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1.
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
2.
Gut ; 69(1): 146-157, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-30723104

RESUMO

OBJECTIVE: We explored the hypothesis that TGR5, the bile acid (BA) G-protein-coupled receptor highly expressed in biliary epithelial cells, protects the liver against BA overload through the regulation of biliary epithelium permeability. DESIGN: Experiments were performed under basal and TGR5 agonist treatment. In vitro transepithelial electric resistance (TER) and FITC-dextran diffusion were measured in different cell lines. In vivo FITC-dextran was injected in the gallbladder (GB) lumen and traced in plasma. Tight junction proteins and TGR5-induced signalling were investigated in vitro and in vivo (wild-type [WT] and TGR5-KO livers and GB). WT and TGR5-KO mice were submitted to bile duct ligation or alpha-naphtylisothiocyanate intoxication under vehicle or TGR5 agonist treatment, and liver injury was studied. RESULTS: In vitro TGR5 stimulation increased TER and reduced paracellular permeability for dextran. In vivo dextran diffusion after GB injection was increased in TGR5-knock-out (KO) as compared with WT mice and decreased on TGR5 stimulation. In TGR5-KO bile ducts and GB, junctional adhesion molecule A (JAM-A) was hypophosphorylated and selectively downregulated among TJP analysed. TGR5 stimulation induced JAM-A phosphorylation and stabilisation both in vitro and in vivo, associated with protein kinase C-ζ activation. TGR5 agonist-induced TER increase as well as JAM-A protein stabilisation was dependent on JAM-A Ser285 phosphorylation. TGR5 agonist-treated mice were protected from cholestasis-induced liver injury, and this protection was significantly impaired in JAM-A-KO mice. CONCLUSION: The BA receptor TGR5 regulates biliary epithelial barrier function in vitro and in vivo through an impact on JAM-A expression and phosphorylation, thereby protecting liver parenchyma against bile leakage.


Assuntos
Sistema Biliar/fisiopatologia , Colestase Intra-Hepática/prevenção & controle , Receptores Acoplados a Proteínas-G/fisiologia , Animais , Bile/metabolismo , Ácidos e Sais Biliares/metabolismo , Moléculas de Adesão Celular/metabolismo , Células Cultivadas , Colestase Intra-Hepática/metabolismo , Impedância Elétrica , Epitélio/fisiopatologia , Ácidos Isonipecóticos/farmacologia , Ácidos Isonipecóticos/uso terapêutico , Camundongos Endogâmicos C57BL , Camundongos Knockout , Oximas/farmacologia , Oximas/uso terapêutico , Permeabilidade , Fosforilação/fisiologia , Receptores de Superfície Celular/metabolismo , Receptores Acoplados a Proteínas-G/agonistas , Transdução de Sinais/fisiologia , Proteínas de Junções Íntimas/metabolismo
3.
J Agric Food Chem ; 68(1): 138-146, 2020 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-31873011

RESUMO

Chronic exposure to minute doses of endotoxin elicits intestinal inflammation and impairs the gut barrier function, potentially resulting in systemic inflammation with elevated concentrations of biomarkers associated with metabolic syndrome. This study aimed to investigate the preventive effects of the Rubus suavissimus S. Lee leaf extract in a model of low-grade systemic inflammation. The predominant compounds found in the leaf extract are gallic acids, ellagic acid, and rubusoside. Results of the present study showed that R. suavissimus leaf extract supplementation could help preserve intestinal barrier integrity by upregulating the expression of the tight junction proteins [e.g., zonula occluden-1 (ZO-1) and junctional adhesion molecule-1 (JAMA)] and mucin (MUC)-4 and also suppress the release of plasmatic proinflammatory cytokines, including tumor necrosis factor (TNF)-α, interleukin (IL)-6, and monocyte chemotactic protein (MCP)-1, while restoring the production of anti-inflammatory adiponectin. We subsequently determined that the leaf extract contributes to restoring glucose metabolic homeostasis through maintaining insulin sensitivity. Furthermore, our mechanistic finding demonstrated that the R. suavissimus leaf extract supplementation prevented systemic inflammation-driven impaired insulin sensitivity in white adipose tissues (WATs) by modulating the expression of peroxisome-proliferator-activated receptor-γ (PPAR-γ) and insulin receptor subset-1 (IRS-1). Altogether, our findings suggest that the above supplementation contributes to restoring immune and metabolic homeostasis to enhance the overall health of the host thereby preventing the early onset of metabolic disorders such as obesity and type 2 diabetes.


