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1.
Adv Exp Med Biol ; 1173: 21-32, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31456203

RESUMO

Iron is an essential trace element in the human body, but excess iron is toxic as it contributes to oxidative damage. To keep iron concentration within the optimal physiologic range, iron metabolism at the cellular level and the whole systemic level are tightly regulated. Balance of iron homeostasis depends on the expression levels and activities of iron carriers, iron transporters, and iron regulatory and storage proteins. Divalent metal transporter 1 (DMT1) at the apical membrane of intestinal enterocyte brings in non-heme iron from the diet, whereas ferroportin 1 (FPN1) at the basal membrane exports iron into the circulation. Plasma transferrin (Tf) then carries iron to various tissues and cells. After binding to transferrin receptor 1 (TfR1), the complex is endocytosed into the cell, where iron enters the cytoplasm via DMT1 on the endosomal membrane. Free iron is either utilized in metabolic processes, such as synthesis of hemoglobin and Fe-S cluster, or sequestered in the cytosolic ferritin, serving as a cellular iron store. Excess iron can be exported from the cell via FPN1. The liver-derived peptide hepcidin plays a major regulatory role in controlling FPN1 level in the enterocyte, and thus controls the whole-body iron absorption. Inside the cells, iron regulatory proteins (IRPs) modulate the expressions of DMT1, TfR1, ferritin, and FPN1 via binding to the iron-responsive element (IRE) in their mRNAs. Both the release of hepcidin and the IRP-IRE interaction are coordinated with the fluctuation of the cellular iron level. Therefore, an adequate and steady iron supplement is warranted for the utilization of cells around the body. Investigations on the molecular mechanisms of cellular iron metabolism and regulation could advance the fields of iron physiology and pathophysiology.


Assuntos
Ferro/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Enterócitos/metabolismo , Ferritinas/metabolismo , Homeostase , Humanos , Sobrecarga de Ferro , Receptores da Transferrina/metabolismo , Fatores de Transcrição/metabolismo , Transferrina/metabolismo
2.
J Dairy Sci ; 102(9): 7684-7696, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31255276

RESUMO

Oxidative stress is the basic reason for aging and age-related diseases. In this study, we investigated the protective effect of 2 strains of lactic acid bacteria (LAB), Lactobacillus rhamnosus GG and L. plantarum J26, against oxidative stress in Caco-2 cells, and gave an overview of the mechanisms of lactic acid bacteria antioxidant activity using digital gene expression profiling. The 2 LAB strains provided significant protection against hydrogen peroxide (H2O2)-induced reduction in superoxide dismutase activity and increase in glutathione peroxidase activity in Caco-2 cells. However, inactive bacteria had little effect on alleviating oxidation stress in Caco-2 cells. Eight genes related to oxidative stress-FOSB, TNF, PPP1R15A, NUAK2, ATF3, TNFAIP3, EGR2, and FBN2-were significantly upregulated in H2O2-induced Caco-2 cells compared with untreated Caco-2 cells. After incubation of the H2O2-induced Caco-2 cells with L. rhamnosus GG and L. plantarum J26, 5 genes (TNF, EGR2, NUAK2, FBN2, and TNFAIP3) and 2 genes (NUAK2 and FBN2) were downregulated, respectively. In addition, the Kyoto Encyclopedia of Genes and Genomes indicated that some signaling pathways associated with inflammation, immune response, and apoptosis, such as Janus kinase/signal transducers and activators of transcription (Jak-STAT), mitogen-activated protein kinase (MAPK), nuclear factor-κB, and tumor necrosis factor, were all negatively modulated by the 2 strains, especially L. rhamnosus GG. In this paper, we reveal the mechanism of LAB in relieving oxidative stress and provide a theoretical basis for the rapid screening and evaluation of new LAB resources.


Assuntos
Enterócitos/metabolismo , Lactobacillus plantarum/fisiologia , Lactobacillus rhamnosus/fisiologia , Estresse Oxidativo/genética , Transcriptoma/genética , Animais , Apoptose/genética , Células CACO-2 , Enterócitos/efeitos dos fármacos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/fisiologia , Humanos , Peróxido de Hidrogênio/farmacologia , Imunidade/genética , Inflamação/genética , Probióticos/farmacologia
3.
Medicine (Baltimore) ; 98(28): e16452, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31305476

