Probiotic Lactobacillus reuteri suppress proinflammatory cytokines via c‐Jun

Background: Differential immunoregulatory capabilities of probiotic Lactobacillus were explored in the context of pediatric Crohn's disease. Experimental strategies addressed molecular mechanisms of tumor necrosis factor (TNF) suppression in activated macrophages by transcriptional regulation. Methods: Secreted factors produced by probiotic Lactobacillus reuteri strains were harvested and tested with human monocytes and macrophages. Quantitative immunoassays and real‐time reverse‐transcriptase polymerase chain reaction (RT‐PCR) were used to examine relative quantities of human cytokines and TNF mRNA, respectively, and reporter assays assessed transcriptional regulation of TNF by probiotics. DNA‐protein macroarrays interrogated probiotic‐mediated effects on transcription factor activation. Finally, enzyme‐linked immunosorbent assays (ELISAs) and immunoblots examined the involvement of the specific transcription factor AP‐1 and its components. Results: Probiotic L. reuteri strain ATCC PTA 6475 demonstrated the ability to potently suppress human TNF production by lipopolysaccharide‐activated monocytes and primary monocyte‐derived macrophages from children with Crohn's disease. Quantities of the chemokine MCP‐1/CCL2 were also reduced by probiotic L. reuteri strain ATCC PTA 6475 in macrophages of children in remission. Quantitative real‐time RT‐PCR and luciferase reporter assays showed that transcriptional regulation of human TNF was a primary mechanism of probiotic‐mediated immunomodulation. Probiotic L. reuteri suppressed TNF transcription by inhibiting activation of MAP kinase‐regulated c‐Jun and the transcription factor, AP‐1. Conclusions: Human TNF and MCP‐1 suppression by probiotic L. reuteri was strain‐dependent, and the activation of c‐Jun and AP‐1 represent primary targets for probiotic‐mediated suppression of TNF transcription. This report emphasizes the clonal nature of immunoprobiosis and delineation of a specific immunomodulatory mechanism for probiotic strain selection in future inflammatory bowel disease‐oriented clinical trials.

[1]  Simon Heath,et al.  Novel Crohn Disease Locus Identified by Genome-Wide Association Maps to a Gene Desert on 5p13.1 and Modulates Expression of PTGER4 , 2007, PLoS genetics.

[2]  O. Dewit,et al.  Multicenter randomized‐controlled clinical trial of probiotics (Lactobacillus johnsonii, LA1) on early endoscopic recurrence of Crohn's disease after ileo‐caecal resection , 2007 .

[3]  Y. Tsao,et al.  ErbB4 (JM-b/CYT-1)-induced expression and phosphorylation of c-Jun is abrogated by human papillomavirus type 16 E5 protein , 2007, Oncogene.

[4]  M. Vatn,et al.  Granulocyte, monocyte/macrophage apheresis for inflammatory bowel disease: The first 100 patients treated in Scandinavia , 2007, Scandinavian journal of gastroenterology.

[5]  G. Reid,et al.  G‐CSF‐mediated inhibition of JNK is a key mechanism for Lactobacillus rhamnosus‐induced suppression of TNF production in macrophages , 2006, Cellular microbiology.

[6]  Judy H. Cho,et al.  A Genome-Wide Association Study Identifies IL23R as an Inflammatory Bowel Disease Gene , 2006, Science.

[7]  J. Versalovic,et al.  Immunoregulation by probiotics , 2006 .

[8]  L. Osiecki,et al.  Novel Therapies for Crohn's Disease: Focus on Immunomodulators and Antibiotics , 2006, The Annals of pharmacotherapy.

[9]  Kazuhiko Nakamura,et al.  Novel strategies for the treatment of inflammatory bowel disease: Selective inhibition of cytokines and adhesion molecules. , 2006, World journal of gastroenterology.

[10]  G. Gazelle,et al.  Risks and benefits of infliximab for the treatment of Crohn's disease. , 2006, Clinical gastroenterology and hepatology : the official clinical practice journal of the American Gastroenterological Association.

[11]  A. Akobeng,et al.  Tumour necrosis factor alpha blocking agents for induction of remission in ulcerative colitis. , 2006, The Cochrane database of systematic reviews.

[12]  K. Madsen,et al.  Bioproduction of conjugated linoleic acid by probiotic bacteria occurs in vitro and in vivo in mice. , 2006, The Journal of nutrition.

