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A. V. Ljubimov, M. Saghizadeh, A. A. Kramerov, Y. Yaghoobzadeh, M. G. Castro; Effects of Adenovirus-Driven Proteinase Overexpression on the Patterns of Diabetic Corneal Markers in Organ-Cultured Normal Human Corneas. Invest. Ophthalmol. Vis. Sci. 2009;50(13):5197.
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Our previous studies have implicated matrix metalloproteinase-10 (MMP-10) and cathepsin F (CTSF) in corneal basement membrane and integrin changes occurring in diabetic corneas. The purpose was to examine the patterns of these markers in normal human organ-cultured corneas upon transduction of MMP-10 and CTSF using recombinant adenoviral (rAV) vectors.
Five pairs of normal organ-cultured autopsy human corneas were transduced with rAV expressing full-length CTSF (rAV-CTSF) or MMP-10 (rAV-MMP-10) genes under cytomegalovirus promoter. Viral constructs at 108 plaque forming units per cornea were added to culture medium for 72 hr. The fellow corneas of each pair received control rAV harboring vector alone. After additional 6 days incubation, corneas were processed for immunohistochemistry or tested for healing of 5-mm circular epithelial wounds caused by topical n-heptanol application.
By immunohistochemistry, all corneas transduced with rAV-CTSF and rAV-MMP-10 had increased epithelial staining of respective proteins compared to fellow corneas transduced with control rAV. Diabetic markers, especially integrin 3β1, and basement membrane components nidogen-1 and nidogen-2, showed decreased and irregular staining upon proteinase transduction. Phosphorylated Akt was decreased in proteinase-transduced corneas, whereas phosphorylated ERK and p38MAPK did not change compared to controls. No change was detected in the expression of tight junction proteins, ZO-1 and claudin-1. There was a tendency to slower wound healing of normal corneas after proteinase overexpression.
The data support the role of MMP-10 and CTSF in abnormal marker patterns and impaired wound healing observed in diabetic corneas. Suppression of proteinase expression in diabetic corneas may provide structural and functional benefit to these corneas and alleviate symptoms of diabetic keratopathy.
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