Assuntos
Diabetes Mellitus Tipo 2/tratamento farmacológico , Inflamação/tratamento farmacológico , Rubus/química , Animais , Citocinas/genética , Citocinas/metabolismo , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Inflamação/induzido quimicamente , Inflamação/genética , Inflamação/imunologia , Lipopolissacarídeos/efeitos adversos , Camundongos , Camundongos Endogâmicos C57BL , PPAR gama/genética , PPAR gama/metabolismo , Folhas de Planta/química , Proteínas de Junções Íntimas/genética , Proteínas de Junções Íntimas/metabolismo
4.
Life Sci ; 238: 116971, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31634462

RESUMO

AIM: High-fat diet (HFD) intake has been associated with changes in intestinal microbiota composition, increased intestinal permeability, and onset of type 2 diabetes mellitus (T2DM). The aim of this work was twofold: 1) to investigate the structural and functional alterations of the tight junction (TJ)-mediated intestinal epithelial barrier of ileum and colon, that concentrate most of the microbiota, after exposure to a HFD for 15, 30 and 60 days, and 2) to assess the effect of in vitro exposure to free fatty acids (FFAs), one of the components of HFD, on paracellular barrier of colon-derived Caco-2 cells. METHODS/KEY FINDINGS: HFD exposure induced progressive metabolic changes in male mice that culminated in prediabetes after 60d. Morphological analysis of ileum and colon mucosa showed no signs of epithelial rupture or local inflammation but changes in the junctional content/distribution and/or cellular content of TJ-associated proteins (claudins-1, -2, -3, and occludin) in intestinal epithelia were seen mainly after a prediabetes state has been established. This impairment in TJ structure was not associated with significant changes in intestinal permeability to FITC-dextran. Exposure of Caco-2 monolayers to palmitic or linoleic acids seems to induce a reinforcement of TJ structure while treatment with oleic acid had a more diverse effect on TJ protein distribution. SIGNIFICANCE: TJ structure in distal intestinal epithelia can be specifically impaired by HFD intake at early stage of T2DM, but not by FFAs in vitro. Since the TJ change in ileum/colon was marginal, probably it does not contribute to the disease onset.


Assuntos
Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/patologia , Dieta Hiperlipídica/efeitos adversos , Mucosa Intestinal/patologia , Estado Pré-Diabético/patologia , Junções Íntimas/patologia , Animais , Células CACO-2 , Diabetes Mellitus Experimental/etiologia , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/etiologia , Diabetes Mellitus Tipo 2/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ocludina , Estado Pré-Diabético/etiologia , Estado Pré-Diabético/metabolismo , Proteínas de Junções Íntimas/metabolismo , Junções Íntimas/metabolismo , Fatores de Tempo
5.
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
6.
Int J Mol Sci ; 20(17)2019 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-31480681

RESUMO

The main function of the skin is to protect the body from the external environment. The barrier function of the skin is mainly provided by the stratum corneum, which consists of corneocytes bound with the corneodesmosomes and lamellar lipids. Skin barrier proteins like loricrin and filaggrin also contribute to the skin barrier function. In various skin diseases, skin barrier dysfunction is a common symptom, and skin irritants like detergents or surfactants could also perturb skin barrier function. Many efforts have been made to develop strategies to improve skin barrier function. Here, we investigated whether the microfluidized lysates of Lactobacillus rhamnosus (LR), one of the most widely used probiotic species for various health benefits, may improve the skin barrier function in a reconstructed human epidermis, Keraskin™. Application of LR lysate on Keraskin™ increased the expression of tight junction proteins; claudin 1 and occludin as determined by immunofluorescence analysis, and skin barrier proteins; loricrin and filaggrin as determined by immunohistochemistry and immunofluorescence analysis and qPCR. Also, the cytotoxicity of a skin irritant, sodium lauryl sulfate (SLS), was alleviated by the pretreatment of LR lysate. The skin barrier protective effects of LR lysate could be further demonstrated by the attenuation of SLS-enhanced dye-penetration. LR lysate also attenuated the destruction of desmosomes after SLS treatment. Collectively, we demonstrated that LR lysate has protective effects on the skin barrier, which could expand the utility of probiotics to skin-moisturization ingredients.


