Date of Award

2-24-2003

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

First Advisor

Jody A Rada

Abstract

The collagenous network of the extracellular matrix (ECM) contributes to the strength and organization of connective tissues. Interspersed among this collagenous network, are a number of tissue specific proteoglycans (PG's) that are thought to guide matrix assembly and organization through protein/protein and/or protein/carbohydrate interactions. In recent literature, attention has been focused on a small leucine rich PG family (SLRPS) found to bind a variety of ECM components via their core proteins. The family is made up of nine members including: decorin, biglycan, lumican, fibromodulin, proline arginine end leucine rich repeat protein (PRELP), keratocan, chondroadherin, epiphycan, and osteoglycin. Recent studies have demonstrated that the decorin, lumican, and fibromodulin interactions with type I collagen have an effect on fibril shape, diameter, and arrangement (Vogel et al., 1984; Hedbom 1993; Rada et al., 1993). These results are reminiscent of changes occurring within the ECM of the myopic sclera, in which type I collagen of highly myopic humans display abnormal morphology diameter, and arrangement (Curtin et al., 1979). We hypothesize that the changes occurring within the ECM of the highly collagenous human sclera are, in part, regulated by SLRP interaction with a variety of ECM components. In order to investigate the role of the SLRPs in the human sclera we characterized the expression levels of SLRP mRNA in the human sclera through semi-quantitative PCR analysis and quantitative competitive PCR analysis. Based upon the results of quantitative PCR analysis, the protein expression and distribution of the SLRP family member, PRELP, was verified in the human sclera through western blot detection and immunohistochemistry. In order to assess the function of SLRPs in the human sclera, recombinant human PRELP and fibromodulin were produced in a baculovirus system and used for in vitro collagen fibrillogenesis and cell attachment assays.

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