Anti-inflammation effect of moxibustion for rats with diarrhea-predominant irritable bowel syndrome based on multiple miRNAs regulating NF-κB signal pathway / 中国针灸

OBJECTIVE@#To observe the effect of moxibustion on the regulation of nuclear factor-kappa B (NF-κB) and inflammatory factors by multiple microRNAs (miRNAs) in rats with diarrhea-predominant irritable bowel syndrome (IBS-D), and to explore the anti-inflammatory mechanism of moxibustion on IBS-D.@*METHODS@#Twelve of 52 newborn rats were randomly selected into a normal group. The remaining rats were made into IBS-D model. A total of 36 rats with successful model were randomly divided into a model group, a medication group and a moxibustion group, 12 rats in each group. The rats in the medication group were intraperitoneally injected with pyrrolidine dithiocarbamate (PDTC). The rats in the moxibustion group were treated with moxibustion at "Tianshu" (ST 25) and "Shangjuxu" (ST 37) for 20 min each time. All the intervention was given once a day for 7 days. Before and after modeling as well as after intervention, the body mass, loose stool rate and the minimum volume threshold of abdominal withdrawal reflex (AWR) were measured. After intervention, the contents of serum tumor necrosis factor α (TNF-α), interleukin (IL)-1β and IL-8 were detected by ELISA method; the morphology of colon tissues was observed by HE staining, and the expressions of miR-155, miR-125b, miR-29b, miR-31, miR-18a and NF-κB p65 mRNA in colon tissues were detected by real-time PCR. The expressions of NF-κB p65, TNF-α, IL-1β and IL-8 protein in colon tissues were detected by immunofluorescence.@*RESULTS@#After modeling, the body mass and the minimum volume threshold of AWR in the model group were lower than those in the normal group (P<0.01); the rates of loose stool in the model group were higher than those in the normal group (P<0.01); after intervention, in the model group, the inflammatory infiltration of colon tissues was obvious, and the serum levels of TNF-α, IL-1 β, IL-8 were higher than those in the normal group (P<0.05); the expression of miR-155, miR-125b, miR-29b, miR-31, miR-18a and NF-κB p65 mRNA in colon tissues was higher than that in the normal group (P<0.05); the protein expression of NF-κB p65, TNF-α, IL-1β, IL-8 was also higher than that in the normal group (P<0.01). After intervention, the body mass and the minimum volume threshold of AWR in the medication group and the moxibustion group were both higher than those in the model group (P<0.05); the loose stool rate in the medication group and the moxibustion group were lower than those in model group (P<0.05); the inflammatory cells infiltration in the colon tissues was less, the serum levels of TNF-α, IL-1β and IL-8 as well as the protein expression of NF-κB p65, TNF-α, IL-1β and IL-8 in the colon tissues in the medication group and the moxibustion group were lower than those in the model group (P<0.05, P<0.01). The expression of miR-125b, miR-31, miR-18a and NF-κB p65 mRNA in the medication group were lower than those in the model group (P<0.05). The expression of miR-155, miR-125b, miR-29b, miR-31, miR-18a and NF-κB p65 mRNA in the moxibustion group were lower than those in the model group (P<0.05). The miR-155, miR-125b, miR-29b, miR-31, miR-18a were positively correlated with NF-κB p65 mRNA (0<r<1, P<0.01).@*CONCLUSION@#The anti-inflammatory mechanism of moxibustion at "Tianshu" (ST 25) and "Shangjuxu" (ST 37) for IBS-D rats may be related to regulating multiple miRNAs to inhibit NF-κB signal pathway and reduce the expression of inflammatory factors.