Assuntos
Epiderme/efeitos dos fármacos , Lactobacillus rhamnosus/metabolismo , Modelos Biológicos , Probióticos/farmacologia , Administração Tópica , Anticorpos/farmacologia , Biomarcadores/metabolismo , Morte Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Desmossomos/efeitos dos fármacos , Desmossomos/metabolismo , Desmossomos/ultraestrutura , Epiderme/patologia , Humanos , Proteínas de Filamentos Intermediários/metabolismo , Irritantes/toxicidade , Proteínas de Membrana/metabolismo , Permeabilidade , Rodaminas/metabolismo , Proteínas de Junções Íntimas/metabolismo
7.
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
8.
Pharm Res ; 36(10): 141, 2019 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-31367840

RESUMO

PURPOSE: The purpose of the present study was to investigate changes of blood-brain barrier (BBB) and brain parenchymal protein expression due to type II diabetes mellitus (T2DM) induced by a high-fat diet (HFD) by using SWATH-based quantitative proteomics. METHODS: Mice were fed a HFD for 2 or 10 weeks, and then SWATH-based quantitative proteomic analysis, western blot analysis, immunohistochemistry and functional transport studies were performed. RESULTS: In brain capillaries, expression levels of BBB transporters (Glut1, P-glycoprotein) and tight-junction proteins (claudin-5, occludin) were significantly reduced in HFD mice at 2 weeks, but recovered to the levels in the normal diet (ND) group at 10 weeks. P-glycoprotein function at the BBB was reduced at 2 weeks. In the cerebral cortex and hippocampus, neurofilament, which is important for neuronal function, was decreased in HFD mice at 2 weeks, but recovered at 10 weeks. CONCLUSION: Our results suggest that changes in the status of insulin resistance influence expression of BBB transporters, which in turn may alter the expression of cognitive function-related proteins.


Assuntos
Encéfalo/metabolismo , Dieta Hiperlipídica/efeitos adversos , Resistência à Insulina , Insulina/metabolismo , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Animais , Barreira Hematoencefálica/metabolismo , Encéfalo/irrigação sanguínea , Capilares/metabolismo , Diabetes Mellitus Tipo 2/etiologia , Diabetes Mellitus Tipo 2/metabolismo , Transportador de Glucose Tipo 1/metabolismo , Filamentos Intermediários/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Proteômica , Proteínas de Junções Íntimas/metabolismo
9.
Int J Mol Sci ; 20(14)2019 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-31330820

RESUMO

Maintaining a robust epithelial barrier requires the accumulation of tight junction proteins, LSR/angulin-1 and tricellulin, at the tricellular contacts. Alterations in the localization of these proteins temporarily cause epithelial barrier dysfunction, which is closely associated with not only physiological differentiation but also cancer progression and metastasis. In normal human endometrial tissues, the endometrial cells undergo repeated proliferation and differentiation under physiological conditions. Recent observations have revealed that the localization and expression of LSR/angulin-1 and tricellulin are altered in a menstrual cycle-dependent manner. Moreover, it has been shown that endometrial cancer progression affects these alterations. This review highlights the differences in the localization and expression of tight junction proteins in normal endometrial cells and endometrial cancers and how they cause functional changes in cells.


Assuntos
Neoplasias do Endométrio/metabolismo , Neoplasias/metabolismo , Receptores de Lipoproteínas/metabolismo , Proteínas de Junções Íntimas/metabolismo , Animais , Células Epiteliais/metabolismo , Feminino , Humanos , Lipólise/fisiologia , Receptores de Lipoproteínas/fisiologia , Proteínas de Junções Íntimas/fisiologia
10.
Pol J Vet Sci ; 22(2): 345-353, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31269354

