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

Cellular & Molecular Immunology 2015年第6期12卷 729-742页

作者： Yang Xiao Fang Liu Jingyi Yang Maohua Zhong Ejuan Zhang Yaoming Li Dihan Zhou Yuan Cao Wei Li Jie Yu Yi Yang Huimin Yan Mucosal immunity Research Group State Key Laboratory of Virology Wuhan institute of Virology Chinese Academy of Sciences Wuhan China

flagellin is a potent activator of a broad range of cell types that are involved in innate and adaptive immunity. Therefore, it is a good adjuvant candidate for vaccines, and it might function as a biological protectant against both major acute radiation syndrome during cancer radiotherapy and a mitigator of radiation emergencies. However, accumulating evidence has implicated flagellin in the occurrence of some inflammatory diseases, such as acute lung inflammation, cardiovascular collapse and inflammatory bowel disease. The aim of this study was to elucidate whether only flagellin-TLR5 signaling activation plays a role in the pathophysiology of liver or whether some other flagellin activity also contributes to liver injury either via bacterial infections or during clinical applications. Recombinant flagellin proteins with or without TLR5-stimulating activity were used to evaluate the role of flagellin-TLR5 signaling in liver injury in wild-type and TLR5 KO mice. Gross lesions and large areas of hepatocellular necrosis were observed in liver tissue 12 h after the intraperitoneal administration of 100 or 200 pg flagellin （FliC） in a dose-and time-dependent manner in wild-type mice, but not in TLR5 KO mice. Deletion of the N-terminal or TLR5 binding domain of flagellin inhibited flagellin-induced inflammatory responses and the subsequent acute liver function abnormality and damage. These data confirmed that flagellin is an essential determinant of liver injury and demonstrated that the over-activation of TLR5 signaling by high-dose flagellin caused acute inflammatory responses, neutrophil accumulation and oxidative stress in the liver, which contributes to the progression and severity of flagellin-induced liver injury.

关键词： flagellin liver injury pathological effect TLR5

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Salmonella flagella confer anti-tumor immunological effect via activating flagellin/TLR5 signalling within tumor microenvironment

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Acta Pharmaceutica Sinica B 2021年第10期11卷 3165-3177页

作者： Jianxiang Chen Yiting Qiao Guo Chen Cunjie Chang Heng Dong Bo Tang Xiawei Cheng Xiufeng Liu Zichun Hua School of Life Sciences Nanjing UniversityNanjing 210023China Changzhou High-Tech Research Institute of Nanjing University and Jiangsu Target Pharma Laboratories Inc. Changzhou 213164China College of Pharmacy School of MedicineHangzhou Normal UniversityHangzhou 311100China Department of Hepatology the Affiliated Hospital of Hangzhou Normal UniversityHangzhou Normal UniversityHangzhou 310003China Division of Hepatobiliary and Pancreatic Surgery Department of Surgerythe First Affiliated HospitalSchool of MedicineZhejiang University School of Biopharmacy China Pharmaceutical UniversityNanjing 211198China

Salmonella:mediated cancer therapy has achieved remarkable anti-tumor effects in experimental animal models,but the detailed mechanism remains *** this report,the active involvement of the host immune response in this process was confirmed by comparing the tumor-suppressive effects of Salmonella in immunocompetent and immunodeficient mice bearing melanoma *** flagella are key inducers of the host immune response during bacterial infection,flagella were genetically disrupted to analyse their involvement in Salmonella-mediated cancer *** results showed that flagellum-deficient strains failed to induce significant anti-tumor effects,even when more bacteria were administered to offset the difference in invasion *** mainly activate immune cells via flagellin/Toll-like receptor 5(TLR5)signalling ***,we showed that exogenous activation of TLR5 signalling by recombinant flagellin and exogenous expression of TLR5 both enhanced the therapeutic efficacy of flagellum-deficient Salmonella against *** study highlighted the therapeutic value of the interaction between Salmonella and the host immune response through flagellin/TLR5 signalling pathway during Salmonella-mediated cancer therapy,thereby suggesting the potential application of TLR5 agonists in the cancer immune therapy.

