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R Montesano

Publications and source records attributed to R Montesano.

At least 91 records · Page 5Linked to original sources
PubMed1995

Modulation of hepatocyte growth factor and c-met in the rat mammary gland during pregnancy, lactation, and involution.

Epithelial tubulogenesis is responsible for the exquisitely intricate organization of functional units of parenchymal organs. We have previously demonstrated that hepatocyte growth factor (HGF--also known as scatter factor) is a stroma-derived epithelial morphogen, which induces tubulogenesis by kidney-derived epithelial cells in vitro. The mammary gland provides a particularly attractive model for the study of epithelial morphogenesis, since its development in postnatal life involves elongation and branching of epithelial tubules. The aim of the present studies was to assess the expression and modulation of HGF and its receptor c-Met in the rat mammary gland during pregnancy, lactation, and involution. By ribonuclease protection assay, we demonstrate that levels of both HGF and c-met transcripts are progressively reduced during pregnancy, are virtually undetectable during lactation, and increase during the phase of involution to prepregnancy levels. The reduction in HGF and c-met expression corresponds to periods in which functions other than tubulogenesis predominate in the mammary gland, namely alveologenesis (mid to late pregnancy) and milk protein synthesis (lactation). Using a murine mammary gland-derived epithelial cell line, we demonstrate that levels of c-met mRNA are significantly reduced by exogenously added prolactin, providing a possible explanation for the reduction in c-met in the rat mammary gland during lactation. The potential significance of down-regulation of HGF/c-met during lactation is discussed.

Amino Acid Sequence↗
PubMed1995

Genistein, a dietary ingested isoflavonoid, inhibits cell proliferation and in vitro angiogenesis.

Consumption of a plant-based diet can prevent the development and progression of chronic diseases that are associated with extensive neovascularization. To determine whether prevention might be associated with dietary derived angiogenesis inhibitors, we have fractionated urine of healthy human subjects consuming a plant-based diet and examined the fractions for their abilities to inhibit the proliferation of vascular endothelial cells. One of the most potent fractions contained several isoflavonoids, which we identified by gas chromatography-mass spectrometry and subsequently synthesized. Of all synthetic compounds, the isoflavonoid genistein was the most potent and inhibited endothelial cell proliferation and in vitro angiogenesis at half maximal concentrations of 5 and 150 mumol/L, respectively. Moreover, genistein inhibited the proliferation of various tumor cells. Genistein excretion in urine of subjects consuming a plant-based diet is in the micromolar range, which is 30-fold higher than that of subjects consuming a traditional Western diet. The high concentrations of genistein in urine of vegetarians and our present results suggest that genistein may contribute to the preventive effect of plant-based diet on chronic diseases, including solid tumors, by inhibiting neovascularization and tumor cell proliferation. Thus genistein may have important applications in the treatment of solid tumors and angiogenic diseases.

Animals↗
PubMed1995

Screening for p53 gene mutations in archived tumors of workers occupationally exposed to carcinogens: examples from analysis of bladder tumors.

Point mutations in the p53 tumor suppressor gene are the most common genetic alterations in human cancers. The nature and location of these mutations can be informative in assessing the importance of putative carcinogenic agents. Potential associations between a given carcinogen and a specific mutation pattern can be substantiated when the exposure history of the patients is known. While the past exposure to environmental risk factors is often difficult to determine, documented occupational exposure to carcinogens presents a unique situation for evaluating this approach. Analysis usually involves working with paraffin-embedded tissues, fixed under conditions suboptimal for genetic analysis and stored for many years, since frozen tissues are not available in sufficient numbers. The particular methodological problems encountered with fixed samples are discussed here, using as illustration an ongoing study of oncogene and tumor suppressor gene mutations in archived bladder tumors of workers exposed to aromatic amines and nonexposed patients.

Base Sequence↗
PubMed1995

Leukemia inhibitory factor (LIF) inhibits angiogenesis in vitro.

