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1.
J Toxicol Sci ; 45(9): 539-548, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32879253

RESUMO

We investigated the mechanism underlying intestinal cadmium (Cd) uptake based on the mediators (metal transporters) of essential elements, such as Fe, Zn, Cu, and Ca, under normal conditions in female rats. These elements interact with Cd uptake from the intestinal tract. Cd concentration at each site of the small intestine (duodenum, jejunum, and ileum) increased as Cd exposure increased. However, Cd concentration was the highest in the duodenum. The gene expression of ZIP14, DMT1, and ATP7A increased with increase in Cd concentration. Further, Cu concentration decreased as Cd concentration increased. In contrast, Fe concentration displayed a decreasing tendency with the increase in Cd concentration. The gene expression levels of ZIP14, DMT1, and ATP7A were positively correlated with Cd concentration. Immunohistochemical staining revealed the positive sites of ZIP14 and DMT1 scattered in the area adjacent to the goblet cells, resorbable epithelial cells, and lamina propria in the duodenum tissue, according to the increase in Cd concentration. Cd is induced to synthesize and bind to metallothionein (MT-I and -II) and accumulate in the intestinal tissues, mainly in the duodenum. Such findings suggest that Cd, a contaminant element, is taken up from the intestinal tract by multiple metal transporters such as Cu, Fe, and Zn, thereby involving in the intestinal Cd absorption.


Assuntos
Cádmio/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Absorção Intestinal/genética , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismo , Animais , Proteínas de Transporte de Cátions/genética , Cobre , ATPases Transportadoras de Cobre/genética , ATPases Transportadoras de Cobre/metabolismo , Duodeno/metabolismo , Feminino , Expressão Gênica , Ferro , Metalotioneína/metabolismo , Ratos , Zinco
2.
PLoS One ; 15(8): e0237101, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32817686

RESUMO

Mutations in the genes encoding for voltage-gated sodium channels cause profound sensory disturbances and other symptoms dependent on the distribution of a particular channel subtype in different organs. Humans with the gain-of-function mutation p.Leu811Pro in SCN11A (encoding for the voltage-gated Nav1.9 channel) exhibit congenital insensitivity to pain, pruritus, self-inflicted injuries, slow healing wounds, muscle weakness, Charcot-like arthropathies, and intestinal dysmotility. As already shown, knock-in mice (Scn11a+/L799P) carrying the orthologous mutation p.Leu799Pro replicate reduced pain sensitivity and show frequent tissue lesions. In the present study we explored whether Scn11a+/L799P mice develop also pruritus, muscle weakness, and changes in gastrointestinal transit time. Furthermore, we analyzed morphological and functional differences in nerves, skeletal muscle, joints and small intestine from Scn11a+/L799P and Scn11a+/+ wild type mice. Compared to Scn11a+/+ mice, Scn11a+/L799P mice showed enhanced scratching bouts before skin lesions developed, indicating pruritus. Scn11a+/L799P mice exhibited reduced grip strength, but no disturbances in motor coordination. Skeletal muscle fiber types and joint architecture were unaltered in Scn11a+/L799P mice. Their gastrointestinal transit time was unaltered. The small intestine from Scn11a+/L799P showed a small shift towards less frequent peristaltic movements. Similar proportions of lumbar dorsal root ganglion neurons from Scn11a+/L799P and Scn11a+/+ mice were calcitonin gene-related peptide (CGRP-) positive, but isolated sciatic nerves from Scn11a+/L799P mice exhibited a significant reduction of the capsaicin-evoked release of CGRP indicating reduced neurogenic inflammation. These data indicate important Nav1.9 channel functions in several organs in both humans and mice. They support the pathophysiological relevance of increased basal activity of Nav1.9 channels for sensory abnormalities (pain and itch) and suggest resulting malfunctions of the motor system and of the gastrointestinal tract. Scn11a+/L799P mice are suitable to investigate the role of Nav1.9, and to explore the pathophysiological changes and mechanisms which develop as a consequence of Nav1.9 hyperactivity.


Assuntos
Mutação com Ganho de Função , Debilidade Muscular/genética , Canal de Sódio Disparado por Voltagem NAV1.9/genética , Prurido/genética , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Feminino , Trânsito Gastrointestinal , Força da Mão , Intestino Delgado/metabolismo , Intestino Delgado/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Movimento , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Canal de Sódio Disparado por Voltagem NAV1.9/metabolismo , Nervo Isquiático/metabolismo , Nervo Isquiático/patologia
3.
Amino Acids ; 52(6-7): 1063-1065, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32627059

RESUMO

Gastrointestinal symptoms are common in COVID-19 patients, especially in younger patients. Our hypothesis was that intestinal SARS-CoV-2 receptor ACE2 expression depends on patients' age. We examined duodenal biopsies from 43 healthy human adults. ACE2 gene expression was directly correlated with age (Spearman's r = 0.317, p = 0.039). With each year, duodenal ACE2 expression increased by 0.083 RU. The higher intestinal ACE2 mRNA expression in older patients may impact on their susceptibility to develop intestinal symptoms.


