DIFFERENTIAL EXPRESSION OF UROKINASE-TYPE PLASMINOGEN-ACTIVATOR (UPA), ITS RECEPTOR (UPA-R), AND INHIBITOR TYPE-2 (PAI-2) DURING DIFFERENTIATION OF KERATINOCYTES IN AN ORGANOTYPIC COCULTURE SYSTEM

Cultured keratinocytes resemble migrating keratinocytes under conditio ns of reepithelialization during wound healing, Such keratinocytes exp ress urokinase-type plasminogen activator (uPA) and its specific recep tor (uPA receptor), Receptor-bound uPA activates plasminogen, thus pro viding plasmin for pericellular proteolysis. uPA is regulated by the p lasminogen activator inhibitors PAI-1 and PAI-2. As indicated by immun ohistology, neither uPA nor uPA receptor is expressed in normal epider mis. Thus, the down-regulation of uPA and uPA-receptor expression in k eratinocytes appears to be an important event in epidermal healing and restoration of a normal epidermal tissue architecture. We have addres sed this matter by using a culture and differentiation system for kera tinocytes in vitro. Keratinocytes were grown in organotypic cocultures for 4, 7, and 14 days, Frozen sections were analyzed with indirect im munofluorescence staining and overlay zymography, the latter detecting activity of plasminogen activators. While tPA and PAI-1 stainings wer e consistently negative over the entire observation period, uPA and uP A receptor were expressed by basal keratinocytes at Days 4 and 7, but not at Day 14. Accordingly, overlay zymography revealed uPA activity a t Days 4 and 7. PAI-2 was found throughout the entire observation peri od, but with varying distribution: at Days 4 and 7 all suprabasal kera tinocytes stained positive for PAI-2, At Day 14, PAI-2-specific staini ngs were confined to the uppermost cells of the stratum spinosum, Our data demonstrate that uPA and uPA receptor, which are up-regulated in cultured keratinocytes, are down-regulated upon restoration of an epid ermis-like structure. The distribution of PAI-2 varied over the observ ation period and at Day 14 resembled the distribution of PAI-2 in norm al epidermis, Taken together, keratinocytes in organotypic coculture b ehave like keratinocytes in healing wounds in vivo with respect to the expression of the plasminogen activator system. (C) 1995 Academic Pre ss, Inc.