title: The presence of lysyl oxidase-like enzymes in human control and keratoconic corneas.
abstract: OBJECTIVE
  Lysyl oxidases, a family comprising lysyl oxidase (LOX) and four LOX-like enzymes (LOXL1-4), catalyse the cross-linking of elastin and collagen fibrils. Keratoconus (KC) is characterized by progressive thinning leading to irregular astigmatism, resulting in significant visual impairment. Although the pathogenesis of KC remains unclear, one of the current hypotheses is based on alterations in the organization and structure of collagen fibrils. To extend existing general knowledge about cross-linking enzymes in the human cornea, in the present study we have focused on the detection of LOXL enzymes.
  
  
  METHODS
  The localization and distribution of LOXL1-4 were assessed in cryosections of 7 control donors (three males and three females; 25-68 years; mean age 46±17.6 years) and 8 KC corneas (5 males and 3 females; 25-46 years; mean age 31.3±7.5 years) using indirect fluorescent immunohistochemistry (IHC). The specimens were examined using an Olympus BX51 microscope (Olympus Co., Tokyo, Japan) at a magnification of 200-1000x. Western blot analysis of 4 control and 4 KC corneas was performed for all tested enzymes.
  
  
  RESULTS
  All four LOX-like enzymes were present in all layers of control corneas as well as in the limbus and conjunctiva. Almost no differences between control and pathological specimens were found for LOXL1. A lower staining intensity of LOXL2 was found using IHC and Western blot analysis in KC specimens. Decreases of the signal and small irregularities in the staining were found in the epithelium, keratocytes and extracellular matrix, where a gradual anterior-posterior weakening of the signal was observed. LOXL3 IHC staining was lower in the corneal stromal extracellular matrix and keratocytes of KC samples. No prominent differences were detected using IHC for LOXL4, but a slight decrease was observed in KC corneas using Western blot analysis.
  
  
  CONCLUSIONS
  We presume that the decrease of LOXL2 in KC corneas is more likely a consequence of the associated pathological processes (activation of stromal cells due to tissue weakening and consequent structural changes) than a direct cause leading to KC development. At this time, we are unable to provide a coherent explanation for the observed decrease of LOXL3 and LOXL4 in KC corneas.
authors.affiliation: Laboratory of the Biology and Pathology of the Eye, Institute of Inherited Metabolic Disorders, First Faculty of Medicine, Charles University in Prague and General University Hospital in Prague, Prague, Czech Republic.
authors.forename: Lubica
authors.initials: L
authors.lastname: Dudakova
authors.affiliation: Department of Biochemistry II, Faculty of Medicine, Oita University, Oita, Japan.
authors.forename: Takako
authors.initials: T
authors.lastname: Sasaki
authors.affiliation: Laboratory of the Biology and Pathology of the Eye, Institute of Inherited Metabolic Disorders and Department of Ophthalmology, First Faculty of Medicine, Charles University in Prague and General Teaching Hospital in Prague, Prague, Czech Republic.
authors.forename: Petra
authors.initials: P
authors.lastname: Liskova
authors.affiliation: Department of Ophthalmology, First Faculty of Medicine, Charles University in Prague and General Teaching Hospital in Prague, Prague, Czech Republic.
authors.forename: Michalis
authors.initials: M
authors.lastname: Palos
authors.affiliation: Laboratory of the Biology and Pathology of the Eye, Institute of Inherited Metabolic Disorders, First Faculty of Medicine, Charles University in Prague and General University Hospital in Prague, Prague, Czech Republic. katerina.jirsova@lf1.cuni.cz.
authors.forename: Katerina
authors.initials: K
authors.lastname: Jirsova
mesh_terms: D000328:Adult; D000368:Aged; D003094:Collagen / Q000378:metabolism / Q000648:ultrastructure; D003228:Conjunctiva / Q000201:enzymology / Q000473:pathology; D003261:Contact Lenses; D003315:Cornea / Q000201:enzymology* / Q000473:pathology; D003319:Corneal Stroma / Q000201:enzymology / Q000473:pathology / Q000648:ultrastructure; D004728:Endothelium, Corneal / Q000201:enzymology / Q000473:pathology / Q000648:ultrastructure; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000378:metabolism; D015603:Keratinocytes / Q000201:enzymology / Q000473:pathology / Q000648:ultrastructure; D007640:Keratoconus / Q000201:enzymology* / Q000473:pathology; D016850:Limbus Corneae / Q000201:enzymology / Q000473:pathology; D008297:Male; D008875:Middle Aged; D008249:Protein-Lysine 6-Oxidase / Q000378:metabolism*
chemical_list: D007527:Isoenzymes; D003094:Collagen; D008249:Protein-Lysine 6-Oxidase