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1.
J Sci Food Agric ; 100(1): 235-244, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31512251

RESUMO

BACKGROUND: This study evaluated the effects of early antibiotic exposure (EAE) on subsequent amino acid (AA) profiles and small intestinal AA transporter and receptor expression level in pigs with different dietary crude protein (CP) levels. Eighteen litters of piglets were fed creep feed diets, either with or without antibiotics while with sow on day 7. The pigs were weaned at day 23 and fed the same diets until day 42, when random pigs within each group were offered a normal- or low-CP diet, thereby creating four groups. On day 120, the pigs were euthanized, and jejunal and ileal mucosa and digesta were collected for gene-expression and AA-concentration analysis. RESULTS: With the normal-CP diet, EAE increased (P < 0.05) the concentrations of six essential amino acids (EAA) and three non-essential amino acids (NEAA) in serum, four EAAs and four NEAAs in jejunal mucosa, one EAA and two NEAAs in ileal mucosa, five EAAs and three NEAAs in jejunal digesta, and three EAAs and two NEAAs in ileal digesta. Early antibiotic exposure upregulated (P < 0.05) CAT1, ASCT2, ATB0,+ , CaSR, T1R1, and T1R3 expression in the jejunum, downregulated PepT1 expression with a normal-CP diet. It upregulated (P < 0.05) the expressions of CAT1, ATB0,+ , ATP1A1, and T1R3 in the ileum with a normal-CP diet. CONCLUSION: These results suggest that EAE has long-term effects on AA profiles, mainly in the jejunum and serum, by increasing AA transporter expression in the intestine, and that these effects may be influenced by dietary CP levels. © 2019 Society of Chemical Industry.


Assuntos
Sistemas de Transporte de Aminoácidos/genética , Aminoácidos/metabolismo , Antibacterianos/efeitos adversos , Mucosa Intestinal/efeitos dos fármacos , Receptores Acoplados a Proteínas-G/genética , Suínos/metabolismo , Sistemas de Transporte de Aminoácidos/metabolismo , Aminoácidos/química , Ração Animal/análise , Animais , Antibacterianos/administração & dosagem , Proteínas na Dieta/análise , Proteínas na Dieta/metabolismo , Feminino , Expressão Gênica/efeitos dos fármacos , Mucosa Intestinal/crescimento & desenvolvimento , Mucosa Intestinal/metabolismo , Masculino , Distribuição Aleatória , Receptores Acoplados a Proteínas-G/metabolismo , Suínos/genética , Suínos/crescimento & desenvolvimento , Fatores de Tempo
2.
Zhonghua Nei Ke Za Zhi ; 59(1): 40-46, 2020 Jan 01.
Artigo em Chinês | MEDLINE | ID: mdl-31887835

RESUMO

Objective: To investigate the association between adherens junction proteins E-cadherin and ß-catenin and tight junction protein claudin-2 and clinical symptoms in patients with diarrhea predominant irritable bowel syndrome (IBS-D). Methods: Cecal biopsy tissues were collected from IBS-D patients (n=26) according to Rome Ⅲ criterion and healthy controls (n=26). The duration of symptoms, abdominal pain score and mean weekly bowel movements were recorded. Colorectal dilatation combined with restraint stress were applied to establish visceral hypersensitivity rat model. Abdominal contraction reflex (AWR) was applied to assess the visceral sensitivity in rats. The stool frequency within 1 hour was recorded after establishing the rat model. The expression of E-cadherin、ß-catenin and claudin-2 were assessed by Western blot and immunofluorescence microscopy. Intercellular ultrastructure was observed by transmission electron microscopy. Results: Compared with the healthy controls, the protein expression of E-cadherin and ß-catenin in cecal epithelium in IBS-D patients were significantly lower (P=0.015 and P=0.005, respectively), while claudin-2 was significantly higher (P=0.000). Reduced E-cadherin and ß-catenin expression was associated high abdominal pain score (r=-0.463, P=0.017 and r=-0.407, P=0.039). The lower expression of ß-catenin was associated with longer duration of symptoms (r=-0.458, P=0.019). The protein expression of E-cadherin and ß-catenin in the cecal epithelium of the visceral hypersensitivity rats were significantly lower (P=0.004 and P=0.003, respectively), while claudin-2 was significantly higher (P=0.008). Reduced E-cadherin and ß-catenin expression was associated high visceral sensitivity in IBS-D rats (r=-0.639, P=0.047 and r=-0.888, P=0.001). Conclusions: Intercellular ultrastructure alterations well as cecal ß-catenin and E-cadherin protein expression decrease and are associated with high abdominal pain score in IBS-D patients and hypersensitivity rats. ß-catenin is further associated with prolonged duration of symptoms in IBS-D patients. The expression of E-cadherin and ß-catenin may play a vital role in visceral sensitivity and intestinal barrier dysfunction in IBS-D.