[13]  A. Gasbarrini,et al.  Efficacy of Lactobacillus GG in maintaining remission of ulcerative colitis , 2006, Alimentary pharmacology & therapeutics.

[14]  C. Pothoulakis,et al.  Saccharomyces boulardii produces a soluble anti-inflammatory factor that inhibits NF-kappaB-mediated IL-8 gene expression. , 2006, Biochemical and biophysical research communications.

[15]  R. Baldassano,et al.  Risk factors for initial surgery in pediatric patients with Crohn's disease. , 2006, Gastroenterology.

[16]  M. Sata,et al.  Pro-inflammatory signaling by Jun-N-terminal kinase in inflammatory bowel disease. , 2006, International journal of molecular medicine.

[17]  K. Rioux,et al.  Probiotics in the treatment of inflammatory bowel disease. , 2006, Journal of clinical gastroenterology.

[18]  P. Rutgeerts,et al.  Human anti-tumor necrosis factor monoclonal antibody (adalimumab) in Crohn's disease: the CLASSIC-I trial. , 2006, Gastroenterology.

[19]  J. Mary,et al.  Ineffectiveness of Lactobacillus johnsonii LA1 for prophylaxis of postoperative recurrence in Crohn’s disease: a randomised, double blind, placebo controlled GETAID trial , 2005, Gut.

[20]  G. Courtois,et al.  Yersinia Outer Protein P Suppresses TGF-β-Activated Kinase-1 Activity to Impair Innate Immune Signaling in Yersinia enterocolitica-Infected Cells1 , 2005, The Journal of Immunology.

[21]  D. Fan,et al.  Correlation between a gene polymorphism of tumor necrosis factor and inflammatory bowel disease. , 2005, Chinese journal of digestive diseases.

[22]  R. Baldassano,et al.  A Randomized, Double‐blind Trial of Lactobacillus GG Versus Placebo in Addition to Standard Maintenance Therapy for Children with Crohn's Disease , 2005, Inflammatory bowel diseases.

[23]  P. Rutgeerts,et al.  Infliximab induces apoptosis of monocytes and T lymphocytes in a human-mouse chimeric model. , 2005, Clinical immunology.

[24]  Å. Danielsson,et al.  Infliximab as rescue therapy in severe to moderately severe ulcerative colitis: a randomized, placebo-controlled study. , 2005, Gastroenterology.

[25]  G. Ramadori,et al.  Quantitative measurement of cytokine mRNA in inflammatory bowel disease: relation to clinical and endoscopic activity and outcome , 2005, European journal of gastroenterology & hepatology.

[26]  P. Nickerson,et al.  The Role of Cytokine Gene Polymorphisms in Determining Disease Susceptibility and Phenotype in Inflammatory Bowel Disease , 2005, The American Journal of Gastroenterology.

[27]  A. Cohen,et al.  Tumor Necrosis Factor Receptor Gene Polymorphisms in Crohn's Disease: Association with Clinical Phenotypes , 2005, The American Journal of Gastroenterology.

[28]  D. Hommes,et al.  Infliximab induced T lymphocyte apoptosis in Crohn's disease. , 2005, The Journal of rheumatology. Supplement.

[29]  J. Versalovic,et al.  Probiotic Lactobacillus spp. Diminish Helicobacter hepaticus-Induced Inflammatory Bowel Disease in Interleukin-10-Deficient Mice , 2005, Infection and Immunity.

[30]  B. Yakir,et al.  A Polymorphism in the TNF-α Promoter Gene is Associated with Pediatric Onset and Colonic Location of Crohn's Disease , 2005, The American Journal of Gastroenterology.

[31]  J. Cnota,et al.  Lactobacillus Sepsis Associated With Probiotic Therapy , 2005, Pediatrics.

[32]  T. Matsui,et al.  Adsorptive granulocyte and monocyte apheresis for refractory Crohn’s disease: an open multicenter prospective study , 2004, Journal of Gastroenterology.

[33]  J. Madara.  Building an intestine--architectural contributions of commensal bacteria. , 2004, The New England journal of medicine.

[34]  Marie Joseph,et al.  NF-κB-mediated repression of growth arrest- and DNA-damage-inducible proteins 45α and γ is essential for cancer cell survival , 2004 .