RESUMO

Endotoxemia often occurs in patients with gram-positive infections. The possible mechanism is thought to be bacterial translocation after enterocyte hypoperfusion injury. However, the association between endotoxemia and enterocyte injury among patients with gram-positive septic shock has never been assessed. The aim of this study was to evaluate the association between endotoxemia and enterocyte injury in gram-positive septic shock patients and to evaluate the association among endotoxemia, subsequent clinical course, and other related factors.This was a posthoc analysis of a prospective observational study that evaluated the capability of intestinal fatty acid-binding protein (I-FABP), an indicator of enterocyte injury, to predict mortality. Among 57 patients in septic shock, those whose causative microorganisms were gram positive were included. The correlation between endotoxin activity (EA), which indicates endotoxemia, and I-FABP levels upon admission to the intensive care unit (ICU), the clinical course, and other related factors were evaluated.A total of 21 patients were examined. One-third of the patients presented with high EA levels at the time of ICU admission. However, there was no significant correlation between EA and I-FABP levels (Spearman ρ = 0.002, P = .993). Additionally, high EA levels were not associated with abdominal complications after ICU admission or mortality. Similarly, high EA levels were not associated with severity scores, inotropic scores, or lactate levels upon ICU admission, which were previously reported to be factors related to high EA levels.In this posthoc analysis, no correlation was observed between endotoxemia and enterocyte injury among patients in gram-positive septic shock. Additionally, high EA levels were not associated with the clinical course and reported factors related to endotoxemia. Although our results need to be validated in a large prospective cohort study, hypoperfusion enterocyte injury might not be a cause of endotoxemia in these patients. Thus, if there is no correlation between EA and I-FABP levels, other mechanisms that induce high EA levels among patients with gram-positive septic shock should be elucidated.


Assuntos
Endotoxemia/sangue , Proteínas de Ligação a Ácido Graxo/sangue , Infecções por Bactérias Gram-Positivas/sangue , Infecções por Bactérias Gram-Positivas/mortalidade , Choque Séptico/sangue , Choque Séptico/mortalidade , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/sangue , Cuidados Críticos , Progressão da Doença , Endotoxemia/mortalidade , Endotoxemia/terapia , Enterócitos/metabolismo , Feminino , Infecções por Bactérias Gram-Positivas/terapia , Humanos , Masculino , Prognóstico , Estudos Prospectivos , Choque Séptico/terapia
4.
Biol Pharm Bull ; 42(6): 989-995, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31155596

RESUMO

An Intact form of lactoferrin (LF) is known to be absorbed from the small intestine and transported into the blood circulation. We reevaluated the cellular uptake and release of LF using an enterocyte model of human small intestinal cells derived from the Caco-2 cell line. In contrast to a previous report, we observed that intact bovine LF was taken up into seven and 21 d-cultured Caco-2 cells and successfully released back into the culture medium, even though the human intestinal LF receptor, intelectin-1, was not immunochemically detectable. Similar observations were made for human LF and its derivatives (the N-terminal half of LF designated N-lobe and Fc fusions). These observations regarding the uptake and release of intact LF in Caco-2 cells were consistent with in vivo observations. Therefore, we propose that the uptake and release of intact LF by Caco-2 cells should be assessed as a potential in vitro model of in vivo LF absorption in human intestines.


Assuntos
Enterócitos/efeitos dos fármacos , Intestinos/citologia , Lactoferrina/farmacologia , Animais , Células CHO , Células CACO-2 , Cricetulus , Enterócitos/metabolismo , Humanos , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Proteínas Recombinantes/metabolismo
5.
J Dairy Sci ; 102(8): 6863-6875, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31178173

RESUMO

Some Lactobacillus strains have been reported to have antioxidative activity. In our previous work, we screened Lactobacillus plantarum Y44 for its antioxidative activity. In this study, we further studied the antioxidative activities of L. plantarum Y44 using chemical antioxidant methods, including the 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) free radical scavenging assays, the ferric reducing antioxidant power test, and oxygen radical absorbance capacity test, and we assessed damage caused by 2,2'-azobis(2-methylpropionamidine) dihydrochloride (ABAP) in a Caco-2 cell model. The results of the chemical antioxidant assays confirmed the antioxidative activity of L. plantarum Y44, which was consistent with the protection of Caco-2 cells against ABAP injury by L. plantarum Y44. We also found that L. plantarum Y44 significantly promoted expression of Nrf2 pathway-associated proteins, downregulated expression of inflammatory-related cytokines IL-8 and tumor necrosis factor-α in ABAP-damaged Caco-2 cells, and enhanced expression of the tight junction proteins ß-catenin and E-cadherin. We determined that L. plantarum Y44 exerted antioxidative effects by quenching oxygen free radicals and activating the Nrf2 signaling pathway in Caco-2 cells.


Assuntos
Amidinas/farmacologia , Antioxidantes/fisiologia , Enterócitos/efeitos dos fármacos , Lactobacillus plantarum/fisiologia , Animais , Células CACO-2 , Enterócitos/metabolismo , Humanos , Fator 2 Relacionado a NF-E2/metabolismo , Espécies Reativas de Oxigênio/química , Transdução de Sinais/fisiologia , Fator de Necrose Tumoral alfa/metabolismo
6.
Molecules ; 24(9)2019 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-31072069

RESUMO

Nandina domestica (Berberidaceae) has been used in traditional medicine for the treatment of cough. This plant is distributed in Korea, Japan, China, and India This study aimed to investigate the anti-inflammatory phytochemicals obtained from the N. domestica fruits. We isolated a biflavonoid-type phytochemical, robustaflavone (R), from N. domestica fruits through bioactivity-guided fractionation based on its capacity to inhibit inflammation. The anti-inflammatory mechanism of R isolated from N. domestica has not yet been studied. In the present study, we evaluated the anti-inflammatory activities of R using lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. We have shown that R reduces the production of nitric oxide (NO), pro-inflammatory cytokine interleukin-1 beta (IL-1ß), and IL-6. Western blot analysis showed that R suppresses the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and downregulates the expression of LPS-induced nuclear factor-kappa B (NF-κB) and the phosphorylation of extracellular-regulated kinases (pERK 1/2). Moreover, R inhibited IL-8 release in LPS-induced human colonic epithelial cells (HT-29). These results suggest that R could be a potential therapeutic candidate for inflammatory bowel disease (IBD).