Using an in vitro model in which endothelial cells can be induced to invade a three-dimensional collagen gel to form capillary-like tubular structures, we demonstrate that leukemia inhibitory factor (LIF) inhibits angiogenesis in vitro. The inhibitory effect was observed on both bovine aortic endothelial (BAE) and bovine microvascular endothelial (BME) cell, and occurred irrespective of the angiogenic stimulus, which included basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), the synergistic effect of the two in combination, or the tumor promoter phorbol myristate acetate. LIF inhibited bFGF- and VEGF-induced proliferation in BAE and BME cells. In addition, LIF inhibited BAE but not BME cell migration in a conventional two-dimensional assay. Finally, LIF decreased the proteolytic activity of BAE and BME cells and increased their expression of plasminogen activator inhibitor-1. These results demonstrate that LIF inhibits angiogenesis in vitro, an effect that can be correlated with a LIF-mediated decrease in endothelial cell proliferation, migration and extracellular proteolysis.

Adrenal Cortex↗
PubMed1995

Hepatocyte growth factor stimulates extensive development of branching duct-like structures by cloned mammary gland epithelial cells.

Although epithelial-mesenchymal (stromal) interactions are thought to play an important role in embryonic and postnatal development of the mammary gland, the underlying mechanisms are still poorly understood. To address this issue, we assessed the effect of fibroblast-derived diffusible factors on the growth and morphogenetic properties of a clonally derived subpopulation (clone TAC-2) of normal murine mammary gland (NMuMG) epithelial cells embedded in collagen gels. Under control conditions, TAC-2 mammary gland epithelial cells suspended within collagen gels formed either irregularly shaped cell aggregates or short branching cord-like structures. Addition of conditioned medium from Swiss 3T3 or MRC-5 fibroblasts dramatically stimulated cord formation by TAC-2 cells, resulting in the development of an extensive, highly arborized system of duct-like structures, which in appropriate sections were seen to contain a central lumen. The effect of fibroblast conditioned medium was completely abrogated by antibodies against hepatocyte growth factor (also known as scatter factor), a fibroblast-derived polypeptide that we have previously shown induces tubulogenesis by Madin-Darby canine kidney epithelial cells. Addition of exogenous recombinant human hepatocyte growth factor to collagen gel cultures of TAC-2 cells mimicked the tubulogenic activity of fibroblast conditioned medium by stimulating formation of branching duct-like structures in a dose-dependent manner, with a maximal 77-fold increase in cord length at 20 ng/ml. The effect of either fibroblast conditioned medium or hepatocyte growth factor was markedly potentiated by the simultaneous addition of hydrocortisone (1 microgram/ml), which also enhanced lumen formation. These results demonstrate that hepatocyte growth factor promotes the formation of branching duct-like structures by mammary gland epithelial cells in vitro, and suggest that it may act as a mediator of the inducing effect of mesenchyme (or stroma) on mammary gland development.

Animals↗
PubMed1995

FGF-1 but not FGF-4 secreted by carcinoma cells promotes in vitro and in vivo angiogenesis and rapid tumor proliferation.

The progressive growth of solid tumors is dependent on the tumor ability to recruit new blood vessels from the surrounding host tissues. We show here that acidic Fibroblast Growth Factor (FGF-1) produced by a rat bladder carcinoma transfected cell line (NBT-II cells) is a potent inducer of angiogenesis. After injection in nude mice, NBT-II cells transfected with FGF-1 form rapidly growing carcinomas which are highly vascularized, whereas carcinoma cells producing a biologically active form of FGF-4 behave like non-producer cells. The vasculature of the tumors obtained with NBT-II cells producing a secreted form of FGF-1 is dramatically expanded but lacking in some places a complete endothelial lining. Conditioned medium from these cells induce formation of capillary-like structures in vitro, whereas those of FGF-4 and non-secreting FGF-1 producing cells failed to induce such structures. Our results indicate that the expression of FGF-1 may promote tumor growth, at least in part, by inducing angiogenesis, and that the acquired ability of tumor cells to secrete FGF-1 but not FGF-4, may result in aberrant neovascularization of the tumor.