[35]  R. Baldassano,et al.  Patterns of Complementary and Alternative Medicine Use in a Population of Pediatric Patients with Inflammatory Bowel Disease , 2004, Inflammatory bowel diseases.

[36]  R. Whelan,et al.  Infliximab: mechanism of action beyond TNF-α neutralization in inflammatory bowel disease , 2004 .

[37]  K. Terpend,et al.  Lactic acid bacteria secrete metabolites retaining anti-inflammatory properties after intestinal transport , 2004, Gut.

[38]  R Balfour Sartor,et al.  Therapeutic manipulation of the enteric microflora in inflammatory bowel diseases: antibiotics, probiotics, and prebiotics. , 2004, Gastroenterology.

[39]  O. Grip,et al.  Circulating Monocytes and Plasma Inflammatory Biomarkers in Active Crohn's Disease: Elevated Oxidized Low-density Lipoprotein and the Anti-inflammatory Effect of Atorvastatin , 2004, Inflammatory bowel diseases.

[40]  J. Hampe,et al.  Reduction in diversity of the colonic mucosa associated bacterial microflora in patients with active inflammatory bowel disease , 2004, Gut.

[41]  H. Herfarth,et al.  Lactobacillus GG in inducing and maintaining remission of Crohn's disease , 2004, BMC gastroenterology.

[42]  Louis J. Cohen,et al.  Infliximab in Pediatric Ulcerative Colitis: Two-year Follow-up , 2004, Journal of pediatric gastroenterology and nutrition.

[43]  N. Valeur,et al.  Colonization and Immunomodulation by Lactobacillus reuteri ATCC 55730 in the Human Gastrointestinal Tract , 2004, Applied and Environmental Microbiology.

[44]  C. Hawkey,et al.  Lactobacillus plantarum 299: Beneficial in vitro immunomodulation in cells extracted from inflamed human colon , 2004, Journal of gastroenterology and hepatology.

[45]  J. Versalovic,et al.  Genotypic and Phenotypic Studies of Murine Intestinal Lactobacilli: Species Differences in Mice with and without Colitis , 2004, Applied and Environmental Microbiology.

[46]  Ashok Kumar,et al.  Dexamethasone Inhibits IL-12p40 Production in Lipopolysaccharide-Stimulated Human Monocytic Cells by Down-Regulating the Activity of c-Jun N-Terminal Kinase, the Activation Protein-1, and NF-κB Transcription Factors1 , 2004, The Journal of Immunology.

[47]  M. Kamm,et al.  Once daily high dose probiotic therapy (VSL#3) for maintaining remission in recurrent or refractory pouchitis , 2003, Gut.

[48]  J. Versalovic,et al.  Lactobacillus rhamnosus GG decreases TNF‐α production in lipopolysaccharide‐activated murine macrophages by a contact‐independent mechanism , 2003, Cellular microbiology.

[49]  J. Dahlerup,et al.  Response, relapse and mucosal immune regulation after infliximab treatment in fistulating Crohn's disease , 2003, Alimentary pharmacology & therapeutics.

[50]  J. Green-Johnson,et al.  Interactions of lactic acid bacteria with human intestinal epithelial cells: effects on cytokine production. , 2003, Journal of food protection.

[51]  C. Hellerbrand,et al.  Polymorphism of monocyte chemoattractant protein 1 in Crohn's disease , 2003, International Journal of Colorectal Disease.

[52]  Penny A. Johnson,et al.  Chemokine expression in IBD. Mucosal chemokine expression is unselectively increased in both ulcerative colitis and Crohn's disease , 2003, The Journal of pathology.

[53]  B. Ohlsson,et al.  Lactoferrin down-regulates the LPS-induced cytokine production in monocytic cells via NF-κB , 2002 .

[54]  A. Reimold,et al.  TNFalpha as therapeutic target: new drugs, more applications. , 2002, Current drug targets. Inflammation and allergy.

[55]  N. Borruel,et al.  Increased mucosal tumour necrosis factor α production in Crohn’s disease can be downregulated ex vivo by probiotic bacteria , 2002, Gut.

[56]  S. Travis,et al.  Role of tumor necrosis factor in Crohn's disease. , 2002, Current opinion in investigational drugs.

[57]  S. Guandalini.  Use of Lactobacillus-GG in paediatric Crohn's disease. , 2002, Digestive and liver disease : official journal of the Italian Society of Gastroenterology and the Italian Association for the Study of the Liver.

