Abstract
Purpose: :
Previous proteomic studies from our lab showed that the matricellular protein osteopontin (OPN) was found in human aqueous humor (AH) at levels comparable to or higher than in serum. To investigate the expression pattern of OPN in the human eye, we examined OPN levels in AH and human eye tissues from primary open angle glaucoma (POAG) and control specimens.
Methods: :
OPN was quantified in AH samples by ELISA. Values were expressed as mean ± standard deviation. AH was obtained from 1) donor eyes and 2) surgical samples collected from POAG or elective cataract removal (control) patients. OPN expression and localization in eye tissue sections and cultured trabecular meshwork cells were studied by immunohistochemistry, Western blot, and real-time PCR. The glycosylation pattern of OPN was studied using sialidase and several glycosidases.
Results: :
OPN levels were significantly reduced in donor glaucoma AH when compared to control (0.54 ± 0.18 ng/µg, n=8, vs. 0.77 ± 0.23 ng/µg, n=9, p=0.0388). A similar trend was observed in surgical AH (1.05 ± 0.31 ng/µg, n=20, vs. 1.43 ± 0.88 ng/µg, n=20, p=0.0826). OPN was present in the trabecular meshwork, corneal epithelium and endothelium, iris, ciliary body, retina, vitreous humor, and anterior and posterior optic nerve. Individual eye tissues showed unique OPN fingerprints with differences noted between normal and POAG. Digestion with sialidase and glycosidases showed that OPN is extensively glycosylated in AH but not in tissue lysates from optic nerve or cell lysates from cultured trabecular meshwork cells.
Conclusions: :
OPN protein levels are decreased in POAG AH. OPN is strongly expressed in the eye, with some tissues having unique OPN profiles between POAG and normal. Variation in OPN protein profile in AH was due in part to glycosylation.
Keywords: aqueous • trabecular meshwork • immunohistochemistry