RESUMO

Tight junction proteins are important for the maintenance and repair of the intestinal mucosal barrier. The present study investigated relationships among tight junction protein gene expression, porcine epidemic diarrhea virus (PEDV) infection, and intestinal mucosal morphology in piglets. We compared the expression of six tight junction proteins (ZO-1, ZO-2, Occludin, Claudin-1, Claudin-4, and Claudin-5) between seven-day-old piglets infected with PEDV and normal piglets, as well as in PEDV-infected porcine intestinal epithelial cells (IPEC-J2). We also evaluated differences in mucosal morphology between PEDV-infected and normal piglets. The expression of six tight junction protein genes was lower in PEDV-infected piglets than in the normal animals. The expression of ZO-1, ZO-2, Occludin, and Claudin-4 in the intestine tissue was significantly lower (p⟨0.05) in PEDV-infected than in normal piglets. The expression of Claudin-5 in the jejunum was significantly lower in PEDV-infected piglets than in the normal animals (p⟨0.01). The expression of Claudin-1 and Claudin-5 genes in the ileum was significantly higher in PEDV-infected piglets than in normal piglets (p⟨0.01). Morphologically, the intestinal mucosa in PEDV-infected piglets exhibited clear pathological changes, including breakage and shedding of intestinal villi. In PEDV-infected IPEC-J2 cells, the mRNA expression of the six tight junction proteins showed a downward trend; in particular, the expression of the Occludin and Claudin-4 genes was significantly lower (p⟨0.01). These data suggest that the expression of these six tight junction proteins, especially Occludin and Claudin-4, plays an important role in maintaining the integrity of the intestinal mucosal barrier and resistance to PEDV infection in piglets.


Assuntos
Infecções por Coronavirus/veterinária , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos/virologia , Proteínas de Junções Íntimas/metabolismo , Animais , Linhagem Celular , Infecções por Coronavirus/virologia , Regulação da Expressão Gênica , Mucosa Intestinal/patologia , Suínos , Doenças dos Suínos/patologia , Proteínas de Junções Íntimas/genética
11.
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
12.
Fish Shellfish Immunol ; 92: 621-628, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31260736

RESUMO

Intestine in fish is a complex multifunctional organ, not only plays roles in digestion and absorption of nutrient, but also has critical role in immunity. The present study evaluated the effects of different levels of dietary sodium butyrate [Butirex® C4 (Butirex)] on intestinal immune-,antioxidant-and tight junction-related gene expression injuvenile rainbow trout(Oncorhynchusmykiss). 240 healthy rainbow trout were dispensed in 12 fiberglass tanks appointed to four treatments [0 (control), 1.5 (B1.5), 2.5 (B2.5) and 5 (B5)g Butirex per kg diet]. After a 45-day feeding trial, the fish fed with the Butirex-supplemented diets showed higher intestinal lysozyme (LYZ), complement(ACH50) and bactericidal activities; the elevations in ACH50 and bactericidal activities depended on Butirex levels (P < 0.05). The Butirex-supplemented groups, particularly the B2.5 group, had significantly higher LYZ gene expression compared to the control group (P < 0.05). Butirex at 2.5 and 5 g/kg levels led to significantly higher IL-1ß gene expression. B2.5 and B5 had significantly lower and higher TNF-α gene expression compared to the control group (P < 0.05). The B2.5 group had significantly higher TGF-B, and significantly lower IL-8 compared to the control group (P < 0.05). The B1.5 and B2.5 group had significantly higher IL-10 gene expression compared to the control group (P < 0.05). The B2.5 and B5 groups had significantly higher SOD gene expression compared to the other groups; the highest expression was related to the B2.5 group (P < 0.05). Dietary Butirex supplementation significantly up-regulated CAT and GPx genes expression compared to the control group; the highest expression as related to the B2.5 and B5 groups (P < 0.05). The B2.5 group had significantly lower CLD12 gene expression compared to the control group (P < 0.05). The B2.5 and B5 groups had significantly higher CLD3, OCLD and ZO-1 gene expression compared to the control. The highest CLD3, ZO-1 gene expressions was related to the B2.5, and B5 groups respectively (P < 0.05). After challenge with Streptococcus iniae, B2.5 and B5 had significantly higher survival compared to the control group (55.6 ±â€¯7.70 and 68.9 ±â€¯10.2 vs. 33.3 ±â€¯6.67). In conclusion, Butirex is efficient immune stimulant and health booster in rainbow trout, which augments the fish resistance to disease. Modulation of immune components, cytokines, antioxidant system and intestinal integrity might involve in improving disease resistance in Butirex-treated fish. Although most of the examined genes were modulated by 2.5 g/kg Butirex under normal conditions, 5 g/kg level is recommended under pathogenic state to mitigate mortality.