[58]  C. Prantera,et al.  Probiotics and Crohn's disease. , 2002, Digestive and liver disease : official journal of the Italian Society of Gastroenterology and the Italian Association for the Study of the Liver.

[59]  P. Gionchetti,et al.  Treatment of mild to moderate ulcerative colitis and pouchitis , 2002, Alimentary pharmacology & therapeutics.

[60]  Philip Rosenstiel,et al.  p38 Mitogen-Activated Protein Kinase Is Activated and Linked to TNF-α Signaling in Inflammatory Bowel Disease1 , 2002, The Journal of Immunology.

[61]  Thomas D. Schmittgen,et al.  Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. , 2001, Methods.

[62]  N. Tsuji,et al.  Tumor necrosis factor-alpha in serum of patients with inflammatory bowel disease as measured by a highly sensitive immuno-PCR. , 2001, Clinical chemistry.

[63]  D. Nicolae,et al.  A frameshift mutation in NOD2 associated with susceptibility to Crohn's disease , 2001, Nature.

[64]  G. Reuter.  The Lactobacillus and Bifidobacterium microflora of the human intestine: composition and succession. , 2001, Current issues in intestinal microbiology.

[65]  N. Mackman,et al.  LPS induction of gene expression in human monocytes. , 2001, Cellular signalling.

[66]  A. Hoffmann,et al.  Regulation of IL-6 and IL-8 Expression in Rheumatoid Arthritis Synovial Fibroblasts: the Dominant Role for NF-κB But Not C/EBPβ or c-Jun1 , 2000, The Journal of Immunology.

[67]  S. Targan,et al.  Tumor necrosis factor: biology and therapeutic inhibitors. , 2000, Gastroenterology.

[68]  B. Kirschner,et al.  Is Lactobacillus GG Helpful in Children With Crohn’s Disease? Results of a Preliminary, Open-Label Study , 2000, Journal of pediatric gastroenterology and nutrition.

[69]  Hyung-Sik Kang,et al.  IFN-γ Up-Regulates IL-18 Gene Expression Via IFN Consensus Sequence-Binding Protein and Activator Protein-1 Elements in Macrophages1 , 2000, The Journal of Immunology.

[70]  A. Poustka,et al.  Differential transcriptional regulation of the monocyte-chemoattractant protein-1 (MCP-1) gene in tumorigenic and non-tumorigenic HPV 18 positive cells: The role of the chromatin structure and AP-1 composition , 2000, Oncogene.

[71]  P. Sassone-Corsi,et al.  The bacterial protein YopJ abrogates multiple signal transduction pathways that converge on the transcription factor CREB , 2000, Cellular microbiology.

[72]  M. Birrer,et al.  TNF-α Gene Expression in Macrophages: Regulation by NF-κB Is Independent of c-Jun or C/EBPβ1 , 2000, The Journal of Immunology.

[73]  P. Brigidi,et al.  Prophylaxis of pouchitis onset with probiotic therapy: A double‐blind, placebo‐controlled trial , 2000, Gastroenterology.

[74]  R. Macdermott.  Chemokines in the Inflammatory Bowel Diseases , 1999, Journal of Clinical Immunology.

[75]  S. Hanauer.  Review article: safety of infliximab in clinical trials , 1999, Alimentary pharmacology & therapeutics.

[76]  S. Orenstein.  Pediatric inflammatory bowel disease , 1999, Current gastroenterology reports.

[77]  P. Rutgeerts,et al.  Infliximab for the treatment of fistulas in patients with Crohn's disease. , 1999, The New England journal of medicine.

[78]  J. Belaiche,et al.  Tumour necrosis factor (TNF) gene polymorphism influences TNF‐α production in lipopolysaccharide (LPS)‐stimulated whole blood cell culture in healthy humans , 1998, Clinical and experimental immunology.

[79]  R. Macdermott,et al.  The Central Role of Chemokines (Chemotactic Cytokines) in the Immunopathogenesis of Ulcerative Colitis and Crohn's Disease , 1998, Inflammatory bowel diseases.

[80]  N. Mackman,et al.  Lipopolysaccharide Induction of the Tumor Necrosis Factor-α Promoter in Human Monocytic Cells , 1997, The Journal of Biological Chemistry.

[81]  S. V. van Deventer.  Tumour necrosis factor and Crohn's disease. , 1997, Gut.