Association of il-8 -251t>a (rs4073) polymorphism with susceptibility to gastric cancer: a systematic review and meta-analysis based on 33 case-control studies / Associação de polimorfismo IL-8 -251T> A (rs4073) com suscetibilidade ao câncer gástrico: uma revisão sistemática e meta-análise com base em 33 estudos caso controle

ABSTRACT BACKGROUND: The role of -251A>T polymorphism in the anti-inflammatory cytokine interleukin-8 (IL-8) gene in gastric cancer was intensively evaluated, but the results of these studies were inconsistent. OBJECTIVE: Therefore, we performed a meta-analysis to provide a comprehensive data on the association of IL-8 -251T>A polymorphism with gastric cancer. METHODS: All eligible studies were identified in PubMed, Web of Science, EMBASE, Wanfang and CNKI databases before September 01, 2019. The pooled odds ratios (ORs) with 95% confidence intervals (CIs) were derived from a fixed effect or random effect model. RESULTS: A total of 33 case-control studies with 6,192 cases and 9,567 controls were selected. Overall, pooled data showed that IL-8 -251T>A polymorphism was significantly associated with an increased risk of gastric cancer under all five genetic models, i.e., allele (A vs T: OR=1.189, 95% CI 1.027-1.378, P=0.021), homozygote (AA vs TT: OR=1.307, 95% CI 1.111-1.536, P=0.001), heterozygote (AT vs TT: OR=1.188, 95% CI 1.061-1.330, P=0.003), dominant (AA+AT vs TT: OR=1.337, 95% CI 1.115-1.602, P=0.002) and recessive (AA vs AT+TT: OR=1.241, 95% CI 1.045-1.474, P=0.014). The stratified analysis by ethnicity revealed an increased risk of gastric cancer in Asians and mixed populations, but not in Caucasians. Moreover, stratified by country found a significant association in Chinese, Korean and Brazilian, but not among Japanese. CONCLUSION: This meta-analysis suggests that the IL-8 -251T>A polymorphism is associated with an increased risk of gastric cancer, especially by ethnicity (Asian and mixed populations) and country (Chinese, Korean and Brazilian).

RESUMO CONTEXTO: O papel do polimorfismo -251A>T no gene anti-inflamatório citocina interleucina-8 (IL-8) no câncer gástrico foi intensamente avaliado, mas os resultados desses estudos foram inconsistentes. OBJETIVO: Portanto, realizamos uma meta-análise para fornecer dados abrangentes sobre a associação de IL-8 -251T>A polimorfismo com câncer gástrico. MÉTODOS: Todos os estudos elegíveis foram identificados nos bancos de dados PubMed, Web of Science, EMBASE, Wanfang e CNKI antes de 01 de setembro de 2019. As relações de probabilidades agrupadas (ORs) com intervalos de confiança de 95% (IC) foram derivadas de um modelo de efeito fixo ou efeito aleatório. RESULTADOS: Foram selecionados 33 estudos de controle de caso com 6.192 casos e 9.567 controles. No geral, dados agrupados mostraram que o polimorfismo IL-8 -251T>A foi significativamente associado a um risco aumentado de câncer gástrico em todos os cinco modelos genéticos, isto é, alelo (A vs T: OR=1,189; 95% CI 1,027-1,378; P=0,021), homozigoto (AA vs TT: OR=1,307; 95% CI 1,111-1,536; P=0,001), heterozigoto (AT vs TT: OR=1,188; 95% CI 1,061-1,330; P=0,003), dominante (AA+AT vs TT: OR=1,337; 95% CI 1,115-1,602; P=0,002) e recessivo (AA vs AT+TT: OR=1,241; 95% CI 1,045-1,474; P=0,014). A análise estratificada por etnia revelou um risco aumentado de câncer gástrico em asiáticos e populações mistas, mas não em caucasianos. Além disso, estratificado por país. Encontrou-se uma associação significativa em chineses, coreanos e brasileiros, mas não entre os japoneses. CONCLUSÃO: Esta meta-análise sugere que o polimorfismo IL-8 -251T>A está associado a um risco aumentado de câncer gástrico, especialmente por etnia (populações asiáticas e mistas) e por país (chinês, coreano e brasileiro).