Assuntos
Ácido Butírico/metabolismo , Resistência à Doença/efeitos dos fármacos , Doenças dos Peixes/imunologia , Imunidade Inata/efeitos dos fármacos , Oncorhynchus mykiss/imunologia , Transcriptoma/efeitos dos fármacos , Ração Animal/análise , Animais , Antioxidantes/metabolismo , Ácido Butírico/administração & dosagem , Dieta/veterinária , Suplementos Nutricionais/análise , Relação Dose-Resposta a Droga , Proteínas de Peixes/metabolismo , Intestinos/efeitos dos fármacos , Oncorhynchus mykiss/metabolismo , Distribuição Aleatória , Sódio na Dieta/administração & dosagem , Infecções Estreptocócicas/imunologia , Streptococcus iniae/fisiologia , Proteínas de Junções Íntimas/metabolismo , Transcriptoma/imunologia
13.
BMC Dev Biol ; 19(1): 15, 2019 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-31277570

RESUMO

BACKGROUND: Prostate androgen-regulated mucin-like protein 1 (PARM1) is a pro-proliferative and anti-apoptotic glycoprotein involved in the endoplasmic reticulum (ER) stress response. A single nucleotide polymorphism in the coding region of PARM1 has been associated with competence of bovine embryos to develop to the blastocyst stage. Here we tested the importance of PARM1 for development by evaluating consequences of reducing PARM1 mRNA abundance on embryonic development and differentiation, gene expression and resistance to ER stress. RESULTS: Knockdown of PARM1 using an anti-PARM1 GapmeR did not affect competence of embryos to develop into blastocysts but decreased the number of trophectoderm (TE) cells in the blastocyst and tended to increase the number of cells in the blastocyst inner cell mass (ICM). Treatment of embryos with anti-PARM1 GapmeR affected expression of 4 and 3 of 90 genes evaluated at the compact-morula and blastocyst stage of development at days 5.5 and 7.5 after fertilization, respectively. In morulae, treatment increased expression of DAB2, INADL, and STAT3 and decreased expression of CCR2. At the blastocyst stage, knockdown of PARM1 increased expression of PECAM and TEAD4 and decreased expression of CCR7. The potential role of PARM1 in ER stress response was determined by evaluating effects of knockdown of PARM1 on development of embryos after exposure to heat shock or tunicamycin and on expression of ATF6, DDIT3 and EIF2AK3 at the compact morula and blastocyst stages. Both heat shock and tunicamycin reduced the percent of embryos becoming a blastocyst but response was unaffected by PARM1 knockdown. Similarly, there was no effect of knockdown on steady-state amounts of ATF6, DDIT3 or EIF2AK3. CONCLUSION: PARM1 participates in formation of TE and ICM cells in early embryonic development but there is no evidence for the role of PARM1 in the ER stress response.


Assuntos
Proteína de Ligação a Androgênios/genética , Blastocisto/citologia , Desenvolvimento Embrionário/genética , Estresse do Retículo Endoplasmático/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Animais , Bovinos , Diferenciação Celular/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Polimorfismo de Nucleotídeo Único/genética , RNA Mensageiro/genética , Receptores CCR2/metabolismo , Receptores CCR7/metabolismo , Fator de Transcrição STAT3/metabolismo , Proteínas de Junções Íntimas/metabolismo , Tunicamicina/farmacologia , Proteínas Ativadoras de ras GTPase/metabolismo
14.
Discov Med ; 27(149): 197-200, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31361982

RESUMO

Ulcerative colitis is a life-long, chronic, relapsing and remitting inflammatory disease of the large intestine with an unpredictable course characterized by debilitating gastrointestinal symptoms accompanied by healthcare and emotional burdens that reduce the quality of life and the ability to work, attend school, and be productive. Ulcerative colitis affects millions of people worldwide and is now considered a global disease. Although some form of primary immune abnormality is thought to underlie this illness, extensive laboratory research conducted since the mid-20th century has largely failed to definitively establish a primary antecedent immune abnormality in individuals with ulcerative colitis or their family members. An alternative approach employing a systems pathogenesis analysis has implicated a causal role for colonocyte-generated hydrogen peroxide in the pathogenesis of this illness. Significantly elevated levels of hydrogen peroxide in non-inflamed colonic mucosa have been demonstrated in individuals with ulcerative colitis, implying a build-up prior to the onset of inflammation and supporting a causal role for colonocyte hydrogen peroxide in the development of this disease. Hydrogen peroxide's unique properties of cell membrane permeability, long life, potent oxidizing potential, and the ability to attract white blood cells combine to promote oxidative disintegration of colonic epithelial tight junctional proteins while attracting white blood cells into the colonic epithelium, both of which lead to colonic inflammation and eventual ulcerative colitis.


