sódio/genética , Transportador 2 de Glucose-Sódio/metabolismo , Molécula 1 de Adesão de Célula Vascular
4.
J Biochem Mol Toxicol ; 33(11): e22397, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31557363

RESUMO

Fumonisins (Fums) are mycotoxins widely distributed in crops and feed, and ingestion of Fums-contaminated crops is harmful to animal health. The purpose of this study is to explore the effect of Fum B1 (FB1 ) on barrier functions of porcine intestinal epithelial cells, IPEC-J2, to clarify the intestinal toxicity of Fums in pigs. The results showed that the persistent treatment of FB1 significantly decreased the viability of IPEC-J2. Moreover, the expressions of Claudin 1, Occludin, Zonula Occluden-1 (ZO-1) on the messenger RNA (mRNA), and protein levels and MUC1 on the mRNA level were significantly inhibited after FB1 treatment, while the mRNA relative expression level of MUC2 was clearly increased. FB1 also enhanced the monolayer cell permeability of IPEC-J2. Importantly, FB1 promoted the expression of phosphorylated extracellular regulated protein kinase (p-ERK1/2 ). These data suggest that long-term treatment of FB1 can suppress IPEC-J2 proliferation, damage tight junctions of IPEC-J2, and regulate expression of mucins to induce the damage of barrier functions of porcine intestinal epithelial cells, which may be associated with the ERK1/2 phosphorylation pathway.


Assuntos
Células Epiteliais/metabolismo , Fumonisinas/farmacologia , Mucosa Intestinal/citologia , Micotoxinas/farmacologia , Permeabilidade/efeitos dos fármacos , Junções Íntimas/efeitos dos fármacos , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Fusarium/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Mucina-1/genética , Mucina-1/metabolismo , Mucina-2/genética , Mucina-2/metabolismo , Fosforilação/efeitos dos fármacos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/efeitos dos fármacos , Suínos , Proteínas de Junções Íntimas/genética , Proteínas de Junções Íntimas/metabolismo , Junções Íntimas/metabolismo
5.
Eur J Pharm Biopharm ; 144: 40-49, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31505225

RESUMO

AIM: The aim of the present study was to develop zeta potential changing self-emulsifying drug delivery systems (SEDDS) via a flip-flop mechanism in order to improve their mucus permeating and cellular uptake properties. METHODS: Phosphorylated serine-oleylamine (p-Ser-OA) conjugates were synthesized and incorporated into SEDDS at a concentration of 1% (v/v). Cytotoxic potential of p-Ser-OA and p-Ser-OA loaded SEDDS was investigated on Caco-2 cells. Phosphate release was evaluated using isolated as well as cell-associated intestinal alkaline phosphatase (AP). In parallel, change in zeta potential and amino group concentration on the surface of SEDDS was determined. Furthermore, mucus permeation and cellular uptake studies were performed. RESULTS: p-Ser-OA was synthesized by covalent attachment of serine (Ser) to oleylamine (OA) via a carbodiimide-mediated reaction followed by phosphorylation using phosphorous pentoxide (P2O5) and phosphoric acid (H3PO4). The chemical structure of p-Ser-OA was confirmed via FT-IR, 1H NMR, 13C NMR, 31P NMR and mass spectroscopic analysis. p-Ser-OA loaded SEDDS exhibited a droplet size and zeta potential of 46.42 ±â€¯0.35 nm and -11.53 mV, respectively. A significant amount of phosphate was released after incubation with isolated as well as cell-associated AP within 6 h and zeta potential raised up to -2.04 mV. p-Ser-OA loaded SEDDS showed improved mucus permeation in comparison to p-Ser-OA loaded SEDDS treated with AP. Moreover, cellular uptake increased almost 2-fold after phosphate cleavage using AP. CONCLUSION: Findings of this study show that SEDDS changing their zeta potential via a flip-flop mechanism exhibit both high mucus permeating and high cellular uptake properties.


