Our long-term objective in this proposal is to evaluate the hypothesis that the sialated macromolecules of the extracellular matrix in the trabecular meshwork/Schlemm's canal (TM/SC) system have a major role in determining the resistance to the outflow of aqueous humor in normal and glaucomatous eyes.
The specific aims are: (1) To document changes, as a function of age, in the amounts and glycoforms of fibronectin, thrombospondin, and a 140 kD glycoprotein (140 kD-GP); these are 3 of 7 sialated molecules that we have identified in the TM/SC system. (2) To investigate the effect of fibronectin and thrombospondin on cytokinesis of trabecular cells in vitro as function of the amount and activity of tissue plasminogen activator (tPA) produced by these cells. (3) To characterize the recently discovered 56 kD sialated glycoprotein (Cort-GP) induced by the administration of -corticosteroids to trabecular cells in vitro. (4) To document the changes that occur in sialated macromolecules of the aqueous outflow pathway in eyes with. glaucomatous disease, with particular emphasis on primary open-angle glaucoma (POAG). The experimental design is to use microanalytical methods that will enable us to study selected macromolecules in the TM/SC system. We will utilize the observed changes in sialic acid moieties in aging eyes and in eyes with POAG as a means for obtaining more information about the structure and expression of specific polypeptides and RNA molecules that are involved in the normal and abnormal functioning of the TM/SC system. The methods to be used are: (1) isoelectric focusing, lectin and antibody affinities, PCR amplification and quantitative analysis to detect changes in the amounts and glycoforms of fibronectin, thrombospondin, and 140 KD-GP; (2) cell culture, ELISA, and a chromogenic substrate assay to determine the effect of fibronectin and thrombospondin on trabecular cells as a function of the amount of tPA produced by these cells; (3) SDS-PAGE, lectin affinity, as well as isolation of polysomes and mRNA to characterize Cort-GP; (4) immunohistochemistry and immunoblotting, with antisera against fibronectin, thrombospondin, type VI collagen, and Cort-GP to document changes that occur in these glycoproteins in tissue from glaucomatous eyes.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
2R01EY003747-09A1
Application #
3258177
Study Section
Visual Sciences A Study Section (VISA)
Project Start
1981-07-01
Project End
1994-08-31
Budget Start
1991-09-01
Budget End
1992-08-31
Support Year
9
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of Chicago
Department
Type
Schools of Medicine
DUNS #
225410919
City
Chicago
State
IL
Country
United States
Zip Code
60637
Li, J; Tripathi, B J; Tripathi, R C (2000) Modulation of pre-mRNA splicing and protein production of fibronectin by TGF-beta2 in porcine trabecular cells. Invest Ophthalmol Vis Sci 41:3437-43
Tripathi, R C; Borisuth, N S; Li, J et al. (1997) Quantitative characterization of high- and low-affinity binding sites for basic fibroblast growth factor on trabecular cells of the eye. Exp Eye Res 64:335-41
Tripathi, B J; Li, T; Li, J et al. (1997) Age-related changes in trabecular cells in vitro. Exp Eye Res 64:57-66
Gong, H; Tripathi, R C; Tripathi, B J (1996) Morphology of the aqueous outflow pathway. Microsc Res Tech 33:336-67
Tripathi, B J; Hansen, M; Li, J et al. (1994) Identification of type VI collagen in the trabecular meshwork and expression of its mRNA by trabecular cells. Exp Eye Res 58:181-7
Tripathi, B J; Tripathi, R C; Yang, C et al. (1991) Synthesis of a thrombospondin-like cytoadhesion molecule by cells of the trabecular meshwork. Invest Ophthalmol Vis Sci 32:181-8
Tripathi, R C; Borisuth, N S; Tripathi, B J (1991) Mapping of Fc gamma receptors in the human and porcine eye. Exp Eye Res 53:647-56
Raja, S C; Tripathi, B J; Tripathi, R C (1991) A method for the rapid detection of single base polymorphisms in genomic DNA. Biotechniques 10:725
Tripathi, B J; Tripathi, R C; Wong, P et al. (1990) Expression of HLA by the human trabecular meshwork and corneal endothelium. Exp Eye Res 51:269-76
Tripathi, R C; Millard, C B; Tripathi, B J et al. (1990) Tau fraction of transferrin is present in human aqueous humor and is not unique to cerebrospinal fluid. Exp Eye Res 50:541-7

Showing the most recent 10 out of 31 publications