Assuntos
Colite Ulcerativa , Colo , Mucosa Intestinal , Proteínas de Junções Íntimas , Colite Ulcerativa/imunologia , Colite Ulcerativa/metabolismo , Colite Ulcerativa/patologia , Colite Ulcerativa/terapia , Colo/imunologia , Colo/metabolismo , Colo/patologia , Feminino , Humanos , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Masculino , Proteínas de Junções Íntimas/imunologia , Proteínas de Junções Íntimas/metabolismo
15.
Mediators Inflamm ; 2019: 5796491, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31354386

RESUMO

Escherichia coli Nissle 1917 (EcN), a kind of probiotic, has been reported to have a protective effect on the intestinal barrier function and can ameliorate certain gastrointestinal disorders. In this study, the potential protective effect of EcN on the intestinal barrier function in a septic mouse model induced by cecal ligation and puncture (CLP) operation was investigated. FITC-Dextran 4,000 Da (FD-4) flux and the expression levels of tight junction (TJ) proteins were measured to evaluate the protective effect of EcN on the intestinal barrier function. Then, Caco-2 monolayers were utilized to further investigate the protective effect of the EcN supernatant (EcNsup) on the barrier dysfunction induced by TNF-α and IFN-γ in vitro; the plasma level of both the cytokines increased significantly during sepsis. Transepithelial electrical resistance (TEER) and FD-4 transmembrane flux were measured, and the localization of ZO-1 and Occludin was investigated by immunofluorescence. The expression of MLCK and the phosphorylation of MLC were detected by western blot. The activation of NF-κB was explored by immunofluorescence, and CHIP assays were performed to investigate the conjunction of NF-κB with the promoter of MLCK. The results indicated that EcN protected the intestinal barrier function in sepsis by ameliorating the altered expression and localization of TJ proteins and inhibiting the NF-κB-mediated activation of the MLCK-P-MLC signaling pathway which might be one of the mechanisms underlying the effect of EcN.


Assuntos
Escherichia coli/fisiologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , NF-kappa B/metabolismo , Animais , Western Blotting , Células CACO-2 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ocludina/metabolismo , Fosforilação/fisiologia , Sepse/metabolismo , Transdução de Sinais , Proteínas de Junções Íntimas/metabolismo , Junções Íntimas/metabolismo , Fator de Transcrição RelA/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
16.
Life Sci ; 231: 116529, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31173781

RESUMO

AIMS: The present study investigated the effect of Escherichia coli Nissle 1917 (EcN) on irinotecan-induced intestinal barrier dysfunction and gut microbial dysbiosis in a mouse model and in the human colonic cells lines Caco-2. MATERIALS AND METHODS: Male BALB/c mice received irinotecan intraperitoneal injection with or without EcN administration intragastrically. Body weight, diarrhea severity, intestinal permeability and histopathological analysis of ileum epithelia of mice from different groups were assessed. The expression and localization of tight junction proteins were examined using western blot and immunofluorescence. Gut microbiota structure and diversity were measured with 16 S rRNA sequencing. Caco-2 monolayers were incubated with EcN culture supernatant (EcNsup) or SN-38 and the monolayer barrier function was assessed by transepithelial electrical resistance (TER) and FITC-dextran 4000 Da (FD-4) flux. KEY FINDINGS: Pretreatment with EcN significantly attenuated irinotecan-induced weight loss and diarrhea in mice. In addition, EcN inhibited the increased intestinal permeability and decreased Claudin-1 expression in irinotecan-treated mice. Furthermore, irinotecan treatment decreased the diversity of gut microbiota and increased the relative abundance of Proteobacteria compared to control group. EcN administration ameliorated the gut microbiota dysbiosis. In Caco-2 monolayers, EcNsup ameliorated the decreased TER and increased FD-4 flux elicited by SN-38. Moreover, EcNsup attenuated SN-38-induced altered localization and distribution of Claudin-1 in Caco-2 monolayers. SIGNIFICANCE: Our results indicated that the administration of EcN protected against irinotecan-induced intestinal injury by regulating intestinal barrier function and gut microbiota.


