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Loss of E-cadherin/catenin mediated cell-cell adhesion and overexpression of matrix metalloproteinases (MMPs) are largely involved in tumor invasion. It has been recently shown that high levels of a soluble 80 kDa fragment of E-cadherin, resulting from a cleavage by MMPs, are found in serum and in urine from cancer patients. Additionally, this soluble E-cadherin (sE-CAD) promotes cell invasion into chick heart and into collagen type I gels. The aim of our study was to examine the mechanism of sE-CAD-induced cell invasion. Since MMPs play a crucial role in invasion, we looked for induction of MMPs by sE-CAD in noninvasive human lung tumor cells 16HBE. An induction of MMP-2, MMP-9 and MT1-MMP expression was observed both at the mRNA and at the protein level in the presence of sE-CAD (in conditioned medium form or in E-cadherin HAV peptide form). No induction of MMP-1, -3 and -7 or variation of the levels of their inhibitors, TIMP-1 and TIMP-2, were detected. The biologic relevance of the sE-CAD-induced MMP upregulation was tested by demonstrating that sE-CAD promotes in vitro cell invasion in a modified Boyden chamber assay. These data provide new insight into mechanisms of tumor invasion by ectodomain shedding of the cell-cell adhesion molecule E-cadherin. © 2003 Wiley-Liss, Inc. Key words: cancer progression; soluble E-cadherin; MMPs; cell invasionTumor cell invasion in the surrounding host tissue is a key event of the metastatic progression. This process requires dispersion of tumor cells from the primary tumor and infiltration of the stromal compartment.Tumor cell dispersion largely relies on the loss of homotypic cell-cell adhesion. The E-cadherin/catenin complex plays a crucial role in epithelial cell-cell adhesion and in the maintenance of tissue architecture. Modifications in the expression or function of this complex result in a decrease of adhesive properties of the complex and, thus, E-cadherin is considered as an important invasion suppressor. 1-3 Indeed, several in vitro studies have shown a strong correlation between the defect of E-cadherin/catenin complex expression at the level of the cell surface and loss of both an epithelial phenotype and enhanced cell invasiveness. 4 -9 In addition, many in vivo studies have described a correlation between loss of E-cadherin or catenin expression or dysfunction of the complex and dedifferentiation, malignancy, tumor aggressiveness, metastasis, or a poor survival rate in several types of cancer including breast, gastric, liver, bladder, prostate, lung and colon carcinomas. 1,2,10 -12 Invasion through basement membranes and interstitial extracellular matrix requires the action of proteolytic enzymes. Among these enzymes, matrix metalloproteinases (MMPs) are able to degrade almost all of the extracellular matrix (ECM) components. Most MMPs are secreted as latent proenzymes that are extracellularly activated through proteolytic cleavage. The MMP activity is inhibited by TIMPs (tissue inhibitors of metalloproteinases), their major physiologic specific inhibitors...

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SummaryMultivariate data sets are now produced in several types of microscopy. Multivariate statistical methods are necessary in order to extract the useful information contained in such (image or spectrum) series. In this review, linear and nonlinear multivariate methods are described and illustrated with examples related both to the segmentation of microanalytical maps and to the study of variability in the images of unit cells in high-resolution transmission electron microscopy. Concerning linear multivariate statistical analysis, emphasis is put on the need to go beyond the classical orthogonal decomposition already routinely performed through principal components analysis or correspondence analysis. It is shown that oblique analysis is often necessary when quantitative results are expected. Concerning nonlinear multivariate analysis, several methods are first described for performing the mapping of data from a high-dimensional space to a space of lower dimensionality. Then, automatic classification methods are described. These methods, which range from classical methods (hard and fuzzy C-means) to neural networks through clustering methods which do not make assumptions concerning the shape of classes, can be used for multivariate image segmentation and image classification and averaging.