Assuntos
Junções Aderentes/metabolismo , Caderinas/metabolismo , Síndrome do Intestino Irritável/metabolismo , Junções Íntimas/metabolismo , Dor Abdominal/metabolismo , Dor Abdominal/fisiopatologia , Animais , Biópsia , Estudos de Casos e Controles , Ceco/metabolismo , Claudina-2 , Diarreia/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Síndrome do Intestino Irritável/patologia , Ratos , beta Catenina
3.
Gut ; 69(1): 62-73, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-30923071

RESUMO

OBJECTIVE: The intestinal lumen contains several proteases. Our aim was to determine the role of faecal proteases in mediating barrier dysfunction and symptoms in IBS. DESIGN: 39 patients with IBS and 25 healthy volunteers completed questionnaires, assessments of in vivo permeability, ex vivo colonic barrier function in Ussing chambers, tight junction (TJ) proteins, ultrastructural morphology and 16 s sequencing of faecal microbiota rRNA. A casein-based assay was used to measure proteolytic activity (PA) in faecal supernatants (FSNs). Colonic barrier function was determined in mice (ex-germ free) humanised with microbial communities associated with different human PA states. RESULTS: Patients with IBS had higher faecal PA than healthy volunteers. 8/20 postinfection IBS (PI-IBS) and 3/19 constipation- predominant IBS had high PA (>95th percentile). High-PA patients had more and looser bowel movements, greater symptom severity and higher in vivo and ex vivo colonic permeability. High-PA FSNs increased paracellular permeability, decreased occludin and increased phosphorylated myosin light chain (pMLC) expression. Serine but not cysteine protease inhibitor significantly blocked high-PA FSN effects on barrier. The effects on barrier were diminished by pharmacological or siRNA inhibition of protease activated receptor-2 (PAR-2). Patients with high-PA IBS had lower occludin expression, wider TJs on biopsies and reduced microbial diversity than patients with low PA. Mice humanised with high-PA IBS microbiota had greater in vivo permeability than those with low-PA microbiota. CONCLUSION: A subset of patients with IBS, especially in PI-IBS, has substantially high faecal PA, greater symptoms, impaired barrier and reduced microbial diversity. Commensal microbiota affects luminal PA that can influence host barrier function.


Assuntos
Síndrome do Intestino Irritável/fisiopatologia , Serina Proteases/fisiologia , Adulto , Animais , Biópsia , Células CACO-2 , Estudos de Casos e Controles , Colo/patologia , Disbiose/enzimologia , Fezes/enzimologia , Feminino , Microbioma Gastrointestinal , Humanos , Absorção Intestinal/fisiologia , Mucosa Intestinal/metabolismo , Síndrome do Intestino Irritável/enzimologia , Síndrome do Intestino Irritável/microbiologia , Síndrome do Intestino Irritável/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Permeabilidade , Estudos Prospectivos , Proteólise , Índice de Gravidade de Doença , Proteínas de Junções Íntimas/metabolismo
4.
Cell Physiol Biochem ; 53(5): 851-864, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31714043

RESUMO

BACKGROUND/AIMS: The growth promoting effect of lysine and betaine as well as the expression of candidate genes reflecting their efficacy, such as ghrelin, leptin, Growth Hormone Secretagogue Receptor (GHS-R), Insulin like Growth Factor (IGF- 1) and Growth Hormone Releasing Hormone (GHRH) was examined in Labeo rohita fingerlings. METHODS: One hundred eighty healthy juveniles from a homologous population were randomly distributed to 15 rectangular tanks of 150 litres capacity. The experiment was carried out for 60 days with five treatment groups consisting T1 (0.25% Betaine), T2 (0.5% Betaine), T3 (0.75% Lysine) and T4 (1.5% Lysine) and control group. The experiment was carried out for 60 days with five treatment groups consisting T1 (0.25% Betaine), T2 (0.5% Betaine), T3 (0.75% Lysine) and T4 (1.5% Lysine) and control group. At the end of trial, the growth parameters such as weight gain, SGR, PER were estimated from the weight of the triplicate groups. The digestive, metabolic and antioxidant enzymes were analysed using spectrophotometric methods. The intestine, brain and liver were sampled from the treatments and expression of different genes ghrelin, leptin, GHSR, IGF-1 and GHRH was also performed by realtime PCR. RESULTS: A significant (P<0.05) increase in weight gain, SGR, PER and lowest FCR was found in T4 group which was significantly (p < 0.05) different from other experimental groups. The highest mRNA expression levels of expression were found in T4 group which was similar to that of ghrelin gene mRNA of T2 group. The significantly (p<0.05) highest GHSR, GHRH and IGF-1 gene expression levels were found in T4 treatment group compared to other groups. CONCLUSION: The present study reveals that the lysine and betaine stimulate growth and expression of ghrelin GHRH, GHS-R and IGF-1 genes. The increase of IGF-I mRNA expression with lysine and betaine supplementation revealed that these compounds act as growth modulators. However, lysine was found to be a more potent modulator of growth compared to betaine.