Assuntos
Microbioma Gastrointestinal/efeitos dos fármacos , Mucosa Intestinal/efeitos dos fármacos , Animais , Células CACO-2 , Claudina-1/metabolismo , Diarreia/patologia , Disbiose/tratamento farmacológico , Escherichia coli/metabolismo , Gastroenteropatias/metabolismo , Microbioma Gastrointestinal/genética , Humanos , Enteropatias/metabolismo , Mucosa Intestinal/metabolismo , Intestinos/efeitos dos fármacos , Irinotecano/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Probióticos/farmacologia , Proteínas de Junções Íntimas/metabolismo
17.
Food Funct ; 10(7): 4124-4133, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31237597

RESUMO

Protopanaxatriol saponin (PPT) has excellent anti-cancer, anti-diabetes, and anti-anemia effects, but its effect on intestinal bacteria has rarely been studied. In this study, we investigated whether PPT has the ability to improve intestinal health in antibiotic-treated mice. Model mice were constructed using a broad-spectrum antibiotic, cephalosporin. The composition of the gut microbiota and relative concentration of short-chain fatty acids (SCFAs), short-chain fatty acid receptor proteins (GPR41, GPR43 and GPR109A), tight junction components (ZO-1 and occludin) and pro-inflammatory cytokines (TNF-α, IL-6, IL-1ß, IL-22 and IFN-γ) were determined. The results showed that PPT improved the composition of the gut microbiota, increased the concentration of SCFAs as well as receptor proteins and tight junction proteins, and decreased the pro-inflammatory cytokines. These findings indicate that PPT has a protective effect on intestinal microbes and enhances the integrity of the intestinal barrier as well as alleviates colonic inflammation in antibiotic-treated mice.


Assuntos
Antibacterianos/efeitos adversos , Microbioma Gastrointestinal/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Sapogeninas/farmacologia , Saponinas/farmacologia , Animais , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Peso Corporal , Cefalosporinas/farmacologia , Colo/metabolismo , Colo/patologia , Citocinas/metabolismo , Disbiose/prevenção & controle , Ácidos Graxos Voláteis/sangue , Ácidos Graxos Voláteis/metabolismo , Fezes/química , Microbioma Gastrointestinal/fisiologia , Inflamação/prevenção & controle , Intestinos/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Ocludina/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Proteínas de Junções Íntimas/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
18.
Mol Biol Cell ; 30(16): 1938-1960, 2019 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-31188739

RESUMO

During morphogenesis, cells must change shape and move without disrupting tissue integrity. This requires cell-cell junctions to allow dynamic remodeling while resisting forces generated by the actomyosin cytoskeleton. Multiple proteins play roles in junctional-cytoskeletal linkage, but the mechanisms by which they act remain unclear. Drosophila Canoe maintains adherens junction-cytoskeletal linkage during gastrulation. Canoe's mammalian homologue Afadin plays similar roles in cultured cells, working in parallel with ZO-1 proteins, particularly at multicellular junctions. We take these insights back to the fly embryo, exploring how cells maintain epithelial integrity when challenged by adherens junction remodeling during germband extension and dorsal closure. We found that Canoe helps cells maintain junctional-cytoskeletal linkage when challenged by the junctional remodeling inherent in mitosis, cell intercalation, and neuroblast invagination or by forces generated by the actomyosin cable at the leading edge. However, even in the absence of Canoe, many cells retain epithelial integrity. This is explained by a parallel role played by the ZO-1 homologue Polychaetoid. In embryos lacking both Canoe and Polychaetoid, cell junctions fail early, with multicellular junctions especially sensitive, leading to widespread loss of epithelial integrity. Our data suggest that Canoe and Polychaetoid stabilize Bazooka/Par3 at cell-cell junctions, helping maintain balanced apical contractility and tissue integrity.


