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      Matrix Metalloproteinase 2 Activation in Cultured Corneas

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          Abstract

          Purpose: Corneas that are maintained in tissue culture medium shed their epithelial cells and repopulation following graft surgery is an essential facet of the healing process. Failure to do so may be a result of structural damage to the epithelial basement membrane of a donor cornea. The purpose of the present investigation was to ascertain whether MMP-2, the matrix metalloproteinase produced by corneal keratocytes, may be activated during storage and hence cleave the type IV collagen component of the epithelial cell basement membrane. Methods: Fresh and transplant rejected corneas that had been stored in culture medium for varying time periods and of known donor age were collected. The soluble protein fractions of these corneas were obtained. Their MMP-2 proteins were visualised by zymography on SDS gelatin polyacrylamide gels and assayed for activity against nitrophenyl acetate and denatured [<sup>3</sup>H]type I collagen. Results: The stromal tissue of fresh, normal corneas produced inactive MMP-2 of M<sub>r</sub> 66,000. Although the cultured corneas did not up-regulate MMP-2 production, they contained additional MMP-2 activities of M<sub>r</sub> 62,000 and M<sub>r</sub> 43,000. The appearance of these additional MMP-2 activities correlated with corneal culture time but not donor age. The ability to cleave denatured [<sup>3</sup>H]type I collagen correlated with the appearance of the M<sub>r</sub> 43,000 activity but not the M<sub>r</sub> 62,000 activity. Conclusion: Activated MMP-2 is produced in cultured corneas. For this reason the corneas donated for all graft procedures should not be held in culture medium for periods exceeding 4 weeks.

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          The Soluble Catalytic Domain of Membrane Type 1 Matrix Metalloproteinase Cleaves the Propeptide of Progelatinase A and Initiates Autoproteolytic Activation

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            Reorganization of polymerized actin: a possible trigger for induction of procollagenase in fibroblasts cultured in and on collagen gels

             Z Werb,  EN Unemori (1986)
            Changes in cell shape are postulated to modulate gene expression during differentiation of a number of cell types, including rabbit synovial fibroblasts, which are inducible for expression of the zymogen form of the metalloendopeptidase, collagenase. In the work presented here, fibroblasts cultured on and within hydrated collagen gels were allowed to contract by release of the gels from the sides of the culture dish. Within 24 h of cell release, synthesis and secretion of procollagenase was initiated in the absence of any chemical manipulation. Fibroblasts grown in and on collagen also responded to 12-O-tetradecanoylphorbol-13- acetate and cytochalasin B with morphologic change and induced procollagenase. However, colchicine, which altered morphology to varying degrees in cells on plastic, on collagen, and within collagen gels, did not induce procollagenase expression. In all cases, the enzyme was induced only after reorganization of polymerized actin, rather than after a change in cellular morphology per se. As a first approach to identifying other aspects of the stimulated phenotype that could affect collagen turnover, the expression of collagen and endogenous metalloproteinase inhibitors in relation to procollagenase secretion was investigated. Collagen secretion by fibroblasts decreased when procollagenase secretion was induced by the pharmacologic agents, but not when cells were stimulated by contraction on or within collagen gels. The expression of two endogenous inhibitors was not coordinately regulated with induction of procollagenase. Therefore, the extracellular matrix and the cellular actin cytoskeleton may transduce signals that modulate the tissue remodeling phenotype of fibroblasts.
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              Author and article information

              Journal
              ORE
              Ophthalmic Res
              10.1159/issn.0030-3747
              Ophthalmic Research
              S. Karger AG
              0030-3747
              1423-0259
              2001
              February 2001
              13 December 2000
              : 33
              : 1
              : 1-6
              Affiliations
              Division of Ophthalmology, Bristol Eye Hospital, University of Bristol, UK
              Article
              55634 Ophthalmic Res 2001;33:1–6
              10.1159/000055634
              11114598
              © 2001 S. Karger AG, Basel

              Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher. Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug. Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.

              Page count
              Figures: 2, Tables: 3, References: 21, Pages: 6
              Categories
              Original Paper

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