scholarly journals Lactobacillus rhamnosus GG attenuates interferon-γ and tumour necrosis factor-α-induced barrier dysfunction and pro-inflammatory signalling

Microbiology ◽  
2010 ◽  
Vol 156 (11) ◽  
pp. 3288-3297 ◽  
Author(s):  
Kevin A. Donato ◽  
Mélanie G. Gareau ◽  
Yu Jing Jenny Wang ◽  
Philip M. Sherman
Keyword(s):  
Epithelial Cells ◽  
Inflammatory Cytokines ◽  
Time Course ◽  
Nuclear Translocation ◽  
Inflammatory Responses ◽  
Cell Signalling ◽  
Lactobacillus Rhamnosus ◽  
Basal Medium ◽  
Barrier Dysfunction ◽  
Pro Inflammatory Cytokines

The intestinal epithelium forms a protective barrier against luminal contents and the external environment, mediated via intercellular tight junctions (TJs). The TJ can be disrupted via cell signalling induced by either enteric pathogens or pro-inflammatory cytokines, thereby contributing to various intestinal disorders ranging from acute infectious diarrhoea to chronic inflammatory bowel diseases. Probiotics, such as Lactobacillus rhamnosus GG (LGG), are reported to confer beneficial effects on epithelial cells, including antagonizing infections and reducing overt pro-inflammatory responses, but the underlying mechanisms of these observed effects require further characterization. We hypothesized that probiotics preserve barrier function by interfering with pro-inflammatory cytokine signalling. Caco-2bbe cells were seeded into Transwells to attain polarized monolayers with intercellular TJs. Monolayers were inoculated apically with the probiotic LGG 3 h prior to the addition of IFN-γ (100 ng ml−1) to the basolateral medium overnight. The monolayers were then placed in fresh basal medium±TNF-α (10 ng ml−1) and transepithelial electrical resistance (TER) measurements were taken over the time-course of TNF-α stimulation. To complement the TER findings, cells were processed for zona occludens-1 (ZO-1) immunofluorescence staining. As a measure of TNF-α downstream signalling, cells were immunofluorescently stained for NF-κB p65 subunit and CXCL-8 mRNA was quantified by qRT-PCR. Basal cell culture medium was collected after overnight TNF-α stimulation to measure secreted chemokines, including CXCL-8 (interleukin-8) and CCL-11 (eotaxin). Following LGG inoculation, IFN-γ priming and 24 h TNF-α stimulation, epithelial cells maintained TER and ZO-1 distribution. LGG diminished the nuclear translocation of p65, demonstrated by both immunofluorescence and CXCL-8 mRNA expression. CXCL-8 and CCL-11 protein levels were decreased in LGG-inoculated, cytokine-challenged cells. These findings indicate that LGG alleviates the effects of pro-inflammatory cytokines on epithelial barrier integrity and inflammation, mediated, at least in part, through inhibition of NF-κB signalling.

scholarly journals Cystine reduces tight junction permeability and intestinal inflammation induced by oxidative stress in Caco-2 cells

Amino Acids ◽  
2021 ◽  
Author(s):  
Tatsuya Hasegawa ◽  
Ami Mizugaki ◽  
Yoshiko Inoue ◽  
Hiroyuki Kato ◽  
Hitoshi Murakami
Keyword(s):  
Oxidative Stress ◽  
Tight Junction ◽  
Mrna Expression ◽  
Inflammatory Cytokines ◽  
Inflammatory Responses ◽  
Intestinal Barrier ◽  
Barrier Dysfunction ◽  
Whole Cells ◽  
Intestinal Barrier Dysfunction ◽  
Pro Inflammatory Cytokines

