RESUMEN
Oxidative stress can lead to intestinal cell injury as well as the induction of inflammation. It is not clear whether inflammation is an important factor leading to cell injury caused by oxidative stress. The purpose of this study was to investigate the role of inflammation in intestinal injury caused by hydrogen peroxide (H2O2). Our results revealed that H2O2 stimulation significantly decreased the viability of intestinal porcine epithelial cells (IPEC-1), increased lactate dehydrogenase (LDH) activity, and disrupted the distribution of the tight junction protein claudin-1. H2O2 significantly increased the mRNA expression of interleukin-6 (IL-6), IL-8, and tumor necrosis factor-α (TNF-α). H2O2 stimulation also led to increased phosphorylation of p38 and jun N-terminal kinase (JNK), and p65 NF-κB protein translocation into the nucleus of IPEC-1 cells. Cells treated with the NF-κB inhibitor (BAY11-7082), the p38 inhibitor (SB202190), or the JNK inhibitor (PD98059) significantly decreased mRNA and protein expression of IL-6, IL-8, and TNF-α. However, treatment with mitogen-activated protein kinase (MAPK) or NF-κB inhibitors did not prevent the damage effect on cell viability, LDH activity, or the distribution of claudin-1 in cells challenged with H2O2. In summary, our data demonstrate that activation of the NF-κB and MAPK signaling pathways can contribute to the inflammatory response, but not cell injury, in IPEC-1 cells challenged with H2O2.
Asunto(s)
Peróxido de Hidrógeno/toxicidad , Inflamación/patología , Sistema de Señalización de MAP Quinasas , FN-kappa B/metabolismo , Animales , Línea Celular , Supervivencia Celular/efectos de los fármacos , Fluorescencia , Inflamación/genética , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , L-Lactato Deshidrogenasa/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Fosforilación/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Porcinos , Proteínas de Uniones Estrechas/metabolismo , Factor de Transcripción ReIA/metabolismoRESUMEN
This study was conducted to investigate whether medium-chain triglycerides (MCTs) attenuated lipopolysaccharide (LPS)-induced liver injury by down-regulating necroptotic and inflammatory signaling pathways. A total of 24 pigs were randomly allotted to four treatments in a 2 × 2 factorial design including diet (0 and 4% MCTs) and immunological challenge (saline and LPS). After three weeks of feeding with or without 4% MCTs, pigs were challenged with saline or LPS. MCTs led to a significant increase in eicosapentaenoic acid, docosahexaenoic acid and total (n-3) polyunsaturated fatty acid concentrations. MCTs attenuated LPS-induced liver injury as indicated by an improvement in liver histomorphology and ultrastructural morphology of hepatocytes, a reduction in serum alanine aminotransferase and alkaline phosphatase activities as well as an increase in claudin-1 protein expression. In addition, MCTs also reduced serum tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß and IL-6 concentrations, liver TNF-α and IL-1ß mRNA expression and protein concentrations and enhanced liver heat shock protein 70 protein expression in LPS-challenged pigs. Moreover, MCTs decreased mRNA expression of receptor-interacting serine/threonine-protein kinase (RIP) 3, mixed-lineage kinase domain-like protein (MLKL) and phosphoglycerate mutase 5 and inhibited MLKL phosphorylation in the liver. Finally, MCTs decreased liver mRNA expression of toll-like receptor (TLR) 4, nucleotide-binding oligomerization domain protein (NOD) 1 and multiple downstream signaling molecules. MCTs also suppressed LPS-induced p38 mitogen-activated protein kinase (MAPK) phosphorylation and increased extracellular signal-related kinase 1/2 phosphorylation in the liver. These results indicated that MCTs are capable of attenuating LPS-induced liver damage by suppressing hepatic necroptotic (RIP1/RIP3/MLKL) and inflammatory (TLR4/NOD1/p38 MAPK) signaling pathways.
Asunto(s)
Antiinflamatorios/uso terapéutico , Apoptosis , Hepatopatías/tratamiento farmacológico , Sistema de Señalización de MAP Quinasas , Triglicéridos/uso terapéutico , Alanina Transaminasa/sangre , Fosfatasa Alcalina/sangre , Animales , Antiinflamatorios/administración & dosificación , Citocinas/genética , Citocinas/metabolismo , Suplementos Dietéticos , Ácidos Grasos Omega-3/metabolismo , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Lipopolisacáridos/toxicidad , Hepatopatías/etiología , Proteína Adaptadora de Señalización NOD1/genética , Proteína Adaptadora de Señalización NOD1/metabolismo , Distribución Aleatoria , Porcinos , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismo , Triglicéridos/administración & dosificaciónRESUMEN
This study was conducted to evaluate whether medium-chain TAG (MCT) could alleviate Escherichia coli lipopolysaccharide (LPS)-induced intestinal injury by regulating intestinal epithelial inflammatory response, as well as necroptosis. A total of twenty-four weanling piglets were randomly allotted to one of four treatments in a 2×2 factorial arrangement including diet type (5 % maize oil v. 4 % MCT+1 % maize oil) and immune stress (saline v. E. coli LPS). The piglets were fed diets containing maize oil or MCT for 21 d. On 21 d, piglets were injected intraperitoneally with saline or LPS. The blood and intestinal samples were collected at 4 h post injection. Supplementation with MCT improved intestinal morphology, digestive and barrier function, indicated by increased jejunal villus height, increased jejunal and ileal disaccharidases (sucrase and maltase) activities, as well as enhanced protein expression of claudin-1. Furthermore, the protein expression of heat-shock protein 70 in jejunum and the concentration of TNF-α in plasma were reduced in the piglets fed diets supplemented with MCT. In addition, MCT down-regulated the mRNA expression of toll-like receptor 4 (TLR4) and nucleotide-binding oligomerisation domain proteins (NOD) signalling-related genes in jejunum and ileum. Finally, MCT inhibited jejunal and ileal enterocyte necroptosis indicated by suppressed mRNA expression of the receptor-interacting protein 3 and mixed-lineage kinase domain-like protein. These results indicate that MCT supplementation may be closely related to inhibition of TLR4, NOD and necroptosis signalling pathways and concomitant improvement of intestinal integrity under an inflammatory condition.