Assuntos
Emulsões/química , Células Epiteliais/metabolismo , Muco/metabolismo , Células CACO-2 , Linhagem Celular Tumoral , Sistemas de Liberação de Medicamentos/métodos , Emulsificantes/química , Humanos , Permeabilidade/efeitos dos fármacos , Fosfatos/química , Ácidos Fosfóricos/química
6.
Mater Sci Eng C Mater Biol Appl ; 104: 109964, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31499990

RESUMO

Choroidal neovascularization (CNV) is the pathological growth of new blood vessels in the sub-retinal pigment epithelial (RPE) space from the choroid through a break in the Bruch's membrane (BM). Despite its importance in studying biological processes and drug discovery, the development of an in vitro CNV model that achieves the physiological structures of native RPE-BM-choroidal capillaries (CC) is still challenging. Here, we develop a novel 3D RPE-BM-CC complex biomimetic system on an ultra-thin, free-standing nanofiber membrane. The thickness of the pristine nanofiber membrane is 2.17 ±â€¯0.81 µm, and the Matrigel-coated nanofiber membrane attains a permeability coefficient of 2.95 ±â€¯0.25 × 10-6 cm/s by 40 kDa FITC-dextran, which is similar to the physiological value of the native BM. On the in vitro 3D RPE-BM-CC complex system, we demonstrate endothelial cell invasion across the 3D RPE-BM-CC complex and the mechanism of the invasion by imposing a hypoxic condition, which is thought to be the major pathological cause of CNV. Furthermore, alleviation of the invasion is achieved by treating with chrysin and anti-VEGF antibody. Thus, the in vitro 3D RPE-BM-CC complex biomimetic system can recapitulate essential features of the pathophysiological environment and be employed for the screening of drug candidates to reduce the number of costly and time-consuming in vivo tests or clinical trials.


Assuntos
Lâmina Basilar da Corioide/patologia , Neovascularização de Coroide/patologia , Hipóxia/patologia , Nanofibras/química , Biomimética/métodos , Linhagem Celular , Colágeno/química , Combinação de Medicamentos , Células Endoteliais/patologia , Flavonoides/química , Humanos , Laminina/química , Permeabilidade/efeitos dos fármacos , Proteoglicanas/química , Epitélio Pigmentado da Retina/patologia
7.
Environ Pollut ; 254(Pt A): 112960, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31394344

RESUMO

That an alteration of the intestinal permeability is associated with gut barrier function has been increasingly evident, which plays an important role in human and animal health. Bisphenol A (BPA), an industrial compound used worldwide, has recently been classified as an environmental pollutant. One of our earlier studies has demonstrated that BPA disrupts the intestinal barrier function by inducing apoptosis and inhibiting cell proliferation in the human colonic epithelial cells line. In this study, we investigated the effects of dietary BPA uptake on the colonic barrier function in mice, as well as the intestinal permeability. Dietary BPA uptake was observed to destroy the morphology of the colonic epithelium and increase the pathology score. The levels of endotoxin, diamine peroxidase, D-lactate, and zonulin were found to have been significantly elevated in both plasma and colonic mucosa. A decline in the number of intestinal goblet cells and in mucin 2 gene expression was observed in the mice belonging to the BPA group. The results of immunohistochemistry revealed that the expression of tight junction proteins (ZO-1, occludin, and claudin-1) in colonic epithelium of BPA mice decreased significantly, and their gene abundance was also inhibited. Moreover, dietary BPA uptake was also found to have significantly reduced colonic microbial diversity and altered microbial structural composition. The functional profiles of colonic bacterial community exhibited adverse effects of dietary BPA intake on the endocrine and digestive systems, as well as the transport and catabolism functions. Collectively, our study highlighted that dietary BPA increased the colonic permeability, and this effect was closely related to the disruption of intestinal chemistry and physical and biological barrier functions.


Assuntos
Compostos Benzidrílicos/toxicidade , Substâncias Perigosas/toxicidade , Intestinos/efeitos dos fármacos , Fenóis/toxicidade , Animais , Apoptose/efeitos dos fármacos , Colo , Humanos , Mucosa Intestinal/metabolismo , Masculino , Camundongos , Mucina-2 , Ocludina , Permeabilidade/efeitos dos fármacos
8.
Animal ; 13(11): 2727-2735, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31407650

RESUMO

The gut is composed of a single layer of intestinal epithelial cells and plays important roles in the digestion and absorption of nutrients, immune and barrier functions and amino acid metabolism. Weaning stress impairs piglet intestinal epithelium structural and functional integrities, which results in reduced feed intake, growth rates and increased morbidity and mortality. Several measures are needed to maintain swine gut development and growth performance after weaning stress. A large body of evidence indicates that, in weaning piglets, glutamine, a functional amino acid, may improve growth performance and intestinal morphology, reduce oxidative damage, stimulate enterocyte proliferation, modulate cell survival and death and enhance intestinal paracellular permeability. This review focuses on the effects of glutamine on intestinal health in piglets. The aim is to provide evidentiary support for using glutamine as a feed additive to alleviate weaning stress.


