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H Ando, S S Twining, B Y Yue, X Zhou, M E Fini, T Kaiya, E J Higginbotham, J Sugar; MMPs and proteinase inhibitors in the human aqueous humor.. Invest. Ophthalmol. Vis. Sci. 1993;34(13):3541-3548.
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PURPOSE: This study was performed to examine the gelatinolytic and caseinolytic activities and the levels of two proteinase inhibitors, alpha 1-proteinase inhibitor (alpha 1-antitrypsin) and alpha 2-macroglobulin, in the human aqueous humor. METHODS: Aqueous humor samples were collected during elective surgery in patients with cataracts. Zymography with gelatin- and casein-containing gels was performed. The inhibitors were examined by Western blot analyses, enzyme-linked immunosorbent assay, and dot blot assays. RESULTS: The aqueous humor contained a major band of gelatinolytic activity at a molecular weight of 66 kD and minor bands at 125, 95, and 62 kD. These gelatinases were inhibited by 10 mM ethylenediaminetetraacetic acid (EDTA) or 1,10-phenanthroline. After extended incubation (48 hours), zymography on casein-containing gels showed proteinase bands with molecular weights in the 80- to 84-kD range. Additional bands at 68 and 48 kD also were observed. All the caseinase activities were inhibited by 10 mM phenylmethylsulfonyl fluoride and 1 microgram/ml aprotinin. No inhibition was observed with 5 mM EDTA, 5 microM E-64, or 1 microM pepstatin. These results indicated that the caseinases are serine proteinases. Western blot analysis showed a 53-kD alpha 1-proteinase inhibitor band in the aqueous humor. The concentration was 32.2 +/- 9.9 micrograms/ml, constituting approximately 15% of the total protein. A 360-kD protein band immunoreactive to anti-alpha 2-macroglobulin also was detected. Its level in the aqueous humor was 3.2 +/- 1.3 micrograms/ml. CONCLUSIONS: The gelatinases, serine-like proteinases, and proteinase inhibitors found in the aqueous humor may participate in the remodeling of extracellular matrices in the trabecular meshwork and other tissues bordering the anterior chamber.
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