Variants in the interleukin 8 gene and the response to inhaled bronchodilators in cystic fibrosis / Variantes no gene da interleucina 8 e a resposta a broncodilatadores inalatórios na fibrose cística

Abstract Objective: Interleukin 8 protein promotes inflammatory responses, even in airways. The presence of interleukin 8 gene variants causes altered inflammatory responses and possibly varied responses to inhaled bronchodilators. Thus, this study analyzed the interleukin 8 variants (rs4073, rs2227306, and rs2227307) and their association with the response to inhaled bronchodilators in cystic fibrosis patients. Methods: Analysis of interleukin 8 gene variants was performed by restriction fragment length polymorphism of polymerase chain reaction. The association between spirometry markers and the response to inhaled bronchodilators was evaluated by Mann-Whitney and Kruskal-Wallis tests. The analysis included all cystic fibrosis patients, and subsequently patients with two mutations in the cystic fibrosis transmembrane conductance regulator gene belonging to classes I to III. Results: This study included 186 cystic fibrosis patients. There was no association of the rs2227307 variant with the response to inhaled bronchodilators. The rs2227306 variant was associated with FEF50% in the dominant group and in the group with two identified mutations in the cystic fibrosis transmembrane conductance regulator gene. The rs4073 variant was associated with spirometry markers in four genetic models: co-dominant (FEF25-75% and FEF75%), dominant (FEV1, FEF50%, FEF75%, and FEF25-75%), recessive (FEF75% and FEF25-75%), and over-dominant (FEV1/FVC). Conclusions: This study highlighted the importance of the rs4073 variant of the interleukin 8 gene, regarding response to inhaled bronchodilators, and of the assessment of mutations in the cystic fibrosis transmembrane conductance regulator gene.

Resumo Objetivo: A proteína interleucina 8 promove respostas inflamatórias, o que inclui sua atuação nas vias aéreas. A presença de variantes no gene da interleucina 8 causa respostas inflamatórias alteradas e possivelmente respostas variadas ao uso de broncodilatadores inalatórios. Assim, este estudo analisou as variantes da interleucina 8 (rs4073, rs2227306, rs2227307) e sua associação à resposta a broncodilatadores inalatórios em pacientes com fibrose cística. Métodos: Foi feita análise das variantes genéticas da interleucina 8 por restriction fragment length polymorphism da reação em cadeia da polimerase. A associação entre os marcadores da espirometria e a resposta a broncodilatadores inalatórios foi feita pelos testes de Mann-Whitney e Kruskal-Wallis. A análise incluiu todos os pacientes com fibrose cística e posteriormente pacientes com duas mutações no gene cystic fibrosis transmembrane conductance regulator pertencentes às Classes I a II. Resultados: Este estudo incluiu 186 pacientes com fibrose cística. Não houve associação da variante rs2227307 à resposta a broncodilatadores inalatórios. A variante rs2227306 foi associada a FEF50% no grupo dominante e no grupo com duas mutações identificadas no gene cystic fibrosis transmembrane conductance regulator. A variante rs4073 foi associada a marcadores da espirometria em quatro modelos genéticos: codominante (FEF25-75% e FEF75%), dominante (VEF1, FEF50%, FEF75% e FEF25-75%), recessivo (FEF75% e FEF25-75%) e overdominante (VEF1/CVF). Conclusões: Este estudo destaca, principalmente, a importância da variante rs4073 do gene da interleucina 8, na resposta a broncodilatadores inalatórios, concomitantemente ao genótipo das mutações no gene cystic fibrosis transmembrane conductance regulator.

Influence of IL-6, IL-8, and TGF-β 1 gene polymorphisms on the risk of human papillomavirus-infection in women from Pernambuco, Brazil