Assuntos
Suplementos Nutricionais/análise , Glutamina/farmacologia , Suínos/fisiologia , Aminoácidos/farmacologia , Ração Animal/análise , Animais , Dieta/veterinária , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Permeabilidade/efeitos dos fármacos , Estresse Fisiológico/efeitos dos fármacos , Suínos/crescimento & desenvolvimento , Desmame
9.
Mar Drugs ; 17(8)2019 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-31370332

RESUMO

The effect of collagen peptides (CPs) in intestinal mucosal protection has been approved in both cell and animal models. However, its structure-activity relationship and efficient peptide sequences are unclear, which hinders the in-depth study of its action mechanism and relative nutraceuticals and pharmaceuticals development. In this work, size exclusion chromatography, cation-exchange chromatography, and RP-HPLC were used to separate Alaska pollock skin-derived collagen hydrolysates based on their molecular weight, charge property, and hydrophobicity. The intestinal epithelial barrier function (IEBF) protective effect of separated peptide fractions were evaluated by tumor necrosis factor (TNF)-α-induced Caco-2 cell model. Results indicated that lower molecular weight (500-1000 Da) and higher hydrophilicity of CPs were related to better IEBF protective effect. Two high-efficiency IEBF protective peptide sequences, GPSGPQGSR and GPSGLLGPK with the corresponding molecular weights of 841.41 Da and 824.38 Da, were subsequently identified by UPLC-QToF-MS/MS. Their IEBF protective ability are comparable or even better than the currently used intestinal health supplements glutamine and arginine. The present findings suggested that the hydrophilic CPs, with molecular weight between 500 Da to 1000 Da, should be preferred in IEBF protective peptides preparation. GPSGPQGSR and GPSGLLGPK might have the potential of being IEBF protective ingredients used in intestinal health supplements and drugs.


Assuntos
Colágeno/farmacologia , Gadiformes , Mucosa Intestinal/efeitos dos fármacos , Peptídeos/farmacologia , Alaska , Animais , Células CACO-2 , Colágeno/química , Colágeno/isolamento & purificação , Suplementos Nutricionais , Humanos , Interações Hidrofóbicas e Hidrofílicas , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Peso Molecular , Peptídeos/química , Peptídeos/isolamento & purificação , Permeabilidade/efeitos dos fármacos , Relação Estrutura-Atividade , Espectrometria de Massas em Tandem
10.
AAPS PharmSciTech ; 20(7): 298, 2019 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-31456109

RESUMO

We aimed to investigate the absorption-enhancing effect (AEE) of caproyl-modified G2 PAMAM dendrimer (G2-AC) on peptide and protein drugs via the pulmonary route. In this study, G2 PAMAM dendrimer conjugates modified with caproic acid was synthesized, the pulmonary absorption of insulin as models with or without G2-AC were evaluated. The results indicated that G2-AC6 exhibited a greatest AEE for insulin in various caproylation levels of G2-AC. G2-AC6 could significantly enhance the absorption of insulin, and the AEE of G2-AC6 was concentration-dependent. In toxicity tests, G2-AC6 displayed no measurable cytotoxicity to the pulmonary membranes over a concentration range from 0.1% (w/v) to 1.0% (w/v). Measurements of the TEER and permeability showed that G2-AC6 significantly reduced the TEER value of CF and increased its Papp value. The results suggested that G2-AC6 could cross epithelial cells by means of a combination of paracellular and transcellular pathways. These findings suggested G2-AC6 at lower concentrations (below 1.0%, w/v) might be promising absorption enhancers for increasing the pulmonary absorption of peptide and protein drugs.