Assuntos
Betacoronavirus/metabolismo , Intestino Delgado/metabolismo , Peptidil Dipeptidase A/genética , Receptores Virais/genética , Adulto , Fatores Etários , Idoso , Feminino , Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Peptidil Dipeptidase A/metabolismo , RNA Mensageiro/metabolismo , Receptores Virais/metabolismo , Adulto Jovem
4.
Proc Natl Acad Sci U S A ; 117(28): 16292-16301, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601208

RESUMO

Notch pathway signaling is implicated in several human cancers. Aberrant activation and mutations of Notch signaling components are linked to tumor initiation, maintenance, and resistance to cancer therapy. Several strategies, such as monoclonal antibodies against Notch ligands and receptors, as well as small-molecule γ-secretase inhibitors (GSIs), have been developed to interfere with Notch receptor activation at proximal points in the pathway. However, the use of drug-like small molecules to target the downstream mediators of Notch signaling, the Notch transcription activation complex, remains largely unexplored. Here, we report the discovery of an orally active small-molecule inhibitor (termed CB-103) of the Notch transcription activation complex. We show that CB-103 inhibits Notch signaling in primary human T cell acute lymphoblastic leukemia and other Notch-dependent human tumor cell lines, and concomitantly induces cell cycle arrest and apoptosis, thereby impairing proliferation, including in GSI-resistant human tumor cell lines with chromosomal translocations and rearrangements in Notch genes. CB-103 produces Notch loss-of-function phenotypes in flies and mice and inhibits the growth of human breast cancer and leukemia xenografts, notably without causing the dose-limiting intestinal toxicity associated with other Notch inhibitors. Thus, we describe a pharmacological strategy that interferes with Notch signaling by disrupting the Notch transcription complex and shows therapeutic potential for treating Notch-driven cancers.


Assuntos
Receptores Notch/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Ativação Transcricional/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Sítios de Ligação , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Drosophila , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Células HeLa , Humanos , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/química , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/genética , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/metabolismo , Camundongos , Mutação , Fenótipo , Multimerização Proteica , Transdução de Sinais/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/uso terapêutico
6.
Int J Exp Pathol ; 101(3-4): 80-86, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32567731

RESUMO

ApcMin/+ mice are regarded as a standard animal model of colorectal cancer (CRC). Tensin4 (TNS4 or Cten) is a putative oncogene conferring features of stemness and promoting motility. Our objective was to assess TNS4 expression in intestinal adenomas and determine whether TNS4 is upregulated by Wnt signalling. ApcMin/+ mice (n = 11) were sacrificed at approximately 120 days old at the onset of anaemia signs. Small intestines were harvested, and Swiss roll preparations were tested for TNS4 expression by immunohistochemistry (IHC). Individual polyps were also separately collected (n = 14) and tested for TNS4 mRNA expression and Kras mutation. The relationship between Wnt signalling and TNS4 expression was tested by Western blotting in the human CRC cell line HCT116 after inhibition of ß-catenin activity with MSAB or its increase by transfection with a Flag ß-catenin expression vector. Overall, 135/148 (91.2%) of the total intestinal polyps were positive for TNS4 expression by IHC, whilst adjacent normal areas were negative. RT-qPCR analysis showed approximately 5-fold upregulation of TNS4 mRNA in the polyps compared to adjacent normal tissue and no Kras mutations were detected. In HCT116, ß-catenin inhibition resulted in reduced TNS4 expression, and conversely, ß-catenin overexpression resulted in increased TNS4 expression. In conclusion, this is the first report linking aberrant Wnt signalling to upregulation of TNS4 both during initiation of intestinal adenomas in mice and in in vitro models. The exact contribution of TNS4 to adenoma development remains to be investigated, but the ApcMin/+ mouse represents a good model to study this.


Assuntos
Pólipos Adenomatosos/metabolismo , Genes APC , Neoplasias Intestinais/metabolismo , Intestino Delgado/metabolismo , Tensinas/metabolismo , Via de Sinalização Wnt , Pólipos Adenomatosos/genética , Pólipos Adenomatosos/patologia , Animais , Modelos Animais de Doenças , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Humanos , Neoplasias Intestinais/genética , Neoplasias Intestinais/patologia , Intestino Delgado/patologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Tensinas/genética , Regulação para Cima , beta Catenina/metabolismo
7.
Life Sci ; 255: 117838, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32450168