Assuntos
Berberidaceae/química , Biflavonoides/isolamento & purificação , Biflavonoides/farmacologia , Regulação para Baixo , Mediadores da Inflamação/metabolismo , Animais , Biflavonoides/química , Morte Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fracionamento Químico , Ciclo-Oxigenase 2/metabolismo , Regulação para Baixo/efeitos dos fármacos , Enterócitos/efeitos dos fármacos , Enterócitos/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Células HT29 , Humanos , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Interleucina-8/biossíntese , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , NF-kappa B/metabolismo , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase Tipo II/metabolismo , Fosforilação/efeitos dos fármacos , Compostos Fitoquímicos/química , Compostos Fitoquímicos/isolamento & purificação , Compostos Fitoquímicos/farmacologia , Extratos Vegetais/farmacologia , Células RAW 264.7
7.
Methodist Debakey Cardiovasc J ; 15(1): 70-76, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31049152

RESUMO

Besides the well-known hepatobiliary pathway of cholesterol excretion into the feces, transintestinal cholesterol excretion (TICE) is a second major pathway through which cholesterol is disposed from the body. In the process of TICE, cholesterol is taken up from lipoprotein particles at the basolateral side of the enterocyte and translocates towards the apical side of the enterocyte. At the apical side, the ATP-binding cassette transporters G5 and G8 form a heterodimer that transports cholesterol into the intestinal lumen. A substantial amount of the secreted cholesterol is likely reabsorbed by the cholesterol influx transporter Niemann-Pick C1-Like 1 (NPC1L1) since recent data indicate that inhibition of NPC1L1 increases the efficacy of TICE for disposal of cholesterol via the feces. The pathways and proteins involved in intracellular cholesterol trafficking in the enterocyte have not yet been identified. Therefore, in addition to discussing known mediators of TICE, this review will also examine potential candidates involved in cholesterol translocation in the enterocyte. Both the cholesterol reuptake and efflux pathways can be influenced by pharmaceutical means; thus, the TICE pathway is a very attractive target to increase cholesterol excretion from the body and prevent or mitigate atherosclerotic cardiovascular disease.


Assuntos
Anticolesterolemiantes/uso terapêutico , Doenças Cardiovasculares/prevenção & controle , Colesterol/sangue , Dislipidemias/tratamento farmacológico , Enterócitos/efeitos dos fármacos , Eliminação Intestinal/efeitos dos fármacos , Proteínas de Membrana Transportadoras/efeitos dos fármacos , Animais , Anticolesterolemiantes/efeitos adversos , Biomarcadores/sangue , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/epidemiologia , Regulação para Baixo , Dislipidemias/sangue , Dislipidemias/diagnóstico , Dislipidemias/epidemiologia , Enterócitos/metabolismo , Fezes/química , Humanos , Proteínas de Membrana Transportadoras/metabolismo
8.
Nat Genet ; 51(5): 777-785, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30988513

RESUMO

BMP/SMAD signaling is a crucial regulator of intestinal differentiation1-4. However, the molecular underpinnings of the BMP pathway in this context are unknown. Here, we characterize the mechanism by which BMP/SMAD signaling drives enterocyte differentiation. We establish that the transcription factor HNF4A acts redundantly with an intestine-restricted HNF4 paralog, HNF4G, to activate enhancer chromatin and upregulate the majority of transcripts enriched in the differentiated epithelium; cells fail to differentiate on double knockout of both HNF4 paralogs. Furthermore, we show that SMAD4 and HNF4 function via a reinforcing feed-forward loop, activating each other's expression and co-binding to regulatory elements of differentiation genes. This feed-forward regulatory module promotes and stabilizes enterocyte cell identity; disruption of the HNF4-SMAD4 module results in loss of enterocyte fate in favor of progenitor and secretory cell lineages. This intersection of signaling and transcriptional control provides a framework to understand regenerative tissue homeostasis, particularly in tissues with inherent cellular plasticity5.