Assuntos
Materiais Biocompatíveis/metabolismo , Dendrímeros/metabolismo , Insulina/metabolismo , Nanopartículas/metabolismo , Absorção pelo Trato Respiratório/fisiologia , Animais , Materiais Biocompatíveis/administração & dosagem , Materiais Biocompatíveis/química , Dendrímeros/administração & dosagem , Dendrímeros/química , Insulina/administração & dosagem , Insulina/química , Masculino , Nanopartículas/administração & dosagem , Nanopartículas/química , Permeabilidade/efeitos dos fármacos , Ratos , Ratos Wistar , Absorção pelo Trato Respiratório/efeitos dos fármacos
11.
Eur J Pharm Biopharm ; 143: 98-105, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31425857

RESUMO

Oral delivery of peptides is challenging due to their low uptake through the small intestinal epithelium. Tight junctions, connecting the enterocytes, impede permeability, often necessitating the use of permeation enhancers in the formulation. Loading of peptide and permeation enhancer into micro-scale devices, such as microcontainers, can potentially confine the effective absorptive area through unidirectional release and thereby enhance absorption. This concept is investigated by in vitro permeation studies of insulin across Caco-2 cell and Caco-2/HT29-MTX-E12 co-culture monolayers mimicking the intestinal absorption barrier. The importance of proximity between the microcontainers and the barrier is assessed, by keeping the amounts of insulin and sodium caprate fixed throughout all experiments, while collectively orienting the unidirectional release towards the cell monolayers. Increasing the distance is observed to have a negative effect on insulin permeation matching a one-phase exponential decay function, while no difference in insulin transport is observed between Caco-2 and co-culture monolayers. Although there are no signs of cytotoxicity caused by the microcontainer material, reversible cell deterioration, as a consequence of high local concentrations of sodium caprate, becomes evident upon qualitative assessment of the cell monolayers. These results both suggest a potential of increasing oral bioavailability of peptides by the use of microcontainers, while simultaneously visualising the ability of regaining monolayer integrity upon local permeation enhancer induced toxicity.


Assuntos
Insulina/administração & dosagem , Insulina/química , Permeabilidade/efeitos dos fármacos , Administração Oral , Disponibilidade Biológica , Transporte Biológico/efeitos dos fármacos , Células CACO-2 , Linhagem Celular Tumoral , Técnicas de Cocultura/métodos , Humanos , Absorção Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Peptídeos/administração & dosagem , Peptídeos/química , Junções Íntimas/metabolismo
12.
Int J Pharm ; 569: 118567, 2019 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-31352051

RESUMO

The aim of this research was to formulate a novel nano-micellar complex carrier with intrinsically enhanced intestinal permeability for rosuvastatin calcium (RSV); as a model of BCS class III active pharmaceutical ingredients (APIs). The model drug is used primarily for treating hypercholesterolemia. Three phospholipid types with different degrees of saturation were chosen for the study. The saturation degree of the phospholipids was calculated accurately by proton NMR. A D-optimal statistical design was utilized to correlate the saturation degree of the phospholipids with the physico-chemical characteristics of the prepared nano-micellar carrier. The nature of the interaction between the phospholipids and the model drug was studied by proton NMR, photon correlation spectroscopy (PCS) and transmission electron microscopy (TEM). Molecular docking and molecular dynamics simulations were performed to understand the formation mechanism of the complex micelles on a molecular level. The results demonstrated that the interaction of the hydrophilic drug molecule with the polar head of a saturated phospholipid induces an intramolecular self-coiling of phospholipid saturated acyl chain leading to a structural transformation from a two-tailed cylindrical configuration into a one-tailed, surfactant-like configuration owing to the flexibility of the saturated chains. This transformation leads to the construction of a novel nano-micellar structure in which the drug has lower water solubility but higher lipophilicity than in traditional micelles. Permeability studies conducted on Caco-2 cells demonstrated that the novel nano-micellar carrier had superior permeability to that of the un-complexed hydrophilic drug. The optimized nano-micellar formulation showed significantly (P < 0.5) superior bioavailability in rats to that of the aqueous drug solution in terms of both the rate and extent of drug absorption. Overall, the results confirmed that the formation of the phospholipid nano-micellar complex increased the permeability of the hydrophilic BCS class III drug and converted it to a class BCS I drug by a simple and effective formulation technique.