RESUMO

AIMS: Dysregulation of iron homeostasis in the body causes a variety of diseases. Iron deficiency leads to anemia, whereas iron overload aggravates cellular oxidative stress. Nuclear factor erythroid-2-related factor 2 (Nrf2) is a protein that is activated in the nucleus and turns on the production of antioxidant enzymes, protecting cell against oxidative damage. This study aimed to investigate whether Nrf2 gene knockout influences iron homeostasis in aging mice. MATERIALS AND METHODS: Iron content and iron metabolism-related proteins were assessed in different organs and blood serum of the 18 month-old Nrf2 knockout (Nrf2-/-) mice in comparison with the wild-type (WT) mice. KEY FINDINGS: Results showed that the iron contents in spleen and liver all increased, and expression levels of iron transporters were altered in Nrf2-/- mice. In particularly, we found that the expression of iron export protein ferroportin 1 (Fpn1) in liver, spleen and small intestine all decreased in Nrf2-/- mice, which might account for the deposition of iron in different organs and the increased ROS. Surprisingly, we found that the serum iron level of Nrf2-/- mice did not decrease, but increased significantly even when the iron absorption at small intestine decreased. Our further investigation revealed that the increase of serum iron was due to the release of iron from the hemolysis of erythrocytes, which caused by the increased ROS level in red blood cells of the Nrf2-/- mice. SIGNIFICANCE: These findings provide a more comprehensive understanding of the important role of Nrf2 in the regulation of systemic iron metabolism.


Assuntos
Hemólise/fisiologia , Ferro/metabolismo , Fator 2 Relacionado a NF-E2/genética , Espécies Reativas de Oxigênio/metabolismo , Envelhecimento/fisiologia , Animais , Eritrócitos/citologia , Homeostase/fisiologia , Intestino Delgado/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Baço/metabolismo
8.
PLoS One ; 15(5): e0233863, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32470053

RESUMO

Adaptive regulation of epithelial transporters to nutrient intake is essential to decrease energy costs of their synthesis and maintenance, however such regulation is understudied. Previously we demonstrated that the transport function of the basolateral amino acid uniporter LAT4 (Slc43a2) is increased by dephosphorylation of serine 274 (S274) and nearly abolished by dephosphorylation of serine 297 (S297) when expressed in Xenopus oocytes. Phosphorylation changes in the jejunum of food-entrained mice suggested an increase in LAT4 transport function during food expectation. Thus, we investigated further how phosphorylation, expression and localization of mouse intestinal LAT4 respond to food-entrained diurnal rhythm and dietary protein content. In mice entrained with 18% protein diet, LAT4 mRNA was not submitted to diurnal regulation, unlike mRNAs of luminal symporters and antiporters. Only in duodenum, LAT4 protein expression increased during food intake. Concurrently, S274 phosphorylation was decreased in all three small intestinal segments, whereas S297 phosphorylation was increased only in jejunum. Interestingly, during food intake, S274 phosphorylation was nearly absent in ileum and accompanied by strong phosphorylation of mTORC1 target S6. Entraining mice with 8% protein diet provoked a shift in jejunal LAT4 localization from the cell surface to intracellular stores and increased S274 phosphorylation in both jejunum and ileum during food anticipation, suggesting decreased transport function. In contrast, 40% dietary protein content led to increased LAT4 expression in jejunum and its internalization in ileum. Ex vivo treatments of isolated intestinal villi fraction demonstrated that S274 phosphorylation was stimulated by protein kinase A. Rapamycin-sensitive insulin treatment and amino acids increased S297 phosphorylation, suggesting that the response to food intake might be regulated via the insulin-mTORC1 pathway. Ghrelin, an oscillating orexigenic hormone, did not affect phosphorylation of intestinal LAT4. Overall, we show that phosphorylation, expression and localization of intestinal mouse LAT4 responds to diurnal and dietary stimuli in location-specific manner.


Assuntos
Sistema y+ de Transporte de Aminoácidos/metabolismo , Ritmo Circadiano , Proteínas na Dieta/farmacologia , Alimentos , Intestinos/fisiologia , Aminoácidos/metabolismo , Animais , Antiporters/metabolismo , Ritmo Circadiano/efeitos dos fármacos , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Grelina/administração & dosagem , Grelina/farmacologia , Insulina/metabolismo , Intestino Delgado/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos Endogâmicos C57BL , Microvilosidades/efeitos dos fármacos , Microvilosidades/metabolismo , Fosforilação/efeitos dos fármacos , Fosfosserina/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Frações Subcelulares/metabolismo , Simportadores/metabolismo , Serina-Treonina Quinases TOR/metabolismo
9.
Am J Physiol Gastrointest Liver Physiol ; 318(6): G1070-G1087, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32390462