AbstractIntestinal oxidative stress produces pro-inflammatory cytokines, which increase tight junction (TJ) permeability, leading to intestinal and systemic inflammation. Cystine (Cys2) is a substrate of glutathione (GSH) and inhibits inflammation, however, it is unclear whether Cys2 locally improves intestinal barrier dysfunction. Thus, we investigated the local effects of Cys2 on oxidative stress-induced TJ permeability and intestinal inflammatory responses. Caco-2 cells were cultured in a Cys2-supplemented medium for 24 h and then treated with H2O2 for 2 h. We assessed TJ permeability by measuring transepithelial electrical resistance and the paracellular flux of fluorescein isothiocyanate–dextran 4 kDa. We measured the concentration of Cys2 and GSH after Cys2 pretreatment. The mRNA expression of pro-inflammatory cytokines was assessed. In addition, the levels of TJ proteins were assessed by measuring the expression of TJ proteins in the whole cells and the ratio of TJ proteins in the detergent-insoluble fractions to soluble fractions (IS/S ratio). Cys2 treatment reduced H2O2-induced TJ permeability. Cys2 did not change the expression of TJ proteins in the whole cells, however, suppressed the IS/S ratio of claudin-4. Intercellular levels of Cys2 and GSH significantly increased in cells treated with Cys2. Cys2 treatment suppressed the mRNA expression of pro-inflammatory cytokines, and the mRNA levels were significantly correlated with TJ permeability. In conclusion, Cys2 treatment locally reduced oxidative stress-induced intestinal barrier dysfunction possively due to the mitigation of claudin-4 dislocalization. Furthermore, the effect of Cys2 on the improvement of intestinal barrier function is related to the local suppression of oxidative stress-induced pro-inflammatory responses.

scholarly journals MiR-200c-3p inhibits LPS-induced M1 polarization of BV2 cells by targeting RIP2

2022 ◽  
Author(s):  
Lei Zhao ◽  
Xiaosong Liu ◽  
Jiankai Yang ◽  
Xiaoliang Wang ◽  
Xiaomeng Liu ◽  
...  
Keyword(s):  
Inflammatory Cytokines ◽  
Nuclear Translocation ◽  
Inflammatory Responses ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
M1 Polarization ◽  
Bv2 Cells ◽  
M1 Phenotype ◽  
Lps Treatment ◽  
Pro Inflammatory Cytokines

Abstract Background Microglia are important immune cells, which can be induced by lipopolysaccharide (LPS) into M1 phenotype that express pro-inflammatory cytokines. Some studies have shown that microRNAs play critical roles in microglial activation. Objective This study was designed to investigate the role of miR-200c-3p in regulating inflammatory responses of LPS-treated BV2 cells. Methods The expression of miR-200c-3p in BV2 cells was detected by real-time PCR. Receptor-interacting protein 2 (RIP2) was predicted as a target gene of miR-200c-3p. Their relationship was verified by dual-luciferase reporter assay. The function of miR-200c-3p and RIP2 in microglial polarization and NF-κB signaling was further evaluated. Results LPS treatment reduced miR-200c-3p expression in a dose-dependent and time-dependent manner in BV2 cells. LPS treatment increased the expression of M1 phenotype markers inducible nitric oxide synthase (iNOS) and major histocompatibility complex class (MHC)-II, promoted the release of pro-inflammatory cytokines interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α, and enhanced the nuclear translocation and phosphorylation of nuclear factor-kappaB (NF-κB) p65. Reversely, miR-200c-3p mimics down-regulated the levels of these inflammatory factors. Furthermore, RIP2 was identified to be a direct target of miR-200c-3p. RIP2 knockdown had a similar effect to miR-200c-3p mimics. Overexpression of RIP2 eliminated the inhibitory effect of miR-200c-3p on LPS-induced M1 polarization and NF-κB activation in BV2 cells. Conclusions MiR-200c-3p mimics suppressed LPS-induced microglial M1 polarization and NF-κB activation by targeting RIP2. MiR-200c-3p/RIP2 might be a potential therapeutic target for the treatment of neuroinflammation-associated diseases.

scholarly journals Oral feeding with probiotic Lactobacillus rhamnosus attenuates cigarette smoke-induced COPD in C57Bl/6 mice: Relevance to inflammatory markers in human bronchial epithelial cells

2019 ◽  
Author(s):  
J.L. Carvalho ◽  
M. Miranda ◽  
A.K. Fialho ◽  
H. Castro-Faria-Neto ◽  
E. Anatriello ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cigarette Smoke ◽  
Inflammatory Cytokines ◽  
Lactobacillus Rhamnosus ◽  
Bronchial Epithelial Cells ◽  
Human Bronchial Epithelial Cells ◽  
Bronchial Epithelial ◽  
Cytokines And Chemokines ◽  
Anti Inflammatory ◽  
Pro Inflammatory Cytokines

