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R Montesano

Publications and source records attributed to R Montesano.

At least 55 records · Page 3Linked to original sources
PubMed1998

Inhibition of leukocyte adherence and transendothelial migration in cultured human liver vascular endothelial cells by prostaglandin E1.

Primary graft dysfunction is a major complication of orthotopic liver transplantation, and hepatic ischemic reperfusion injury is considered to be its major determinant cause. Although oxygen free radicals play an important role, leukocytes, cytokines, and adhesion molecules also contribute to hepatic ischemic reperfusion injury. Prostaglandin E1 (PGE1) has been shown to protect against impairment and dysfunction of transplanted livers in various experimental models as well as in clinical liver transplantation. In this study, the role of PGE1 on leukocyte adherence and transendothelial migration was investigated in cultured human liver vascular endothelial cells (HLVEC). Our results indicated that stimulated, but not resting, leukocytes exhibited high adhesion and transmigration capacity. HLVEC incubated with tumor necrosis factor (TNF) promoted leukocyte adherence and transendothelial migration. PGE1 inhibited leukocyte adherence to HLVEC when it was preincubated with either HLVEC or leukocytes. Moreover, PGE1 also suppressed stimulated leukocyte transendothelial migration in a dose-dependent manner. The inhibitory activity of PGE1 was further investigated on both HLVEC and leukocytes with attention to adhesion molecules. On HLVEC, PGE1 down-regulated TNF-induced expression of endothelial cell leukocyte adhesion molecule 1 and vascular adhesion molecule 1, but not intercellular adhesion molecule 1. On leukocytes, PGE1 inhibited expression of CD11a/CD18 and membrane-bound TNF on PHA-stimulated leukocytes. PGE1 also suppressed TNF release from the stimulated leukocytes. These results indicated that inhibition of leukocyte adherence and transendothelial migration is one of the mechanisms by which PGE1 protects liver grafts.

Alprostadil↗
PubMed1998

Isolation of EpH4 mammary epithelial cell subpopulations which differ in their morphogenetic properties.

EpH4 is a nontumorigenic cell line derived from spontaneously immortalized mouse mammary gland epithelial cells (Fialka et al., 1996). When grown in collagen gels, EpH4 cells give rise to different types of structures, e.g., solid cords or branching tubes. By removing and subsequently dissociating single three-dimensional colonies of defined morphology, we have isolated six clonal subpopulations of EpH4 cells which display distinct morphogenetic properties in collagen gel cultures. Thus, cells from the H1B clone form branching cords devoid of a central lumen, K3A3 cells form cords enclosing small multifocal lumina, and J3B1 cells form large cavitary structures containing a wide lumen. I3G2 cells form either cords or tubes, depending on the type of serum added to the culture medium. Finally, when grown in serum-free medium, Bela cells form spherical cysts, whereas Be4a cells form long, extensively branched tubes. In additional assays of morphogenesis, i.e., cell sandwiching between two collagen gels or culture on a thick layer of Matrigel (a laminin-rich extracellular matrix), all clones form epithelial-cell lined cavitary structures, except H1B cells which are unable to generate lumina under these conditions. The EpH4 sublines we have isolated provide an in vitro system for studying the mechanisms responsible for lumen formation and branching morphogenesis, as well as for identifying the factors which subvert these developmental processes during mammary carcinogenesis.

Animals↗
PubMed1998

Phytoestrogens and inhibition of angiogenesis.

The consumption of a plant-based diet can prevent the development and progression of chronic diseases associated with extensive neovascularization, including the progression and growth of solid malignant tumours. We have previously shown that the plant-derived isoflavonoid genistein is a potent inhibitor of cell proliferation and in vitro angiogenesis. Moreover, the concentration of genistein in the urine of subjects consuming a plant-based diet is 30-fold higher than that in subjects consuming a traditional Western diet. We have also reported that certain structurally related flavonoids are more potent inhibitors than genistein. Indeed, 3-hydroxyflavone, 3',4'-dihydroxyflavone, 2',3'-dihydroxyflavone, fisetin, apigenin and luteolin inhibit the proliferation of normal and tumour cells as well as in vitro angiogenesis at half-maximal concentrations in the lower micromolar range. The wide distribution of isoflavonoids and flavonoids in the plant kingdom, together with their anti-angiogenic and anti-mitotic properties, suggest that these phytoestrogens may contribute to the preventive effect of a plant-based diet on chronic diseases, including solid tumours.