Assuntos
Portadores de Fármacos , Micelas , Fosfolipídeos , Animais , Disponibilidade Biológica , Células CACO-2 , Portadores de Fármacos/administração & dosagem , Portadores de Fármacos/química , Portadores de Fármacos/farmacocinética , Humanos , Absorção Intestinal/efeitos dos fármacos , Masculino , Simulação de Acoplamento Molecular , Permeabilidade/efeitos dos fármacos , Fosfolipídeos/administração & dosagem , Fosfolipídeos/química , Fosfolipídeos/farmacocinética , Ratos Wistar
13.
J Dairy Sci ; 102(9): 7697-7706, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31326167

RESUMO

Nutrition plays a crucial role in human gut health through the improvement of gut barrier functionality. Donkey milk represents an interesting source of natural antimicrobial factors such as lysozyme. Recently, anti-inflammatory properties of donkey milk lysozyme activity were described in a mouse model of ileitis. The current increase of donkey milk consumption highlights the necessity to propose a healthy milk compliant with microbiological standards. This study aims to define a heat treatment of donkey milk, retaining its high lysozyme activity, and to evaluate its beneficial effects on a gut barrier impairment model due to chronic stress in mice. To perform this experiment, samples of raw donkey milk were collected in 15 distinct French farms. Microbiological analysis and lysozyme content and activity were evaluated for each sample. Then, several heat treatments were carried out to define a time and temperature combination that allowed for both a reduction in the number of total micro-organisms, increasing the shelf-life of the product, and preservation of lysozyme activity. The beneficial effect of heated donkey milk on the gut barrier of mice was evaluated and compared with raw donkey milk. We found that samples of raw donkey milk showed low total mesophilic microbial counts, and no pathogens were detected. Among the different heat-treatment procedures tested, a 2-min, 72°C combination was determined to be the most optimal time and temperature combination to preserve lysozyme activity and increase the shelf-life of donkey milk. Oral administration of this heat-treated donkey milk in mice counteracted chronic stress-induced intestinal damage, illustrated by gut hyper-permeability and low-grade inflammation, similar to raw donkey milk. We have demonstrated for the first time that oral intervention with donkey milk, optimally heat-treated to retain enzymatic lysozyme activity, improves intestinal barrier damage linked to psychological stress in mice.


Assuntos
Equidae , Temperatura Alta , Mucosa Intestinal/fisiologia , Leite/enzimologia , Muramidase/metabolismo , Estresse Fisiológico/fisiologia , Animais , Anti-Inflamatórios , Aprendizagem da Esquiva , Manipulação de Alimentos/métodos , Humanos , Mucosa Intestinal/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Leite/microbiologia , Muramidase/farmacologia , Permeabilidade/efeitos dos fármacos , Água
14.
Drug Dev Ind Pharm ; 45(10): 1635-1645, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31342792

RESUMO

Objective: The aim of the study was to formulate, cyclodextrin (CD)-polyanhydride (PA) nanoparticles (CPNs) with rosuvastatin calcium (RCa) in order to enhance the poor oral bioavailability. Methods: CPNs containing RCa/CD complexes were prepared by a modified solvent displacement method and morphological analyses, particle size (PS), polydispersity index (PDI), zeta potential (ZP), encapsulation efficiency (EE), DSC, FT-IR, XRD, 1H-NMR analyses were performed. In vitro release properties, release kinetics, cytotoxicity, in vitro permeability and pharmacokinetic studies were also studied. The stability of the formulations were evaluated during the storage period of 3 months. Results: The physicochemical studies showed that the RCa/CD complexes were well incorporated into CPNs resulted in nanosized particles (215.22 and 189.13 nm) with homogenous size distribution (PDI: 0.203 and 0.182) with relatively high incorporation capacity (76.11 and 68.18%) for the CPN1 and CPN2 formulations respectively. Sustained release of RCa from CPNs were achieved. The cytotoxicity values showed that the safety of the formulations. According to permeability studies, pure RCa had lowest permeability data (3.08 × 10-7 cm⋅s-1 Papp value) while CPNs gained higher permeability data (1.36 × 10-5 and 1.12 × 10-5 cm⋅s-1 Papp values) for the CPN1 and CPN2 formulations respectively. CPN2 formulation was selected for pharmacokinetic studies and analyses results demonstrated that approximately 8-fold relative oral bioavailability enhancement compared to the pure RCa was achieved. Conclusion: Considering the analyses results of the study, CPNs can be regarded as suitable, safe, functional oral delivery systems for RCa with enhanced oral bioavailability.