关键词： Bacteria-mediated cancer therapy Salmonella VNP20009 Flagellum flagellin TLR5 NF-kB Cancer immune therapy

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Salmonella enterica flagellin Is Recognized via FLS2 and Activates PAMP-Triggered Immunity in Arabidopsis thaliana

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Molecular Plant 2014年第4期7卷 657-674页

作者： Ana Victoria Garcia Amelie Charrier Adam Schikora Jean Bigeard Stephanie Pateyron Marie-Ludivine de Tauzia-Moreau Alexandre Evrard Axel Mithofer Marie Laure Martin-Magniette Isabelle Virlogeux-Payant Heribert Hirt Unite de Recherche en Genomique Vegetale (URGV) UMR INRA/CNRS/Universite d'Evry Val d'Essonne 91057 Evry France Institute for Plant Pathology and Applied Zoology Research Centre for BioSystems Land Use and Nutrition JL University Giessen Heinrich-Buff-Ring 26-32 35392 Giessen Germany Transcriptomic Platform Unite de Recherche en Genomique Vegetale (URGV) 91057 Evry France Department of Bioorganic Chemistry Max Planck Institute for Chemical Ecology Hans-Knoll-Str. 8 07745 Jena Germany INRA UMR 518 MIA 75005 Paris France AgroParisTech UMR 518 MIA 75005 Paris France INRA UMR 1282 infectiologie et Sante Publique 37380 Nouzilly France

Infections with Salmonella enterica belong to the most prominent causes of food poisoning and infected fruits and vegetables represent important vectors for salmonellosis. Recent evidence indicates that plants recognize S. enterica and raise defense responses. Nonetheless, the molecular mechanisms controlling the interaction of S. enterica with plants are still largely unclear. Here, we show that flagellin from S. enterica represents a prominent pathogenassociated molecular pattern （PAMP） in Arabidopsis thaliana, which induces PAMP-triggered immunity （PTI） via the recognition of the fig22 domain by the receptor kinase FLS2. The Arabidopsis fls2 mutant shows reduced though not abolished PTI activation, indicating that plants rely also on recognition of other S. enterica PAMPs. Interestingly, the S. enterica type III secretion system （T3SS） mutant prgH- induced stronger defense gene expression than wild-type bacteria in Arabidopsis, suggesting that T3SS effectors are involved in defense suppression. Furthermore, we observe that S. enterica strains show variation in the fig22 epitope, which results in proteins with reduced PTI-inducing activity. Altogether, these results show that S. enterica activates PTI in Arabidopsis and suggest that, in order to accomplish plant colonization, S. enterica evolved strategies to avoid or suppress PTI.

关键词： Salmonella enterica Arabidopsis thaliana flagellin FLS2 immunity.

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flagellin of Pseudomonas aeruginosa induces transforming growth factor beta 1 expression in normal bronchial epithelial cells through mitogen activated protein kinase cascades

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Chinese Medical Journal 2011年第4期124卷 599-605页

作者： YANG Jing-jing WANG Dan-dan SUN Tie-ying Department of Respiratory Diseases Peking University Fifth School of Clinical Medicine Beijing Hospital Beijing 100730China

Background Acute lung infection due to Pseudomonas aeruginosa （P. Aeruginosa） is a serious problem, especially in patients with structural lung conditions or immune compromised hosts, leading to an overwhelming threat with a high risk of morbidity and mortality. As an outcome of infection, fibrosis can be linked with chronic lung diseases. But some fibrotic manifestations, such as an irreversible decrease of lung function and fibrous bands seen on chest imaging, have been found after an acute infection with P. Aeruginosa. Fibrogenesis/remodeling resulting from acute lung infection by *** is rarely reported. This study was designed to explore the relation between fibrogenesis/remodeling and acute infection by P. Aeruginosa in vitro. We used flagellin protein from P. Aeruginosa, a key initiator of acute *** lung infection, to elucidate mechanisms by which acute lung infection with P. Aeruginosa can cause fibrogenesis/*** We studied the effect of flagellin from P. Aeruginosa （flagellin for short） on the transforming growth factor beta 1 （TGF-β1） and interleukin-8 （IL-8） expression, and the possible involvement of the signaling pathway, tumor necrosis factor receptor-associated factor 6 （TRAF6）/mitogen activated protein kinase （MAPK） pathway. flagellin was purified from the P. Aeruginosa standard strain, PAO1. Normal bronchial epithelial cells BEAS-2B were challenged with different concentrations of flagellin, and cell viability assessment was performed by cell counting kit-8. BEAS-2B cells were incubated with flagellin with the specific MAPK inhibitors or TRAF6 siRNA. Cell lysates and the cultured supernatant were collected. The level of TGF-β1 and IL-8 were detected by enzyme-linked immunosorbant assay （ELISA）. Western blotting was used to detect the protein levels of MAPK signal proteins p38, c-Jun NH2-terminal kinase （JNK） and extracellular regulated kinase （ERK）.Results Expression of TGF-β1 in BEAS-2B cells was elevated by flagellin

关键词： flagellin Pseudomonas aeruginosa transforming growth factor beta 1 mitogen activated protein kinase tumor necrosis factor receptor-associated factor 6

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

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Cellular & Molecular Immunology 2016年第4期13卷 514-523页

作者： Wei Li Jingyi Yang Ejuan Zhang Maohua Zhong Yang Xiao Jie Yu Dihan Zhou Yuan Cao Yi Yang Yaoming Li Huimin Yan Mucosal Immunity Research Group State Key Laboratory of Virology Wuhan Institute of Virology Chinese Academy of Sciences Wuhan 430071 People's Republic of China

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.