Cell Division↗
PubMed1998

Roles of hepatocyte growth factor/scatter factor and transforming growth factor-beta1 in mammary gland ductal morphogenesis.

Epithelial-mesenchymal interactions are responsible for the unique pattern of ductal branching morphogenesis characteristic of the mammary gland. To investigate the factors which control the elongation and branching of lactiferous ducts, we developed an in vitro model of ductal morphogenesis in which clonal mouse mammary epithelial cells (TAC-2 cells) are grown in collagen gels. In this experimental system, fibroblast conditioned medium (CM)3 stimulates the formation of extensively arborized tubules. The molecule responsible for this tubulogenic effect was identified as hepatocyte growth factor/scatter factor (HGF/SF). To determine whether HGF/SF plays a role in mammary gland morphogenesis in vivo, the expression of HGF/SF and its receptor, c-Met, were analyzed in the rat mammary gland during pregnancy, lactation, and involution. Levels of HGF/SF and c-Met transcripts were progressively reduced during pregnancy, were virtually undetectable during lactation, and increased again during involution. Collectively, these in vitro and in vivo findings suggest that HGF/SF is a paracrine mediator of mammary gland ductal morphogenesis. We subsequently investigated the effect of another multifunctional cytokine, namely TGF-beta1, on branching morphogenesis of TAC-2 cells. TGF-beta1 had a striking biphasic effect: whereas relatively high concentrations of this cytokine inhibited colony formation, lower concentrations stimulated extensive elongation and branching of epithelial cords. Taken together, these studies indicate that HGF/SF is a stromal-derived paracrine mediator of mammary ductal morphogenesis, and that when present at low concentrations, TGF-beta1 can contribute to this process.

Animals↗
PubMed1998

Analysis of p53, p16MTS, p21WAF1 and H-ras in archived bladder tumours from workers exposed to aromatic amines.

Exposure to aromatic amines is considered a major risk factor for the development of bladder cancer. In this study, we have analysed the pattern of point mutations in several tumour genes in 21 cases of bladder cancer arising among western European workers exposed to aromatic amines in an attempt to determine whether this exposure may be associated with a unique spectrum of mutations. Of the four genes analysed (p53, p16MTS1, p21WAF1 and H-ras), only p53 showed a high frequency of mutations (in 8 out of 21 cases, 38%). Two mutations were found in p16, one in H-ras and none in p21 exon 3. All mutations were at G:C base pairs, mostly at non-CpG residues. This spectrum of mutations, which is highly suggestive of an involvement of exogenous carcinogens, is however identical to the spectrum of p53 mutations detected in bladder cancers of the general population. In exposed workers, p53 mutations were associated with tumour grade and with high occupational and tobacco exposure. Taken together, our data suggest that the same carcinogens may be responsible for the development of bladder cancers in workers exposed to aromatic amines and in the general population.

Adolescent↗
PubMed1998

FGF-2 stimulates migration of Kaposi's sarcoma-like vascular cells by HGF-dependent relocalization of the urokinase receptor.

The spindle-shaped cell line TTB was recently isolated from highly vascularized skin lesions of BKV/HIV-1 tat transgenic mice and shown to possess an autocrine loop for hepatocyte growth factor (HGF). We show that fibroblast growth factor-2 (FGF-2) stimulates TTB cell migration and promotes polarization of uPAR at the leading edge of migrating cells. FGF-stimulated TTB cells presented the typical migratory phenotype, with a triangular cell shape and concomitant breakdown of actin stress fibers and smooth muscle-specific actin isoform. FGF-2-stimulated migration was blocked by antibodies against urokinase-type plasminogen activator (uPA) or uPA receptor (uPAR) and by neutralizing anti-HGF antibodies. The latter also inhibited uPAR relocalization at the cell surface of FGF-2-treated TTB cells. This points to a crosstalk between FGF-2 and HGF that might mediate TTB cell migration by modulating the localization of cell surface uPAR.