Human papillomavirus (HPV) infections are strongly associated with the development of cervical intraepithelial neoplasias and invasive cervical cancer. Polymorphisms in cytokine-encoding genes and behavioural cofactors could play an important role in protecting an individual against viral infections and cancer. Here, we investigated whether IL-6 -174 G>C, IL-8 +396 G>T, and TGF-β1 +869 G>C and +915 G>C polymorphisms were associated with susceptibility to HPV infection in women from north-east (Pernambuco) Brazil. We analysed 108 healthy uninfected women (HC) and 108 HPV-positive women with cervical lesions. Genetic polymorphisms were assessed using Sanger sequencing and polymerase chain reaction-restriction fragment length polymorphism. Comparison of the distribution of the genotypic and allelic frequencies of the IL-18 +396 T>G polymorphism between HPV infected woman an uninfected controls showed that the GG genotype and G allele were both more frequent in the HC group, and were associated with protection from HPV infection (p = 0.0015; OR = 0.29 CI95% = 0.13-0.61; p = 0.0005; OR = 0.45 CI95% 0.29-0.7, respectively). Individuals from the control group could have previously had HPV infection that was spontaneously eliminated; however, it was undetectable at the time of sample collection. Based on our findings, we hypothesize that the IL-8 +396 G>T polymorphism could interfere with susceptibility to HPV infection, by modulating the ability of immune system to fight the virus.

alpha-Lipoic Acid Inhibits Expression of IL-8 by Suppressing Activation of MAPK, Jak/Stat, and NF-kappaB in H. pylori-Infected Gastric Epithelial AGS Cells

The epithelial cytokine response, associated with reactive oxygen species (ROS), is important in Helicobacter pylori (H. pylori)-induced inflammation. H. pylori induces the production of ROS, which may be involved in the activation of mitogen-activated protein kinases (MAPK), janus kinase/signal transducers and activators of transcription (Jak/Stat), and oxidant-sensitive transcription factor, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kappaB), and thus, expression of interleukin-8 (IL-8) in gastric epithelial cells. alpha-lipoic acid, a naturally occurring thiol compound, is a potential antioxidant. It shows beneficial effects in treatment of oxidant-associated diseases including diabetes. The present study is purposed to investigate whether alpha-lipoic acid inhibits expression of inflammatory cytokine IL-8 by suppressing activation of MAPK, Jak/Stat, and NF-kappaB in H. pylori-infected gastric epithelial cells. Gastric epithelial AGS cells were pretreated with or without alpha-lipoic acid for 2 h and infected with H. pylori in a Korean isolate (HP99) at a ratio of 300:1. IL-8 mRNA expression was analyzed by RT-PCR analysis. IL-8 levels in the medium were determined by enzyme-linked immunosorbent assay. NF-kappaB-DNA binding activity was determined by electrophoretic mobility shift assay. Phospho-specific and total forms of MAPK and Jak/Stat were assessed by Western blot analysis. ROS levels were determined using dichlorofluorescein fluorescence. As a result, H. pylori induced increases in ROS levels, mRNA, and protein levels of IL-8, as well as the activation of MAPK [extracellular signal-regulated kinase 1/2 (ERK1/2), c-Jun NH2-terminal kinase 1/2 (JNK1/2), p38], Jak/Stat (Jak1/2, Stat3), and NF-kappaB in AGS cells, which was inhibited by alpha-lipoic acid. In conclusion, alpha-lipoic acid may be beneficial for prevention and/or treatment of H. pylori infection-associated gastric inflammation.

Jak1/Stat3 Is an Upstream Signaling of NF-kappaB Activation in Helicobacter pylori-Induced IL-8 Production in Gastric Epithelial AGS Cells

Helicobacter pylori (H. pylori) induces the activation of nuclear factor-kB (NF-kappaB) and cytokine expression in gastric epithelial cells. The Janus kinase/signal transducers and activators of transcription (Jak/Stat) cascade is the inflammatory signaling in various cells. The purpose of the present study is to determine whether H. pylori-induced activation of NF-kappaB and the expression of interleukin-8 (IL-8) are mediated by the activation of Jak1/Stat3 in gastric epithelial (AGS) cells. Thus, gastric epithelial AGS cells were infected with H. pylori in Korean isolates (HP99) at bacterium/cell ratio of 300:1, and the level of IL-8 in the medium was determined by enzyme-linked immonosorbent assay. Phospho-specific and total forms of Jak1/Stat3 and IkappaBalpha were assessed by Western blot analysis, and NF-kappaB activation was determined by electrophoretic mobility shift assay. The results showed that H. pylori induced the activation of Jak1/Stat3 and IL-8 production, which was inhibited by a Jak/Stat3 specific inhibitor AG490 in AGS cells in a dose-dependent manner. H. pylori-induced activation of NF-kappaB, determined by phosphorylation of IkappaBalpha and NF-kappaB-DNA binding activity, were inhibited by AG490. In conclusion, Jak1/Stat3 activation may mediate the activation of NF-kappaB and the expression of IL-8 in H. pylori-infected AGS cells. Inhibition of Jak1/Stat3 may be beneficial for the treatment of H. pylori-induced gastric inflammation, since the activation of NF-kappaB is inhibited and inflammatory cytokine expression is suppressed.