The disorganization of E-cadherin/catenin complexes and the overexpression of matrix metalloproteinases (MMPs) are frequently involved in the capacity of epithelial cells to acquire an invasive phenotype. The functional link between E-cadherin and MMPs was studied by transfecting invasive bronchial BZR tumor cells with human E-cadherin cDNA. Using different in vitro (cell dispersion, modified Boyden chamber) and in vivo assays (human airway epithelial xenograft), we showed that E-cadherin-positive clones displayed a decrease of invasive abilities. As shown by immunoprecipitation, the re-expressed E-cadherin was able to sequestrate one part of free cytoplasmic beta-catenin in BZR cells. The decrease of beta-catenin transcriptional activity in E-cadherin-transfected clones was demonstrated using the TOP-FLASH reporter construct. Finally, we observed a decrease of MMP-1, MMP-3, MMP-9, and MT1-MMP, both at the mRNA and at the protein levels, in E-cadherin-positive clones whereas no changes in MMP-2, TIMP-1, or TIMP-2 were observed when compared with control clones. Moreover, zymography analysis revealed a loss of MMP-2 activation ability in E-cadherin-positive clones treated with the concanavalin A lectin. These data demonstrate a direct role of E-cadherin/catenin complex organization in the regulation of MMPs and suggest an implication of this regulation in the expression of an invasive phenotype by bronchial tumor cells.

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This study was designed to investigate the molecular mechanisms of Staphylococcus aureus adherence to human airway epithelium. Using a humanized bronchial xenograft model in the nude mouse and primary cultures of human airway epithelial cells (HAEC), we showed that S. aureus adhered mainly to undifferentiated HAEC whereas weak adherence (11- to 20-fold lower) to differentiated HAEC was observed (P < 0.01). A fibronectin (FN)-binding protein (FnBP)-deficient strain of S. aureus had a fivefold-lower adherence level to undifferentiated HAEC than did the parental strain (P < 0.005), suggesting that S. aureus FN-binding capacity is involved in the adherence to HAEC. We also showed that 97% of 32 S. aureus clinical strains, isolated from the airway secretions of cystic fibrosis patients (n = 18) and patients with nosocomial pneumonia (n = 14), possessed the two fnb genes. The strains from pneumonia patients had a significantly (P < 0.05) higher FN-binding capacity than did the strains from CF patients. This result was confirmed by the expression of FnBPs, investigated by Western ligand affinity blotting. Our results suggest a major role of FnBPs in the colonization of the airways by S. aureus and point to the importance of the adhesin regulatory pathways in the staphylococcal infectious process.

AGEs (advanced glycation end-products) accumulate in collagen molecules during uraemia and diabetes, two diseases associated with high susceptibility to bacterial infection. Because neutrophils bind to collagen during their locomotion in extravascular tissue towards the infected area we investigated whether glycoxidation of collagen (AGE-collagen) alters neutrophil migration. Type I collagen extracted from rat tail tendons was used for in vitro glycoxidation (AGE-collagen). Neutrophils were obtained from peripheral blood of healthy adult volunteers and were used for the in vitro study of adhesion and migration on AGE- or control collagen. Glycoxidation of collagen increased adhesion of neutrophils to collagen surfaces. Neutrophil adhesion to AGE-collagen was inhibited by a rabbit anti-RAGE (receptor for AGEs) antibody and by PI3K (phosphoinositide 3-kinase) inhibitors. No effect was observed with ERK (extracellular-signal-regulated kinase) or p38 MAPK (mitogen-activated protein kinase) inhibitors. AGE-collagen was able to: (i) induce PI3K activation in neutrophils, and (ii) inhibit chemotaxis and chemokinesis of chemoattractant-stimulated neutrophils. Finally, we found that blocking RAGE with anti-RAGE antibodies or inhibiting PI3K with PI3K inhibitors restored fMLP (N-formylmethionyl-leucyl-phenylalanine)-induced neutrophil migration on AGE-collagen. These results show that RAGE and PI3K modulate adhesion and migration rate of neutrophils on AGE-collagen. Modulation of adhesiveness may account for the change in neutrophil migration rate on AGE-collagen. As neutrophils rely on their ability to move to perform their function as the first line of defence against bacterial invasion, glycoxidation of collagen may participate in the suppression of normal host defence in patients with diabetes and uraemia.