Over-activation of TLR5 signaling by high-dose flagellin induces liver injury in mice

Yang, Xiao; Liu, Fang; Yang, Jingyi; Zhong, Ma; Zhang, Ejuan; Li, Yaoming; Zhou, Demin; Cao, Yuan; Li, Wei; Yu, Jie; Yang, Yi; Yan, Huimin

doi:10.1038/cmi.2014.110

Cited by 68 publications

(73 citation statements)

References 49 publications

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“…27 Briefly, spleens were obtained from 6-to 8-week-old female C57BL/6 mice, smashed using syringe pistons in phosphate-buffered saline Stimulating macrophages with flagellin It had been reported that LPS pretreatment of macrophages allowed a robust IL-1b released. So, macrophages were pretreated with 50 ng/ml LPS for 3 h, and then stimulated with flagellin using or no using DOTAP transfection (Roche Diagnostics, BASEL, Switzerland).…”

Section: Animalsmentioning

confidence: 99%

Activation of NLRC4 downregulates TLR5-mediated antibody immune responses against flagellin

Yang

Zhang

et al. 2015

Cell Mol Immunol

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Bacterial flagellin is a unique pathogen-associated molecular pattern (PAMP), which can be recognized by surface localized Toll-like receptor 5 (TLR5) and the cytosolic NOD-like receptor (NLR) protein 4 (NLRC4) receptors. Activation of the TLR5 and/or NLRC4 signaling pathways by flagellin and the resulting immune responses play important roles in anti-bacterial immunity. However, it remains unclear how the dual activities of flagellin that activate the TLR5 and/or NLRC4 signaling pathways orchestrate the immune responses. In this study, we assessed the effects of flagellin and its mutants lacking the ability to activate TLR5 and NLRC4 alone or in combination on the adaptive immune responses against flagellin. Flagellin that was unable to activate NLRC4 induced a significantly higher antibody response than did wild-type flagellin. The increased antibody response could be eliminated when macrophages were depleted in vivo. The activation of NLRC4 by flagellin downregulated the flagellin-induced and TLR5-mediated immune responses against flagellin. Cellular & Molecular ImmunologyKeywords: flagellin; NLRC4; TLR5; macrophage; adaptive immunity INTRODUCTION Bacterial flagellin is one of a small number of protein pathogenassociated molecular patterns (PAMPs), which can be recognized by cell surface Toll-like receptor 5 (TLR5) 1 and the cytosolic NOD-like receptor protein 4 (NLRC4) inflammasome receptor NAIP5.2,3 Flagellin consists of two highly conserved domains (D0 and D1) and one central hypervariable domain (D2/D3). [4][5][6] The conserved D0 and D1 domains are required for the immune activity of flagellin as a PAMP. The Nterminal amino acids 90-97 (QRVRELAV) of D1 form a highly conserved motif that is essential for both high-affinity binding and signaling to TLR5. 7,8 The C-terminal 35 amino acids of flagellin in the D0 domain are responsible for the interaction between flagellin and NLRC4. The substitution of three conserved leucine residues L502, L504, and L505 for alanine in the 35 C-terminal amino acids could abolish the ability of flagellin to activate NLRC4.9 Flagellin-mediated activation of TLR5 activates inflammatory genes via the MyD88 pathway, whereas flagellin-activated NLRC4 triggers the assembly of the inflammasome, which culminates in caspase-1 activation, IL-1b/IL-18 secretion, and cellular pyroptosis.

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Section: Animalsmentioning

confidence: 99%

Activation of NLRC4 downregulates TLR5-mediated antibody immune responses against flagellin

Yang

Zhang

et al. 2015

Cell Mol Immunol

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“…Additionally, Neely et al showed that administering a single low dose of flagellin in a murine model of burn injury and infection restored neutrophil response increasing bacterial clearance [12]. In contrast, others studies have shown that flagellin can induce pathophysiological inflammation that causes tissue damage [5, 9, 10, 13], a condition we have termed as flagellemia. Neutralization of flagellin can attenuate systemic inflammatory responses [14].…”

Section: Introductionmentioning

confidence: 99%

“…Neutralization of flagellin can attenuate systemic inflammatory responses [14]. We, as well as others, have reported that flagellin is an active contributor to pathophysiological responses such as sepsis [5, 9, 10, 12, 15, 16], liver injury [13], pulmonary inflammation and cardiovascular damage [9, 16, 17, 18]. We recently reported that luminal flagellin is internalized by human IECs [6], followed by dissemination into the systemic circulation under various pathophysiological conditions including burn injury [19, 20] and inflammatory bowel disease (IBD) [21–24].…”

