title: Alteration of galectin-3 in tears of patients with dry eye disease.
abstract: OBJECTIVE
  To investigate the expression, release, and proteolytic degradation of galectin-3 in patients with dry eye disease.
  
  
  METHODS
  Observational case series with a comparison group.
  
  
  METHODS
  Tear washes and conjunctival impression cytology specimens were collected through standard procedures from 16 patients with dry eye and 11 age-matched healthy subjects. Galectin-3 content in tears was analyzed by quantitative Western blot, using recombinant galectin-3 protein to generate a calibration curve. The relative expression of galectin-3 and matrix metalloproteinase 9 (MMP9) was evaluated by quantitative polymerase chain reaction. The cleavage of galectin-3 was studied in vitro using activated recombinant MMP9 and protease inhibitors.
  
  
  RESULTS
  The concentration of galectin-3 protein in tears, but not galectin-3 expression in conjunctival epithelium, was significantly higher in tears of patients with dry eye (0.38 ng/μg total protein, range 0.04-1.36) compared to healthy subjects (0.12 ng/μg total protein, range 0.00-0.41) (P < .01). By Western blot, an intact (∼28.0 kDa) galectin-3 band was identified in tear samples from healthy subjects, whereas 50% of the dry eye samples were characterized by the additional presence of a partially degraded form (∼25.4 kDa). In our experiments, elevated expression of MMP9 in dry eye subjects correlated with the ability of active MMP9 to cleave galectin-3 from recombinant origin. Interestingly, cleavage of endogenous galectin-3 in tear samples was impaired using a broad-spectrum proteinase inhibitor cocktail, but not the pan-specific MMP inhibitor GM6001, suggesting the presence of proteases other than MMPs in promoting galectin-3 degradation in dry eye.
  
  
  CONCLUSIONS
  Our results indicate that release of cellular galectin-3 into tears is associated with epithelial dysfunction in dry eye, and that galectin-3 proteolytic cleavage may contribute to impaired ocular surface barrier function.
authors.affiliation: Schepens Eye Research Institute and Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Uchino
authors.affiliation: Schepens Eye Research Institute and Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Jerome
authors.initials: J
authors.lastname: Mauris
authors.affiliation: Schepens Eye Research Institute and Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Ashley M
authors.initials: AM
authors.lastname: Woodward
authors.affiliation: Schepens Eye Research Institute and Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Julia
authors.initials: J
authors.lastname: Dieckow
authors.affiliation: Schepens Eye Research Institute and Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Francisco
authors.initials: F
authors.lastname: Amparo
authors.affiliation: Schepens Eye Research Institute and Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Reza
authors.initials: R
authors.lastname: Dana
authors.affiliation: Campus Bio Medico University of Rome, Rome, Italy.
authors.forename: Flavio
authors.initials: F
authors.lastname: Mantelli
authors.affiliation: Schepens Eye Research Institute and Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts. Electronic address: pablo_argueso@meei.harvard.edu.
authors.forename: Pablo
authors.initials: P
authors.lastname: Argüeso
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001798:Blood Proteins; D015153:Blotting, Western; D003228:Conjunctiva / Q000378:metabolism; D015352:Dry Eye Syndromes / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004847:Epithelial Cells / Q000378:metabolism; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D005260:Female; D037502:Galectin 3 / Q000235:genetics / Q000378:metabolism*; D037161:Galectins; D006801:Humans; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D013666:Tears / Q000378:metabolism*
chemical_list: D001798:Blood Proteins; D005136:Eye Proteins; D037502:Galectin 3; D037161:Galectins; C428445:LGALS3 protein, human; D012333:RNA, Messenger; C042434:tear proteins; C579270:MMP9 protein, human; D020780:Matrix Metalloproteinase 9