AbstractCOPD is a prevalent lung disease with significant impacts on public health. Affected airways exhibit pulmonary neutrophilia and consequent secretion of pro-inflammatory cytokines and proteases, which result in lung emphysema. Probiotics act as nonspecific modulators of the innate immune system that improve several inflammatory responses. To investigate the effect of Lactobacillus rhamnosus (Lr) on cigarette smoke (CS)-induced COPD C57Bl/6 mice were treated with Lr during the week before COPD induction and three times/week until euthanasia. For in vitro assays, murine bronchial epithelial cells as well as human bronchial epithelial cells exposed to cigarette smoke extract during 24 hours were treated with Lr 1 hour before CSE addition. Lr treatment attenuated the inflammatory response both in the airways and lung parenchyma, reducing neutrophilic infiltration and the production of pro-inflammatory cytokines and chemokines. Also, Lr-treated mice presented with lower metalloproteases in lung tissue and lung remodeling. In parallel to the reduction in the expression of TLR2, TLR4, TLR9, STAT3, and NF-κB in lung tissue, Lr increased the levels of IL-10 as well as SOCS3 and TIMP1/2, indicating the induction of an anti-inflammatory environment. Similarly, murine bronchial epithelial cells as well as human bronchial epithelial cells (BEAS) exposed to CSE produced pro-inflammatory cytokines and chemokines, which were inhibited by Lr treatment in association with the production of anti-inflammatory molecules. Moreover, the presence of Lr also modulated the expression of COPD-associated transcription found into BALF of COPD mice group, i.e., Lr downregulated expression of NF-κB and STAT3, and inversely upregulated increased expression of SOCS3. Thus, our findings indicate that Lr modulates the balance between pro- and anti-inflammatory cytokines in human bronchial epithelial cells upon CS exposure and it can be a useful tool to improve the lung inflammatory response associated with COPD.

scholarly journals Mitochondrial DNA leakage induces odontoblast inflammation via the cGAS-STING pathway

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Lu Zhou ◽  
Yi-Fei Zhang ◽  
Fu-Hua Yang ◽  
Han-Qing Mao ◽  
Zhi Chen ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Real Time ◽  
Inflammatory Cytokines ◽  
Real Time Pcr ◽  
Nuclear Translocation ◽  
Inflammatory Responses ◽  
Immunofluorescent Staining ◽  
Bacterial Invasion ◽  
Sting Pathway ◽  
Pro Inflammatory Cytokines

Abstract Background Mitochondrial DNA (mtDNA) is a vital driver of inflammation when it leaks from damaged mitochondria into the cytosol. mtDNA stress may contribute to cyclic GMP-AMP synthase (cGAS) stimulator of interferon genes (STING) pathway activation in infectious diseases. Odontoblasts are the first cells challenged by cariogenic bacteria and involved in maintenance of the pulp immune and inflammatory responses to dentine-invading pathogens. In this study, we investigated that mtDNA as an important inflammatory driver participated in defending against bacterial invasion via cGAS-STING pathway in odontoblasts. Methods The normal tissues, caries tissues and pulpitis tissues were measured by western blotting and immunohistochemical staining. Pulpitis model was built in vitro to evaluated the effect of the cGAS-STING pathway in odontoblast-like cell line (mDPC6T) under inflammation. Western blot and real-time PCR were performed to detect the expression of cGAS-STING pathway and pro-inflammatory cytokines. The mitochondrial function was evaluated reactive oxygen species (ROS) generated by mitochondria using MitoSOX Red dye staining. Cytosolic DNA was assessed by immunofluorescent staining and real-time PCR in mDPC6T cells after LPS stimulation. Furthermore, mDPC6T cells were treated with ethidium bromide (EtBr) to deplete mtDNA or transfected with isolated mtDNA. The expression of cGAS-STING pathway and pro-inflammatory cytokines were measured. Results The high expression of cGAS and STING in caries and pulpitis tissues in patients, which was associated with inflammatory progression. The cGAS-STING pathway was activated in inflamed mDPC6T. STING knockdown inhibited the nuclear import of p65 and IRF3 and restricted the secretion of the inflammatory cytokines CXCL10 and IL-6 induced by LPS. LPS caused mitochondrial damage in mDPC6T, which promoted mtDNA leakage into the cytosol. Depletion of mtDNA inhibited the cGAS-STING pathway and nuclear translocation of p65 and IRF3. Moreover, repletion of mtDNA rescued the inflammatory response, which was inhibited by STING knockdown. Conclusion Our study systematically identified a novel mechanism of LPS-induced odontoblast inflammation, which involved mtDNA leakage from damaged mitochondria into the cytosol stimulating the cGAS-STING pathway and the inflammatory cytokines IL-6 and CXCL10 secretion. The mtDNA-cGAS-STING axis could be a potent therapeutic target to prevent severe bacterial inflammation in pulpitis. Graphic abstract