Section: Introductionmentioning

confidence: 99%

Intraluminal Flagellin Differentially Contributes to Gut Dysbiosis and Systemic Inflammation following Burn Injury

et al. 2016

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Burn injury is associated with a loss of gut barrier function, resulting in systemic dissemination of gut-derived bacteria and their products. The bacterial protein and TLR5 agonist, flagellin, induces non-specific innate immune responses. Because we detected flagellin in the serum of burn patients, we investigated whether gut-derived flagellin was a primary or secondary contributor to intestinal dysfunction and systemic inflammation following burn injury. The apical surface of polarized human intestinal epithelial cells (IECs), Caco-2BBe, were exposed to 50 or 500 ng of purified flagellin and 1 x 105 of an intestinal E. coli (EC) isolate as follows: 1) flagellin added 30 min prior to EC, 2) flagellin and EC added simultaneously, or 3) EC added 30 min prior to flagellin. Our results showed that luminal flagellin and EC modulated each other's biological actions, which influenced their ability to induce basolateral secretion of inflammatory cytokines and subsequent translocation of bacteria and their products. A low dose of flagellin accompanied by an enteric EC in the lumen, tempered inflammation in a dose- and time-dependent manner. However, higher doses of flagellin acted synergistically with EC to induce both intestinal and systemic inflammation that compromised barrier integrity, increasing systemic inflammation following burn injury, a process we have termed flagellemia. In a murine model of burn injury we found that oral gavage of flagellin (1 μg/mouse) significantly affected the gut microbiome after burn injury. In these mice, flagellin disseminated out of the intestine into the serum and to distal organs (mesenteric lymph nodes and lungs) where it induced secretion of monocyte chemoattractant protein (MCP-1) and CXCL1/KC (mouse equivalent of human IL-8) at 24 and 48h post-burn. Our results illustrated that gut-derived flagellin alone or accompanied by a non-pathogenic enteric EC strain can function as an initiator of luminal and systemic inflammation following burn injury.

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“…27 Briefly, spleens were obtained from 6-to 8-week-old female C57BL/6 mice, smashed using syringe pistons in phosphate-buffered saline NLRC4 activating reduces the immune responses against FliC W Li et al…”

Section: Animalsmentioning

confidence: 99%

“…So, macrophages were pretreated with 50 ng/ml LPS for 3 h, and then stimulated with flagellin using or no using DOTAP transfection (Roche Diagnostics, BASEL, Switzerland). 27,29 Briefly, 15 ml of DOTAP liposomal transfection reagent was incubated with 5 mg of purified flagellin for 20 min in serum-free media, and then 1 ml of RPMI 1640 medium was added. For no DOTAP transfection, 100 nM flagellin was added to macrophages directly in a 96-well plate, and supernatants were collected after 6 h to measure IL-1b and IL-18 levels.…”

Section: Stimulating Macrophages With Flagellinmentioning

confidence: 99%

Activation of NLRC4 downregulates TLR5-mediated antibody immune responses against flagellin

Li¹,

Yang²,

Zhang³

et al. 2015

Cell Mol Immunol

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Bacterial flagellin is a unique pathogen-associated molecular pattern (PAMP), which can be recognized by surface localized Toll-like receptor 5 (TLR5) and the cytosolic NOD-like receptor (NLR) protein 4 (NLRC4) receptors. Activation of the TLR5 and/or NLRC4 signaling pathways by flagellin and the resulting immune responses play important roles in anti-bacterial immunity. However, it remains unclear how the dual activities of flagellin that activate the TLR5 and/or NLRC4 signaling pathways orchestrate the immune responses. In this study, we assessed the effects of flagellin and its mutants lacking the ability to activate TLR5 and NLRC4 alone or in combination on the adaptive immune responses against flagellin. Flagellin that was unable to activate NLRC4 induced a significantly higher antibody response than did wild-type flagellin. The increased antibody response could be eliminated when macrophages were depleted in vivo. The activation of NLRC4 by flagellin downregulated the flagellin-induced and TLR5-mediated immune responses against flagellin. Cellular & Molecular ImmunologyKeywords: flagellin; NLRC4; TLR5; macrophage; adaptive immunity INTRODUCTION Bacterial flagellin is one of a small number of protein pathogenassociated molecular patterns (PAMPs), which can be recognized by cell surface Toll-like receptor 5 (TLR5) 1 and the cytosolic NOD-like receptor protein 4 (NLRC4) inflammasome receptor NAIP5. 2,3 Flagellin consists of two highly conserved domains (D0 and D1) and one central hypervariable domain (D2/D3). [4][5][6] The conserved D0 and D1 domains are required for the immune activity of flagellin as a PAMP. The Nterminal amino acids 90-97 (QRVRELAV) of D1 form a highly conserved motif that is essential for both high-affinity binding and signaling to TLR5. 7,8 The C-terminal 35 amino acids of flagellin in the D0 domain are responsible for the interaction between flagellin and NLRC4. The substitution of three conserved leucine residues L502, L504, and L505 for alanine in the 35 C-terminal amino acids could abolish the ability of flagellin to activate NLRC4. 9 Flagellin-mediated activation of TLR5 activates inflammatory genes via the MyD88 pathway, whereas flagellin-activated NLRC4 triggers the assembly of the inflammasome, which culminates in caspase-1 activation, IL-1b/IL-18 secretion, and cellular pyroptosis. 10,11

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Over-activation of TLR5 signaling by high-dose flagellin induces liver injury in mice

Cited by 68 publications

References 49 publications

Activation of NLRC4 downregulates TLR5-mediated antibody immune responses against flagellin

Activation of NLRC4 downregulates TLR5-mediated antibody immune responses against flagellin

Intraluminal Flagellin Differentially Contributes to Gut Dysbiosis and Systemic Inflammation following Burn Injury

Activation of NLRC4 downregulates TLR5-mediated antibody immune responses against flagellin

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