Assuntos
Betaína/farmacologia , Cyprinidae/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Lisina/farmacologia , Ração Animal , Animais , Catalase/metabolismo , Cyprinidae/crescimento & desenvolvimento , Grelina/genética , Grelina/metabolismo , Hormônio Liberador de Hormônio do Crescimento/genética , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Fator de Crescimento Insulin-Like I/genética , Fator de Crescimento Insulin-Like I/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Leptina/genética , Leptina/metabolismo , Fígado/enzimologia , Fígado/metabolismo , Receptores de Grelina/genética , Receptores de Grelina/metabolismo , Superóxido Dismutase/metabolismo
5.
J Agric Food Chem ; 67(45): 12481-12495, 2019 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-31630515

RESUMO

Biochanin A is a dietary isoflavone with multiple biological functions. Owing to a lack of comprehensive studies of biochanin A metabolism, this study was designed to further clarify the processes involved in biochanin A metabolism. In this study, ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS/MS) was utilized to characterize the metabolism of biochanin A in vivo and in vitro. As a result, 43 metabolites in rats, 22 metabolites in liver microsomes, and 18 metabolites in intestinal flora were elucidated, and 5 metabolites were identified by comparison with standards. Oxidation, demethylation, hydrogenation, internal hydrolysis, conjugation (e.g., glucuronidation, sulfonation, glucose conjugation, methylation, and acetylation), and their composite reactions were determined to be major processes involved in biochanin A biotransformation. The results contribute to a better understanding of the pharmacological mechanism of biochanin A and provide a basis for comprehension of the safety and toxicity of biochanin A.


Assuntos
Genisteína/metabolismo , Isoflavonas/metabolismo , Animais , Biotransformação , Cromatografia Líquida de Alta Pressão , Microbioma Gastrointestinal , Genisteína/química , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Isoflavonas/química , Masculino , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Ratos , Ratos Sprague-Dawley , Espectrometria de Massas em Tandem
6.
Eur J Pharm Biopharm ; 145: 76-84, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31639417

RESUMO

Oral drug delivery is a preferred administration route due to its low cost, high patient compliance and fewer adverse events compared to intravenous administration. However, many pharmaceuticals suffer from poor solubility and low oral bioavailability. One major factor that contributes to low bioavailability are efflux transporters which prevent drug absorption through intestinal epithelial cells. P-glycoprotein (P-gp) and Breast Cancer Resistance Protein (BCRP) are two important efflux transporters in the intestine functioning to prevent toxic materials from entering systemic circulation. However, due to its broad substrate specificity, P-gp limits the absorption of many therapeutics, including chemotherapeutics and antibacterial agents. Methods to inhibit P-gp with competitive inhibitors have not been clinically successful. Here, we show that micron scale devices (microdevices) made from a commonly used biomaterial, polyethylene glycol (PEG), inhibit P-gp through a biosimilar mucus in Caco-2 cells and that transporter function is restored when the microdevices are removed. Microdevices were shown to inhibit P-gp mediated transport of calcein AM, doxorubicin, and rhodamine 123 (R123) and BCRP mediated transport of BODIPY-FL-prazosin. When in contact with Caco-2 cells, microdevices decrease the cell surface amount of P-gp without affecting the passive transport. Moreover, there was an increase in mucosal to serosal transport of R123 with microdevices in an ex-vivo mouse model and increased absorption in vivo. This biomaterial-based approach to inhibit efflux transporters can be applied to a range of drug delivery systems and allows for a nonpharmacologic method to increase intestinal drug absorption while limiting toxic effects.


Assuntos
Transporte Biológico/efeitos dos fármacos , Hidrogéis/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Disponibilidade Biológica , Compostos de Boro/metabolismo , Células CACO-2 , Linhagem Celular Tumoral , Humanos , Absorção Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Intestinos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Polietilenoglicóis/química , Prazosina/análogos & derivados , Prazosina/metabolismo , Rodamina 123/metabolismo , Solubilidade/efeitos dos fármacos
7.
J Agric Food Chem ; 67(46): 12786-12795, 2019 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-31657926

