title: Pro-angiogenic activity of TLRs and NLRs: a novel link between gut microbiota and intestinal angiogenesis.
abstract: OBJECTIVE
  In intestinal inflammation the gut microbiota induces an innate immune response by activating epithelial and immune cells that initiate or maintain inflammation. We investigated whether the microbiota also can activate local microvascular cells and induce angiogenesis.
  
  
  METHODS
  Human intestinal microvascular endothelial cells (HIMEC) and human intestinal fibroblasts (HIF) were exposed to bacterial ligands specific for Toll-like receptor (TLR)2/6 and 4, and NOD1 and NOD2, and cell proliferation, migration, transmigration, tube formation, and production of pro-angiogenic factors were measured. The ability of the ligands to induce ex vivo vessel sprouting in an aortic ring assay and in vivo angiogenesis using a collagen gel assay also were assessed.
  
  
  RESULTS
  Bacterial ligands induced proliferation, migration, transmigration, tube formation of HIMEC, vessel sprouting, and in vivo angiogenesis; they also stimulated production of angiogenic factors from HIMEC and HIF, and HIF-derived angiogenic factors promoted HIMEC proliferation. To various degrees, all ligands induced angiogenic responses, but these were ligand- and cell type-dependent. Responses were mediated through receptor interacting protein-2 (RIP2)- and tumor necrosis factor receptor-associated factor 6 (TRAF6)-dependent signaling, involved the mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) pathways and the up-regulation of vascular endothelial growth factor receptor 2 (VEGF-R2) and focal adhesion kinase (FAK). Knockdown of RIP2 and TRAF6 by RNA interference and neutralization of interleukin-8, basic fibroblast growth factor, and vascular endothelial growth factor inhibited TLR-/NOD-like receptor-induced HIMEC angiogenesis.
  
  
  CONCLUSIONS
  The gut microbiota can selectively activate mucosal endothelial and mesenchymal cells to promote specific angiogenic responses in a TLR- and NOD-like receptor-dependent fashion. This innate immunity-mediated response may expand the mucosal microvascular network, foster immune cell recruitment, and contribute to chronic intestinal inflammation.
authors.affiliation: Department of Pathobiology, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, Ohio, USA.
authors.forename: Anja
authors.initials: A
authors.lastname: Schirbel
authors.forename: Sean
authors.initials: S
authors.lastname: Kessler
authors.forename: Florian
authors.initials: F
authors.lastname: Rieder
authors.forename: Gail
authors.initials: G
authors.lastname: West
authors.forename: Nancy
authors.initials: N
authors.lastname: Rebert
authors.forename: Kewal
authors.initials: K
authors.lastname: Asosingh
authors.forename: Christine
authors.initials: C
authors.lastname: McDonald
authors.forename: Claudio
authors.initials: C
authors.lastname: Fiocchi
mesh_terms: D049109:Cell Proliferation / Q000187:drug effects; D017077:Culture Media, Conditioned / Q000494:pharmacology; D005347:Fibroblasts / Q000502:physiology; D006801:Humans; D007422:Intestines / Q000098:blood supply* / Q000382:microbiology*; D054892:Metagenome / Q000502:physiology*; D016328:NF-kappa B / Q000378:metabolism; D018919:Neovascularization, Physiologic; D053474:Nod1 Signaling Adaptor Protein / Q000378:metabolism*; D053473:Nod2 Signaling Adaptor Protein / Q000378:metabolism*; D034622:RNA Interference; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2 / Q000235:genetics / Q000378:metabolism; D048029:TNF Receptor-Associated Factor 6 / Q000378:metabolism; D051193:Toll-Like Receptors / Q000378:metabolism*; D057705:Transendothelial and Transepithelial Migration / Q000187:drug effects; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
chemical_list: D017077:Culture Media, Conditioned; D016328:NF-kappa B; C119067:NOD1 protein, human; C429018:NOD2 protein, human; D053474:Nod1 Signaling Adaptor Protein; D053473:Nod2 Signaling Adaptor Protein; D048029:TNF Receptor-Associated Factor 6; D051193:Toll-Like Receptors; D040301:Vascular Endothelial Growth Factor Receptor-2; C506253:RIPK2 protein, human; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2