Cell Line↗
PubMed1998

Molecular precursor lesions in oesophageal cancer.

Oesophageal cancer is the fifth most frequent cause of cancer death world wide and most of these cancers occur in developing countries. The survival rate for SCC or ADCs of the oesophagus is equally poor, mainly due to their late detection and the poor efficacy of the therapy. A short review of the natural history of these cancers, and in particular the occurrence of genetic and cellular alterations associated with cancer progression, is presented and discussed in the context of the relevance to aetiology and pathogenesis. SCCs and ADCs show a distinct pattern of TP53 mutations, namely a high prevalence of G > A transitions at CpG sites in ADCs whereas in SCCs a higher prevalence of G to T transversions and mutations at A:T base pairs is present. In both types of cancers TP53 mutations occur very early and are followed by the accumulation of other genetic alterations during the process of oesophageal carcinogenesis. The value of these genetic alterations in assessing the multifocal monoclonal origin of oesophageal cancer is also addressed.

Adenine↗
PubMed1998

Differential effects of hepatocyte growth factor isoforms on epithelial and endothelial tubulogenesis.

Hepatocyte growth factor (HGF)/scatter factor (SF) is a pleiotropic cytokine that acts as a mitogen, motogen, and morphogen for a variety of cell types. HGF/NK1 and HGF/NK2 are two naturally occurring truncated variants of HGF/SF, which extend from the NH2 terminus through the first and second kringle domain, respectively. Although these variants have been reported to have agonistic or antagonistic activity relative to HGF/SF in assays of cell proliferation and motility, their potential morphogenic activity has not been investigated. To address this issue, we assessed the ability of HGF/NK1 and HGF/NK2 to induce tube formation by (a) MCF-10A mammary epithelial cells grown within collagen gels and (b) human umbilical vein endothelial (HUVE) cells grown on Matrigel. We found that HGF/NK1 stimulated tubulogenesis by both MCF-10A and HUVE cells, whereas HGF/NK2 did not stimulate tubulogenesis, but efficiently antagonized the morphogenic effect of full-length HGF/SF. HGF/NK1 and HGF/NK2 also had agonistic and antagonistic effects, respectively, on MCF-10A cell proliferation and HUVE cell migration. These results demonstrate that HGF/NK1, which only consists of the NH2-terminal hairpin and first kringle domain, is sufficient to activate the intracellular signaling pathways required to induce morphogenic responses in epithelial and endothelial cells. In contrast, HGF/NK2, which differs from HGF/ NK1 by the presence of the second kringle domain, is devoid of intrinsic activity but opposes the effects of HGF/SF. The differential properties of the two HGF/SF isoforms provide a basis for the design of more potent HGF/SF agonists and antagonists.

Cell Division↗
PubMed1997

Hepatocellular carcinoma: from gene to public health.