RESUMO

Lipopolysaccharides (LPS) are potent pro-inflammatory molecules that enter the systemic circulation from the intestinal lumen by uncertain mechanisms. We investigated these mechanisms and the effect of exogenous glucagon-like peptide-2 (GLP-2) on LPS transport in the rodent small intestine. Transmucosal LPS transport was measured in Ussing-chambered rat jejunal mucosa. In anesthetized rats, the appearance of fluorescein isothiocyanate (FITC)-LPS into the portal vein (PV) and the mesenteric lymph was simultaneously monitored after intraduodenal perfusion of FITC-LPS with oleic acid and taurocholate (OA/TCA). In vitro, luminally applied LPS rapidly appeared in the serosal solution only with luminal OA/TCA present, inhibited by the lipid raft inhibitor methyl-ß-cyclodextrin (MßCD) and the CD36 inhibitor sulfosuccinimidyl oleate (SSO), or by serosal GLP-2. In vivo, perfusion of FITC-LPS with OA/TCA rapidly increased FITC-LPS appearance into the PV, followed by a gradual increase of FITC-LPS into the lymph. Rapid PV transport was inhibited by the addition of MßCD or by SSO, whereas transport into the lymph was inhibited by chylomicron synthesis inhibition. Intraveous injection of the stable GLP-2 analog teduglutide acutely inhibited FITC-LPS transport into the PV, yet accelerated FITC-LPS transport into the lymph via Nω-nitro-l-arginine methyl ester (l-NAME)- and PG97-269-sensitive mechanisms. In vivo confocal microscopy in mouse jejunum confirmed intracellular FITC-LPS uptake with no evidence of paracellular localization. This is the first direct demonstration in vivo that luminal LPS may cross the small intestinal barrier physiologically during fat absorption via lipid raft- and CD36-mediated mechanisms, followed by predominant transport into the PV, and that teduglutide inhibits LPS uptake into the PV in vivo.NEW & NOTEWORTHY We report direct in vivo confirmation of transcellular lipopolysaccharides (LPS) uptake from the intestine into the portal vein (PV) involving CD36 and lipid rafts, with minor uptake via the canonical chylomicron pathway. The gut hormone glucagon-like peptide-2 (GLP-2) inhibited uptake into the PV. These data suggest that the bulk of LPS absorption is via the PV to the liver, helping clarify the mechanism of LPS transport into the PV as part of the "gut-liver" axis. These data do not support the paracellular transport of LPS, which has been implicated in the pathogenesis of the "leaky gut" syndrome.


Assuntos
Gorduras/metabolismo , Intestino Delgado/metabolismo , Lipopolissacarídeos/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Transporte Biológico/fisiologia , Fármacos Gastrointestinais/farmacologia , Células HEK293 , Humanos , Intestino Delgado/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos/química , Peptídeos/farmacologia , Ratos , Ratos Sprague-Dawley , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo
10.
Rev Esp Enferm Dig ; 112(5): 383-388, 2020 05.
Artigo em Inglês | MEDLINE | ID: covidwho-148632

RESUMO

Although SARS-CoV-2 may primarily enter the cells of the lungs, the small bowel may also be an important entry or interaction site, as the enterocytes are rich in angiotensin converting enzyme (ACE)-2 receptors. The initial gastrointestinal symptoms that appear early during the course of Covid-19 support this hypothesis. Furthermore, SARS-CoV virions are preferentially released apically and not at the basement of the airway cells. Thus, in the setting of a productive infection of conducting airway epithelia, the apically released SARS-CoV may be removed by mucociliary clearance and gain access to the GI tract via a luminal exposure. In addition, post-mortem studies of mice infected by SARS-CoV have demonstrated diffuse damage to the GI tract, with the small bowel showing signs of enterocyte desquamation, edema, small vessel dilation and lymphocyte infiltration, as well as mesenteric nodes with severe hemorrhage and necrosis. Finally, the small bowel is rich in furin, a serine protease which can separate the S-spike of the coronavirus into two "pinchers" (S1 and 2). The separation of the S-spike into S1 and S2 is essential for the attachment of the virion to both the ACE receptor and the cell membrane. In this special review, we describe the interaction of SARS-CoV-2 with the cell and enterocyte and its potential clinical implications.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/metabolismo , Enterócitos/virologia , Gastroenteropatias/virologia , Intestino Delgado/virologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/metabolismo , Betacoronavirus/metabolismo , Infecções por Coronavirus/virologia , Enterócitos/metabolismo , Gastroenteropatias/metabolismo , Humanos , Intestino Delgado/citologia , Intestino Delgado/metabolismo , Pandemias , Pneumonia Viral/virologia , Receptores de Angiotensina/metabolismo , Mucosa Respiratória/fisiologia , Mucosa Respiratória/virologia
11.
Nat Commun ; 11(1): 1936, 2020 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-32321913

RESUMO

The intestinal epithelium is a structured organ composed of crypts harboring Lgr5+ stem cells, and villi harboring differentiated cells. Spatial transcriptomics have demonstrated profound zonation of epithelial gene expression along the villus axis, but the mechanisms shaping this spatial variability are unknown. Here, we combine laser capture micro-dissection and single cell RNA sequencing to uncover spatially zonated populations of mesenchymal cells along the crypt-villus axis. These include villus tip telocytes (VTTs) that express Lgr5, a gene previously considered a specific crypt epithelial stem cell marker. VTTs are elongated cells that line the villus tip epithelium and signal through Bmp morphogens and the non-canonical Wnt5a ligand. Their ablation is associated with perturbed zonation of enterocyte genes induced at the villus tip. Our study provides a spatially-resolved cell atlas of the small intestinal stroma and exposes Lgr5+ villus tip telocytes as regulators of the epithelial spatial expression programs along the villus axis.