Assuntos
Enterócitos/citologia , Enterócitos/metabolismo , Fator 4 Nuclear de Hepatócito/metabolismo , Proteína Smad4/metabolismo , Animais , Sítios de Ligação/genética , Células CACO-2 , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Elementos Facilitadores Genéticos , Fator 4 Nuclear de Hepatócito/deficiência , Fator 4 Nuclear de Hepatócito/genética , Humanos , Camundongos , Camundongos Knockout , Transdução de Sinais , Proteína Smad4/deficiência , Proteína Smad4/genética
9.
Immunopharmacol Immunotoxicol ; 41(2): 285-291, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30892107

RESUMO

Background: The majority of children undergoing allogenic hematopoietic stem cell transplantation (HSCT) experience severe pain due to chemotherapy-induced gastrointestinal toxicity. Inter-individual differences in pain perceived and opioid consumption remain unexplained, limiting the possibility for individualized pain control. The aim of this study was to investigate the associations between opioid consumption and markers of gastrointestinal toxicity (plasma citrulline) and systemic inflammation (plasma CRP and IL-6) in these patients. Methods: We retrospectively included 38 children undergoing HSCT in Denmark in 2010-2012. Opioids doses on days 0-21 post-HSCT were registered as intravenous morphine equivalents (MEs). CRP was measured daily on days 0-21. IL-6 was measured on day 7. Citrulline was measured before conditioning, on days 7 and 21. Results: Out of 38 children, 37 (97%) received opioids during days 0-21. CRP level and ME dose peaked on days 9-10 while citrulline level reached a nadir on day 7 indicating maximum enterocyte loss. CRP was associated with ME dose, with an estimated increase of 0.030 mg/kg (95% CI 0.024-0.035) in ME for a 50% increase in CRP level on the same day (p < .001). IL-6 was correlated with ME on day 7 (rho = 0.55, p = .002). Citrulline did not correlate with ME. Conclusions: Opioid consumption in the early post-HSCT period is associated with the degree of chemotherapy-induced systemic inflammation and not with the extent of enterocyte loss. These findings contribute to our understanding of mucositis-related pain and may be of interest for future studies on therapeutic strategies.


Assuntos
Gastroenteropatias/tratamento farmacológico , Transplante de Células-Tronco Hematopoéticas , Morfina/administração & dosagem , Dor/tratamento farmacológico , Adolescente , Aloenxertos , Proteína C-Reativa/metabolismo , Criança , Pré-Escolar , Citrulina/sangue , Enterócitos/metabolismo , Enterócitos/patologia , Feminino , Gastroenteropatias/sangue , Gastroenteropatias/genética , Gastroenteropatias/patologia , Humanos , Lactente , Inflamação/sangue , Inflamação/tratamento farmacológico , Inflamação/etiologia , Interleucina-6/sangue , Masculino , Dor/sangue , Dor/etiologia , Dor/patologia
10.
Nat Commun ; 10(1): 1029, 2019 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-30833673

RESUMO

Enteroendocrine cells are specialised sensory cells located in the intestinal epithelium and generate signals in response to food ingestion. Whilst traditionally considered hormone-producing cells, there is evidence that they also initiate activity in the afferent vagus nerve and thereby signal directly to the brainstem. We investigate whether enteroendocrine L-cells, well known for their production of the incretin hormone glucagon-like peptide-1 (GLP-1), also release other neuro-transmitters/modulators. We demonstrate regulated ATP release by ATP measurements in cell supernatants and by using sniffer patches that generate electrical currents upon ATP exposure. Employing purinergic receptor antagonists, we demonstrate that evoked ATP release from L-cells triggers electrical responses in neighbouring enterocytes through P2Y2 and nodose ganglion neurones in co-cultures through P2X2/3-receptors. We conclude that L-cells co-secrete ATP together with GLP-1 and PYY, and that ATP acts as an additional signal triggering vagal activation and potentially synergising with the actions of locally elevated peptide hormone concentrations.


Assuntos
Trifosfato de Adenosina/metabolismo , Enterócitos/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Intestinos , Neurônios Aferentes/metabolismo , Vias Aferentes , Animais , Linhagem Celular , Ingestão de Alimentos , Células Enteroendócrinas/metabolismo , Feminino , Cistos Glanglionares/metabolismo , Cistos Glanglionares/patologia , Incretinas/metabolismo , Mucosa Intestinal/inervação , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/patologia , Gânglio Nodoso/metabolismo , Gânglio Nodoso/patologia , Peptídeo YY/metabolismo , Receptores Purinérgicos P2X2/metabolismo , Receptores Purinérgicos P2X3/metabolismo , Nervo Vago/metabolismo
11.
PLoS Genet ; 15(3): e1007931, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30822306

RESUMO

Dipteran insects transmit serious diseases to humans, often in the form of trypanosomatid parasites. To accelerate research in more difficult contexts of dipteran-parasite relationships, we studied the interaction of the model dipteran Drosophila melanogaster and its natural trypanosomatid Herpetomonas muscarum. Parasite infection reduced fecundity but not lifespan in NF-κB/Relish-deficient flies. Gene expression analysis implicated the two NF-κB pathways Toll and Imd as well as STAT signalling. Tissue specific knock-down of key components of these pathways in enterocytes (ECs) and intestinal stem cells (ISCs) influenced initial numbers, infection dynamics and time of clearance. Herpetomonas triggered STAT activation and proliferation of ISCs. Loss of Relish suppressed ISCs, resulting in increased parasite numbers and delayed clearance. Conversely, overexpression of Relish increased ISCs and reduced uptake. Finally, loss of Toll signalling decreased EC numbers and enabled parasite persistence. This network of signalling may represent a general mechanism with which dipteran respond to trypanosomatids.