RESUMO

EPA-enriched phosphoethanolamine plasmalogens (EPA-pPE), widely present in marine creatures, is a unique glycerophospholipid with EPA at the sn-2 position of the glycerol backbone. EPA-pPE has been reported to exhibit numerous distinctive bioactivities. However, the digestion, absorption, and metabolism characteristics of EPA-pPE in vivo are not clear, which restrict the molecular mechanism analysis related to its distinctive activities. The aim of the present study was to illustrate the digestion, absorption, and metabolism characteristics of EPA-pPE by lipid analysis in serum, intestinal wall, and content after oral administration of EPA-pPE emulsion. Results showed the EPA percentage of total fatty acids in serum was increasing over time, with two peaks at 5 and 10 h by 1.89 ± 0.2 and 2.58 ± 0.27, respectively, and then fell from 1.89 ± 0.17 at 10 h to 1.35 ± 0.16 at 16 h. In small intestinal content, EPA-pPE was hydrolyzed to lyso-phospholipids and EPA by phospholipases A2 and the vinyl ether bond was retained at the sn-1 position. The released EPA could be quickly taken up into the enterocytes and enter circulation. The comparison of simulated digestion in vitro showed that the distinct digestion and absorption process of EPA-pPE was a unique phenomenon. EPA could be retained in serum at a high level for a substantial period of time, which suggested that EPA-pPE was not just a short-lived circulating molecule.


Assuntos
Ácido Eicosapentaenoico/metabolismo , Etanolaminas/metabolismo , Mucosa Intestinal/metabolismo , Plasmalogênios/metabolismo , Animais , Digestão , Ácido Eicosapentaenoico/química , Etanolaminas/química , Absorção Intestinal , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fosfolipases A2/metabolismo , Plasmalogênios/química
8.
Pharm Res ; 36(12): 172, 2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31659456

RESUMO

PURPOSE: Caco-2 monolayers are the most common model of the intestinal epithelium and are critical to the development of oral drug delivery strategies and gastrointestinal disease treatments. However, current monolayer systems are cost- and/or time-intensive, hampering progress. This study evaluates two separate methods to reduce resource input: FB Essence as a fetal bovine serum (FBS) alternative and a new, 3-day Caco-2 system deemed "thrifty, rapid intestinal monolayers" (TRIM). METHODS: Caco-2 cells were cultured with FB Essence and compared to cells in 10% FBS for proliferation and monolayer formation. TRIM were compared to commonly-used 21-day and Corning® HTS monolayer systems, as well as mouse intestines, for permeability behavior, epithelial gene expression, and tight junction arrangement. RESULTS: No amount of FB Essence maintained Caco-2 cells beyond 10 passages. In contrast, TRIM compared favorably in permeability and gene expression to intestinal tissues. Furthermore, TRIM cost $109 and required 1.3 h of time per 24-well plate, compared to $164 and 3.7 h for 21-day monolayers, and $340 plus 1.0 h for the HTS system. CONCLUSIONS: TRIM offer a new approach to generating Caco-2 monolayers that resemble the intestinal epithelium. They are anticipated to accelerate the pace of in vitro intestinal experiments while easing financial burden.


Assuntos
Mucosa Intestinal/metabolismo , Administração Oral , Animais , Células CACO-2 , Proliferação de Células , Células Cultivadas , Colágeno/química , Dextranos/metabolismo , Liberação Controlada de Fármacos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Permeabilidade , Propriedades de Superfície , Junções Íntimas/metabolismo
9.
Nature ; 574(7779): 532-537, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31645730

RESUMO

The colorectal adenoma-carcinoma sequence has provided a paradigmatic framework for understanding the successive somatic genetic changes and consequent clonal expansions that lead to cancer1. However, our understanding of the earliest phases of colorectal neoplastic changes-which may occur in morphologically normal tissue-is comparatively limited, as for most cancer types. Here we use whole-genome sequencing to analyse hundreds of normal crypts from 42 individuals. Signatures of multiple mutational processes were revealed; some of these were ubiquitous and continuous, whereas others were only found in some individuals, in some crypts or during certain periods of life. Probable driver mutations were present in around 1% of normal colorectal crypts in middle-aged individuals, indicating that adenomas and carcinomas are rare outcomes of a pervasive process of neoplastic change across morphologically normal colorectal epithelium. Colorectal cancers exhibit substantially increased mutational burdens relative to normal cells. Sequencing normal colorectal cells provides quantitative insights into the genomic and clonal evolution of cancer.


Assuntos
Colo/citologia , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Mutação , Sintomas Prodrômicos , Reto/citologia , Adenoma/genética , Adenoma/patologia , Idoso , Proteína Axina/genética , Carcinoma/genética , Carcinoma/patologia , Transformação Celular Neoplásica , Células Clonais/citologia , Células Clonais/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Variações do Número de Cópias de DNA , Análise Mutacional de DNA , Feminino , Humanos , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Masculino , Pessoa de Meia-Idade , Células-Tronco/citologia , Células-Tronco/metabolismo
10.
Eur J Pharm Biopharm ; 144: 125-131, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31526841

