title: Regulated expression of collagenases MMP-1, -8, and -13 and stromelysins MMP-3, -10, and -11 by human corneal epithelial cells.
abstract: OBJECTIVE
  This study investigated the regulated expression of collagenases (MMP-1, -8, and -13) and stromelysins (MMP-3, -10, and -11) by human corneal epithelial cells treated with IL-1 beta, TNF-alpha, and doxycycline, a medication used to treat ocular surface diseases.
  
  
  METHODS
  Primary human corneal epithelial cell cultures were treated with IL-1 beta or TNF-alpha, with or without their corresponding inhibitors. Total RNA extracted from cells treated for 4 to 24 hours was subjected to semiquantitative RT-PCR and Northern hybridization. Conditioned media from 24-hour-treated cultures were evaluated for MMP production by ELISA and activity assays.
  
  
  RESULTS
  Semiquantitative RT-PCR and Northern hybridization revealed that the mRNAs of MMP-1, -13, -3, -10, and -11 were dose dependently upregulated by IL-1 beta and TNF-alpha, whereas MMP-8 and -14 and tissue inhibitor of metalloproteinase (TIMP)-1 were not altered, in corneal epithelial cells. MMP ELISA and activity assays confirmed this dose-dependent increase in MMP-1, -13, -3, and -10 protein production in conditioned media by IL-1 beta and TNF-alpha. This stimulated production was inhibited by their neutralizing antibodies and by IL-1 receptor antagonist. Doxycycline suppressed stimulated MMP-1, -10, and -13 production at both the mRNA and protein levels.
  
  
  CONCLUSIONS
  This study demonstrated that IL-1 beta and TNF-alpha upregulate collagenases (MMP-1, -13) and stromelysins (MMP-3, -10, and -11) in human corneal epithelial cells. Doxycycline suppresses stimulated MMP-1, -13, and -10 at the mRNA and protein levels, which suggests that collagenases and stromelysins may play a role in the pathogenesis of sterile corneal ulceration and other ocular surface diseases.
authors.affiliation: Ocular Surface Center, Cullen Eye Institute, Department of Ophthalmology, Baylor College of Medicine, Houston, Texas 77030, USA. dequanl@bcm.tmc.edu
authors.forename: De Quan
authors.initials: DQ
authors.lastname: Li
authors.forename: Tie Yan
authors.initials: TY
authors.lastname: Shang
authors.forename: Hyun-Seung
authors.initials: HS
authors.lastname: Kim
authors.forename: Abraham
authors.initials: A
authors.lastname: Solomon
authors.forename: Balakrishna L
authors.initials: BL
authors.lastname: Lokeshwar
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Pflugfelder
mesh_terms: D015152:Blotting, Northern; D002478:Cells, Cultured; D017364:Collagenases / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004318:Doxycycline / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D019573:Epithelium, Corneal / Q000187:drug effects / Q000201:enzymology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism; D053506:Matrix Metalloproteinase 10; D053507:Matrix Metalloproteinase 11; D053509:Matrix Metalloproteinase 13; D019278:Matrix Metalloproteinase 3 / Q000235:genetics / Q000378:metabolism; D020784:Matrix Metalloproteinase 8 / Q000235:genetics / Q000378:metabolism; D008666:Metalloendopeptidases / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation
chemical_list: D007375:Interleukin-1; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D017364:Collagenases; C507079:MMP13 protein, human; D053507:Matrix Metalloproteinase 11; D053509:Matrix Metalloproteinase 13; D008666:Metalloendopeptidases; D019278:Matrix Metalloproteinase 3; D053506:Matrix Metalloproteinase 10; D020784:Matrix Metalloproteinase 8; D020781:Matrix Metalloproteinase 1; D004318:Doxycycline