Assuntos
Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Interações Hospedeiro-Parasita/genética , Fatores de Transcrição/genética , Trypanosomatina/genética , Animais , Proliferação de Células/genética , Drosophila melanogaster/parasitologia , Enterócitos/metabolismo , Enterócitos/parasitologia , Fertilidade/genética , Regulação da Expressão Gênica/genética , Humanos , Intestinos/parasitologia , Fatores de Transcrição STAT/genética , Transdução de Sinais/genética , Células-Tronco/metabolismo , Receptores Toll-Like/genética , Fator de Transcrição RelA/genética , Trypanosomatina/patogenicidade
12.
Int J Mol Med ; 43(5): 2086-2102, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30864680

RESUMO

The purpose of the present study was to investigate whether bone marrow mesenchymal stem cells (BMMSCs) modified by CXC­chemokine receptor type 3 (CXCR3) and heme oxygenase­1 (HO­1) genes can repair damaged intestinal epithelial cells in vitro, and the role of the p38 mitogen­activated protein kinase (p38­MAPK) pathway in this process. A model of intestinal epithelial crypt cell line­6 (IEC­6) damage was created, and BMMSCs were transfected with either the CXCR3 and/or HO­1 gene in vitro. There were nine experimental groups in which the damaged IEC­6 cells were co­cultured with differentially­treated BMMSCs and lymphocytes for 24 h. Reverse transcription­quantitative polymerase chain reaction analysis, immunohistochemistry and a western blot analysis were performed to detect stem cell transfection, the repair of damaged intestinal epithelial cells and the expression of related molecules in the P38­MAPK pathway, respectively. Crystal violet staining and live cell imaging were used to detect the chemotaxis of BMMSCs. Flow cytometry was used to detect T lymphocyte activity and the surface markers expressed on BMMSCs. An ELISA was used to quantify cytokine production. The adenovirus (Ad)­CXCR3/MSCs exhibited the characteristics of stem cells and exhibited chemotaxis. The Ad­CXCR3/MSCs and Ad­(CXCR3 + HO)/MSCs exhibited increased expression of tight junction protein zonula occludens­1 (ZO­1) and anti­proliferating cell nuclear antigen in the damaged IEC­6 cells, and apoptosis of the damaged IEC­6 cells was decreased. BMMSCs inhibited the phosphorylation of p38, in addition to downstream molecules of the p38MAPK signaling pathway. The Ad­CXCR3/MSCs and Ad­(CXCR3 + HO)/MSCs exhibited significantly decreased expression levels of downstream molecules, including phosphorylated (p)­p38, p­activated transcription factor 2, p­C/EBP homologous protein­10, and p­myocyte enhancer factor 2C, and target molecules (e.g., apoptotic bodies). The effects of Ad­(CXCR3 + HO)/MSCs on the repair of the damaged intestinal tract and inhibition of the p38­MAPK pathway was more marked than those in other groups on day 7 post­surgery in the rejection model for small bowel transplantation. BMMSCs modified by the CXCR3 and HO­1 genes exhibited superior ability to repair damaged intestinal epithelial cells and served this role via the p38­MAPK pathway.


Assuntos
Enterócitos/metabolismo , Enterócitos/patologia , Heme Oxigenase-1/genética , Sistema de Sinalização das MAP Quinases , Células-Tronco Mesenquimais/metabolismo , Substâncias Protetoras/metabolismo , Receptores CXCR3/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Apoptose , Linhagem Celular , Sobrevivência Celular , Quimiotaxia , Heme Oxigenase-1/metabolismo , Intestino Delgado/transplante , Linfócitos/metabolismo , Masculino , Transplante de Células-Tronco Mesenquimais , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Endogâmicos BN , Ratos Endogâmicos Lew , Reprodutibilidade dos Testes , Proteínas de Junções Íntimas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
13.
Int J Mol Sci ; 20(6)2019 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-30917504

RESUMO

Na-amino acid co-transporters (NaAAcT) are uniquely affected in rabbit intestinal villus cell brush border membrane (BBM) during chronic intestinal inflammation. Specifically, Na-alanine co-transport (ASCT1) is inhibited secondary to a reduction in the affinity of the co-transporter for alanine, whereas Na-glutamine co-transport (B0AT1) is inhibited secondary to a reduction in BBM co-transporter numbers. During chronic intestinal inflammation, there is abundant production of the potent oxidant peroxynitrite (OONO). However, whether OONO mediates the unique alteration in NaAAcT in intestinal epithelial cells during chronic intestinal inflammation is unknown. In this study, ASCT1 and B0AT1 were inhibited by OONO in vitro. The mechanism of inhibition of ASCT1 by OONO was secondary to a reduction in the affinity of the co-transporter for alanine, and secondary to a reduction in the number of co-transporters for B0AT1, which were further confirmed by Western blot analyses. In conclusion, peroxynitrite inhibited both BBM ASCT1 and B0AT1 in intestinal epithelial cells but by different mechanisms. These alterations in the villus cells are similar to those seen in the rabbit model of chronic enteritis. Therefore, this study indicates that peroxynitrite may mediate the inhibition of ASCT1 and B0AT1 during inflammation, when OONO levels are known to be elevated in the mucosa.