RESUMO

AIM: The aim of this study was to evaluate the potential of chitosan/tripolyphosphate (TPP) nanoparticles to provide a targeted release of ß-galactosidase behind the intestinal mucus gel barrier. METHODS: Nanoparticles were prepared by ionic gelation of chitosan and TPP in the presence of ß-galactosidase. Particles were characterized regarding size, polydispersity index and drug load. Target mediated hydrolysis of the TPP cross-linker followed by particle degradation and release of ß-galactosidase was investigated during incubation with isolated as well as cell and tissue associated intestinal alkaline phosphatase (IAP). Phosphate content in the media was quantified via malachite assay, whereas particle disintegration was monitored in parallel by measuring the decrease in particle size as well as in optical density at 600 nm. The released amount of ß-galactosidase was either determined utilizing bicinchoninic acid (BCA) protein detection or via an enzymatic activity assay with 2-nitrophenyl ß-D-galactopyranoside (ONPG) as substrate. Protection towards tryptic degradation was verified by ONPG assay. RESULTS: The size of nanoparticles was 573 ±â€¯34 nm and a payload of 376 ±â€¯18 µg ß-galactosidase per mg particles was achieved. Degradation studies with isolated IAP revealed a maximum phosphate cleavage of 118 ±â€¯1 µg/mg particles, a size decrease up to 38 ±â€¯7 % and a release of 58 ±â€¯0.5 % ß-galactosidase. Release of 94 ±â€¯6 % of the incorporated initial amount of ß-galactosidase was proven after 3 h incubation on porcine mucosa. Furthermore a protection against tryptic degradation was attained resulting in a 3-fold higher residual enzymatic activity of encapsulated ß-galactosidase compared to a control of free enzyme. CONCLUSION: Chitosan/TPP nanoparticles seem to be qualified as a suitable carrier for a targeted delivery of active ingredients to mucosal tissues expressing alkaline phosphatase.


Assuntos
Quitosana/análogos & derivados , Géis/química , Mucosa Intestinal/metabolismo , Muco/metabolismo , Nanopartículas/química , beta-Galactosidase/química , beta-Galactosidase/metabolismo , Animais , Células CACO-2 , Linhagem Celular Tumoral , Quitosana/química , Sistemas de Liberação de Medicamentos/métodos , Humanos , Intestinos , Tamanho da Partícula , Suínos
11.
Toxicol Lett ; 316: 109-118, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31472180

RESUMO

Lithocholic acid (LCA) is both a secondary bile acid and a vitamin D receptor (VDR) ligand. The VDR is activated by 1,25-dihydroxy vitamin D3 and plays an important role in maintaining integrity of the intestinal mucosal barrier. LCA can also substitute for vitamin D to carry out the in vivo functions of vitamin D. However, it is unclear whether activation of the VDR by LCA affects mucosal barrier function. In the present study, we researched the protective effect of LCA on tumor necrosis factor-alpha (TNF-α)-induced intestinal epithelial barrier dysfunction in Caco-2 cells of the human epithelial intestinal adenocarcinoma cell line. Caco-2 cell monolayers were pretreated with LCA and then exposed to 100 ng/mL TNF-α. The results showed that LCA alleviated the decrease in transepithelial electrical resistance and the increase in FITC-Dextran flux induced by TNF-α. LCA ameliorated the TNF-α-induced decrease in protein expression and distribution of ZO-1, E-cadherin, Occludin, and Claudin-1, which are tight junction markers. Additionally, the LCA treatment effectively counteracted TNF-α-mediated downregulation of silent information regulator 1 (SIRT1), nuclear factor erythroid2-related factor 2 (Nrf2), and heme oxygenase-1, which are related to oxidative stress. Increases in NF-κB p-p65 and p-IκB-α induced by TNF-α were significantly inhibited by LCA. Considering all these, the present study indicates that LCA has a significant protective effect on TNF-α-induced injury of intestinal barrier function through the VDR and suggests that suppressing NF-κB signaling and activating the SIRT1/Nrf2 pathway might be one of the mechanisms underlying the protective effect of LCA.


Assuntos
Células Epiteliais/efeitos dos fármacos , Absorção Intestinal/efeitos dos fármacos , Mucosa Intestinal/efeitos dos fármacos , Ácido Litocólico/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Receptores de Calcitriol/agonistas , Sirtuína 1/metabolismo , Fator de Necrose Tumoral alfa/toxicidade , Células CACO-2 , Citoproteção , Impedância Elétrica , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Permeabilidade , Receptores de Calcitriol/genética , Receptores de Calcitriol/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas de Junções Íntimas/metabolismo , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo , Junções Íntimas/patologia
12.
J Agric Food Chem ; 67(39): 10871-10879, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31517482