Animals↗
PubMed1994

Mutations in the conserved regions of p53 are infrequent in betel-associated oral cancers from Papua New Guinea.

Seventy-five % of the world's oral cancers arise in developing countries. In high incidence areas of Southeast Asia such as Papua New Guinea (PNG) the major region-specific risk exposure is betel quid chewing. While it has been shown that p53 gene mutations in the conserved midregion (exons 5-9) are a common feature of oral cancers in the developed world, there is no information on this type of genetic lesion in betel quid-associated oral cancers. We examined 50 oral squamous cell carcinomas, 20 from Baltimore, MD and 30 from PNG, for mutations in exons 5-9 of the p53 gene. DNA extracted from frozen biopsies was amplified by polymerase chain reaction, the purified product was sequenced directly, and mutations were confirmed by repeating the entire procedure. Mutations were found in 3 of 30 tumors from PNG (10%), whereas 9 mutations were detected among the 20 tumors (45%) from Baltimore, MD. This difference in frequency is statistically significant (P < 0.01) by chi 2 analysis. Nuclear accumulation of p53 protein, determined by immunohistochemistry with the CM-1 antiserum, was observed in the PNG cases harboring a missense mutation of the p53 gene. In agreement with the low number of PNG cancers with mutations, only 17% of the cases from PNG were immunostain positive. To explore whether less conserved regions of the gene are preferential targets for alterations in this patient group, sequence analysis in tumors from PNG was extended to outlying regions of the gene (all of exons 4 and 10, and splice sites), but mutations in only two additional tumors were identified. The presence of human papillomavirus DNA in PNG cases was examined with a polymerase chain reaction-based procedure, and viral sequences (human papillomavirus strains 11/16) were detected in two tumors. Human papillomavirus-triggered degradation of the tumor suppressor protein is thus unlikely to be a typical pathway to p53 dysfunction in tumors from PNG.

Adult↗
PubMed1994

The endogenous oestrogen metabolite 2-methoxyoestradiol inhibits angiogenesis and suppresses tumour growth.

The formation of new blood vessels (angiogenesis) is critical for the growth of tumours and is a dominant feature in various angiogenic diseases such as diabetic retinopathy, arthritis, haemangiomas and psoriasis. Recognition of the potential therapeutic benefits of controlling pathological angiogenesis has led to a search for angiogenesis inhibitors. Here we report that 2-methoxyoestradiol, an endogenous oestrogen metabolite of previously unknown function, is a potent inhibitor of endothelial cell proliferation and migration as well as angiogenesis in vitro. Moreover, when administered orally in mice, it strongly inhibits the neovascularization of solid tumors and suppresses their growth. Unlike the angiostatic steroids of corticoid structure, it does not require the co-administration of heparin or sulphated cyclodextrins for activity. Thus, 2-methoxyoestradiol is the first steroid to have high antiangiogenic activity by itself. Our results suggest that this compound may have therapeutic potential in cancer and other angiogenic diseases.

2-Methoxyestradiol↗
PubMed1994

Modulation of bovine microvascular endothelial cell proteolytic properties by inhibitors of angiogenesis.