Assuntos
Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Enterócitos/metabolismo , Proteínas de Transporte de Glutamato da Membrana Plasmática/metabolismo , Microvilosidades/metabolismo , Simportadores/metabolismo , Animais , Linhagem Celular , Enterócitos/efeitos dos fármacos , Enterócitos/patologia , Inflamação/metabolismo , Ácido Peroxinitroso/toxicidade , Ratos
14.
Cell Mol Gastroenterol Hepatol ; 7(2): 313-337, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30704982

RESUMO

BACKGROUND & AIMS: The small intestine regulates plasma triglyceride (TG) concentration. Within enterocytes, dietary TGs are packaged into chylomicrons (CMs) for secretion or stored temporarily in cytoplasmic lipid droplets (CLDs) until further mobilization. We and others have shown that oral and intravenous glucose enhances CM particle secretion in human beings, however, the mechanisms through which this occurs are incompletely understood. METHODS: Two separate cohorts of participants ingested a high-fat liquid meal and, 5 hours later, were assigned randomly to ingest either a glucose solution or an equivalent volume of water. In 1 group (N = 6), plasma and lipoprotein TG responses were assessed in a randomized cross-over study. In a separate group (N = 24), duodenal biopsy specimens were obtained 1 hour after ingestion of glucose or water. Ultrastructural and proteomic analyses were performed on duodenal biopsy specimens. RESULTS: Compared with water, glucose ingestion increased circulating TGs within 30 minutes, mainly in the CM fraction. It decreased the total number of CLDs and the proportion of large-sized CLDs within enterocytes. We identified 2919 proteins in human duodenal tissue, 270 of which are related to lipid metabolism and 134 of which were differentially present in response to glucose compared with water ingestion. CONCLUSIONS: Oral glucose mobilizes TGs stored within enterocyte CLDs to provide substrate for CM synthesis and secretion. Future studies elucidating the underlying signaling pathways may provide mechanistic insights that lead to the development of novel therapeutics for the treatment of hypertriglyceridemia.


Assuntos
Glucose/administração & dosagem , Intestinos/química , Triglicerídeos/metabolismo , Administração Oral , Adulto , Biópsia , Quilomícrons/metabolismo , Dieta Hiperlipídica , Duodeno/patologia , Enterócitos/metabolismo , Enterócitos/ultraestrutura , Jejum , Feminino , Ontologia Genética , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/ultraestrutura , Metabolismo dos Lipídeos/genética , Lipoproteínas VLDL/metabolismo , Masculino , Pessoa de Meia-Idade , Triglicerídeos/sangue
15.
J Agric Food Chem ; 67(10): 2782-2793, 2019 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-30785738

RESUMO

To investigate effects of dietary nitrogen level on nutrient absorption and utilization in small intestinal enterocyte of piglets, weaned piglets were fed for 10 days with diets containing 20%, 17%, or 14% crude protein (CP) with supplementation to meet requirements for essential amino acids in vivo, and IPEC-1 cells were cultured with different nitrogen levels (NL) in a culture medium (70%, 85%, and 100%) in vitro by monocultured and cocultured intestinal porcine epithelial cells (IPEC-1) and human gastric epithelial cells (GES-1). The results showed the following: (1) In animal trial, decreased dietary CP reduced transcript abundance of nutrient transporters like CAT1, PepT1, GLUT2, and SGLT-1 in jejunal mucosa (0.09 ± 0.03, P < 0.0001; 0.40 ± 0.04, P = 0.0087; 0.20 ± 0.07, P = 0.0003; 0.35 ± 0.02, P = 0.0001), but 17% CP diet did not affect jejunal protein synthesis. (2) The transcript abundance of nutrient transporters displayed similarly effective tendency in jejunal mucosa and cocultured IPEC-1 rather than that in monocultured IPEC-1. (3) Decreased nitrogen levels reduced expressive abundance of PI3K, Class 3 PI3K, TSC2, and 4E-BP1 in monocultured IPEC-1, but 85% nitrogen level did not affect expressive abundance of PI3K, TSC2, mTORC1, 4E-BP1, and S6K1 in cocultured IPEC-1. In general, decreased 3% CP or 15% nitrogen level reduced relative transcript expression of nutrient transporters, but did not affect protein synthesis in jejunal mucosa and cocultured IPEC-1. Therefore, decreased 3% dietary CP increased utilized and synthetic efficiency of nitrogen resource in small intestine and was beneficial in saving the dietary nitrogen resource.


