title: SuperSILAC Quantitative Proteome Profiling of Murine Middle Ear Epithelial Cell Remodeling with NTHi.
abstract: BACKGROUND
  Chronic Otitis Media with effusion (COME) develops after sustained inflammation and is characterized by secretory middle ear epithelial metaplasia and effusion, most frequently mucoid. Non-typeable Haemophilus influenzae (NTHi), the most common acute Otitis Media (OM) pathogen, is postulated to promote middle ear epithelial remodeling in the progression of OM from acute to chronic. The goals of this study were to examine histopathological and quantitative proteomic epithelial effects of NTHi challenge in a murine middle ear epithelial cell line.
  
  
  METHODS
  NTHi lysates were generated and used to stimulate murine epithelial cells (mMEEC) cultured at air-liquid interface over 48 hours- 1 week. Conditional quantitative Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC) of cell lysates was performed to interrogate the global protein production in the cells, using the SuperSILAC technique. Histology of the epithelium over time was done to measure bacterial dependent remodeling.
  
  
  RESULTS
  Mass spectrometry analysis identified 2,565 proteins across samples, of which 74 exhibited differential enrichment or depletion in cell lysates (+/-2.0 fold-change; p value<0.05). The key molecular functions regulated by NTHi lysates exposure were related to cell proliferation, death, migration, adhesion and inflammation. Finally, chronic exposure induced significant epithelial thickening of cells grown at air liquid interface.
  
  
  CONCLUSIONS
  NTHi lysates drive pathways responsible of cell remodeling in murine middle ear epithelium which likely contributes to observed epithelial hyperplasia in vitro. Further elucidation of these mediators will be critical in understanding the progression of OM from acute to chronic at the molecular level.
authors: Sheikh Zayed Center for Pediatric Surgical Innovation, Children's National Health System, Washington, DC, United States of America.
authors: Stéphanie
authors: S
authors: Val
authors: Sheikh Zayed Center for Pediatric Surgical Innovation, Children's National Health System, Washington, DC, United States of America.
authors: Katelyn
authors: K
authors: Burgett
authors: Center for Genetic Medicine Research, Children's National Health System, Washington, DC, United States of America.
authors: Kristy J
authors: KJ
authors: Brown
authors: Sheikh Zayed Center for Pediatric Surgical Innovation, Children's National Health System, Washington, DC, United States of America.|Division of Pediatric Otolaryngology, Children's National Health System, Washington, DC, United States of America.
authors: Diego
authors: D
authors: Preciado
mesh_terms: D000818:Animals; D002460:Cell Line; D004432:Ear, Middle / Q000378:metabolism / Q000473:pathology; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology; D006192:Haemophilus Infections / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology*; D006193:Haemophilus influenzae / Q000145:classification; D051379:Mice; D010034:Otitis Media with Effusion / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology*; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods
chemical_list: D020543:Proteome