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Rossen M Hazarbassanov, Frans Eberth Costa Andrade, Joyce Covre, Myrna Serapião dos Santos, Mauro Silveira de Queiroz Campos, Jose Alvaro Pereira Gomes, Cristiane Damas Gil; Galectin-1 and -3 as prosprective biomarkers in keratoconus patients. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3909.
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© ARVO (1962-2015); The Authors (2016-present)
To evaluate the expression of galectin-1 (Gal-1) and -3 (Gal-3) in the keratoconus (KC) diagnosed patients and riboflavin and ultraviolet light effect on human keratocytes cultivated in vitro.
Tear fluid and conjunctival impression cytology specimens from control and KC patients with and without atopy (n = 10/group) were used to evaluate Gal-1 and Gal-3 expression by immunofluorescence and ELISA. Primary human keratocytes were isolated by digestion in collagenase from surgically removed corneas of five normal human corneal buttons and cultured in DMEM/Ham's F12 medium supplemented with 2 % fetal bovine serum. These cells were evaluated under two experimental conditions: control and submitted to the application of UVA light and riboflavin 0.1% for 30 minutes called cross-linking (CXL). After 24 hours of CXL, expression of Gal-1 and Gal-3 in the and their supernatants was performed using immunofluorescence and ELISA; multiplex assay was also done to detect inflammatory biomarkers in supernatants. Student’s t-test was used for statistical analysis.
Atopic KC patients exhibited increased levels of Gal-1 in the tear fluids (14.5±3 ng/mL, p>0.05) and epithelial cells of bulbar conjunctiva (130±3 arbitrary units, p<0.001) compared to control (9.8±0.5 and 87±1.6, respectively). On the other hand, non-atopic KC patients exhibited increased levels of Gal-3 in the tear fluid (70±16 ng/mL, p<0.05), but decreased endogenous expression in conjunctival epithelial cells. In in vitro experiments, keratocytes exhibited intense nuclear Gal-1 expression while Gal-3 was localized especially in the cytoplasm and both were strongly diminished after 24 hours of CXL. Further, CXL induced significant release of Gal-1 in the cell supernatants (116±18 ng/mL, p<0.05) and decreased inflammatory biomarkers as IL-6, IL-8, MMP-2 and MMP-9. Gal-3 levels was not detected in the keratocyte supernatants
Gal-1 and Gal-3 represent new interesting biomarkers as revealed by their different expression patterns in atopic and non-atopic KC patients. CXL has immunosuppressive effect on keratocytes by reducing the release of cytokines and MMPs and increased anti-inflammatory protein Gal-1.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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