RESUMO

This study evaluated the effect of triterpenoids from edible mushroom Poria cocos on intestinal epithelium integrity and revealed the transcriptional regulatory pathways that underpin restorative mechanisms in the gut. Based on computational docking studies, transcriptional activation experiments and glucocorticoid receptor (GR) protein immunofluorescence localization assays in cultured cells, 16α-hydroxytrametenolic acid (HTA) was discovered as a novel GR agonist in this study. HTA ameliorates TNF-α-induced Caco-2 monolayer intestinal epithelial barrier damage and suppressed activation of phosphatidylinositol 3-kinase (PI3K) and protein kinase B (Akt), which attenuated downstream IκB and nuclear factor kappa-B (NF-κB) phosphorylation through GR activation. Moreover, HTA prevented NF-κB translocation into the nucleus and binding to its cis-element and suppressed lipopolysaccharide-induced downstream NO production and pro-inflammatory cytokines at both protein and mRNA expression levels. In conclusion, HTA from P. cocos improves intestinal barrier function through a GR-mediated PI3K/Akt/NF-κB signaling pathway and may be potentially exploited as a supportive dietary therapeutic strategy for restoring gut health.


Assuntos
Mucosa Intestinal/efeitos dos fármacos , NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Extratos Vegetais/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Glucocorticoides/metabolismo , Triterpenos/farmacologia , Wolfiporia/química , Células CACO-2 , Humanos , Proteínas I-kappa B/genética , Proteínas I-kappa B/metabolismo , Mucosa Intestinal/metabolismo , Simulação de Acoplamento Molecular , NF-kappa B/genética , Fosfatidilinositol 3-Quinase/genética , Fosforilação , Extratos Vegetais/química , Proteínas Proto-Oncogênicas c-akt/genética , Receptores de Glucocorticoides/genética , Transdução de Sinais/efeitos dos fármacos , Triterpenos/química , Verduras/química
13.
Anticancer Res ; 39(9): 4965-4970, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31519602

RESUMO

BACKGROUND/AIM: Colonic crypts with normal epithelium albeit with corrupted shapes (CCS) were previously found beneath nonpolypoid adenomas (NPA). This study aimed to analyze the distribution of proliferating cells (PC) and p53-up-regulated cells in CCS. MATERIALS AND METHODS: Sections from 48 NPA were immunostained with the proliferating-marker Ki67 and against the tumor-suppressor protein p53. RESULTS: Asymmetric-haphazardly distributed PC were found in 87.5% of the NPA, continuous PC-domains in 8.3%, asymmetric-haphazardly distributed single PC in 4.2% and p53-up-regulated cells in 29.2%. In 12 controls, the normal-shaped crypts revealed symmetrically-distributed PC-domains in their lower thirds, and no p53-up-regulated cells. CONCLUSION: The normal epithelium that lines the CCS below NPA, thrives with relocated PC-domains, and with occasional p53-up-regulated cells. These findings strongly suggest that the normal epithelium of CCS beneath NPA might harbor somatic mutations. The accretion of putative mutated CCS might play an important role in the evolution of nonpolypoid adenomas in the human colon.


Assuntos
Adenoma/genética , Neoplasias do Colo/genética , Regulação Neoplásica da Expressão Gênica , Mucosa Intestinal/metabolismo , Proteína Supressora de Tumor p53/genética , Adenoma/metabolismo , Adenoma/patologia , Proliferação de Células , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Humanos , Fenótipo , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima
14.
J Agric Food Chem ; 67(42): 11638-11649, 2019 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-31532204

RESUMO

Naturally occurring cinnamon compounds such as cinnamaldehyde (CAL) and structurally related constituents have been associated with antiobesity activities, although studies regarding the impact on intestinal fatty acid uptake are scarce. Here, we demonstrate the effects of CAL and structural analogues cinnamyl alcohol (CALC), cinnamic acid (CAC), and cinnamyl isobutyrate on mechanisms regulating intestinal fatty acid uptake in differentiated Caco-2 cells. CAL, CALC, and CAC (3000 µM) were found to decrease fatty acid uptake by 58.0 ± 8.83, 19.4 ± 8.98, and 21.9 ± 6.55%, respectively. While CAL and CALC at a concentration of 300 µM increased serotonin release 14.9 ± 3.00- and 2.72 ± 0.69-fold, respectively, serotonin alone showed no effect on fatty acid uptake. However, CAL revealed transient receptor potential channel A1-dependency in the decrease of fatty acid uptake, as well as in CAL-induced serotonin release. Overall, CAL was identified as the most potent of the cinnamon constituents tested.