Jak1/Stat3 Is an Upstream Signaling of NF-kappaB Activation in Helicobacter pylori-Induced IL-8 Production in Gastric Epithelial AGS Cells

Helicobacter pylori (H. pylori) induces the activation of nuclear factor-kB (NF-kappaB) and cytokine expression in gastric epithelial cells. The Janus kinase/signal transducers and activators of transcription (Jak/Stat) cascade is the inflammatory signaling in various cells. The purpose of the present study is to determine whether H. pylori-induced activation of NF-kappaB and the expression of interleukin-8 (IL-8) are mediated by the activation of Jak1/Stat3 in gastric epithelial (AGS) cells. Thus, gastric epithelial AGS cells were infected with H. pylori in Korean isolates (HP99) at bacterium/cell ratio of 300:1, and the level of IL-8 in the medium was determined by enzyme-linked immonosorbent assay. Phospho-specific and total forms of Jak1/Stat3 and IkappaBalpha were assessed by Western blot analysis, and NF-kappaB activation was determined by electrophoretic mobility shift assay. The results showed that H. pylori induced the activation of Jak1/Stat3 and IL-8 production, which was inhibited by a Jak/Stat3 specific inhibitor AG490 in AGS cells in a dose-dependent manner. H. pylori-induced activation of NF-kappaB, determined by phosphorylation of IkappaBalpha and NF-kappaB-DNA binding activity, were inhibited by AG490. In conclusion, Jak1/Stat3 activation may mediate the activation of NF-kappaB and the expression of IL-8 in H. pylori-infected AGS cells. Inhibition of Jak1/Stat3 may be beneficial for the treatment of H. pylori-induced gastric inflammation, since the activation of NF-kappaB is inhibited and inflammatory cytokine expression is suppressed.

Nucleotide Binding Oligomerization Domain 1 Is an Essential Signal Transducer in Human Epithelial Cells Infected with Helicobacter pylori That Induces the Transepithelial Migration of Neutrophils

BACKGROUND/AIMS: The cytosolic host protein nucleotide binding oligomerization domain 1 (Nod1) has emerged as a key pathogen recognition molecule for innate immune responses in epithelial cells. The purpose of the study was to elucidate the mechanism by which Helicobacter pylori infection leads to transepithelial neutrophil migration in a Nod1-mediated manner. METHODS: Human epithelial cell lines AGS and Caco-2 were grown and infected with H. pylori. Interleukin (IL)-8 mRNA expression and IL-8 secretion were assessed, and nuclear factor kappaB (NF-kappaB) activation was determined. Stable transfections of AGS and Caco-2 cells with dominant negative Nod1 were generated. Neutrophil migration across the monolayer was quantified. RESULTS: Cytotoxin-associated gene pathogenicity island (cagPAI)(+) H. pylori infection upregulated IL-8 mRNA expression and IL-8 secretion in AGS and Caco-2 cells compared with controls. NF-kappaB activation, IL-8 mRNA expression and IL-8 secretion by cagPAI knockdown strains were reduced compared with those infected with the wild-type strain. NF-kappaB activation, IL-8 mRNA expression and IL-8 secretion in dominant-negative (DN)-Nod1 stably transfected cells were reduced compared with the controls. The transepithelial migration of neutrophils in DN-Nod1 stably transfected cells was reduced compared with that in controls. CONCLUSIONS: Signaling through Nod1 plays an essential role in neutrophil migration induced by the upregulated NF-kappaB activation and IL-8 expression in H. pylori-infected human epithelial cells.