Assuntos
Enterócitos/metabolismo , Mucosa Intestinal/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Animais , Enterócitos/citologia , Mucosa Intestinal/citologia , Intestino Delgado/citologia , Intestino Delgado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores Acoplados a Proteínas-G/genética , Células Estromais/metabolismo , Proteína Wnt-5a/metabolismo
12.
Rev Esp Enferm Dig ; 112(5): 383-388, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32343593

RESUMO

Although SARS-CoV-2 may primarily enter the cells of the lungs, the small bowel may also be an important entry or interaction site, as the enterocytes are rich in angiotensin converting enzyme (ACE)-2 receptors. The initial gastrointestinal symptoms that appear early during the course of Covid-19 support this hypothesis. Furthermore, SARS-CoV virions are preferentially released apically and not at the basement of the airway cells. Thus, in the setting of a productive infection of conducting airway epithelia, the apically released SARS-CoV may be removed by mucociliary clearance and gain access to the GI tract via a luminal exposure. In addition, post-mortem studies of mice infected by SARS-CoV have demonstrated diffuse damage to the GI tract, with the small bowel showing signs of enterocyte desquamation, edema, small vessel dilation and lymphocyte infiltration, as well as mesenteric nodes with severe hemorrhage and necrosis. Finally, the small bowel is rich in furin, a serine protease which can separate the S-spike of the coronavirus into two "pinchers" (S1 and 2). The separation of the S-spike into S1 and S2 is essential for the attachment of the virion to both the ACE receptor and the cell membrane. In this special review, we describe the interaction of SARS-CoV-2 with the cell and enterocyte and its potential clinical implications.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/metabolismo , Enterócitos/virologia , Gastroenteropatias/virologia , Intestino Delgado/virologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/metabolismo , Betacoronavirus/metabolismo , Infecções por Coronavirus/virologia , Enterócitos/metabolismo , Gastroenteropatias/metabolismo , Humanos , Intestino Delgado/citologia , Intestino Delgado/metabolismo , Pandemias , Pneumonia Viral/virologia , Receptores de Angiotensina/metabolismo , Mucosa Respiratória/fisiologia , Mucosa Respiratória/virologia
13.
Am J Physiol Gastrointest Liver Physiol ; 318(5): G980-G987, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32308039

RESUMO

Glucagon-like peptide (GLP)-1 and -2-secreting L cells have been shown to express the bile acid receptor Takeda G protein-receptor-5 (TGR5) and increase secretion upon receptor activation. Previous studies have explored GLP-1 secretion following acute TGR5 activation, but chronic activation and GLP-2 responses have not been characterized. In this study, we aimed to investigate the consequences of pharmacological TGR5 receptor activation on L cell hormone production in vivo using the specific TGR5 agonist RO5527239 and the GLP-2 receptor knockout mouse. Here, we show that 1) TGR5 receptor activation led to increased GLP-1 and GLP-2 content in the colon, which 2) was associated with an increased small intestinal weight that 3) was GLP-2 dependent. Additionally, we report that TGR5-mediated gallbladder filling occurred independently of GLP-2 signaling. In conclusion, we demonstrate that pharmacological TGR5 receptor activation stimulates L cells, triggering GLP-2-dependent intestinal adaption in mice.NEW & NOTEWORTHY Using the specific Takeda G protein-receptor-5 (TGR5) agonist RO5527239 and GLP-2 receptor knockout mice, we show that activation of TGR5 led to the increase in colonic GLP-1 and GLP-2 concomitant with a GLP-2 dependent growth response in the proximal portion of the small intestine.


Assuntos
Proliferação de Células/efeitos dos fármacos , Células Enteroendócrinas/efeitos dos fármacos , Peptídeo 2 Semelhante ao Glucagon/metabolismo , Intestino Delgado/efeitos dos fármacos , Ácidos Isonipecóticos/farmacologia , Oximas/farmacologia , Receptores Acoplados a Proteínas-G/agonistas , Animais , Colo/efeitos dos fármacos , Colo/crescimento & desenvolvimento , Colo/metabolismo , Células Enteroendócrinas/metabolismo , Feminino , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 2/genética , Receptor do Peptídeo Semelhante ao Glucagon 2/metabolismo , Intestino Delgado/crescimento & desenvolvimento , Intestino Delgado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores Acoplados a Proteínas-G/metabolismo , Transdução de Sinais
14.
Georgian Med News ; (298): 128-132, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32141865