Assuntos
Ciclodextrinas/química , Ciclodextrinas/farmacocinética , Nanopartículas/química , Polianidridos/química , Polianidridos/farmacocinética , Rosuvastatina Cálcica/química , Rosuvastatina Cálcica/farmacocinética , Animais , Disponibilidade Biológica , Células CACO-2 , Linhagem Celular Tumoral , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacocinética , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Humanos , Masculino , Tamanho da Partícula , Permeabilidade/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley
15.
Eur J Pharm Biopharm ; 142: 387-395, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31306752

RESUMO

Oral administration of drug products is the preferred administration route. In recent decades there has been an increase in drug candidates with low solubility and/or low permeability. To increase the possibility of oral administration for the poorly permeating drugs, the use of absorption modifying excipients (AMEs) has been proposed. These types of AMEs may also affect the regulatory assessment of a novel drug delivery system if they affect the absorption of a drug from any of the four BCS classes. The effects of AMEs have previously been investigated in various animal models, including the single-pass intestinal perfusion (SPIP) in rats. To further improve the biorelevance and the in vivo predictiveness of the SPIP model, four compounds (atenolol, enalaprilat, ketoprofen, metoprolol) were perfused in fasted or fed state simulated intestinal fluid (FaSSIF or FeSSIF) together with the AMEs N-acetyl-cysteine, caprate, or sodium dodecyl sulfate. For the highly soluble and poorly permeating compounds enalaprilat and atenolol (BCS class III), the flux was increased the most by the addition of SDS in both FaSSIF and FeSSIF. For ketoprofen (BCS class II), the flux decreased in the presence of all AMEs in at least one of the perfusion media. The flux of metoprolol (BCS class I) was not affected by any of the excipients in none of simulated prandial states. The changes in magnitude in the absorption of the compounds were in general smaller in FeSSIF than in FaSSIF. This may be explained by a reduced free concentration AMEs in FeSSIF. Further, the results in FeSSIF were similar to those from intrajejunal bolus administration in rat in a previous study. This suggests that the biorelevance of the SPIP method may be increased when investigating the effects of AMEs, by the addition of intraluminal constituents representative to fasted and/or fed state to the inlet perfusate.


Assuntos
Excipientes/química , Jejum/metabolismo , Absorção Intestinal/efeitos dos fármacos , Jejuno/metabolismo , Preparações Farmacêuticas/química , Preparações Farmacêuticas/metabolismo , Administração Oral , Animais , Disponibilidade Biológica , Líquidos Corporais/metabolismo , Mucosa Intestinal/metabolismo , Intestinos/fisiologia , Masculino , Permeabilidade/efeitos dos fármacos , Ratos , Ratos Wistar , Dodecilsulfato de Sódio/química , Solubilidade/efeitos dos fármacos
16.
Eur J Pharm Biopharm ; 142: 353-363, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31278992

RESUMO

Tight junction (TJ) modulation is a promising approach for improving drug bioavailability by enhancing the absorption of active pharmaceutical ingredients. However, the application of many different test methods to determine the efficacy of new TJ modulators (TJMs) or to assess different compounds is accompanied by a lack of comparable results, reducing the rational evaluation and commercial marketing of these pharmaceutical excipients. The establishment of unified testing methods can fill this gap and offers the opportunity to compare results from different laboratories. Furthermore, the calculation of a TJ modulation score allows the objective comparison of TJ modulators and facilitates the selection of appropriate candidates. In this study, eight well-known TJ modulators were tested with a focus on four different in vitro bioassays carried out with MDCK cells. The extent of TJ modulation was determined by transepithelial electric resistance (TEER) measurements and permeability studies with mannitol. To evaluate tolerability, cell viability (MTT) and cytotoxicity (CellTox™ Green) assays were performed, and TEER regeneration was monitored for 24 h after exposure. With the exception of labradimil, seven TJ modulators caused significant TEER reduction of up to 100 %. For five compounds, an enhancement of mannitol permeation was observed. As expected, first-generation enhancers exhibited lower cell compatibility than mechanism-based modulators. Based on the experimental results of this study, for the first time, an evaluation system (tight junction modulator scoring system, TJMSS) is presented that provides a ranking of the tested modulators depending on weighted parameters. Such a system offers the possibility of rational formulation development for drugs requiring improved absorption.