Asunto(s)
Intestinos/efectos de los fármacos , Porcinos/fisiología , Receptor Toll-Like 4/metabolismo , Triglicéridos/farmacología , Alimentación Animal/análisis , Animales , Dieta/veterinaria , Suplementos Dietéticos , Regulación hacia Abajo , Regulación de la Expresión Génica/efectos de los fármacos , Intestinos/patología , Intestinos/fisiología , Lipopolisacáridos/efectos adversos , Masculino , ARN Mensajero/genética , ARN Mensajero/metabolismo , Distribución Aleatoria , Porcinos/metabolismo , Receptor Toll-Like 4/genética , Triglicéridos/administración & dosificación , Factor de Necrosis Tumoral alfa/sangreRESUMEN
SCOPE: Flaxseed oil is a rich source of α-linolenic acid (ALA), which is the precursor of the long-chain n-3 polyunsaturated fatty acids (PUFAs), including docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). This study investigates the protective effect of flaxseed oil against intestinal injury induced by lipopolysaccharide (LPS). MATERIALS AND RESULTS: Twenty-four weaned pigs were used in a 2 × 2 factorial experiment with dietary treatment (5% corn oil vs 5% flaxseed oil) and LPS challenge (saline vs LPS). On day 21 of the experiment, pigs were administrated with LPS or saline. At 2 h and 4 h post-administration, blood samples were collected. After the blood harvest at 4 h, all piglets were slaughtered and intestinal samples were collected. Flaxseed oil supplementation led to the enrichment of ALA, EPA, and total n-3 PUFAs in intestine. Flaxseed oil improved intestinal morphology, jejunal lactase activity, and claudin-1 protein expression. Flaxseed oil downregulated the mRNA expression of intestinal necroptotic signals. Flaxseed oil also downregulated the mRNA expression of intestinal toll-like receptors 4 (TLR4) and its downstream signals myeloid differentiation factor 88 (MyD88), nuclear factor kappa B (NF-κB), and nucleotide-binding oligomerization domain proteins 1, 2 (NOD1, NOD2) and its adapter molecule, receptor-interacting protein kinase 2 (RIPK2). CONCLUSION: These results suggest that dietary addition of flaxseed oil enhances intestinal integrity and barrier function, which is involved in modulating necroptosis and TLR4/NOD signaling pathways.
Asunto(s)
Apoptosis , Enterocolitis Necrotizante/prevención & control , Mucosa Intestinal/metabolismo , Aceite de Linaza/uso terapéutico , Transducción de Señal , Receptor Toll-Like 4/metabolismo , Ácido alfa-Linolénico/uso terapéutico , Animales , Apoptosis/efectos de los fármacos , Peso Corporal , Cruzamientos Genéticos , Enterocolitis Necrotizante/sangre , Enterocolitis Necrotizante/inducido químicamente , Enterocolitis Necrotizante/metabolismo , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/crecimiento & desarrollo , Mucosa Intestinal/inmunología , Intestinos/efectos de los fármacos , Intestinos/crecimiento & desarrollo , Intestinos/inmunología , Aceite de Linaza/efectos adversos , Lipopolisacáridos/toxicidad , Masculino , Proteína Adaptadora de Señalización NOD1/genética , Proteína Adaptadora de Señalización NOD1/metabolismo , Proteína Adaptadora de Señalización NOD2/genética , Proteína Adaptadora de Señalización NOD2/metabolismo , Orquiectomía/veterinaria , Sustancias Protectoras/efectos adversos , Sustancias Protectoras/uso terapéutico , Distribución Aleatoria , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor/genética , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor/metabolismo , Sus scrofa , Receptor Toll-Like 4/genética , Destete , Ácido alfa-Linolénico/efectos adversos , Ácido alfa-Linolénico/metabolismoRESUMEN
Infection and inflammation can result in the rapid loss of muscle mass and myofibrillar proteins (muscle atrophy). In addition, aspartate (Asp) is necessary for protein synthesis in mammalian cells. We hypothesized that Asp could attenuate LPS-induced muscle atrophy in a piglet model. Twenty-four weaning piglets were allotted to four treatments, including non-challenged control, LPS challenged control, LPS+0.5% Asp and LPS+1.0% Asp. On d 21, the piglets were injected with i.p. injection of LPS (100 ug/kg BM) or saline. At 4 h post-injection, blood, gastrocnemius and longissimus dorsi muscles samples were collected for biochemical and molecular analyses. Asp decreased the concentrations of cortisol and glucagon in plasma. In addition, Asp increased protein and RNA contents in muscles, and decreased mRNA expression of muscle atrophy F-box (MAFbx) and muscle RING finger 1 (MuRF1). Moreover, Asp decreased phosphorylation of AMPKα but increased phosphorylation of Akt and Forkhead Box O (FOXO) 1 in the muscles. Our results indicate that Asp suppresses LPS-induced MAFbx and MuRF1 expression via activation of Akt signaling, and inhibition of AMPKα and FOXO1 signaling.