title: Fibronectin-integrin signaling is required for L-glutamine's protection against gut injury.
abstract: BACKGROUND
  Extracellular matrix (ECM) stabilization and fibronectin (FN)-Integrin signaling can mediate cellular protection. L-glutamine (GLN) is known to prevent apoptosis after injury. However, it is currently unknown if ECM stabilization and FN-Integrin osmosensing pathways are related to GLN's cell protective mechanism in the intestine.
  
  
  METHODS
  IEC-6 cells were treated with GLN with or without FN siRNA, integrin inhibitor GRGDSP, control peptide GRGESP or ERK1/2 inhibitors PD98059 and UO126 under basal and stressed conditions. Cell survival measured via MTS assay. Phosphorylated and/or total levels of cleaved caspase-3, cleaved PARP, Bax, Bcl-2, heat shock proteins (HSPs), ERK1/2 and transcription factor HSF-1 assessed via Western blotting. Cell size and F-actin morphology quantified by confocal fluorescence microscopy and intracellular GLN concentration by LC-MS/MS.
  
  
  RESULTS
  GLN's prevention of FN degradation after hyperthermia attenuated apoptosis. Additionally, inhibition of FN-Integrin interaction by GRGDSP and ERK1/2 kinase inhibition by PD98059 inhibited GLN's protective effect. GRGDSP attenuated GLN-mediated increases in ERK1/2 phosphorylation and HSF-1 levels. PD98059 and GRGDSP also decreased HSP levels after GLN treatment. Finally, GRGDSP attenuated GLN-mediated increases in cell area size and disrupted F-actin assembly, but had no effect on intracellular GLN concentrations.
  
  
  CONCLUSIONS
  Taken together, this data suggests that prevention of FN degradation and the FN-Integrin signaling play a key role in GLN-mediated cellular protection. GLN's signaling via the FN-Integrin pathway is associated with HSP induction via ERK1/2 and HSF-1 activation leading to reduced apoptosis after gut injury.
authors.affiliation: Department of Anesthesiology, University of Colorado, Aurora, Colorado, United States of America. Stefanie.Niederlechner@ucdenver.edu
authors.forename: Stefanie
authors.initials: S
authors.lastname: Niederlechner
authors.forename: Jelena
authors.initials: J
authors.lastname: Klawitter
authors.forename: Christine
authors.initials: C
authors.lastname: Baird
authors.forename: Alyssa R
authors.initials: AR
authors.lastname: Kallweit
authors.forename: Uwe
authors.initials: U
authors.lastname: Christians
authors.forename: Paul E
authors.initials: PE
authors.lastname: Wischmeyer
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D019610:Cytoprotection; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D005353:Fibronectins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D020868:Gene Silencing; D005973:Glutamine / Q000494:pharmacology*; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006358:Hot Temperature; D016023:Integrins / Q000235:genetics / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism; D059748:Proteolysis; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects*; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: C494828:Bax protein, rat; D004791:Enzyme Inhibitors; D005353:Fibronectins; D018840:HSP70 Heat-Shock Proteins; D016023:Integrins; D034741:RNA, Small Interfering; D051028:bcl-2-Associated X Protein; D005973:Glutamine; D011065:Poly(ADP-ribose) Polymerases; D048049:Extracellular Signal-Regulated MAP Kinases; D053148:Caspase 3