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AM McDermott, RJ Proske, HM Woo, S Campbell, CJ Murphy; Rat Beta-Defensin-2 is Upregulated During Corneal Re-epithelialisation In Vivo . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4198.
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Purpose: Defensins are antimicrobial peptides which may also act as signalling molecules, regulating events such as cell migration and proliferation. Previously (Curr. Eye Res. 2001, 22, 64-67) we have shown that human ß-defensin (hBD)-1 is constitutively expressed whilst hBD-2 is upregulated in regenerating corneal epithelium in organ culture. Here we have investigated if similar events occur in vivoby studying the expression of the rat defensin homologues rat ß-defensin (rBD)-1 and rBD-2 during corneal epithelial wound healing. Methods: 12 male SD rats were anaesthetised and the corneal epithelium was removed limbus to limbus from each eye and snap frozen (original epithelial samples). At 24, 48, 72 hrs and 1 week after the injury 3 rats were killed and the regrown epithelium collected from each eye by scraping (regrown samples). Preliminary studies showed no difference in defensin expression between OS and OD so samples were pooled, RNA was extracted and RT-PCR performed using specific primers for rBD-1, rBD -2 and the house keeping gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH). RNA from rat kidney and lung acted as positive controls for rBD-1 and rBD-2 respectively. Results: rBD-1 mRNA was detected in all epithelial samples. Its expression was consistent and not significantly different between original and regrown samples. rBD-2 mRNA was detected in all control samples although its expression was low and variable compared to that of rBD-1. All regrown epithelial samples showed a marked upregulation in rBD-2 expression. Semiquantitative analysis relative to GAPDH showed a 103%, 104%, 117% and 78% increase in rBD-2 mRNA expression at 24, 48, 72 hrs and 1 week respectively. Conclusion: rBD-1 is constitutively expressed by the rat corneal epithelium whereas rBD-2 is normally present at low levels but is markedly upregulated in the regenerating epithelium. These data support our previous in vitro observations on human ß-defensins. Inducible ß-defensins likely give greater protection against infection after injury and may directly influence the wound healing process.
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