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D.J. Brown, B. Holguin, N. Udar, M.C. Kenney; Distinct Splice Variants Of Nrg-1 and Sod1 Associated with Keratoconus . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1306.
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Purpose: Keratoconus (KC) is a leading indication for corneal transplantation affecting 1 in 2,000 individuals. Our previous studies suggested that both EGF signaling pathways and oxidative stress play a role in the disease process. In this study, we investigated the expression levels of several genes in these pathways. Methods: RNA from normal and KC stromal keratocyte cultures and/or tissues were assessed by RT-PCR for a number of gene products known to participate in EGF signaling pathways and oxidative stress. Two of these, representing heregulin alpha (an EGF family member derived from the NRG-1 gene) and superoxide dismutase (SOD1) displayed multiple specific products distinct from the normal samples. Primers were designed to encompass the coding regions of the mRNA, long accurate RT-PCR performed, and the products subcloned into plasmids. Direct sequencing of the plasmids was performed to characterize the transcripts. Results: Three distinct splice variants of SOD1 were isolated and identified from KC tissue, wild type SOD1, SOD1 lacking exon 2, and SOD1 lacking exons 2 and 3. Four variants derived from the NRG-1 gene were identified. Each of these were unique in that they have not been previously characterized. Conclusions: These data demonstrate that unusual splicing of NRG-1 occurs in corneal tissue and some of these variants are selectively expressed in KC tissue. SOD1 variants are also expressed in KC tissue. Currently, it is not clear if any of these variants are translated into protein in KC patients or whether these variants have biologic consequences. Using in vitro systems, some of these questions are being explored. Support: NIH EY06807; Schoellerman Charitable Foundation; Discovery Fund for Eye Research; Skirball Molecular Ophthalmology Program, and the Guenther Foundation.
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