Corneal scars exhibit a disruption of stroma collagen fibril diameter and spacing that contributes to loss of transparency. These alterations are closely associated with changes in stromal glycosaminoglycans, acidic polysaccharides that form bridges between adjacent collagen fibrils. This project will examine the hypothesis that altered transcription of genes involved in elongation and sulfation of glycosaminoglycans generates the abnormal glycosaminoglycans of scar tissue and is directly responsible for disruption of stromal transparency. In the past two years genes coding for the enzymes involved in elongation and sulfation of corneal glycosaminoglycans have been identified. At the same time, we have developed and characterized cultures of primary keratocytes that maintain a glycosaminoglycan expression profile similar to that of normal cornea but which can be stimulated to secrete glycosaminoglycans resembling those of scar tissue. These developments present the opportunity to directly test the role of glycosaminoglycans in stromal transparency. We will address the hypothesis in three stages: (1) Identify the genes involved in generating normal and fibrotic stromal glycosaminoglycans. (2) Demonstrate that altered glycosaminoglycan biosynthesis results from transcriptional regulation of biosynthetic genes under the control of gene-specific promoter sequences. (3) Characterize the effects of corneal-specific overexpression of chondroitin sulfotransferase and synthase in vivo.
This third aim will generate transgenic mice in which corneal chondroitin sulfate overexpression is inducible. Corneal transparency cellularity and collagen fibril organization of the mice will be documented. Together these three aims will establish which genes are involved in mediating the corneal glycosaminoglycan phenotype and how these genes are controlled, and will show that inappropriate expression of these genes leads to tissue changes similar to those in scars. These experiments will link the long held idea of the importance of glycosaminoglycan in corneal transparency with the expression patterns of specific genes. These results may be relevant to development of therapeutic approaches in the cornea as well as other tissues.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY009368-14
Application #
7104959
Study Section
Visual Sciences A Study Section (VISA)
Program Officer
Shen, Grace L
Project Start
1993-08-01
Project End
2008-03-31
Budget Start
2006-08-01
Budget End
2008-03-31
Support Year
14
Fiscal Year
2006
Total Cost
$238,078
Indirect Cost
Name
University of Pittsburgh
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Syed-Picard, Fatima N; Du, Yiqin; Hertsenberg, Andrew J et al. (2018) Scaffold-free tissue engineering of functional corneal stromal tissue. J Tissue Eng Regen Med 12:59-69
Syed-Picard, Fatima N; Du, Yiqin; Lathrop, Kira L et al. (2015) Dental pulp stem cells: a new cellular resource for corneal stromal regeneration. Stem Cells Transl Med 4:276-85
Wu, Jian; Du, Yiqin; Mann, Mary M et al. (2014) Corneal stromal stem cells versus corneal fibroblasts in generating structurally appropriate corneal stromal tissue. Exp Eye Res 120:71-81
Wu, Jian; Rnjak-Kovacina, Jelena; Du, Yiqin et al. (2014) Corneal stromal bioequivalents secreted on patterned silk substrates. Biomaterials 35:3744-55
Karamichos, Dimitrios; Funderburgh, Martha L; Hutcheon, Audrey E K et al. (2014) A role for topographic cues in the organization of collagenous matrix by corneal fibroblasts and stem cells. PLoS One 9:e86260
Chan, Audrey A; Hertsenberg, Andrew J; Funderburgh, Martha L et al. (2013) Differentiation of human embryonic stem cells into cells with corneal keratocyte phenotype. PLoS One 8:e56831
Roh, Danny S; Du, Yiqin; Gabriele, Michelle L et al. (2013) Age-related dystrophic changes in corneal endothelium from DNA repair-deficient mice. Aging Cell 12:1122-31
Boote, Craig; Du, Yiqin; Morgan, Sian et al. (2012) Quantitative assessment of ultrastructure and light scatter in mouse corneal debridement wounds. Invest Ophthalmol Vis Sci 53:2786-95
Roh, Danny S; Funderburgh, James L (2011) Rapid changes in connexin-43 in response to genotoxic stress stabilize cell-cell communication in corneal endothelium. Invest Ophthalmol Vis Sci 52:5174-82
Du, Yiqin; Roh, Danny S; Funderburgh, Martha L et al. (2010) Adipose-derived stem cells differentiate to keratocytes in vitro. Mol Vis 16:2680-9

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