Assuntos
Preparações Farmacêuticas/química , Junções Íntimas/efeitos dos fármacos , Animais , Disponibilidade Biológica , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cães , Excipientes/química , Células Madin Darby de Rim Canino , Manitol/química , Permeabilidade/efeitos dos fármacos
17.
Food Funct ; 10(7): 4350-4360, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31276135

RESUMO

Cinnamon is known to have several physiological effects; the effects of Cinnamomum japonicum Sieb. on anti-inflammation and tight junctions were investigated in the cellular intestinal inflammation model. Cinnamon subcritical water extract (CSWE) significantly down-regulated the protein and expression levels of nitrite, prostaglandin E2 (PGE2), interleukin (IL)-6, IL-8, tumor necrosis factor (TNF)-α, nuclear factor kappa B (NF-κB) activity, and the phosphorylation of the factors of the NF-κB pathway. It also significantly decreased the permeability but increased the transepithelial electrical resistance (TEER) value and the protein and expression levels of tight junction proteins (i.e., zonula occludens (ZO)-1, occludin, and claudin-1). Furthermore, cinnamic acid and cinnamaldehyde, the major components of C. japonicum, inhibited the phosphorylation of the NF-κB pathway and increased the tight junction protein expression. CSWE from C. japonicum may improve intestinal health by enhancing tight junctions and inhibiting inflammation of the intestines.


Assuntos
Cinnamomum zeylanicum/química , Inflamação/tratamento farmacológico , Intestinos/efeitos dos fármacos , Extratos Vegetais/farmacologia , Junções Íntimas/efeitos dos fármacos , Animais , Células CACO-2 , Claudina-1/metabolismo , Técnicas de Cocultura , Dinoprostona , Humanos , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Camundongos , NF-kappa B/metabolismo , Nitritos/metabolismo , Ocludina/metabolismo , Permeabilidade/efeitos dos fármacos , Fosforilação , Células RAW 264.7 , Proteínas de Junções Íntimas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Água/química
18.
Eur J Pharm Biopharm ; 142: 364-376, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31283980

RESUMO

Intestinal permeability is a key biopharmaceutical variable in pharmaceutical research and development, and regulatory assessment. In situ rat models are often used to predict the corresponding human intestinal permeability data. The rat single-pass intestinal perfusion (SPIP) and intestinal closed loop (ICL) models are commonly applied. The primary objective of this study was to collect, summarize, and evaluate all the available intestinal permeability data for drugs that have been obtained using these two in-situ rat models. The permeability data were also investigated for variability between the experimental designs. The literature survey found 635 permeability determinations for 90 drugs. The studies were performed on the jejunum (n = 284), whole small intestine (n = 111), colon (n = 108), ileum (n = 101), and duodenum (n = 30). All the SPIP (n = 484) and ICL (n = 147) permeability values were summarized in an easily accessible database. There was wide variability in the intestinal permeability to each drug between studies, which was unrelated to the permeability class of the drug. There was no relationship between rat intestinal permeability and luminal pH, luminal drug concentration, rat strain, experimental method, or intestinal region. There was, however, a correlation between permeability values determined in the same laboratory. This report showed that the SPIP and ICL methods are important in situ models for understanding and predicting intestinal drug absorption. However, conclusions based on permeability values sourced from different laboratories may not be reliable. Because each permeability study is unique and because between- and even within-laboratory variability can be substantial, data from individual studies should preferably be interpreted separately.


