title: Microbiota, innate immune system, and gastrointestinal muscle: ongoing studies.
abstract: OBJECTIVE
  To test the activities of culture-extracted or commercially available toll-like receptors (TLRs) ligands to establish their direct impact on target gastrointestinal motor cells.
  
  
  METHODS
  Short-term and long-term effects of Shigella flexneri M90T and Escherichia coli K-2 strains-extracted lipopolysaccharides (LPS), commercially highly purified LPS (E. coli O111:B4 and EH100), and Pam2CSK4 and Pam3CSK4, which bind TLR2/6 and TLR1/2 heterodimers, respectively, have been assessed on pure primary cultures of colonic human smooth muscle cells (HSMC).
  
  
  RESULTS
  Pathogenic Shigella-LPS and nonpathogenic E. coli K-2-LPS induced a time-dependent decrease of resting cell length and acetylcholine-induced contraction, with both alterations occurring rapidly and being more pronounced in response to the former. However, their effects differed, prolonging HSMC exposure with Shigella-LPS effects maintained throughout the 4 hours of observation compared with E. coli K-2-LPS, which disappeared after 60 minutes of incubation. Similar differences in magnitude and time dependency of myogenic effects were observed between pure TLR4 and TLR2/1 or TLR2/6 ligands. The specific activation of TLR4 with LPS from pathogen or nonpathogen E. coli, O111:B4 and EH100, respectively, induced smooth muscle alterations that progressively increased, prolonging incubation, whereas TLR2 ligands induced short-term alterations, of a lesser magnitude, which decreased over time. The real-time polymerase chain reaction analysis showed that HSMC express mRNA for TLR1, 2, 4, and 6, substantiating a direct effect of TLR ligands on human colonic smooth muscle.
  
  
  CONCLUSIONS
  This study highlights that bacterial products can directly affect gastrointestinal motility and that TLRs subtypes may differ in their cellular activity.
authors.affiliation: Department of Internal Medicine and Medical Specialties, University La Sapienza, Rome, Italy.
authors.forename: Ivan
authors.initials: I
authors.lastname: Tattoli
authors.forename: Chiara
authors.initials: C
authors.lastname: Petitta
authors.forename: Annunziata
authors.initials: A
authors.lastname: Scirocco
authors.forename: Francesca
authors.initials: F
authors.lastname: Ammoscato
authors.forename: Alessia
authors.initials: A
authors.lastname: Cicenia
authors.forename: Carola
authors.initials: C
authors.lastname: Severi
mesh_terms: D002478:Cells, Cultured; D003106:Colon / Q000166:cytology / Q000276:immunology / Q000378:metabolism / Q000503:physiopathology; D004926:Escherichia coli / Q000276:immunology* / Q000378:metabolism; D005769:Gastrointestinal Motility / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D007113:Immunity, Innate; D008024:Ligands; D008070:Lipopolysaccharides / Q000494:pharmacology*; D054892:Metagenome; D009119:Muscle Contraction; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D012763:Shigella flexneri / Q000276:immunology* / Q000378:metabolism; D051195:Toll-Like Receptor 2 / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D051193:Toll-Like Receptors / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*
chemical_list: D008024:Ligands; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors