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J.M. Johnson, J.A. Rada; SLRP Expression and Function in Human Sclera . Invest. Ophthalmol. Vis. Sci. 2003;44(13):415.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: We have previously shown that several members of the SLRP (small leucine rich proteoglycan) family, including decorin, PRELP (proline arginine rich end leucine-rich protein), fibromodulin, biglycan, chondroadherin, and lumican are expressed in the human sclera. The focus of the present study is to further characterize the expression and function of scleral PRELP and fibromodulin. Methods: Total RNA was extracted from the human sclera and mRNA levels of PRELP, decorin and fibromodulin were compared using competitive PCR analysis. Additionally, protein levels of PRELP were assessed in the sclera of human donors aged 2 – 84 years using western blot analysis with antibodies specific for human PRELP. In order to understand the function of the SLRPs in the human sclera recombinant PRELP and fibromodulin were expressed in a baculovirus system and used in in-vitro collagen fibrillogenesis assays. Results: Competitive PCR analysis confirmed the expression of PRELP to be 5 fold higher than the expression of decorin. By comparison, the expression of fibromodulin was approximately 70% lower than the expression of decorin. Western blot analysis of PRELP showed a decrease of PRELP protein concentration in the sclera from ages 2 –19 years followed by a relative increase in concentration from ages 19-84 years. Full length recombinant fibromodulin and PRELP have been generated using the baculovirus system. Preliminary in-vitro collagen fibrillogenesis studies have shown that purified recombinant fibromodulin inhibits type I collagen fibrillogenesis by 55% as compared to fibrillogenesis of collagen alone, indicating that these recombinant proteins retain their native biological activities. Studies involving recombinant PRELP are underway. Conclusions: These results suggest that SLRP proteoglycans are expressed at varying levels in the human sclera. The abundance of PRELP mRNA and protein in the human sclera, and the observed age-related variation in scleral PRELP suggests that this SLRP may play a critical role in the scleral extracellular matrix. Furthermore, the baculovirus system can be used to express functional human recombinant protein which will greatly help to elucidate the role of these SLRPs in scleral extracellular matrix structure and organization.
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