Influência dos polimorfismos genéticos (IL10/CXCL8/CXCR2/ NF?B) na susceptibilidade das doenças reumatológicas autoimunes / Influence of genetic polymorphisms (IL-10/CXCL8/CXCR2/ NF?B) on the susceptibility of autoimmune rheumatic diseases

As doenças reumatológicas autoimunes, na maioria das vezes, possuem uma via genética comum para a autoimunidade. Vários genes foram associados com as doenças reumatológicas, para tanto iremos analisar somente alguns genes nos quais há várias evidências da existência de associação com risco ou proteção de doença autoimune. O fator de transcrição nuclear kappa B (NF-kappa B), o qual regula as respostas imunes e inflamatórias, está associado com esclerose sistêmica (ES), artrite reumatoide (AR) e lúpus eritematoso sistêmico (LES), assim como os genes CXCR2 e CXCL8. Já a interleucina 10 (IL-10), que é uma citocina anti-inflamatória, está associada com quase todas as doenças reumatológicas. Neste artigo, revisamos os potenciais papéis desses genes no sistema imunológico e em diversas doenças reumatológicas. Com relação à IL-10, diversos estudos foram realizados, porém em sua maioria contraditórios - alguns encontraram ausência de associação e outros encontraram associação em diferentes polimorfismos do genes. Já em relação ao NF-kappa B, somente foi estudado em AR e LES, e não foram observadas análises significativas relevantes. Os polimorfismos genéticos do gene CXCR2 foram associados com ES, mas não estão associados com AR e LES. Já os polimorfismos genéticos do gene CXCL8 não estão associados com ES, mas estão associados com AR.

The autoimmune rheumatologic disorders mostly have a common genetic path to the autoimmunity. Several genes have been associated with rheumatologic disorders; therefore, we are analyzing just the ones in those containing several evidences of the existence of association with the risk or protection from autoimmune disorder. The nuclear factor kappa beta (NF-kappa B), which regulates the autoimmune and anti-inflammatory responses, is associated with systemic sclerosis (SS), rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE), just as the CXCR2 e CXCL8 genes. On the other hand, the interleukin-10 (IL-10), which is an anti-inflammatory cytokine, is associated with almost all rheumatologic disorders. In this article, we are reviewing the potential roles of these genes in the immune system and in several rheumatologic disorders. In relation to IL-10, several studies have been carried out, but most of them are controversial - some detected the absence of association, and others found association in different genetic polymorphisms. Conversely, in relation to NF-kappa B, it was studied just in RA and SLE, and no relevant significant analyses were observed. The genetic polymorphisms of the CXCR2 gene were associated with SS, but not with RA e SLE. On the other side, the genetic polymorphisms of the CXCL8 gene are not associated with SS, but with RA.

The role of adiponectin in the production of IL-6, IL-8, VEGF and MMPs in human endothelial cells and osteoblasts: implications for arthritic joints

This study was performed to evaluate the contribution of adiponectin to the production of interleukin (IL)-6, IL-8, vascular endothelial growth factor (VEGF), matrix metalloproteinase (MMP)-1 and MMP-13 in human endothelial cells and osteoblasts in arthritic joints. Cultured human umbilical vascular endothelial cells (HUVECs) and osteoblasts were stimulated with adiponectin (1 or 10 mug ml-1) or IL-1beta (0.1 ng ml-1) in the presence or absence of hypoxia for 24 h. The protein expression patterns were examined by analyzing culture supernatants using the enzyme-linked immunosorbent assay (ELISA). Adiponectin significantly stimulated the production of VEGF, MMP-1 and MMP-13 in osteoblasts but not in endothelial cells, whereas it significantly stimulated the production of IL-6 and IL-8 in both endothelial cells and osteoblasts. The increase in VEGF production induced by adiponectin was significantly greater than that induced by IL-1beta. The production of IL-6 and IL-8 in adiponectin-stimulated endothelial cells was approximately 10-fold higher than that in IL-1beta-stimulated endothelial cells; in osteoblasts, adiponectin-induced IL-6 and IL-8 secretion was approximately twofold higher than that induced by IL-1beta. In addition, IL-8 production in endothelial cells was approximately sevenfold higher than in osteoblasts. However, IL-6 levels were similar between the two cell types, suggesting that adiponectin may be involved in the production of IL-8 in endothelial cells, which may have an important role in neutrophil recruitment to arthritic joints. Furthermore, the increases in protein expression induced by adiponectin were differentially regulated by hypoxia. In conclusion, adiponectin has a more important role than does IL-1beta in the production of mediators that drive synovitis and joint destruction in endothelial cells and osteoblasts at physiological concentrations.