Liver diseases associated with chronic hepatitis B virus (HBV) infection, including hepatocellular carcinoma, account for more than 1 million deaths annually worldwide. In addition to HBV infection, other risk factors are involved in the etiology of hepatocellular carcinoma and, among these, dietary exposure to the carcinogenic aflatoxins is of particular importance in certain regions of southeast Asia and sub-Saharan Africa. The relative contributions of these two risk factors and the mechanism of the interaction between them in the pathogenesis of hepatocellular carcinoma are still poorly understood. The recently developed individual biochemical and molecular markers of aflatoxin exposure, i.e., aflatoxin-albumin adducts in blood and a specific GC to TA transversion mutation in codon 249 of the p53 gene (249ser p53 mutation) in hepatocellular carcinomas, permit a better quantitative estimation of aflatoxin exposure in different populations of the world. A comprehensive summary of the data from our laboratory and the literature, based on a large number (>1000) of individual cases of hepatocellular carcinoma, is presented here and shows the following: 1) A high level and high prevalence of exposure to aflatoxins occur in West Africa, Mozambique, and some regions of China; 2) a high prevalence of the 249ser p53 mutation is detected in these countries; and 3) hepatocellular carcinomas from countries with low or no exposure to aflatoxins show a very low prevalence of the 249ser p53 mutation and distinctly different p53 mutation spectra, probably indicating different etiologies. Experimental and epidemiologic studies demonstrate an interaction between HBV infection and aflatoxins in hepatocarcinogenesis. The relevance of the biochemical/molecular markers of aflatoxin exposure, HBV vaccination, and the reduction of aflatoxin exposure, in addition to the interaction between HBV infection and other risk factors in liver carcinogenesis, are discussed with regard to the implementation of measures for primary prevention.

Aflatoxins↗
PubMed1997

Flavonoids, dietary-derived inhibitors of cell proliferation and in vitro angiogenesis.

Consumption of a plant-based diet can prevent the development and progression of chronic diseases associated with extensive neovascularization, including solid malignant tumors. In previous studies, we have shown that the plant-derived isoflavonoid genistein is a potent inhibitor of cell proliferation and in vitro angiogenesis. In the present study, we report that certain structurally related flavonoids are more potent inhibitors than genistein. Indeed, 3-hydroxyflavone, 3',4'-dihydroxyflavone, 2',3'-dihydroxyflavone, fisetin, apigenin, and luteolin inhibited the proliferation of normal and tumor cells, as well as in vitro angiogenesis, at half-maximal concentrations in the low micromolar range. We have previously demonstrated that genistein concentrations in the urine of subjects consuming a plant-based diet is 30-fold higher than in subjects consuming a traditional Western diet. The wider distribution and the more abundant presence of flavonoids in the plant kingdom, together with the present results, suggest that flavonoids may contribute to the preventive effect of a plant-based diet on chronic diseases, including solid tumors.

Animals↗
PubMed1997

Polyoma middle T-induced vascular tumor formation: the role of the plasminogen activator/plasmin system.

The middle T antigen of murine Polyomavirus (PymT) rapidly transforms endothelial cells, leading to the formation of vascular tumors in newborn mice. Transformed endothelial (End.) cell lines established from such tumors exhibit altered proteolytic activity as a result of increased expression of urokinase-type plasminogen activator (uPA) and are capable of inducing vascular tumors efficiently when injected into adult mice. In this study we have used mice lacking components of the PA/plasmin system to analyze the role of this system in the transformation process and in tumor growth. We found that the proteolytic status of the host is not a critical determinant for PymT-induced vascular tumor formation. In addition, the lack of either uPA or tissue-type PA (tPA) activity is not limiting for the establishment and proliferation of End. cells in vitro, although the combined loss of both PA activities leads to a marked reduction in proliferation rates. Furthermore, the in vitro morphogenetic properties of mutant End. cells in fibrin gels could only be correlated with an altered proteolytic status in cells lacking both uPA and tPA. However, in contrast with tumors induced by PymT itself, the tumorigenic potential of mutant and wild-type End. cell lines was found to be highly dependent on the proteolytic status of both the tumor cells and the host. Thus, genetic alterations in the PA/plasmin system affect vascular tumor development, indicating that this system is a causal component in PymTmediated oncogenesis.

Animals↗
PubMed1997

p53 gene mutation pattern in rat liver tumors induced by vinyl chloride.

