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MS Katar, Z Dong, RS Berk; Expression of Tumor Necrosis Factor-alpha Convertase in Mouse Corneas . Invest. Ophthalmol. Vis. Sci. 2002;43(13):36.
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Purpose: This study was designed to demonstrate the expression of tumor necrosis factor-alpha convertase (TACE, ADAM-17) in uninfected corneas and corneas infected with Pseudomonas aeruginosa. Of the 30 sheddases that are known, only TACE has been studied enzymatically. It is a membrane bound disintegrin metalloproteinase that processes the membrane-associated proTNF-a to a soluble form and plays a role in inflammatory processes. Methods: C57BL/6J mice were intracorneally infected with P. aeruginosa. Corneal samples were harvested, washed and pooled for various time samples ranging from 0 to 8 days post-infection. Semi-quantitative RT-PCR was used to demonstrate TACE gene expression and immunoblot analysis was used to demonstrate protein expression. Enzyme activity was detected by use of the fluorogenic substrates Q45 and Q48. Results: Semi-quantitative RT-PCR for TACE gene expression indicated an upregulated expression of TACE over a 0 to 8 day time period with maximum expression by day 8 while corneas from immunized mice exhibited a lower and shorter expression. Immunoblotting indicated the expression of IL-1, IL-1 receptor, IL-6, IL-6 receptor, TNF-a, TNF-a receptors 1 and 2 (P80 and P60) and trance. Enzymatic activity was detected fluorometrically using the fluorogenic TACE substrates Q45 and Q48. Conclusion: TACE activity was constitutively expressed in uninfected mouse corneas and was upregulated during corneal infection with P. aeruginosa.
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