关键词： flagellin NLRC4 TLR5 macrophage adaptive immunity

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Relationship between genotype and phenotype of flagellin C in Salmonella

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World Journal of Gastroenterology 2001年第6期7卷 864-867页

作者： Wan-Sheng Ji~1 Jia-Lu Hu~1 Jun-Wen Qiu~1 Bo-Rong Pan~1 Dao-Rong Peng~2 Bing-Long Shi~1 Shao-Juan Zhou~1 Kai-Chun Wu~1 Dai-Ming Fan~1 1 Chinese PLA Institute of Digestive Diseases2 Department of Bacteriology,Xijing Hospital,Fourth Military Medical University,Xi’an 710032,Shaanxi Province,China Chinese PLA Institute of Digestive Diseases Xijing Hospital Fourth Military Medical University Xi'an 710032 Shaanxi Province China.

AIM: To discover the relationship between the genotype and antigen serotype of flagellin C among Salmonella strains. METHODS: Fragment of Salmonella flagellin C in plasmid pLS408 was cloned, sequenced and compared with the corresponding sequence in other strains. Salmonella strains including two typhi strains, one paratyphoid strain, one enteritidis and one typhimurium strain were isolated from outpatients. Genome DNA was purified respectively from these clinical isolates, then the corresponding flagellin C fragment was amplified by polymerase chain reaction,and the amplification products were analyzed by agarose gel electrophoresis. RESULTS: The cloned fragment includes 582 nucleotides encoding the variable region and partial conservative region of Salmonella flagellin C in plasmid pLS408. With comparison to the corresponding sequences reported previously, there is only a little difference from other strains with the same flagellar serotype in both nucleotide and amino acid level. Specific PCR products were amplified in Salmonella strains with flagellar serotype H-1-d including S. muenchen, typhi and typhimurium, but not in S. paratyphoid C or S. enteritidis strains. CONCLUSION: In this experiment, the specificity of nucleotide sequence could be found in flagellin C central variable regions as it exists in flagellar serotypes in Salmonella. It may be helpful to developing a rapid, sensitive, accurate and PCR-based method to detect Salmonella strains with serotype H-1-d.

关键词： Amino Acid Sequence Base Sequence flagellin Genotype Molecular Sequence Data Phenotype Protein Isoforms Salmonella Species Specificity

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Calcium phosphate nanoparticles show an effective activation of the innate immune response in vitro and in vivo after functionalization with flagellin

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Virologica Sinica 2014年第1期29卷 33-39页

作者： Diana Kozlova Viktoriya Sokolova Maohua Zhong Ejuan Zhang Jingyi Yang Wei Li Yi Yang Jan Buer Astrid Maria Westendorf Matthias Epple Huimin Yan Institute of Inorganic Chemistry and Center for Nanointegration Duisburg-Essen(CeNIDE) University of Duisburg-Essen Mucosal Immunity Research Group State Key Laboratory of VirologyWuhan Institute of VirologyChinese Academy of Sciences Institute of Medical Microbiology University Hospital EssenUniversity of Duisburg-Essen

For subunit vaccines,adjuvants play a key role in shaping the magnitude,persistence and form of targeted antigen-specific immune *** is a potent immune activator by bridging innate inflammatory responses and adaptive immunity and an adjuvant candidate for clinical *** phosphate nanoparticles are efficient carriers for different biomolecules like DNA,RNA,peptides and ***-functionalized calcium phosphate nanoparticles were prepared and their immunostimulatory effect on the innate immune system,*** cytokine production,was *** induced the production of the proinflammatory cytokines IL-8 （Caco-2 cells） and IL-1β（bone marrow-derived macrophages; BMDM） in vitro and IL-6 in vivo after intraperitoneal injection in *** immunostimulation was more pronounced than with free flagellin.

关键词： calcium phosphate nanoparticle immunostimulation innate immune system flagellin adjuvants

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Pyrimidine Triazole Thioether Derivatives as Toll-Like Receptor 5 (TLR5)/flagellin Complex Inhibitors

Pyrimidine Triazole Thioether Derivatives as Toll-Like Recep...

