In the integrin family, the collagen receptors form a structurally and functionally distinct subgroup. Two members of this subgroup, ␣ 1 ␤ 1 and ␣ 2 ␤ 1 integrins, are known to bind to monomeric form of type I collagen. However, in tissues type I collagen monomers are organized into large fibrils immediately after they are released from cells. Here, we studied collagen fibril recognition by integrins. By an immunoelectron microscopy method we showed that integrin ␣ 2 I domain is able to bind to classical D-banded type I collagen fibrils. However, according to the solid phase binding assay, the collagen fibril formation appeared to reduce integrin ␣ 1 I and ␣ 2 I domain avidity to collagen and to lower the number of putative ␣I domain binding sites on it. Respectively, cellular ␣ 1 ␤ 1 integrin was able to mediate cell spreading significantly better on monomeric than on fibrillar type I collagen matrix, whereas ␣ 2 ␤ 1 integrin appeared still to facilitate both cell spreading on fibrillar type I collagen matrix and also the contraction of fibrillar type I collagen gel. Additionally, ␣ 2 ␤ 1 integrin promoted the integrin-mediated formation of long cellular projections typically induced by fibrillar collagen. Thus, these findings suggest that ␣ 2 ␤ 1 integrin is a functional cellular receptor for type I collagen fibrils, whereas ␣ 1 ␤ 1 integrin may only effectively bind type I collagen monomers. Furthermore, when the effect of soluble ␣I domains on type I collagen fibril formation was tested in vitro, the observations suggest that integrin type collagen receptors might guide or even promote pericellular collagen fibrillogenesis.A fibril-forming type I collagen, a ubiquitous protein in all vertebrates, is known to provide mechanical stability for tissues and serve as a functional environment for cells (1). Depending on the physical properties of the tissue, type I collagen fibrils are arranged with different suprafibrillar architectures and diameters. Thus, narrow fibrils (ϳ20 nm) in highly ordered arrangement occur in the cornea, where optical transparency is important, whereas large diameter fibrils (ϳ500 nm) provide high tensile strength in mature tendon (2).The mechanism of type I collagen fibril formation has been under extensive research for decades. In tissues, type I collagen is synthesized as a monomeric precursor, which is secreted by exocytosis into the extracellular space. In addition to the triple helical collagenous domain, the precursor contains noncollagenous C-and N-propeptides, which are linked to the triple helical domain by short sequences called telopeptides (3). After the enzymatic removal of propeptides, the solubility of collagen monomers decreases, and they spontaneously form fibrils, assisted by remaining nonhelical telopeptides (1, 4). Evidently, collagen molecules themselves contain all the information needed for fibril assembly. Therefore, in physiological conditions, acid-solubilized collagen monomers form tissue-type long fibrils with characteristic axial periodic structure also in v...

Controlled degradation of extracellular matrix (ECM)1 is essential in physiological situations involving connective tissue remodeling, such as tissue morphogenesis, repair, and angiogenesis. On the other hand, excessive breakdown of connective tissue components plays an important role in destruction of functional tissue architecture, e.g. in rheumatoid arthritis, osteoarthritis, atherosclerosis, periodontitis, autoimmune blistering disorders of skin, and dermal photoaging as well as in invasion and metastasis of tumor cells (see Refs. 1-3). Matrix metalloproteinases (MMPs) are a family of structurally related zinc-dependent endopeptidases collectively capable of degrading essentially all ECM components, and they are implicated in ECM remodeling in the physiologic and pathologic situations mentioned above. At present, 18 human members of the MMP family have been characterized, and most of them can be divided into subgroups of collagenases, gelatinases, stromelysins, and membrane-type MMPs based on their substrate specificity and structure (1-3).Collagenase-1 (MMP-1), collagenase-2 (MMP-8), and collagenase-3 (MMP-13) are the principal neutral proteinases capable of degrading native fibrillar collagens in the extracellular space. They all cleave type I, II, and III collagens at a specific site, generating 3 ⁄4 N-terminal and 1 ⁄4 C-terminal fragments, which denature in physiological temperature and are further degraded by other MMPs, e.g. gelatinases (see Refs. 1-3). MMP-13 also cleaves type I collagen at N-terminal nonhelical telopeptide (4). MMP-1 cleaves type III collagen and MMP-8 type I collagen most effectively (1-3). MMP-13, in turn, cleaves fibrillar collagens with preference to type II collagen over type I and III collagens and displays 40-fold stronger gelatinase activity than MMP-1 and MMP-8 (5-7). In addition, MMP-13 degrades type IV, X, and XIV collagens, tenascin, fibronectin, and aggrecan core protein (8 -9). Apparently due to its exceptionally wide substrate specificity, the physiologic expression of MMP-13 is limited to situations in which rapid and effective remodeling of collagenous ECM takes place, i.e. fetal bone development and adult bone remodeling (10, 11). On the other hand, MMP-13 is expressed at sites of excessive degradation of