Suppressive Effects of Britanin, a Sesquiterpene Compound Isolated from Inulae Flos, on Mast Cell-Mediated Inflammatory Responses

2014 ◽  
Vol 42 (04) ◽  
pp. 935-947 ◽  
Author(s):  
Hyo-Hyun Park ◽  
Sun-Gun Kim ◽  
Young Na Park ◽  
Jiean Lee ◽  
Youn Ju Lee ◽  
...  
Keyword(s):  
Mast Cell ◽  
Inflammatory Cytokines ◽  
Nuclear Translocation ◽  
Inflammatory Responses ◽  
Human Mast Cell ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Vitro Assay ◽  
Dose Dependent ◽  
Pro Inflammatory Cytokines

Mast cells are central players in immediate-type hypersensitvity and inflammatory responses. In the present study, the effects of britanin on the passive cutaneous anaphylaxis (PCA) reaction in mice and on the phorbol 12-myristate 13-acetate and calcium ionophore A23187 (PMACI)-induced production of pro-inflammatory cytokines in human mast cell line (HMC-1) were evaluated. The oral administration of britanin (10–20 mg/kg) decreased the mast cell-mediated PCA reaction in IgE-sensitized mice. In the activity and mechanism of britanin in vitro assay, britanin suppressed the gene expression and secretion of pro-inflammatory cytokines in a dose-dependent manner in HMC-1. In addition, britanin attenuated PMACI-induced activation of NF-κB as indicated by the inhibition of the degradation of IκBα, nuclear translocation of NF-κB, NF-κB/DNA binding activity assay, and blocked the phosphorylation of p38 MAP kinase, in a dose-dependent manner. We conclude that britanin may have potential as a treatment for allergic-inflammatory diseases.

scholarly journals Heat-killed probiotic bacteria differentially regulate colonic epithelial cell production of human β-defensin-2: dependence on inflammatory cytokines

2014 ◽  
Vol 5 (4) ◽  
pp. 483-495 ◽  
Author(s):  
N. Habil ◽  
W. Abate ◽  
J. Beal ◽  
A.D. Foey
Keyword(s):  
Epithelial Cells ◽  
Inflammatory Cytokines ◽  
Inflammatory Responses ◽  
Quantitative Polymerase Chain Reaction ◽  
Probiotic Bacteria ◽  
Enzyme Linked Immunosorbent Assay ◽  
Necrosis Factor Alpha ◽  
Tnf Α ◽  
Probiotic Strains ◽  
Pro Inflammatory Cytokines

The inducible antimicrobial peptide human β-defensin-2 (hBD-2) stimulated by pro-inflammatory cytokines and bacterial products is essential to antipathogen responses of gut epithelial cells. Commensal and probiotic bacteria can augment such mucosal defences. Probiotic use in the treatment of inflammatory bowel disease, however, may have adverse effects, boosting inflammatory responses. The aim of this investigation was to determine the effect of selected probiotic strains on hBD-2 production by epithelial cells induced by pathologically relevant pro-inflammatory cytokines and the role of cytokine modulators in controlling hBD-2. Caco-2 colonic intestinal epithelial cells were pre-incubated with heat-killed probiotics, i.e. Lactobacillus casei strain Shirota (LcS) or Lactobacillus fermentum strain MS15 (LF), followed by stimulation of hBD-2 by interleukin (IL)-1β and tumour necrosis factor alpha (TNF-α) in the absence or presence of exogenous IL-10 or anti-IL-10 neutralising antibody. Cytokines and hBD-2 mRNA and protein were analysed by real-time quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. LcS augmented IL-1β-induced hBD-2, whereas LF enhanced TNF-α- and suppressed IL-1β-induced hBD-2. LF enhanced TNF-α-induced TNF-α and suppressed IL-10, whereas augmented IL-1β-induced IL-10. LcS upregulated IL-1β-induced TNF-α mRNA and suppressed IL-10. Endogenous IL-10 differentially regulated hBD-2; neutralisation of IL-10 augmented TNF-α- and suppressed IL-1β-induced hBD-2. Exogenous IL-10, however, suppressed both TNF-α- and IL-1β-induced hBD-2; LcS partially rescued suppression in TNF-α- and IL-1β-stimulation, whereas LF further suppressed IL-1β-induced hBD-2. It can be concluded that probiotic strains differentially regulate hBD-2 mRNA expression and protein secretion, modulation being dictated by inflammatory stimulus and resulting cytokine environment.