Assuntos
Junções Aderentes/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Células Epiteliais/metabolismo , Proteínas de Junções Íntimas/metabolismo , Animais , Forma Celular , Citoesqueleto/metabolismo , Drosophila melanogaster/embriologia , Desenvolvimento Embrionário , Epiderme/metabolismo , Homeostase , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Morfogênese , Mutação/genética , Fenótipo , Pseudópodes/metabolismo
19.
Int J Biol Macromol ; 136: 27-34, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31185242

RESUMO

Lactobacillus acidophilus NCFM, a probiotic generally regarded as safe, carries a proteinaceous surface (S) layer, composed of numerous identical subunits (surface layer protein, Slp). S-layer proteins have been confirmed to possess multiple biological properties, but their role in maintaining the intestinal epithelial barrier is not fully known. We investigated the effects of Slp on tumor necrosis factor (TNF)-α-elicited intestinal barrier dysfunction and explored the underlying molecular mechanism. TNF-α administration markedly induced intestinal epithelial injury and inflammation in Caco-2 cells. Preincubation of Caco-2 cells with Slp at concentrations ranging from 50 to 100 µg/mL for 6 h improved intestinal epithelial cell integrity and permeability, restored ZO-1 and Occludin protein expressions (P < 0.05) and reduced the secretion of interleukin 8 by a maximum of 47.8%. Furthermore, the addition of Slp to Caco-2 cell monolayers attenuated cell apoptosis and inhibited nuclear factor-κB (NF-κB) p65 nucleus translocation by suppressing the activation of NF-κB. Collectively, the ability of Slp to attenuate dysfunction of the intestinal epithelial barrier stimulated by TNF-α and to exert anti-inflammatory effects supports its potential use in the development of functional foods and in the prevention of inflammatory bowel diseases.


Assuntos
Proteínas de Bactérias/farmacologia , Mucosa Intestinal/efeitos dos fármacos , Lactobacillus acidophilus , Glicoproteínas de Membrana/farmacologia , Fator de Necrose Tumoral alfa/farmacologia , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Proteínas de Bactérias/uso terapêutico , Células CACO-2 , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Inflamação/tratamento farmacológico , Interleucina-8/metabolismo , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Glicoproteínas de Membrana/uso terapêutico , Permeabilidade/efeitos dos fármacos , Proteínas de Junções Íntimas/metabolismo , Fator de Transcrição RelA/metabolismo
20.
Eur J Pharmacol ; 857: 172456, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31220438

RESUMO

Intestinal barrier injury is recognized as the main reason for ulcerative colitis (UC) which is a chronic inflammatory disease and the effective therapeutic methods remain limited. In this study, we explored the protection effects of naringenin, nobiletin and hesperetin from Citrus aurantium on colonic mucus layer integrity. The UC mice model was induced by trinitrobenzenesulfonic acid (TNBS). Forty mg/kg/day of naringenin, nobiletin and hesperetin were administrated orally for 7 and 3 days before and after TNBS treatment. Then body weight, disease activity index (DAI) score and colon length were recorded. Colons were collected for immunofluorescence, RT-PCR and western blot assay. Lipopolysaccharide (LPS)-induced cell-co-culture system of Caco-2 and RAW264.7 cells were used to determine activities on protecting intestinal epithelial monolayers damage. Our results showed that Naringenin, nobiletin and hesperetin alleviated body weight loss and colon shortness, decreased DAI score, and up-regulated claudin-2, occludin and zona occludens-1 (ZO-1) expression significantly. Meanwhile, the flavonoids enhanced trans epithelial electric resistance, decreased the permeability and up-regulated occludin and ZO-1 expression in LPS-damaged epithelial monolayers system. Hence, Naringnien, nobiletin and hesperetin show the regulatory effect on UC by protecting colonic mucosa layer integrity. It is suggested the flavonoids may be new supplements for the treatment of UC.


Assuntos
Citrus/química , Colite Ulcerativa/tratamento farmacológico , Colite Ulcerativa/patologia , Colo/efeitos dos fármacos , Flavonoides/farmacologia , Mucosa Intestinal/efeitos dos fármacos , Ácido Trinitrobenzenossulfônico/farmacologia , Animais , Células CACO-2 , Colite Ulcerativa/induzido quimicamente , Colo/patologia , Citoproteção/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Mucosa Intestinal/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células RAW 264.7 , Proteínas de Junções Íntimas/metabolismo
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