Assuntos
Acroleína/análogos & derivados , Cinamatos/farmacologia , Cinnamomum zeylanicum/química , Ácidos Graxos/metabolismo , Extratos Vegetais/farmacologia , Propanóis/farmacologia , Acroleína/química , Acroleína/farmacologia , Transporte Biológico/efeitos dos fármacos , Células CACO-2 , Diferenciação Celular , Cinamatos/química , Humanos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Extratos Vegetais/química , Propanóis/química
15.
J Agric Food Chem ; 67(41): 11464-11473, 2019 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-31532211

RESUMO

The intestinal epithelium is derived from intestinal stem cells (ISCs) and has direct contact with nutrients and toxins. However, whether methionine (Met) or a methionine hydroxyl analogue (2-hydroxy-4-(methylthio)butanoic acid (HMB)) can alleviate deoxynivalenol (DON)-induced intestinal injury remains unknown. Mice were treated orally with Met or HMB on days 1-11 and with DON on days 4-8. On day 12, the mice were sacrificed, and the jejunum was collected for crypt isolation and culture. Mouse enteroids were treated with DON and Met or HMB ex vivo. The results showed that Met and HMB increased the average daily feed intake and average daily gain of the mice. Met and HMB also improved the jejunal structure and barrier integrity and promoted ISC expansion, as indicated by the increased enteroid formation efficiency and area, under DON-induced injury conditions. In addition, DON-induced decreases in ISC activity were rescued Wnt/ß-catenin signaling reactivation by Met or HMB in vivo and ex vivo. Collectively, our findings reveal that Met and HMB alleviated DON-induced intestinal injury by improving ISC expansion and reactivating Wnt/ß-catenin signaling. Our study thus provides a nutritional intervention for intestinal diseases involving Wnt/ß-catenin signaling.


Assuntos
Enteropatias/tratamento farmacológico , Metionina/análogos & derivados , Metionina/administração & dosagem , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Tricotecenos/toxicidade , Via de Sinalização Wnt/efeitos dos fármacos , Animais , Proliferação de Células/efeitos dos fármacos , Humanos , Enteropatias/genética , Enteropatias/metabolismo , Enteropatias/fisiopatologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/lesões , Mucosa Intestinal/metabolismo , Masculino , Camundongos , Células-Tronco/metabolismo , beta Catenina/genética , beta Catenina/metabolismo
16.
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
17.
Vet Immunol Immunopathol ; 216: 109919, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31446207

RESUMO

Mucosal surfaces such as the gastrointestinal tract, and skin are the front line of host defence and immunity against many pathogens. Gamma delta (γδ) T lymphocytes preferentially localize to the mucosal surfaces in several species including cattle, and are thought to play crucial roles in immunosurveillance and host defence, particularly against mycobacteria. Many γδ T cells are present in young calves, which is the period when calves are thought to be initially exposed to Mycobacterium avium subspecies paratuberculosis (Map). The role of mucosal γδ T cells in cattle, especially during host-pathogen interactions during early pre-clinical phases of infectious disease remains unclear. The purposes of this study were to investigate and characterize WC1+ and WC1neg γδ  T cell subsets in various segments of the gastrointestinal (GI) tract of young calves, and then to examine γδ  T cell subsets in the distal small intestine of calves after experimental intestinal Map infection by direct Peyer's patch inoculation. We show that in healthy calves, the relative proportion of γδ T cells is constant throughout the GI mucosa, though the ileum has significantly more γδ T cells. In the distal intestine, γδ T cells are mainly WC1neg and primarily located within the lamina propria of the jejunum and ileum. In Map-infected intestine, there are higher numbers of γδ T cells in the lamina propria and a greater proportion of WC1+ cells within the epithelial layer compared to control calves. While WC1neg γδ T cells preferentially localize to the distal small intestine of healthy calves, WC1+ γδ T cells are increased in the intestinal mucosa during Map infection, which is suggestive of effector cell function. Further, spectral microscopy and flow cytometry in tandem will lead to improved understanding of the functions of these cells during health and disease.


Assuntos
Doenças dos Bovinos/imunologia , Mucosa Intestinal/metabolismo , Glicoproteínas de Membrana/metabolismo , Mycobacterium avium subsp. paratuberculosis , Paratuberculose/imunologia , Subpopulações de Linfócitos T/metabolismo , Animais , Bovinos , Sobrevivência Celular , Células Cultivadas , Regulação da Expressão Gênica/imunologia , Mucosa Intestinal/patologia , Glicoproteínas de Membrana/genética , Paratuberculose/metabolismo
18.
J Agric Food Chem ; 67(34): 9477-9491, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31429552

RESUMO

Lipopolysaccharide (LPS) is a bacterial endotoxin that induces intestine inflammation. Milk exosomes improve the intestine and immune system development of newborns. This study aims to establish the protective mechanisms of porcine milk exosomes on the attenuation of LPS-induced intestinal inflammation and apoptosis. In vivo, exosomes prevented LPS-induced intestine damage and inhibited (p < 0.05) LPS-induced inflammation. In vitro, exosomes inhibited (p < 0.05) LPS-induced intestinal epithelial cells apoptosis (23% ± 0.4% to 12% ± 0.2%). Porcine milk exosomes also decreased (p < 0.05) the LPS-induced TLR4/NF-κB signaling pathway activation. Furthermore, exosome miR-4334 and miR-219 reduced (p < 0.05) LPS-induced inflammation through the NF-κB pathway and miR-338 inhibited (p < 0.05) the LPS-induced apoptosis via the p53 pathway. Cotransfection with these three miRNAs more effectively prevented (p < 0.05) LPS-induced cell apoptosis than these miRNAs individual transfection. The apoptosis percentage in the group cotransfected with the three miRNAs (14% ± 0.4%) was lower (p < 0.05) than that in the NC miRNA group (28% ± 0.5%), and also lower than that in each individual miRNA group. In conclusion, porcine milk exosomes protect the intestine epithelial cells against LPS-induced injury by inhibiting cell inflammation and protecting against apoptosis through the action of exosome miRNAs. The presented results suggest that the physiological amounts of miRNAs-enriched exosomes addition to infant formula could be used as a novel preventative measure for necrotizing enterocolitis.