RESUMO

Experimental modeling of dysbacterioses associated with antibiotics is an urgent issue of morphological studies. The present paper was aimed at the detection and study of the primordial forms of Peyer's patches developed in the small intestine of albino rats after administration of clarithromycin. 30 mature albino male rats weighing 200.0 ± 20.0 g were involved into the experiment. Antibiotic was administered to the rodents as a supplement to food during their two-meals-a-day feeding. Areas of the small intestine with Peyer's patches have been studied. Serial paraffin sections have been analyzed using the "Konus" light microscope. Morphometric characteristics of the tissue structures were obtained using the Sigeta X 1 mm / 100 Div.x0.01mm stage micrometer. Administration of clarithromycin caused a significant increase in the amount of Peyer's patches, the appearance in the mucous membrane of newly formed aggregated lymphoid nodules, being the primordial forms of Peyer's patches, the appearance of which can be explained by only one factor, namely, impaired microbiocenosis in the small intestine under the influence of the broad-spectrum antibacterial drug, clarithromycin, which has immunotropic effect.


Assuntos
Antibacterianos/administração & dosagem , Claritromicina/administração & dosagem , Intestino Delgado/efeitos dos fármacos , Nódulos Linfáticos Agregados , Animais , Antibacterianos/efeitos adversos , Claritromicina/efeitos adversos , Desenvolvimento Embrionário , Intestino Delgado/metabolismo , Masculino , Ratos
15.
PLoS One ; 15(3): e0229463, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32214355

RESUMO

Food and feeds contaminated with mycotoxins have been a threat to the rearing industry by causing some of the most fatal toxic reactions not only in the farm animals but also in humans who consume them. Toxicity to juvenile goats was induced by feed contamination with T-2 toxin (at 10 and 20 ppm dosage; group I and II, respectively). The toxicity impact was assessed on days 15 and 30 post treatment with respect to growth performance, oxidative stress, apoptotic studies and detailed pathomorphology. The study revealed that apart from the obvious clinical toxicosis (weakness, lethargy, and retardation in growth), the toxin fed groups also exhibited significant haematological (reduced hemoglobin, total leukocyte and thrombocyte counts) and biochemical changes (increased levels of oxidative stress markers with concomitant decrease in levels of serum and tissue catalase and superoxide dismutase). The pathomorphological and histological alterations suggested that the liver and intestine were the most affected organs. Ultra-structurally, varying degrees of degeneration, cytoplasmic vacuolations and pleomorphic mitochondria were observed in the hepatocytes and the enterocytes of the intestine. Kidney also revealed extensive degeneration of the cytoplasmic organelles with similar condensation of the heterochromatin whereas the neuronal degeneration was characterized by circular, whirling structures. In addition, the central vein and portal triad of the hepatocytes, cryptic epithelial cells of the intestine, MLNs in the lymphoid follicles, PCT and DCT of the nephronal tissues and the white pulp of the spleen exhibited extensive apoptosis. In this study, it was also observed that the expression of HSPs, pro-apoptotic proteins and pro-inflammatory cytokines were significantly upregulated in response to the toxin treatment. These results suggest that the pathogenesis of T-2 toxicosis in goats employs oxidative, apoptotic and inflammatory mechanisms.


Assuntos
Apoptose , Regulação da Expressão Gênica/efeitos dos fármacos , Cabras/fisiologia , Mediadores da Inflamação/metabolismo , Fígado/patologia , Estresse Oxidativo/efeitos dos fármacos , Toxina T-2/toxicidade , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/patologia , Citocinas/metabolismo , Proteínas de Choque Térmico/metabolismo , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/metabolismo , Intestino Delgado/patologia , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Linfonodos/efeitos dos fármacos , Linfonodos/metabolismo , Linfonodos/patologia , Baço/efeitos dos fármacos , Baço/metabolismo , Baço/patologia
16.
Food Chem ; 318: 126463, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32135421

RESUMO

The stability behaviours of whey-protein-stabilised emulsions under gastric conditions and the effects on the lipolysis of the emulsions were investigated using an in vitro dynamic human gastric simulator and a subsequent small intestinal model. Under gastric conditions, heated whey-protein-stabilised emulsions flocculated to a greater extent and with a larger floc size, whereas unheated emulsions were more prone to coalescence. The greater extent of flocculation delayed the delivery of oil droplets to the small intestine, leading to a lower amount of oil in the emptied gastric digesta from the heated emulsion in the early period of digestion. The differences in oil content, droplet size and interfacial composition led to a greater rate and extent of lipolysis in the subsequent intestinal digestion in the heated emulsion than the unheated emulsion. The results suggest that the lipid digestion of whey-protein-stabilised emulsions in the intestinal stage could be manipulated by thermal treatment.