Anti-inflammatory Mechanism of Lactobacillus rhamnosus GG in Lipopolysaccharide-stimulated HT-29 Cell / 대한소화기학회지

BACKGROUND/AIMS: Probiotics are live non-pathogenic organisms that belong to the resident microflora, and confer health benefits by multiple mechanisms. Lactobacillus rhamnosus GG (LGG) is one of the probiotic bacteria that ameliorates intestinal injury and inflammation caused by various stimuli. We aimed to evaluate the anti-inflammatory effect and mechanism of LGG in lipopolysaccharide (LPS)-stimulated HT-29 cells. METHODS: HT-29 cells were stimulated with interleukin (IL)-1beta (2 ng/mL), tumor necrosis factor (TNF)-alpha (20 ng/mL), and LPS (20 microg/mL) in the presence or absence of LGG (107-109 colony forming units/mL). Production of the pro-inflammatory chemokine IL-8 was measured by ELISA and semi-quantitative PCR. Transcriptional activity of NF-kappaB-responsive gene was evaluated by luciferase assay with reporter gene. Toll-like receptor 4 (TLR4) mRNA expression was assessed by semi-quantitative PCR. The IkappaBalpha degradation was evaluated by western blot and intranuclear translocation of NF-kappaB was determined by western blot and immunofluorescence. RESULTS: LGG did not affect the viability of HT-29 cells. Pretreatment of HT-29 cells with LGG significantly blocked TNF-alpha, and LPS induced IL-8 activation at both mRNA and protein level (p<0.05). Pretreatment of HT-29 cells with LGG attenuated LPS-induced NF-kappaB nuclear translocation and also blocked LPS-induced IkappaBalpha degradation. LGG also down-regulated TLR4 mRNA activated by LPS. CONCLUSIONS: LGG attenuates LPS induced inflammation, and this may be associated with TLR4/NF-kappaB down-regulation.

Repression of TNF-alpha-induced IL-8 expression by the glucocorticoid receptor-beta involves inhibition of histone H4 acetylation

Increased expression of a number of proinflammatory genes, including IL-8, is associated with inflammatory conditions such as asthma. Glucocorticoid receptor (GR)beta, one of the GR isoforms, has been suggested to be upregulated in asthma associated with glucocorticoid insensitivity and to work as a dominant negative inhibitor of wild type GRalpha. However, recent data suggest that GRbeta is not a dominant negative inhibitor of GRalpha in the transrepressive process and has its own functional role. We investigated the functional role of GRbeta expression in the suppressive effect of glucocorticoids on tumor necrosis factor (TNF)-alpha-induced IL-8 release in an airway epithelial cell line. GRbeta expression was induced by treatment of epithelial cells with either dexamethasone or TNF-alpha. GRbeta was able to inhibit glucocorticoid-induced transcriptional activation mediated by binding to glucocorticoid response elements (GREs). The suppressive effect of dexamethasone on TNF-alpha-induced IL-8 transcription was not affected by GRbeta overexpression, rather GRbeta had its own weak suppressive activity on TNF-alpha-induced IL-8 expression. Overall histone deacetylase activity and histone acetyltransferase activity were not changed by GRbeta overexpression, but TNF-alpha-induced histone H4 acetylation at the IL-8 promoter was decreased with GRbeta overexpression. This study suggests that GRbeta overexpression does not affect glucocorticoid-induced suppression of IL-8 expression in airway epithelial cells and GRbeta induces its own histone deacetylase activity around IL-8 promoter site.