Inflammatory cytokines tumor necrosis factor-␣ and interleukin-1 trigger the ceramide signaling pathway, initiated by neutral sphingomyelinase-elicited hydrolysis of cell membrane phospholipid sphingomyelin to ceramide, a new lipid second messenger. Here, we show that triggering the ceramide pathway by sphingomyelinase or C 2 -and C 6 -ceramide enhances collagenase-1 (matrix metalloproteinase-1; MMP-1) gene expression by fibroblasts. C 2 -ceramide activates three distinct mitogen-activated protein kinases (MAPKs) in dermal fibroblasts, i.e. extracellular signal-regulated kinase 1/2 (ERK1/2), stress-activated protein kinase/Jun N-terminal-kinase (SAPK/JNK), and p38. Stimulation of MMP-1 promoter activity by C 2 -ceramide is dependent on the presence of a functional AP-1 cis-element and is entirely inhibited by overexpression of MAPK inhibitor, dual specificity phosphatase CL100 (MAPK phosphatase-1). Activation of MMP-1 promoter by C 2 -ceramide is also effectively inhibited by kinase-deficient forms of ERK1/2 kinase (MEK1/2) activator Raf-1, ERK1 and ERK2, SAPK/ JNK activator SEK1, or SAPK␤. In addition, ceramidedependent induction of MMP-1 expression is potently prevented by PD 98059, a selective inhibitor of MEK1 activation, and by specific p38 inhibitor SB 203580. These results show that triggering the ceramide signaling pathway activates MMP-1 gene expression via three distinct MAPK pathways, i.e. ERK1/2, SAPK/JNK, and p38, and suggest that targeted modulation of the ceramide signaling pathway may offer a novel therapeutic approach for inhibiting collagenolytic activity, e.g. in inflammatory disorders.

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During wound-healing, cells are required to migrate rapidly into the wound site via a proteolytically generated pathway in the provisional matrix, to produce new extracellular matrix, and, subsequently, to remodel the newly formed tissue matrix during the maturation phase. Two classes of molecules cooperate closely to achieve this goal, namely, the matrix adhesion and signaling receptors, the integrins, and matrix-degrading and -processing enzymes, the matrix metalloproteinases (MMPs). There is now substantial experimental evidence that blocking key molecules of either group will prevent or seriously delay wound-healing. It has been known for some time now that cell adhesion by means of the integrins regulates the expression of MMPs. In addition, certain MMPs can bind to integrins or other receptors on the cell surface involved in enzyme activation, thereby providing a mechanism for localized matrix degradation. By proteolytically modifying the existing matrix molecules, the MMPs can then induce changes in cell behavior and function from a state of rest to migration. During wound repair, the expression of integrins and MMPs is simultaneously up-regulated. This review will focus on those aspects of the extensive knowledge of fibroblast and keratinocyte MMPs and integrins in biological processes that relate to wound-healing.