Assuntos
Absorção Intestinal/fisiologia , Mucosa Intestinal/metabolismo , Intestinos/fisiologia , Preparações Farmacêuticas/metabolismo , Animais , Feminino , Masculino , Perfusão/métodos , Permeabilidade/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Ratos Wistar
19.
Appl Microbiol Biotechnol ; 103(16): 6593-6604, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31286166

RESUMO

A novel antimicrobial peptide named NP-6 was identified in our previous work. Here, the mechanisms of the peptide against Escherichia coli (E. coli) were further investigated, as well as the peptide's resistance to temperature, pH, salinity, and enzymes. The transmission electron microscopy (TEM), confocal laser scanning microcopy (CLSM), and flow cytometric (FCM) analysis, combined with measurement of released K+, were performed to evaluate the effect of NP-6 E. coli cell membrane. The influence of NP-6 on bacterial DNA/RNA and enzyme was also investigated. The leakage of K+ demonstrated that NP-6 could increase the permeability of E. coli cell membrane. The ATP leakage, FCM, and CLSM assays suggested that NP-6 caused the disintegration of bacterial cell membrane. The TEM observation indicated that NP-6 could cause the formation of empty cells and debris. Besides, the DNA-binding assay indicated that NP-6 could bind with bacterial genomic DNA in a way that ethidium bromide (EB) did, and suppress the migration of DNA/RNA in gel retardation. Additionally, NP-6 could also affect the activity of ß-galactosidase. Finally, the effect of different surroundings such as heating, pH, ions, and protease on the antimicrobial activity of NP-6 against E. coli was also investigated. Results showed that the peptide was heat stable in the range of 60~100 °C and performed well at pH 6.0~8.0. However, the antimicrobial activity of NP-6 decreased significantly in the presence of Mg2+/Ca2+, and after incubation with trypsin/proteinase K. The results will provide a theoretical support in the further application of NP-6.


Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Membrana Celular/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Permeabilidade/efeitos dos fármacos , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/isolamento & purificação , Membrana Celular/ultraestrutura , DNA Bacteriano/metabolismo , Estabilidade de Medicamentos , Escherichia coli/ultraestrutura , Concentração de Íons de Hidrogênio , Viabilidade Microbiana/efeitos dos fármacos , Ligação Proteica , Salinidade , Sementes/química , Temperatura Ambiente , Zanthoxylum/química , beta-Galactosidase/antagonistas & inibidores
20.
J Agric Food Chem ; 67(33): 9277-9285, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31353906

RESUMO

3,3'-Diindolylmethane (DIM), a digestive metabolite originating from cruciferous vegetables, has dietary potential for the treatment of various human intestinal diseases. Although intestinal permeability dysfunction is closely related to the initiation and progression of human intestinal inflammatory diseases (IBDs), the effect of DIM on intestinal permeability is unclear. We evaluated the effect of DIM on the intestinal permeability of human intestinal cell monolayers and the animal model Caenorhabditis elegans, which were treated with IL-1ß and Pseudomonas aeruginosa, respectively, to mimic IBD conditions. DIM substantially restored the intestinal permeability of differentiated Caco-2 cells by enhancing the expression of tight junction proteins (including occludin and ZO-1). Compared to the IL-1ß single treatment (551.0 ± 49.0 Ω·cm2), DIM (10 µM) significantly increased the transepithelial electrical resistance (TEER) of Caco-2 cell monolayers (919.0 ± 66.4 Ω·cm2, p < 0.001). DIM also ameliorated the impaired intestinal permeability and extended the lifespan of C. elegans fed P. aeruginosa. The mean lifespan of DIM-treated worms (10.8 ± 1.3 days) was higher than that of control-treated worms (9.7 ± 1.1 days, p < 0.01). Thus, DIM is a potential nutraceutical candidate for the treatment of leaky gut syndrome by improving intestinal permeability.


Assuntos
Indóis/administração & dosagem , Doenças Inflamatórias Intestinais/tratamento farmacológico , Mucosa Intestinal/efeitos dos fármacos , Animais , Células CACO-2 , Caenorhabditis elegans/efeitos dos fármacos , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Mucosa Intestinal/metabolismo , Intestinos , Masculino , Ocludina/genética , Ocludina/metabolismo , Permeabilidade/efeitos dos fármacos , Proteína da Zônula de Oclusão-1/genética , Proteína da Zônula de Oclusão-1/metabolismo
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