Assuntos
Ração Animal/análise , Enterócitos/metabolismo , Intestino Delgado/metabolismo , Proteínas de Membrana Transportadoras/genética , Nitrogênio/análise , Biossíntese de Proteínas , Suínos/metabolismo , Animais , Transporte Biológico , Proteínas na Dieta/análise , Proteínas na Dieta/metabolismo , Jejuno/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Nitrogênio/metabolismo , Nutrientes/análise , Nutrientes/metabolismo , Suínos/genética
16.
Eur J Pharm Sci ; 131: 195-207, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30776469

RESUMO

Physiologically-based pharmacokinetic (PBPK) models provide a framework for in vitro-in vivo extrapolation of metabolic drug clearance. Many of the concepts in PBPK can have consequential impact on more mechanistic systems pharmacology models. In the gut wall, turnover of enzymes and enterocytes are typically lumped into one rate constant that describes the time dependent enzyme activity. This assumption may influence predictability of any sustained and dynamic effects such as mechanism-based inhibition (MBI), particularly when considering translation from healthy to gut disease. A novel multi-level systems PBPK model was developed. This model comprised a 'nested enzyme-within enterocyte' (NEWE) turnover model to describe levels of drug-metabolising enzymes. The ability of the model to predict gut metabolism following MBI and gut disease was investigated and compared to the conventional modelling approach. For MBI, the default NEWE model performed comparably to the conventional model. However, when drug-specific spatial crypt-villous absorption was considered, up to approximately 50% lower impact of MBI was simulated for substrates highly metabolised by cytochrome P450 (CYP) 3A4, interacting with potent inhibitors. Further, the model showed potential in predicting the disease effect of gastrointestinal mucositis and untreated coeliac disease when compared to indirect clinical pharmacokinetic parameters. Considering the added complexity of the NEWE model, it does not provide an attractive solution for improving upon MBI predictions in healthy individuals. However, nesting turnover may enable extrapolation to gut disease-drug interactions. The principle detailed herein may be useful for modelling drug interactions with cellular targets where turnover is significant enough to affect this process.


Assuntos
Citocromo P-450 CYP3A/metabolismo , Enterócitos/metabolismo , Enteropatias/metabolismo , Mucosa Intestinal/metabolismo , Modelos Biológicos , Citrus paradisi , Inibidores do Citocromo P-450 CYP3A/farmacologia , Sucos de Frutas e Vegetais , Preparações Farmacêuticas/metabolismo
17.
J Biol Chem ; 294(14): 5666-5676, 2019 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-30760524

RESUMO

Atg101 is an autophagy-related gene identified in worms, flies, mice, and mammals, which encodes a protein that functions in autophagosome formation by associating with the ULK1-Atg13-Fip200 complex. In the last few years, the critical role of Atg101 in autophagy has been well-established through biochemical studies and the determination of its protein structure. However, Atg101's physiological role, both during development and in adulthood, remains less understood. Here, we describe the generation and characterization of an Atg101 loss-of-function mutant in Drosophila and report on the roles of Atg101 in maintaining tissue homeostasis in both adult brains and midguts. We observed that homozygous or hemizygous Atg101 mutants were semi-lethal, with only some of them surviving into adulthood. Both developmental and starvation-induced autophagy processes were defective in the Atg101 mutant animals, and Atg101 mutant adult flies had a significantly shorter lifespan and displayed a mobility defect. Moreover, we observed the accumulation of ubiquitin-positive aggregates in Atg101 mutant brains, indicating a neuronal defect. Interestingly, Atg101 mutant adult midguts were shorter and thicker and exhibited abnormal morphology with enlarged enterocytes. Detailed analysis also revealed that the differentiation from intestinal stem cells to enterocytes was impaired in these midguts. Cell type-specific rescue experiments disclosed that Atg101 had a function in enterocytes and limited their growth. In summary, the results of our study indicate that Drosophila Atg101 is essential for tissue homeostasis in both adult brains and midguts. We propose that Atg101 may have a role in age-related processes.


Assuntos
Proteínas Relacionadas à Autofagia/metabolismo , Proteínas de Drosophila/metabolismo , Enterócitos/metabolismo , Homeostase , Intestinos , Longevidade , Neurônios/metabolismo , Animais , Proteínas Relacionadas à Autofagia/genética , Proteínas de Drosophila/genética , Drosophila melanogaster , Mutação com Perda de Função
18.
Nature ; 566(7742): 115-119, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30700910

RESUMO

The biochemical response to food intake must be precisely regulated. Because ingested sugars and fats can feed into many anabolic and catabolic pathways1, how our bodies handle nutrients depends on strategically positioned metabolic sensors that link the intrinsic nutritional value of a meal with intermediary metabolism. Here we describe a subset of immune cells-integrin ß7+ natural gut intraepithelial T lymphocytes (natural IELs)-that is dispersed throughout the enterocyte layer of the small intestine and that modulates systemic metabolism. Integrin ß7- mice that lack natural IELs are metabolically hyperactive and, when fed a high-fat and high-sugar diet, are resistant to obesity, hypercholesterolaemia, hypertension, diabetes and atherosclerosis. Furthermore, we show that protection from cardiovascular disease in the absence of natural IELs depends on the enteroendocrine-derived incretin GLP-12, which is normally controlled by IELs through expression of the GLP-1 receptor. In this metabolic control system, IELs modulate enteroendocrine activity by acting as gatekeepers that limit the bioavailability of GLP-1. Although the function of IELs may prove advantageous when food is scarce, present-day overabundance of diets high in fat and sugar renders this metabolic checkpoint detrimental to health.