scholarly journals Punicalagin Prevents Inflammation in LPS-Induced RAW264.7 Macrophages by Inhibiting FoxO3a/Autophagy Signaling Pathway

Nutrients ◽  
2019 ◽  
Vol 11 (11) ◽  
pp. 2794 ◽  
Author(s):  
Cao ◽  
Chen ◽  
Ren ◽  
Zhang ◽  
Tan ◽  
...  
Keyword(s):  
Western Blot ◽  
Signaling Pathway ◽  
Inflammatory Cytokines ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Inflammatory Responses ◽  
Raw264.7 Macrophages ◽  
Tnf Α ◽  
Autophagy Inhibition ◽  
Pro Inflammatory Cytokines

Punicalagin, a hydrolysable tannin of pomegranate juice, exhibits multiple biological effects, including inhibiting production of pro-inflammatory cytokines in macrophages. Autophagy, an intracellular self-digestion process, has been recently shown to regulate inflammatory responses. In this study, we investigated the anti-inflammatory potential of punicalagin in lipopolysaccharide (LPS) induced RAW264.7 macrophages and uncovered the underlying mechanisms. Punicalagin significantly attenuated, in a concentration-dependent manner, LPS-induced release of NO and decreased pro-inflammatory cytokines TNF-α and IL-6 release at the highest concentration. We found that punicalagin inhibited NF-κB and MAPK activation in LPS-induced RAW264.7 macrophages. Western blot analysis revealed that punicalagin pre-treatment enhanced LC3II, p62 expression, and decreased Beclin1 expression in LPS-induced macrophages. MDC assays were used to determine the autophagic process and the results worked in concert with Western blot analysis. In addition, our observations indicated that LPS-induced releases of NO, TNF-α, and IL-6 were attenuated by treatment with autophagy inhibitor chloroquine, suggesting that autophagy inhibition participated in anti-inflammatory effect. We also found that punicalagin downregulated FoxO3a expression, resulting in autophagy inhibition. Overall these results suggested that punicalagin played an important role in the attenuation of LPS-induced inflammatory responses in RAW264.7 macrophages and that the mechanisms involved downregulation of the FoxO3a/autophagy signaling pathway.

scholarly journals Fucoxanthin Ameliorates Oxidative Stress and Airway Inflammation in Tracheal Epithelial Cells and Asthmatic Mice

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1311
Author(s):  
Shu-Ju Wu ◽  
Chian-Jiun Liou ◽  
Ya-Ling Chen ◽  
Shu-Chen Cheng ◽  
Wen-Chung Huang
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Airway Inflammation ◽  
Inflammatory Cytokines ◽  
Inflammatory Responses ◽  
Therapeutic Potential ◽  
Eosinophil Infiltration ◽  
Tracheal Epithelial Cells ◽  
Tracheal Epithelial ◽  
And Oxidative Stress

Fucoxanthin is isolated from brown algae and was previously reported to have multiple pharmacological effects, including anti-tumor and anti-obesity effects in mice. Fucoxanthin also decreases the levels of inflammatory cytokines in the bronchoalveolar lavage fluid (BALF) of asthmatic mice. The purpose of the present study was to investigate the effects of fucoxanthin on the oxidative and inflammatory responses in inflammatory human tracheal epithelial BEAS-2B cells and attenuated airway hyperresponsiveness (AHR), airway inflammation, and oxidative stress in asthmatic mice. Fucoxanthin significantly decreased monocyte cell adherence to BEAS-2B cells. In addition, fucoxanthin inhibited the production of pro-inflammatory cytokines, eotaxin, and reactive oxygen species in BEAS-2B cells. Ovalbumin (OVA)-sensitized mice were treated by intraperitoneal injections of fucoxanthin (10 mg/kg or 30 mg/kg), which significantly alleviated AHR, goblet cell hyperplasia and eosinophil infiltration in the lungs, and decreased Th2 cytokine production in the BALF. Furthermore, fucoxanthin significantly increased glutathione and superoxide dismutase levels and reduced malondialdehyde (MDA) levels in the lungs of asthmatic mice. These data demonstrate that fucoxanthin attenuates inflammation and oxidative stress in inflammatory tracheal epithelial cells and improves the pathological changes related to asthma in mice. Thus, fucoxanthin has therapeutic potential for improving asthma.

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