A tightly controlled increase in extracellular proteolysis, restricted both in time and space, is an important component of the angiogenic process, while anti-proteolysis is effective in inhibiting angiogenesis. By focussing on the plasminogen activator (PA)-plasmin system, the objective of the present studies was to assess whether previously described inhibitors of angiogenesis modify bovine microvascular endothelial cell proteolytic properties. We demonstrate that although synthetic angiostatic steroids (U-24067 and U-42129), heparin, suramin, interferon alpha-2a, and retinoic acid are all inhibitors of in vitro angiogenesis, each of these agents has distinct effects on the plasminogen-dependent proteolytic system. Specifically, angiostatic steroids and interferon alpha-2a reduce urokinase-type PA (u-PA) and PA inhibitor-1 activity, while heparin and retinoic acid increase u-PA activity. Suramin reduces cell-associated u-PA activity and greatly increases PAI-1 production at doses which induce monolayer disruption. These findings demonstrate that a spectrum of alterations in extracellular proteolysis is associated with anti-angiogenesis, and that anti-angiogenesis and anti-proteolysis are not necessarily correlated. A reduction in extracellular proteolysis would be expected to reduce invasion, whereas an increase in proteolysis might modulate the activity of inhibitory cytokines, which in turn could reduce endothelial cell proliferation and migration and inhibit angiogenesis. The spectrum of effects on different elements of the PA system observed in response to the agents assessed suggests that the role of modulations in extracellular proteolytic activity in anti-angiogenesis is likely to be varied and complex.

Adrenal Cortex↗
PubMed1994

Induction of specific cytochrome P450s involved in aflatoxin B1 metabolism in hepatitis B virus transgenic mice.

The relative roles of hepatitis B virus (HBV) and aflatoxin and their possible mechanism of interaction in the etiopathogenesis of hepatocellular carcinoma (HCC) are not understood. One hypothesis is that viral infection and associated liver injury alter expression of carcinogen-metabolizing enzymes. We tested this hypothesis in an HBV-transgenic mouse model in which a synergistic interaction occurs between aflatoxin B1 (AFB1) and HBV in the induction of HCC (Sell et al., Cancer Res 51:1278-1285, 1991). In this transgenic mouse lineage, overproduction of the HBV large envelope protein results in progressive liver cell injury, inflammation, and regenerative hyperplasia. Initially, two cytochrome P450s of importance in AFB1 metabolism in the mice were identified, namely Cyp2a-5 and Cyp3a, using specific antibodies and chemical inhibitors. The expression of these P450 isoenzymes and an alpha-class glutathione S-transferase (GST) isoenzyme, YaYa, were examined. Increased expression and altered distribution of Cyp2a-5 were demonstrated, by immunohistochemical analysis, to be associated with the development of liver injury in mice and to increase with age between 1 and 12 months. Cyp3a expression was also increased in HBV-transgenic mice, but the increase was not as clearly related to age. GST YaYa levels were the same in HBV-transgenic mice and their nontransgenic littermates of all ages. These results show that expression of specific cytochrome P450s is altered in association with overexpression of HBV large envelope protein and liver injury in this model. This may have general relevance to human HCC, the etiology of which is associated with a diverse range of liver-damaging agents.

Aflatoxin B1↗
PubMed1994

In vitro angiogenic and proteolytic properties of bovine lymphatic endothelial cells.

The aim of the present study was to determine whether angiogenic cytokines, which induce neovascularization in the blood vascular system, might also be operative in the lymphatic system. In an assay of spontaneous in vitro angiogenesis, endothelial cells isolated from bovine lymphatic vessels retained their histotypic morphogenetic properties by forming capillary-like tubes. In a second assay, in which endothelial cells could be induced to invade a three-dimensional collagen gel within which they formed tube-like structures, lymphatic endothelial cells responded to basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) in a manner similar to what has previously been observed with endothelial cells derived from the blood vascular system. Finally, since angiogenesis is believed to require extracellular proteolytic activity, we investigated the effects of bFGF and VEGF on lymphatic endothelial cell proteolytic properties by focussing on the plasminogen activator (PA) system. bFGF and VEGF increased urokinase, urokinase receptor, and tissue-type PA expression. This was accompanied by an increase in PA inhibitor-1, which is thought to play an important permissive role in angiogenesis by protecting the extracellular matrix against excessive proteolytic degradation. Taken together, these results demonstrate that with respect to in vitro morphogenetic and proteolytic properties, lymphatic endothelial cells respond to the previously described angiogenic factors, bFGF and VEGF, in a manner very similar to what has been described for endothelial cells derived from the blood vascular system.