Vinyl chloride (VC) induces angiosarcomas of the liver (ASL) and hepatocellular carcinomas (HCCs) in humans and rodents. We examined the presence of p53 gene mutations in ASL and HCC induced by VC in Sprague Dawley rats; 25 ASL and eight HCCs were analyzed for point mutations in exons 5-8, using PCR amplification, single-strand conformation polymorphism analysis, and direct DNA sequencing. Mutations were found in 11 (44%) of the ASL and in 1 HCC. A 12-base pair deletion was found in one tumor; all others were base pair substitutions. Nine of the point mutations were observed at A:T base pairs (5 A:T --> T:A; 2 A:T --> G:C, and 2 A:T --> C:G), and of three G:C --> A:T transitions, only one was at a CpG site. In ASL, four mutations were found in exon 5, two in exon 6, and six in exon 7; the base pair substitution found in one HCC was in exon 8. One ASL exhibited two point mutations, including a silent one. Two ASL exhibited the same mutation in codon 203 and two other samples in codon 253. Codon 235 was found to be mutated in three ASL. These data show that p53 is often mutated in ASL induced by VC in rats and, as observed in ASL in humans exposed to VC, the majority of the missense mutations involved A:T base pairs. The characteristic patterns of mutations found suggest that a common mechanism operates in VC-induced p53 mutagenesis in both species, and these mutations are consistent with the formation of DNA etheno adducts by VC in the liver. The A:T --> T:A transversion observed in the first nucleotide of codon 253 in two rat ASL is equivalent to the A:T --> T:A transversion characterized previously in codon 255 in one human ASL associated with VC exposure.

Animals↗
PubMed1997

Inactivation of the p53 protein in cell lines derived from human esophageal cancers.

Alteration of the p53 gene is thought to be important in the early stages of human esophageal cancers, but how this confers a selective advantage to esophageal cancer cells is unknown. In this report, we analyzed 9 cell lines derived from human esophageal cancers (TE-1, TE-3, TE-6, TE-7, TE-9, TE-10, TE-11, TE-13 and TE-15) for mutations in the p53 sequence, p53 protein expression and p53 protein DNA-binding activity. The cell lines could be grouped in 3 categories, including (1) cell lines with mis-sense mutations in the coding sequence and accumulation of mutant proteins (TE-1, TE-6, TE-10 and TE-11); (2) cell lines expressing truncated forms of p53 as a result of frameshift (TE-9) or splice-site (TE-15) mutations; and (3) cell lines with wild-type p53 sequences but with impaired expression of p53 mRNA and protein, suggesting that p53 is inactivated by transcriptional repression (TE-3, TE-7 and TE-13). With the exception of TE-1, none of the cell lines exhibited p53-DNA-binding activity. In TE-1, a mutation at codon 272 (methionine to valine) generated a protein that retains basal DNA-binding activity, but that was not activated in response to DNA damage, suggesting that this mutation prevented p53 induction by genotoxic stress. Thus, p53 activity was impaired in all esophageal cell lines, including those containing wild-type p53 sequences.

Base Sequence↗
PubMed1997

Vascular endothelial growth factor is increased in devascularized rat islets of Langerhans in vitro.

Vascular endothelial growth factor (VEGF) is an endothelial cell-specific mitogen with potent angiogenic and vascular permeability-inducing properties, both of which may be important for the function of islets of Langerhans. In this study, we have examined the expression of VEGF and its tyrosine kinase receptors (flt and flk-1) in isolated rat islets of Langerhans in vitro. When analyzed by in situ hybridization, islet tissue showed a significant 4.6-fold increase in VEGF mRNA expression over time in culture from 0 to 7 days. Islet tissue exposed to hypoxic/anoxic conditions for a period of 8 hr showed a 3.7-fold increase in VEGF mRNA when analyzed by Northern blot hybridization. Reverse transcriptase-polymerase chain reaction revealed the presence of both flt and flk-1 in freshly isolated islets, and two VEGF isoforms, namely VEGF120 and VEGF164. Three rodent beta-cell lines derived from insulinomas (RINm5F-2A, INS-1, and MIN6) were also found to express VEGF by Northern blot hybridization. However, neither hypoxia/anoxia nor low (0.3 g/L)- or high (3.0 g/L)-glucose culture conditions modulated their expression of VEGF. VEGF derived from RINm5F-2A cells was bioactive in a three-dimensional in vitro model of angiogenesis, which assays for endothelial cell invasion and capillary morphogenesis. These findings demonstrate, first, that devascularization increases VEGF expression in isolated islet tissue, and they point to VEGF as a potentially important endogenous angiogenic stimulus for subsequent revascularization in vivo. Second, our observations raise the possibility that survival of transplanted islets may be improved by increasing VEGF expression before transplantation.