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第十届全国化学生物学学术会议

作者： Yan Lei Liang Jiaqi Yao Chenbo Wu Peiyao Zeng Xianfeng Cheng Kui Yin Hang Center of Basic Molecular Science Department of ChemistryTsinghua University

Protein protein interactions have been regarded as "undruggabl|e" despite their importance in many biological *** complex formed between host toll-like receptor 5(TLR5) and flagellin,a globular protein that is the main component of a bacterial flagellum,plays a vital role in a number of pathogen defenses,immunological diseases and *** high-throughput screening,we identified two hits with a common pharmacophore,which were used to successfully develop a series of pyrimidine triazole thioether derivatives as novel inhibitors of flagellin binding to *** a multitude of assays,TH1020 was identified as a potent antagonist of TLR5 signaling with promising activity(IC=0.85±0.12 μM) and ***,TH1020 was shown to repress the expression of downstream TNF-a signaling pathways mediated by the TLR5/flagellin complex *** on molecular docking simulation,TH1020 is suggested to compete with flagellin and disrupt its association with *** the first potent TLR5 inhibitor reported in the literature,TH1020 provides a much-needed molecular probe for studying this important protein-protein interaction and a lead compound for identifying novel therapeutics targeting TLR5.

关键词： TLR5 flagellin protein-protein interactions immunomodulators

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TLR5 Specific Ligand, flagellin, Prolonged Allograft Survival via Activating Regulatory T Cells in MAPKdependent Pathway

TLR5 Specific Ligand, Flagellin, Prolonged Allograft Surviva...

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第八届全国免疫学学术大会

作者： Jing Hao Chao Zhang Ting Liang Jing Song Guihua Hou Key Laboratory for Experimental Teratology of the Ministry of Education and Institute of Experimental Nuclear Medicine School of Medicine Shandong University

BACKG ROUND and Purpose:It has been well documented that Regulatory T cells(Tregs) play an important role in immuno-tolerance keeping, and activating of Tregs would be a practicable way to overcome the *** current research, the effect of recombinant flagellin(fli C) on allograft survival and its mechanisms were explored with murine skin allotransplantation ***:Herein, recombinant flagellin was expressed and purified firstly, then murine skin allotransplantation models were established and the effects of fli C on allograft survival and recipients Treg were studied in vivo and in *** experiments were performed using anti-TLR5 blocking antibody or JNK- / p38 MAPKsignaling inhibitors(SP600125/SB203580).Results:It was shown that the fli C(3mg/kg)-administration in vivo obviously prolonged graft survival(MST 18.4±1.1 day) than control(10±0.7 day), pflagellinTLR5 signaling pathway was JNK- and MAPK- dependent;expression of transcription factor Foxp3 was partially prevented by pharmacological inhibiting MAPK ***:Together, these data demonstrate that flagellin administration could significantly prolong allograft survival through activating and expanding Treg subset via TLR5 pathway specifically, by a mechanism partially dependent on the activation of p38 MAPK;thereby flagellin may become a promising new candidate to down-regulate allograft rejection.

关键词： flagellin TLR5 allotransplantation regulatory T cells

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AEG-1肺归巢域与flagellin融合基因的杆状病毒制备及鉴定

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中国实验诊断学 2018年第10期22卷 1807-1812页

作者：王希王琳张喆龙敏董轲张惠中第四军医大学唐都医院临床实验与检验科陕西西安710038

目的通过重叠PCR方法获得AEG-1C-Flic融合基因,采用Bac-To-Bac杆状病毒表达系统获得重组AEG-1C-Flic杆状病毒,为后续AEG-1C-Flic病毒样颗粒疫苗的制备奠定基础。方法从鼠伤寒沙门菌的基因组中PCR扩增细菌鞭毛蛋白flagellin,采用重叠PCR... 详细信息

目的通过重叠PCR方法获得AEG-1C-Flic融合基因,采用Bac-To-Bac杆状病毒表达系统获得重组AEG-1C-Flic杆状病毒,为后续AEG-1C-Flic病毒样颗粒疫苗的制备奠定基础。方法从鼠伤寒沙门菌的基因组中PCR扩增细菌鞭毛蛋白flagellin,采用重叠PCR的方法将AEG1肺归巢域段基因与flagellin融合,并添加猴免疫缺陷病毒(simian immunodeficiency virus,SIV)包膜蛋白gp41信号肽与跨膜区,构建AEG-1C-Flic pFastBacTM重组表达载体,转座大肠杆菌***10Bac感受态细胞后获得重组杆粒,重组杆粒转染昆虫细胞Tn5,获得AEG-1CFlic重组杆状病毒,并采用Western Blot方法对该病毒进行鉴定。结果构建的AEG-1C-Flic杆状病毒表达载体经双酶切及测序正确,通过转化*** DH10Bac感受态细胞获得重组Bacmid,经PCR鉴定大小正确,Western Blot结果显示Bacmid转染Tn5细胞后成功获得重组杆状病毒rBv AEG-1C-Flic。结论成功构建的AEG-1C-Flic杆状病毒可用于新型AEG1-病毒样颗粒疫苗的制备。

关键词：病毒样颗粒 AEG-1 肺归巢域 flagellin

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