Characterization of Suprachoroidal Fluid in Chick Eyes Recovering From Myopia

Abstract

Abstract: : Purpose:In order to identify mechanisms involved in the regulation of visually guided ocular growth, the composition of suprachoroidal fluid was evaluated in control eyes and eyes recovering from induced myopia. Methods:Form-deprivation myopia was induced in the right eye of newly hatched chicks by the application of translucent ocluders. Following 10 days of deprivation, occluders were removed and chicks were provided unrestricted vision for an additional 3 days (recovery). At the end of the recovery period, chicks were sacrificed and suprachoroidal fluid was removed from enucleated control and recovering eyes with a 10 µl Hamilton syringe with the aid of a dissection microscope. Suprachoroidal fluid was fractioned using molecular weight cut-off filters. Punches of normal chick sclera were incubated with various fractions of suprchoroidal fluid in the presence of DMEM containing ₃₅SO₄ (100 µCi/ml) at 37°C for 24 hours and scleral proteoglycan synthesis was determined by measuring ₃₅SO₄ incorporation into CPC-precipitable glycosaminoglycans. Fractions of suprachoroidal fluid were further analyzed by MALDI-TOF mass spectrometry. Results:Suprachoroidal fluid from recoverying eyes was shown to inhibit proteoglycan synthesis as compared with fluid from control eyes (p < 0.01). Significant differences in proteoglycan inhibition between control and recovering eyes were present in fractions of suprachoroidal fluid < 10 kD, while no differences in activities were observed in fractions ≷ 10 kD. Further fractionation of the < 10 kD sample revealed significant inhibitory activity at < 3 kD in both recovering and control samples. MALDI-TOF analyses of the < 10 kD fraction demonstrated a cluster of peaks at m/z 4717 in recovering samples not present in the control samples, while analyses of the < 3 kD fraction demonstrated peaks at m/z 1551- 2000 in both control and recoverying samples. Conclusion:Our results indicate that suprachoroidal fluid isolated from recovering eyes significantly inhibits scleral proteoglycan synthesis and contains moieties not present in control suprachoroidal fluid. Current studies are underway to identify the nature of the 4717 m/z compound(s) in suprachoroidal fluid of recovering eyes, as they may be responsible for the scleral proteoglycan inhibitory activity.

Keywords: 481 myopia • 345 choroid • 529 proteoglycans/glycosaminoglycans