Animals↗
PubMed1997

Database of p53 gene somatic mutations in human tumors and cell lines: updated compilation and future prospects.

In recent years, there has been an exponential increase in the number of p53 mutations identified in human cancers. The p53 mutation database consists of a list of point mutations in thep53 gene of human tumors and cell lines, compiled from the published literature and made available through electronic media. The database is now maintained at the International Agency for Research on Cancer (IARC) and is updated twice a year. The current version contains records on 5091 published mutations and is expected to surpass the 6000 mark in the January 1997 release. The database is available in various formats through the European Bioinformatics Institute (EBI) ftp server at: ftp://ftp.ebi.ac.uk/pub/databases/p53/ or by request from IARC (p53database@iarc.fr) and will be searchable through the SRS system in the near future. This report provides a description of the criteria for inclusion of data and of the current formats, a summary of the relevance ofp53 mutation analysis to clinical and biological questions, and a brief discussion of the prospects for future developments.

Databases, Factual↗
PubMed1997

Inhibition of angiogenesis by tissue inhibitor of metalloproteinase-3.

PURPOSE: It has been established that Sorsby's fundus dystrophy, a dominantly inherited form of blindness, is caused by mutations in the tissue inhibitor of metalloproteinase-3 (TIMP-3) gene. Because choroidal neovascularization is a prominent feature of Sorsby's fundus dystrophy, the authors have examined whether TIMP-3 protein plays a role in the regulation of angiogenesis. METHODS: Chemotaxis of endothelial cells toward vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) was examined using a modified Boyden chamber assay. Endothelial cells placed in the upper chamber were allowed to migrate through a polycarbonate membrane with 8 microns pores toward VEGF or bFGF present in the lower chamber. Next, the ability of TIMP-3 to inhibit chemotaxis was studied by incubating the cells with varying amounts of TIMP-3 during the assay. Finally, an in vitro angiogenesis assay was performed on collagen gels. Endothelial cells were seeded onto three-dimensional collagen gels. Treatment with bFGF and VEGF induced invasion of the gel and the formation of tube-like structures. TIMPs (1, 2, and 3) were added to the cultures to determine their effect on invasion. An in vivo chorioallantoic membrane (CAM) assay was performed using methylcellulose discs containing bFGF with or without TIMP-3. Induction of new blood vessels was observed with a stereomicroscope. RESULTS: TIMP-3 inhibits chemotaxis of vascular endothelial cells toward VEGF and bFGF, inhibits collagen gel invasion and capillary morphogenesis in vitro, and inhibits bFGF-induced angiogenesis in the CAM assay in vivo. CONCLUSIONS: TIMP-3 has the potential to inhibit angiogenesis. These results allow us to speculate on a possible mechanism by which mutant TIMP-3 protein might contribute to the Sorsby fundus dystrophy phenotype.

Allantois↗
PubMed1997

The use of biomarkers to study pathogenesis and mechanisms of cancer: oesophagus and skin cancer as models.

Recent advances in molecular biology have made it possible to use genetic alterations associated with cancer as biomarkers to study the pathogenesis and mechanisms of cancer. However, the lessons that can be drawn from the analysis of alterations in a particular cancer gene are extremely dependent upon the biological context in which they arise. In this article, we discuss the biological significance of alterations in the p53 tumour suppressor gene in cancers of the oesophagus and of the skin. In both tissues, different forms of cancer occur at high frequency (squamous-cell carcinoma and adenocarcinoma in the oesophagus; squamous-cell carcinoma, basal-cell carcinoma and melanoma in the skin). We show that specific patterns of p53 alteration occur in these various cancers and that analysis of these alterations is useful to make inferences about the etiopathogenesis of cancers of the oesophagus and of the skin.

Adenocarcinoma↗