EXPRESSION OF FIBRONECTIN ISOFORMS IN RAT CORNEA AFTER AN EPITHELIAL-SCRAPE WOUND

The polymerase chain reaction (PCR) technique was used to analyze the presence or absence of the EIIIB and V region fibronectin (FN) mRNA is oforms, generated by alternative splicing of the FN primary mRNA trans cript during epithelial wound healing, in a rat cornea wound model. A central 4mm area of the cornea was denuded of epithelium. At 5, 15, 30 , 45 minutes, 1, 2, 4, 8, 24, 48 hours, and 4 days post-wounding, the rats were euthanized and a 6mm diameter full thickness corneal biopsy was performed. cDNA was synthesized from extracted RNA, and specific F N sequences were amplified by PCR as directed by different sets of 5' and 3' primers specific for the alternatively spliced EIIIB and V regi on domains. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA expr ession was used to normalize the data. PCR products were analyzed by a garose gel electrophoresis and the identity of the bands were confirme d by direct nucleotide sequencing. The alternatively-spliced EIIIB and all three alternatively. spliced isoforms of V domain FN mRNA were ob served post-wounding. Similarly, in normal tissue, EIIIB and all three V region alternatively spliced FN isoforms were expressed. These find ings suggest that in situ synthesis of cellular associated EIIIB FN an d FN V domain may play a role in corneal wound healing.