Assuntos
Digestão , Emulsões/química , Emulsões/farmacocinética , Proteínas do Soro do Leite/química , Floculação , Temperatura Alta , Humanos , Concentração de Íons de Hidrogênio , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/metabolismo , Lipídeos/farmacocinética , Lipólise , Tamanho da Partícula
17.
Endocrinology ; 161(4)2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32147716

RESUMO

Glucagon-like peptide-2 (GLP-2) is an intestinotrophic hormone that promotes intestinal growth and proliferation through downstream mediators, including epidermal growth factor (EGF) and insulin-like growth factor-1 (IGF-1). EGF synergistically enhances the proliferative actions of IGF-1 in intestinal cell lines, and both of these factors are known to be essential for the trophic effects of GLP-2 in vivo. However, whether EGF and IGF-1 interact to mediate the proliferative actions of GLP-2 in vivo remains unknown. Normal and knockout (KO) mice lacking the intestinal epithelial IGF-1 receptor (IE-IGF-1R) were therefore treated chronically with EGF and/or long-acting human hGly2GLP-2, followed by determination of intestinal growth parameters. Intestines from control and IE-IGF-1R KO mice were also used to generate organoids (which lack the GLP-2 receptor) and were treated with EGF and/or IGF-1. Combination treatment with EGF and hGly2GLP-2 increased small intestinal weight and crypt-villus height in C57Bl/6 mice in an additive manner, whereas only hGly2GLP-2 treatment increased crypt cell proliferation. However, although combination treatment also increased small intestinal weight and crypt-villus height in IE-IGF-1R KO mice, the proliferative responses to hGly2GLP-2 alone or with EGF were diminished in these animals. Finally, IGF-1 treatment of organoids undergoing EGF withdrawal was not additive to the effect of EGF replacement on proliferation, but could restore normal proliferation in the absence of EGF. Together, these findings demonstrate that the intestinal proliferative effects of hGly2GLP-2 are augmented by exogenous EGF in a manner that is partially dependent upon IE-IGF-1R signaling.


Assuntos
Fator de Crescimento Epidérmico/farmacologia , Peptídeo 2 Semelhante ao Glucagon/metabolismo , Mucosa Intestinal/metabolismo , Intestino Delgado/efeitos dos fármacos , Receptor IGF Tipo 1/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Intestino Delgado/metabolismo , Camundongos , Camundongos Knockout , Receptor IGF Tipo 1/genética , Transdução de Sinais/efeitos dos fármacos
18.
J Dairy Sci ; 103(5): 4754-4764, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32197854

RESUMO

The development of the small intestine (SI) is important for the health and growth of neonatal calves. This study evaluated the effect of arginine (Arg) and glutamine (Gln) supplementation and 2 levels of milk allowance on the histomorphological development of the SI in preweaning calves. Sixty mixed-sex Friesian × Jersey calves (3-5 d of age) were offered reconstituted whole milk (125 g/L, 26% fat, 26% protein) at either high (20% of arrival body weight/d; HM) or low (10% of arrival body weight/d; LM) milk allowance without (Ctrl) or with supplementary Arg or Gln (at 1% of milk dry matter) in a 2 × 3 factorial design (n = 10/treatment). After 35 d on the diets, all calves were slaughtered to collect tissues for examination of SI development. Calves in the HM group had higher milk intake, total weight gain, and average daily gain compared with LM calves, but no effect of AA supplementation nor an interaction between milk allowance and AA supplementation was observed. For the duodenum, we observed an AA by milk allowance interaction for villus height and width, and goblet cell number per villus (HM-Arg > HM-Gln > HM-Ctrl), and villus height to crypt depth ratio (HM-Arg > HM-Gln = HM-Ctrl), but no effect of AA supplementation in the LM group. Goblet cell numbers per 100 µm of SI were greater in Arg-supplemented calves than in unsupplemented controls, with Gln-supplemented calves intermediate to but not different from the other groups. Epithelium thickness was greater in LM than in HM calves. Villus density, crypt depth, and muscle thickness did not differ between groups. For the jejunum, there was an AA by milk allowance interaction for villus height, villus surface area, and villus height to crypt depth ratio (HM-Arg = HM-Gln > HM-Ctrl), with no effect of AA supplementation in the LM groups. Amino acid supplementation affected goblet cell number per villus (HM-Gln > HM-Ctrl calves, HM-Arg intermediate), and both LM-Arg and LM-Gln calves had greater numbers than LM-Ctrl calves. Villus width, crypt depth, and muscle thickness were greater in HM than LM calves but there was no effect of AA supplementation. Villus density, goblet cell number per 100 µm of SI, and epithelium thickness were unaffected by AA supplementation and milk allowance. Milk allowance and AA supplementation had no effect on SI morphology in the ileum. Increasing milk allowance improved villus height, width, and surface area but only in Arg- or Gln-supplemented calves, not in control calves. The observed changes in development may be important for intestinal functionality, integrity, and barrier function in preweaning calves, potentially through increased cell growth and proliferation or reduced levels of cellular atrophy.