[82]  E. Vellenga,et al.  Effects of IL-10 and IL-4 on LPS-induced transcription factors (AP-1, NF-IL6 and NF-kappa B) which are involved in IL-6 regulation. , 1996, Leukemia.

[83]  A. DeFranco,et al.  Activation of c-Jun N-terminal kinase in bacterial lipopolysaccharide-stimulated macrophages. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[84]  T. Sternsdorf,et al.  Constitutive nuclear NF-kappa B in cells of the monocyte lineage. , 1994, The Biochemical journal.

[85]  Esther Jacobowitz Israel,et al.  Inflammatory bowel disease: diagnosis and treatment. , 1994, Seminars in gastrointestinal disease.

[86]  C. Weber,et al.  Pyrrolidine dithiocarbamate inhibits NF-kappa B mobilization and TNF production in human monocytes. , 1993, Journal of immunology.

[87]  C. Holdsworth,et al.  The assessment of faecal flora in patients with inflammatory bowel disease by a simplified bacteriological technique. , 1991, Journal of medical microbiology.

[88]  J T Boyle,et al.  Development and validation of a pediatric Crohn's disease activity index. , 1991, Journal of pediatric gastroenterology and nutrition.

[89]  D. Savage,et al.  Models for Study of the Specificity by Which Indigenous Lactobacilli Adhere to Murine Gastric Epithelia , 1979, Infection and immunity.

[90]  S. Willich,et al.  Bmc Complementary and Alternative Medicine Use of Complementary and Alternative Medicine in Germany – a Survey of Patients with Inflammatory Bowel Disease , 2006 .

[91]  B. Aggarwal,et al.  TNF blockade: an inflammatory issue. , 2006, Ernst Schering Research Foundation workshop.

[92]  J. Mrukowicz,et al.  Use of Probiotics in Children with Acute Diarrhea , 2005, Paediatric drugs.

[93]  W. Sandborn.  New concepts in anti-tumor necrosis factor therapy for inflammatory bowel disease. , 2005, Reviews in gastroenterological disorders.

[94]  H. Tilg,et al.  Elevated systemic levels of free interleukin-18 (IL-18) in patients with Crohn's disease. , 2005, European cytokine network.

[95]  R. Whelan,et al.  Infliximab: mechanism of action beyond TNF-alpha neutralization in inflammatory bowel disease. , 2004, European journal of gastroenterology & hepatology.

[96]  Marie Joseph,et al.  NF-kappa B-mediated repression of growth arrest- and DNA-damage-inducible proteins 45alpha and gamma is essential for cancer cell survival. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[97]  B. Sands.  Why do anti-tumor necrosis factor antibodies work in Crohn's disease? , 2004, Reviews in gastroenterological disorders.

[98]  Takayuki Matsumoto,et al.  Adsorptive monocyte‐granulocytapheresis (M‐GCAP) for refractory Crohn's disease , 2004, Journal of clinical apheresis.

[99]  Judy H. Cho,et al.  Significant role of genetics in IBD: the NOD2 gene. , 2003, Reviews in gastroenterological disorders.

[100]  Philip Rosenstiel,et al.  p38 mitogen-activated protein kinase is activated and linked to TNF-alpha signaling in inflammatory bowel disease. , 2002, Journal of immunology.

[101]  C. Prantera,et al.  INFLAMMATION AND INFLAMMATORY BOWEL DISEASE Ineffectiveness of probiotics in preventing recurrence after curative resection for Crohn’s disease: a randomised controlled trial with Lactobacillus GG , 2002 .

[102]  M. Birrer,et al.  TNF-alpha gene expression in macrophages: regulation by NF-kappa B is independent of c-Jun or C/EBP beta. , 2000, Journal of immunology.

[103]  P. Rutgeerts,et al.  Tumor necrosis factor alpha antibody (infliximab) therapy profoundly down-regulates the inflammation in Crohn's ileocolitis. , 1999, Gastroenterology.

[104]  N. Mackman,et al.  Lipopolysaccharide induction of the tumor necrosis factor-alpha promoter in human monocytic cells. Regulation by Egr-1, c-Jun, and NF-kappaB transcription factors. , 1997, The Journal of biological chemistry.

[105]  B. Crohn.  Segmental (Granulomatous) colitis. A historical survey. , 1970, Bibliotheca gastroenterologica.