ROLE OF THE MATRIXIN MMP-2 IN MULTICELLULAR ORGANIZATION OF ADIPOCYTES CULTURED IN BASEMENT-MEMBRANE COMPONENTS

Primary rat adipocytes cultured in basement membrane component gels mi grated and organized into large, three-dimensional, multicellular clus ters. Gross morphological changes seen during this reorganization are described. The rate of cluster formation decreased with age of the rat s and was stimulated by insulin in older, but not in younger rats. Ech istatin, a disintegrin, partially inhibited the formation of multicell ular clusters in a concentration-dependent fashion (50% inhibitory con centration approximate to 10 nM). The original extracellular matrix wa s initially remodeled and eventually destroyed by the time large multi cellular clusters were observed. This implied that one or more matrix- degrading protease(s) were being secreted. Adipocyte-conditioned mediu m was found to contain a divalent cation-sensitive gelatinase activity at similar to 72 and/or similar to 62 kDa. The elution profile of thi s activity from gelatin-Sepharose 4B was similar to matrix metalloprot einase 2 (MMP-2, a 72-kDa matrixin with a 62-kDa mature form), and the dimethyl sulfoxide eluant from these columns contained MMP-2 immunore activity. MMP-2 concentration and activity were greater in conditioned medium from young than from older animals; however, insulin did not a ffect the amount of MMP-2 in adipocyte-conditioned media. The matrixin inhibitor 1,10-phenanthroline not only blocked gelatinase activity in zymograms but also prevented extracellular matrix remodeling and dest ruction, as well as adipocyte migration and the formation of cell-cell contacts in adipocyte cultures. These observations are consistent wit h the hypothesis that the matrixin MMP-2 is secreted by adipocytes. Wh ereas matrixin activity alone may not be sufficient for the formation of multicellular clusters, the data indicate that it may have a requis ite role in this process.