title: Demodex-associated Bacillus proteins induce an aberrant wound healing response in a corneal epithelial cell line: possible implications for corneal ulcer formation in ocular rosacea.
abstract: OBJECTIVE
  The aim of the work presented here was to establish the response of a corneal epithelial cell line (hTCEpi) to protein extracted from a bacterium (Bacillus oleronius) previously isolated from a Demodex mite from a rosacea patient.
  
  
  METHODS
  The response of the corneal epithelial cell line to Bacillus proteins was measured in terms of alterations in cell migration and invasiveness. Changes in the expression of metalloproteinase genes and proteins were also assessed.
  
  
  RESULTS
  The results indicated increased cell migration (14.5-fold, P = 0.001) as measured using 8-μm PET inserts (BD Falcon) in a transwell assay and invasiveness (1.7-fold, P = 0.003) as measured using 8-μm Matrigel (BD Biocoat) invasion inserts in a 24-well plate assay format, following exposure to the Bacillus proteins. Cells exposed to the Bacillus protein showed a dose-dependent increase in expression of genes coding for matrix metalloprotease (MMP)-3 (61-fold) and MPP-9 (301-fold). This dose-dependent increase in gene expression was also reflected in elevated levels of MMP-9 protein (1.34-fold, P = 0.033) and increased matrix metalloprotease activity (1.96-fold, P = 0.043) being present in the culture supernatant. Cells also displayed reduced levels of β-integrin (1.25-fold, P = 0.01), indicative of increased motility and elevated levels of vinculin (2.7-fold, P = 0.0009), suggesting altered motility.
  
  
  CONCLUSIONS
  The results indicate that exposure of corneal epithelial cells to Bacillus proteins results in an aberrant wound healing response as visualized using a scratch wound assay. These results suggest a possible link between the high density of Demodex mites on the eyelashes of ocular rosacea patients and the development of corneal ulcers.
authors.affiliation: Department of Biology, NUI Maynooth, Co. Kildare, Ireland.
authors.forename: Niamh
authors.initials: N
authors.lastname: O'Reilly
authors.forename: Clair
authors.initials: C
authors.lastname: Gallagher
authors.forename: Kishore
authors.initials: K
authors.lastname: Reddy Katikireddy
authors.forename: Martin
authors.initials: M
authors.lastname: Clynes
authors.forename: Finbarr
authors.initials: F
authors.lastname: O'Sullivan
authors.forename: Kevin
authors.initials: K
authors.lastname: Kavanagh
mesh_terms: D000818:Animals; D001407:Bacillus / Q000378:metabolism*; D001426:Bacterial Proteins / Q000494:pharmacology*; D015153:Blotting, Western; D002460:Cell Line; D002465:Cell Movement; D003320:Corneal Ulcer / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D004591:Electrophoresis, Polyacrylamide Gel; D019573:Epithelium, Corneal / Q000378:metabolism / Q000382:microbiology / Q000473:pathology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D039641:Integrin beta Chains / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000096:biosynthesis / Q000235:genetics; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis / Q000235:genetics; D008925:Mites / Q000382:microbiology; D012313:RNA / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012393:Rosacea / Q000150:complications* / Q000382:microbiology / Q000473:pathology; D014945:Wound Healing / Q000187:drug effects*
chemical_list: D001426:Bacterial Proteins; D039641:Integrin beta Chains; D012313:RNA; D019278:Matrix Metalloproteinase 3; D020780:Matrix Metalloproteinase 9