Assuntos
Apoptose , Células Epiteliais/citologia , Exossomos/metabolismo , MicroRNAs/metabolismo , Leite/metabolismo , NF-kappa B/metabolismo , Receptor 4 Toll-Like/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Células Epiteliais/metabolismo , Exossomos/genética , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Lipopolissacarídeos/efeitos adversos , Masculino , Camundongos , MicroRNAs/genética , NF-kappa B/genética , Transdução de Sinais , Suínos , Receptor 4 Toll-Like/genética , Proteína Supressora de Tumor p53/genética
19.
Life Sci ; 234: 116778, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31430454

RESUMO

AIMS: To clarify the role of the gut-brain axis in depression. MAIN METHODS: We used the iTRAQ technique to identify differential proteins in the intestine of the rat model of chronic unpredictable mild stress (CUMS)-induced depression. Significant differential proteins were subjected to Gene Ontology (GO) functional annotations and KEGG pathway enrichment analysis. Key proteins were validated at the mRNA and protein levels. The levels of cytokines in the intestine, serum and hypothalamus were examined by ELISA. HPLC-UV was used to detect the levels of amino acids. KEY FINDINGS: In the rat intestine, 349 differential proteins (209 downregulated, 140 upregulated) were identified. GO analysis indicated that "protein complex assembly" was the first-ranked biological process. SNARE complex components, including SNAP23, VAMP3 and VAMP8, were increased at the mRNA levels, while only VAMP3 and VAMP8 were also upregulated at the protein level. TNFα, IL6 and IL1ß were upregulated in the CUMS rat intestine, while TNFα was decreased in the serum and hypothalamus. IL1ß was decreased in the serum. "Protein digestion and absorption" was the most significantly enriched KEGG pathway, involving 5 differential proteins: SLC9A3, ANPEP, LAT1, ASCT2 and B0AT1. Glutamine, glycine and aspartic acid were perturbed in the CUMS rat intestine. SIGNIFICANCE: Our findings suggest that CUMS enhances the adaptive immune response in the intestine through ER-phagosome pathway mediated by SNARE complex and disturb absorption of amino acids. It advances our understanding of the role of gut-brain axis in depression and provides a potential therapeutic target for the disease.


Assuntos
Aminoácidos/análise , Citocinas/análise , Depressão/patologia , Mucosa Intestinal/metabolismo , Intestinos/patologia , Proteínas SNARE/análise , Aminoácidos/metabolismo , Animais , Citocinas/genética , Depressão/etiologia , Depressão/genética , Modelos Animais de Doenças , Regulação da Expressão Gênica , Absorção Intestinal , Masculino , Proteômica , Ratos , Ratos Sprague-Dawley , Proteínas SNARE/genética , Estresse Psicológico/complicações
20.
Acta Cir Bras ; 34(6): e201900610, 2019 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-31433001

RESUMO

PURPOSE: To identify whether the colon mucosa is affected by ten days of gastric restriction in an animal model. METHODS: An experimental model of gastric restriction was devised using rats. The animals were submitted to surgical gastrostomy, and a cylindrical loofah was inserted into the stomach. We studied 30 adult male Wistar rats divided into three groups: the stomach restriction group (R10); the sham group (S10), which underwent the same procedure except for the loofah insertion; and the control group (C10). The expression of neutral and acid mucins was evaluated using histochemical techniques. Goblet cells and protein content were compared between groups using generalized estimation equations (GEEs). Bonferroni's multiple comparison was applied to identify differences between the groups. All tests considered a 5% significance level. RESULTS: There was an increased expression of neutral mucins, acid mucins and goblet cells in the R10 group. Collagen was also enhanced in the R10 group. CONCLUSION: The colon mucosa is affected by ten days of gastric restriction in an animal model, increasing neutral mucins, acid mucins and collagen content with trophic maintenance.


Assuntos
Privação de Alimentos , Mucosa Intestinal/metabolismo , Mucinas/metabolismo , Animais , Colo , Gastrostomia , Mucosa Intestinal/patologia , Masculino , Modelos Animais , Ratos , Ratos Wistar , Fatores de Tempo
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