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1.  A Novel Monoclonal Antibody to Human Laminin α5 Chain Strongly Inhibits Integrin-Mediated Cell Adhesion and Migration on Laminins 511 and 521 
PLoS ONE  2013;8(1):e53648.
Laminins, a large family of αβγ heterotrimeric proteins mainly found in basement membranes, are strong promoters of adhesion and migration of multiple cell types, such as tumor and immune cells, via several integrin receptors. Among laminin α (LMα) chains, α5 displays the widest tissue distribution in adult life and is synthesized by most cell types. Here, we have generated and characterized five novel monoclonal antibodies (mAbs) to the human LMα5 chain to further study the biological relevance of α5 laminins, such as laminins 511 (α5β1γ1) and 521 (α5β2γ1). As detected by ELISA, immunohistochemistry, immunoprecipitation and Western blotting, each antibody displayed unique properties when compared to mAb 4C7, the prototype LMα5 antibody. Of greatest interest, mAb 8G9, but not any other antibody, strongly inhibited α3β1/α6β1 integrin-mediated adhesion and migration of glioma, melanoma, and carcinoma cells on laminin-511 and, together with mAb 4C7, on laminin-521. Accordingly, mAb 8G9 abolished the interaction of soluble α3β1 integrin with immobilized laminins 511 and 521. Binding of mAb 8G9 to laminin-511 was unaffected by the other mAbs to the LMα5 chain but largely hindered by mAb 4E10 to a LMβ1 chain epitope near the globular domain of laminin-511. Thus, mAb 8G9 defines a novel epitope localized at or near the integrin-binding globular domain of the LMα5 chain, which is essential for cell adhesion and migration, and identifies a potential therapeutic target in malignant and inflammatory diseases.
doi:10.1371/journal.pone.0053648
PMCID: PMC3538678  PMID: 23308268
2.  Differences in the Nemosis Response of Normal and Cancer-Associated Fibroblasts from Patients with Oral Squamous Cell Carcinoma 
PLoS ONE  2009;4(9):e6879.
Background
Tumor-stroma reaction is associated with activation of fibroblasts. Nemosis is a novel type of fibroblast activation. It leads to an increased production of growth factors and proinflammatory and proteolytic proteins, while at the same time cytoskeletal proteins are degraded. Here we used paired normal skin fibroblasts and cancer-associated fibroblasts (CAF) and primary and recurrent oral squamous cell carcinoma (SCC) cells to study the nemosis response.
Principal Findings
Fibroblast nemosis was analyzed by protein and gene expression and the paracrine regulation with colony formation assay. One of the normal fibroblast strains, FB-43, upregulated COX-2 in nemosis, but FB-74 cells did not. In contrast, CAF-74 spheroids expressed COX-2 but CAF-43 cells did not. Alpha-SMA protein was expressed in both CAF strains and in FB-74 cells, but not in FB-43 fibroblasts. Its mRNA levels were downregulated in nemosis, but the CAFs started to regain the expression. FSP1 mRNA was downregulated in normal fibroblasts and CAF-74 cells, but not in CAF-43 fibroblasts. Serine protease FAP was upregulated in all fibroblasts, more so in nemotic CAFs. VEGF, HGF/SF and FGF7 mRNA levels were upregulated to variable degree in nemosis. CAFs increased the colony formation of primary tumor cell lines UT-SCC-43A and UT-SCC-74A, but normal fibroblasts inhibited the anchorage-independent growth of recurrent UT-SCC-43B and UT-SCC-74B cells.
Conclusions
Nemosis response, as observed by COX-2 and growth factor induction, and expression of CAF markers α-SMA, FSP1 and FAP, varies between fibroblast populations. The expression of CAF markers differs between normal fibroblasts and CAFs in nemosis. These results emphasize the heterogeneity of fibroblasts and the evolving tumor-promoting properties of CAFs.
doi:10.1371/journal.pone.0006879
PMCID: PMC2730537  PMID: 19721715
3.  Snail1 Protein in the Stroma as a New Putative Prognosis Marker for Colon Tumours 
PLoS ONE  2009;4(5):e5595.
Over-expression of Snail1 gene transcriptional repressor promotes an epithelial-to-mesenchymal transition in epithelial tumour cell lines. Expression of Snail1 RNA has been associated to the pathogenesis of a number of malignancies; however, the lack of good monoclonal antibodies against this protein has precluded a definitive analysis of Snail1 protein. In this study, we aimed to determine the expression of this transcriptional factor in colorectal tumours. Using a Snail1 well-characterized monoclonal antibody developed in our laboratories we have analyzed by immunohistochemistry a cohort of 162 human colorectal tumours. Ninety tumours (56%) showed nuclear expression in the tumoral tissue and the adjacent stroma; in 34 (21%), Snail1 was detected just in the stroma, whereas in only 4 the expression of Snail1 was detected in the tumoral tissue and the stroma was negative. No correlation was found between the presence of Snail1 in the tumour and tumour stage; however, a trend (p = 0.054) was detected when the expression of this factor in the stroma was considered. Snail1 immunoreactivity in this compartment was associated with presence of distant metastasis (p = 0.006). Moreover, expression of Snail1 in the tumor stroma correlated with lower specific survival of cancer patients (p = 0.011). Interestingly, this correlation was also detected in stage I and II tumors. Therefore, our results indicate that the presence of nuclear Snail1 immunoreactive cells in the stroma may be an informative indicator of prognosis of colon tumours especially useful in those corresponding to lower stages and identify a new marker suitable to label activated stroma in colon tumours.
doi:10.1371/journal.pone.0005595
PMCID: PMC2680015  PMID: 19440385

Results 1-3 (3)