Assuntos
Doenças Cardiovasculares/metabolismo , Progressão da Doença , Intestino Delgado/citologia , Linfócitos Intraepiteliais/metabolismo , Animais , Aterosclerose/genética , Aterosclerose/metabolismo , Aterosclerose/prevenção & controle , Doenças Cardiovasculares/genética , Doenças Cardiovasculares/prevenção & controle , Modelos Animais de Doenças , Ingestão de Alimentos , Enterócitos/citologia , Enterócitos/metabolismo , Feminino , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Cadeias beta de Integrinas/genética , Cadeias beta de Integrinas/metabolismo , Masculino , Síndrome Metabólica/genética , Síndrome Metabólica/metabolismo , Síndrome Metabólica/prevenção & controle , Camundongos
19.
Cell Mol Gastroenterol Hepatol ; 7(2): 255-274, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30686779

RESUMO

BACKGROUND & AIMS: Epithelial regeneration is essential for homeostasis and repair of the mucosal barrier. In the context of infectious and immune-mediated intestinal disease, interleukin (IL) 22 is thought to augment these processes. We sought to define the mechanisms by which IL22 promotes mucosal healing. METHODS: Intestinal stem cell cultures and mice were treated with recombinant IL22. Cell proliferation, death, and differentiation were assessed in vitro and in vivo by morphometric analysis, quantitative reverse transcriptase polymerase chain reaction, and immunohistochemistry. RESULTS: IL22 increased the size and number of proliferating cells within enteroids but decreased the total number of enteroids. Enteroid size increases required IL22-dependent up-regulation of the tight junction cation and water channel claudin-2, indicating that enteroid enlargement reflected paracellular flux-induced swelling. However, claudin-2 did not contribute to IL22-dependent enteroid loss, depletion of Lgr5+ stem cells, or increased epithelial proliferation. IL22 induced stem cell apoptosis but, conversely, enhanced proliferation within and expanded numbers of transit-amplifying cells. These changes were associated with reduced wnt and notch signaling, both in vitro and in vivo, as well as skewing of epithelial differentiation, with increases in Paneth cells and reduced numbers of enteroendocrine cells. CONCLUSIONS: IL22 promotes transit-amplifying cell proliferation but reduces Lgr5+ stem cell survival by inhibiting notch and wnt signaling. IL22 can therefore promote or inhibit mucosal repair, depending on whether effects on transit-amplifying or stem cells predominate. These data may explain why mucosal healing is difficult to achieve in some inflammatory bowel disease patients despite markedly elevated IL22 production.


Assuntos
Interleucinas/farmacologia , Receptores Acoplados a Proteínas-G/metabolismo , Receptores Notch/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Biomarcadores/metabolismo , Contagem de Células , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Claudina-2/metabolismo , Enterócitos/citologia , Enterócitos/efeitos dos fármacos , Enterócitos/metabolismo , Intestinos/citologia , Camundongos , Camundongos Endogâmicos C57BL , Organoides/metabolismo , Células-Tronco/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos
20.
Development ; 146(3)2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30658986

RESUMO

A distinct taxon of the Drosophila microbiota, Lactobacillus plantarum, is capable of stimulating the generation of reactive oxygen species (ROS) within cells, and inducing epithelial cell proliferation. Here, we show that microbial-induced ROS generation within Drosophila larval stem cell compartments exhibits a distinct spatial distribution. Lactobacilli-induced ROS is strictly excluded from defined midgut compartments that harbor adult midgut progenitor (AMP) cells, forming a functional 'ROS sheltered zone' (RSZ). The RSZ is undiscernible in germ-free larvae, but forms following monocolonization with L. plantarum L. plantarum is a strong activator of the ROS-sensitive CncC/Nrf2 signaling pathway within enterocytes. Enterocyte-specific activation of CncC stimulated the proliferation of AMPs, which demonstrates that pro-proliferative signals are transduced from enterocytes to AMPs. Mechanistically, we show that the cytokine Upd2 is expressed in the gut following L. plantarum colonization in a CncC-dependent fashion, and may function in lactobacilli-induced AMP proliferation and intestinal tissue growth and development.


Assuntos
Microbioma Gastrointestinal/fisiologia , Lactobacillus plantarum/crescimento & desenvolvimento , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/fisiologia , Nicho de Células-Tronco/fisiologia , Células-Tronco/metabolismo , Animais , Drosophila melanogaster , Enterócitos/citologia , Enterócitos/metabolismo , Oxirredução , Células-Tronco/citologia
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