No abstract

The matrix metalloproteinase stromelysin-2 is expressed in keratinocytes of the epithelial tongue of skin wounds, suggesting a role in keratinocyte migration. Here, we show that stromelysin-2 enhances migration of cultured keratinocytes. To gain insight into the in vivo activities of stromelysin-2 in epithelial repair, we generated transgenic mice expressing a constitutively active stromelysin-2 mutant in keratinocytes. These animals had no alterations in skin architecture, and the healing rate of skin wounds was normal. Histologically, however, we found abnormalities in the organization of the wound epithelium. Keratinocytes at the migrating epidermal tip were scattered in most sections of mice with high expression level, and there was a reduced deposition of new matrix. In particular, the staining pattern of laminin-5 at the wound site was altered. This may be due to proteolytic processing of laminin-5 by stromelysin-2, because degradation of laminin-5 by this enzyme was observed in vitro. The inappropriate matrix contact of keratinocytes was accompanied by aberrant localization of ␤1-integrins and phosphorylated focal adhesion kinase, as well as by increased apoptosis of wound keratinocytes. These results suggest that a tightly regulated expression level of stromelysin-2 is required for limited matrix degradation at the wound site, thereby controlling keratinocyte migration. INTRODUCTIONProteolytic degradation of matrix proteins occurs in a variety of events that require tissue reorganization, such as embryonic development, wound healing, and cancer progression (Vu and Werb, 2000). Besides proteinases from the serine and cysteine family, matrix metalloproteinases (MMPs) are major players in these processes. Acting on specific protein substrates, these enzymes serve numerous and diverse functions. In addition to removing or remodeling extracellular matrix, secreted MMPs regulate cell-cell and cell-matrix signaling. For example, they release, activate or silence growth factors, modify cell surface receptors, and regulate apoptosis and inflammation (McCawley and Matrisian, 2001;Egeblad and Werb, 2002;Parks et al., 2004).Obviously, the activity of enzymes with such a diversity of functions has to be tightly regulated, and this occurs on the transcriptional and posttranscriptional level. Most MMPs are absent in healthy, resting tissue and are only expressed in response to specific stimuli. The transcription of many MMPs is induced by a variety of growth factors and cytokines and by changes in cell-cell and cell-matrix interactions (Sternlicht and Werb, 2001). Most MMPs are secreted as inactive proenzymes, which have to be activated either by cleavage through other proteinases or by induction of autocatalytic processing (Visse and Nagase, 2003).Cutaneous wound repair is an excellent model system to study proteinase function and regulation, because it comprises several processes requiring proteinase action: invasion of inflammatory cells as well as migration of fibroblasts and keratinocytes, angiogenesis, wound cont...

Owing to its potent cytoprotective properties for epithelial cells, keratinocyte growth factor (KGF) is successfully used for the treatment of chemotherapy- and radiotherapy-induced oral mucositis in cancer patients. It is therefore of major interest to determine possible clinical applications of KGF in other organs and in different stress situations and to unravel common and organ-specific mechanisms of KGF action. Here we show that KGF protects human keratinocytes from the toxicity of xenobiotics with electrophilic and oxidative properties and reduces the cell death induced by UV irradiation. In contrast to other cell types, cytoprotection of keratinocytes by KGF is not a direct anti-apoptotic effect but requires de novo protein synthesis. The in vitro findings are clinically relevant because KGF protected keratinocytes in organ-cultured human scalp hair follicles from the toxicity of the xenobiotic menadione. Moreover, injection of KGF into murine back skin markedly reduced cell death in the epidermis after UVB irradiation. This activity is dependent on FGF receptor signaling because it was abrogated in transgenic mice expressing a dominant-negative FGF receptor mutant in keratinocytes. Taken together, our results encourage the use of KGF for skin protection from chemical and physical insults.

The metalloproteinase ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin motifs) is induced under inflammatory conditions, and it is also a potent inhibitor of angiogenesis. Due to these properties, we speculated about the role of ADAMTS1 in cutaneous wound repair. Here we have shown up-regulation of ADAMTS1 expression in wounds of normal and particularly of healing-impaired genetically diabetic mice. Immunofluorescence staining identified macrophages as the source of ADAMTS1 in early wounds, whereas keratinocytes and fibroblasts produce this protein at later stages of wound healing. The distribution of AD-AMTS1 in the normal and wounded epidermis, its regulation in cultured keratinocytes, as well as the skin phenotype of ADAMTS1 knock-out mice suggests a role of this metalloproteinase in keratinocyte differentiation. Furthermore, we provide evidence for a novel dual function of ADAMTS1 in fibroblast migration; although low concentrations of this protein stimulate fibroblast migration via its proteolytic activity, high concentrations inhibit this process because of binding to fibroblast growth factor-2 and subsequent inhibition of its promotogenic activity. Similar effects were also observed with endothelial cells. Taken together, our results suggest a role of ADAMTS1 in keratinocyte differentiation and migration of fibroblasts and endothelial cells in healing skin wounds.Repair of wounded tissue is a complex process that requires temporal and spatial coordination of different events, including inflammation, formation of new tissue, angiogenesis, and matrix remodeling. These processes are regulated by growth factors and cytokines but also by cell-cell and cell-matrix interactions (1-3). Proteolytic processes are required during all phases of cutaneous wound healing. Proteolysis is necessary to allow inflammatory cells to enter the wound site and to degrade the provisional fibrin clot. Proteinases are involved in cell migration, in wound contraction, and in scar remodeling, and the role of serine proteinases, such as plasmin and leukocyte elastase and of matrix metalloproteinases, in wound repair is well established (4 -6). In addition, another proteinase family has recently been identified, whose members are of potential interest for the wound-healing process based on their biological activities. These proteinases, which are designated ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) comprise a family of Zn 2ϩ -dependent metalloproteinases closely related to the matrix metalloproteinases and ADAM family of proteinases. They are structurally characterized by a metalloproteinase domain, a disintegrin-like domain, and several thrombospondin motifs (7). The adamts1 gene was originally cloned from a colon carcinoma cell line (8). Some of the ADAMTS1 substrates have been identified, and they include mainly proteoglycans, such as aggrecan (9, 10) and versican (11). adamts1 knock-out mice are characterized by growth retardation, changes in kidney structure, and impaired female fertil...

Our studies suggest that ProIAPP has typical properties of an amyloidogenic polypeptide but also indicate that the pro-region suppresses the amyloidogenic and cytotoxic potentials of IAPP.