Repression of TNF-alpha-induced IL-8 expression by the glucocorticoid receptor-beta involves inhibition of histone H4 acetylation

Increased expression of a number of proinflammatory genes, including IL-8, is associated with inflammatory conditions such as asthma. Glucocorticoid receptor (GR)beta, one of the GR isoforms, has been suggested to be upregulated in asthma associated with glucocorticoid insensitivity and to work as a dominant negative inhibitor of wild type GRalpha. However, recent data suggest that GRbeta is not a dominant negative inhibitor of GRalpha in the transrepressive process and has its own functional role. We investigated the functional role of GRbeta expression in the suppressive effect of glucocorticoids on tumor necrosis factor (TNF)-alpha-induced IL-8 release in an airway epithelial cell line. GRbeta expression was induced by treatment of epithelial cells with either dexamethasone or TNF-alpha. GRbeta was able to inhibit glucocorticoid-induced transcriptional activation mediated by binding to glucocorticoid response elements (GREs). The suppressive effect of dexamethasone on TNF-alpha-induced IL-8 transcription was not affected by GRbeta overexpression, rather GRbeta had its own weak suppressive activity on TNF-alpha-induced IL-8 expression. Overall histone deacetylase activity and histone acetyltransferase activity were not changed by GRbeta overexpression, but TNF-alpha-induced histone H4 acetylation at the IL-8 promoter was decreased with GRbeta overexpression. This study suggests that GRbeta overexpression does not affect glucocorticoid-induced suppression of IL-8 expression in airway epithelial cells and GRbeta induces its own histone deacetylase activity around IL-8 promoter site.

Probiotics may Reduce Inflammation by Enhancing Peroxisome Proliferator Activated Receptor gamma Activation in HT-29 Cells / 대한소화기학회지

BACKGROUND/AIMS: The peroxisome proliferator-activated receptor gamma (PPARgamma) is a nuclear receptor highly expressed in the colon which plays an anti-inflammatory role through the inhibition of nuclear factor-kappaB (NF-kappaB) pathway. Probiotics have been shown to exert beneficial effects on inflammatory bowel diseases. However, the exact mechanism by which probiotics exert protection against intestinal inflammation is not well understood. The aims of this study were to evaluate the attenuation of inflammatory response by probiotics in intestinal epithelial cells and to study the association between probiotics and PPARgamma. METHODS: HT-29 human epithelial cells were stimulated with LPS (20microgram/mL) and probiotics, Lactobacillus casei (L. casei) (10(5)-10(7) cfu/mL), or with LPS (20microgram/mL) alone for 24 hours. Interleukin-8 (IL-8), cyclooxygenase-2 (COX-2), toll-like receptor-4 (TLR-4) and PPARgamma mRNA expressions were assessed by RT-PCR. IL-8 protein secretion was measured by ELISA. HT-29 cells were transfected with tk promoter-luciferase plasmid containing a peroxisome proliferator response element (PPRE). After stimulation with L. casei or PPARgamma agonist (15d-PGJ2 or ciglitazone), luciferase activities were measured. RESULTS: LPS induced IL-8, COX-2, TLR-4 mRNA expression, and IL-8 protein secretion in HT-29 cells. Treatment with LPS and L. casei in comparison with LPS stimulation alone lowered IL-8, COX-2, TLR-4 mRNA expression, and IL-8 protein secretion. L. casei increased PPARgamma mRNA expression in dose-dependent manner. L. casei activated PPRE in HT-29 cells transfected with PPRE3-tk-luciferase construct. CONCLUSIONS: Probiotics, L. casei, suppresses the expression of inflammatory mediators in intestinal epithelial cells. The anti-inflammatory action of L. casei might be partially related to PPARgamma activation.