Assuntos
Ração Animal , Arginina/farmacologia , Bovinos/crescimento & desenvolvimento , Suplementos Nutricionais , Glutamina/farmacologia , Intestino Delgado/crescimento & desenvolvimento , Leite , Animais , Peso Corporal , Dieta/veterinária , Feminino , Mucosa Intestinal/crescimento & desenvolvimento , Intestino Delgado/metabolismo , Masculino , Ganho de Peso
19.
J Pathol ; 251(2): 160-174, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32222043

RESUMO

The IκB kinase (IKK)-NF-κB signaling pathway plays a multifaceted role in inflammatory bowel disease (IBD): on the one hand, it protects from apoptosis; on the other, it activates transcription of numerous inflammatory cytokines and chemokines. Although several murine models of IBD rely on disruption of IKK-NF-κB signaling, these involve either knockouts of a single family member of NF-κB or of upstream kinases that are known to have additional, NF-κB-independent, functions. This has made the distinct contribution of NF-κB to homeostasis in intestinal epithelium cells difficult to assess. To examine the role of constitutive NF-κB activation in intestinal epithelial cells, we generated a mouse model with a tissue-specific knockout of the direct inhibitor of NF-κB, Nfkbia/IκBα. We demonstrate that constitutive activation of NF-κB in intestinal epithelial cells induces several hallmarks of IBD including increased apoptosis, mucosal inflammation in both the small intestine and the colon, crypt hyperplasia, and depletion of Paneth cells, concomitant with aberrant Wnt signaling. To determine which NF-κB-driven phenotypes are cell-intrinsic, and which are extrinsic and thus require the immune compartment, we established a long-term organoid culture. Constitutive NF-κB promoted stem-cell proliferation, mis-localization of Paneth cells, and sensitization of intestinal epithelial cells to apoptosis in a cell-intrinsic manner. Increased number of stem cells was accompanied by a net increase in Wnt activity in organoids. Because aberrant Wnt signaling is associated with increased risk of cancer in IBD patients and because NFKBIA has recently emerged as a risk locus for IBD, our findings have critical implications for the clinic. In a context of constitutive NF-κB, our findings imply that general anti-inflammatory or immunosuppressive therapies should be supplemented with direct targeting of NF-κB within the epithelial compartment in order to attenuate apoptosis, inflammation, and hyperproliferation. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.


Assuntos
Apoptose , Doenças Inflamatórias Intestinais/metabolismo , Intestino Delgado/metabolismo , Inibidor de NF-kappaB alfa/deficiência , Celulas de Paneth/metabolismo , Células-Tronco/metabolismo , Animais , Células Cultivadas , Citocinas/metabolismo , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/patologia , Intestino Delgado/patologia , Camundongos Knockout , Inibidor de NF-kappaB alfa/genética , Organoides/metabolismo , Organoides/patologia , Celulas de Paneth/patologia , Células-Tronco/patologia , Fator de Transcrição RelA/metabolismo , Via de Sinalização Wnt
20.
J Pharmacol Sci ; 143(1): 30-38, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32151540

RESUMO

The role of nitric oxide (NO) on intestinal mucosal injury induced by single or consecutive administration of methotrexate was investigated in a rodent model. Rats received methotrexate intraperitoneally either as a single administration (50 mg/kg) or as a consecutive administration (12.5 mg/kg/day) for 4 days. NG-nitro-l-arginine methyl ester (L-NAME) was given subcutaneously to inhibit NO synthase (NOS). Ninety-six hours after the first administration of methotrexate, ileal tissues were collected for analysis. Consecutive administration of methotrexate led to decreased body weight and reduced intake of food and water, which were further worsened by L-NAME. Although a slight mucosal injury resulted from single administration of methotrexate, L-NAME had almost no effect. Consecutive administration of methotrexate caused a significant mucosal injury, which was further worsened by L-NAME. Consecutive, but not single, administration of methotrexate induced mRNA expression of inflammatory cytokines in ileal tissue. Consecutive administration of methotrexate significantly induced constitutive NOS expression in ileal tissue. These results suggest that consecutive administration, rather than single administration, of methotrexate aggravates mucosal injury. Potentiation of constitutive NOS expression by consecutive administration might be one of the main reason to antagonize the intestinal mucosal injury as well as lead to a reduction in rat quality of life.


Assuntos
Expressão Gênica , Enteropatias/etiologia , Intestino Delgado/metabolismo , Intestino Delgado/patologia , Metotrexato/administração & dosagem , Metotrexato/efeitos adversos , Óxido Nítrico/efeitos adversos , Óxido Nítrico/metabolismo , Animais , Citocinas/genética , Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Masculino , Modelos Animais , Óxido Nítrico/genética , RNA Mensageiro/metabolismo , Ratos Wistar
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