Animals↗
PubMed1994

Duck hepatitis B virus infection, aflatoxin B1 and liver cancer in domestic Chinese ducks.

The oncogenicity of Duck hepatitis B virus (DHBV) is unclear since hepatocellular carcinomas (HCCs) have been reported only in domestic ducks in Qidong, an area of China where hepatitis B virus (HBV) and aflatoxin B1 (AFB1) are risk factors for liver cancer in man. In order to better define the association between DHBV infection, AFB1 and HCC we analysed a series of 16 duck liver samples collected from local farms in Qidong. HCC was found in eight and cirrhosis in one of these samples. Furthermore bile duct proliferation, characteristic of AFB1 exposure in ducks and other animal species, was found in these ducks. Integration of DHBV DNA into cellular DNA was observed in only one out of four DHBV positive HCCs, indicating that viral integration is not prerequisite for tumour development. In four remaining HCCs the polymerase chain reaction (PCR) failed to show any DHBV DNA suggesting that liver tumours do occur in polymerase chain reaction (PCR) failed to show any DHBV DNA suggesting that liver tumours do occur in these ducks in the absence of DHBV infection. In addition, AFB1-DNA adducts were detected by hplc-immunoassay in one such DHBV-negative tumour. In summary we demonstrate that risk factors other than DHBV, including AFB1 exposure, may be important in duck liver carcinogenesis in Qidong.

Aflatoxin B1↗
PubMed1994

p53 mutations at A:T base pairs in angiosarcomas of vinyl chloride-exposed factory workers.

Mutations in the p53 tumor suppressor gene are commonly found in the major human cancers and the mutational spectrum in some cancer types is consistent with the genotoxic effects of the associated environmental risk factors. Thus far there is little information on p53 mutations in cancers of factory workers with a history of carcinogen exposure in the workplace. Occupational exposure to vinyl chloride causes liver angiosarcomas (ASL) and also increases the risk of several other cancers. Loss of p53 function in osteo- and fibrosarcomas can occur by two different mechanisms, p53 mutation and amplification of the MDM2 gene. We examined tumors from five vinyl chloride-exposed patients, four with ASL and one with hepatocellular carcinoma (HCC), for evidence of MDM2 proto-oncogene amplification or p53 mutation in exons 5-8. Amplification of MDM2 was not found, but in two of the angiosarcomas an A:T to T:A missense mutation was detected. p53 sequence analysis of vinyl chloride associated cancers may provide valuable information on the relationship between carcinogen exposure and DNA damage in cancer-related genes.

Adenine↗
PubMed1994

DNA methylation adduct formation and H-ras gene mutations in progression of N-butyl-N-(4-hydroxybutyl)nitrosamine-induced bladder tumors caused by a single exposure to N-methyl-N-nitrosourea.

After receiving 500 p.p.m. N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) in their drinking water for an initial 10 weeks, rats were given a single i.p. injection of N-methyl-N-nitrosourea (MNU) at a dose of 50 mg/kg body wt at week 20 (at a stage when bladder tumor development had already occurred), and then maintained until they were killed at week 40. Three and six hours after the MNU injection, the DNA methylation adducts, O6-methyldeoxyguanine (O6-medG) and 7-methyldeoxyguanine (7-medG), were immunohistochemically revealed to be markedly more frequent in urothelial preneoplasias or neoplasias than in normal cells. These adducts were rapidly repaired, and although 7-dmeG in tumor cells still persisted after 72 h, they appeared essentially to have returned to normal levels. At the termination, conversion of transitional cell carcinomas (TCC) to squamous cell carcinomas (SCC) of the urinary bladder was significantly increased in the BBN+MNU group. The extent of invasion was also significantly greater with the additional MNU treatment. Expression of p21 protein, detected by immunohistochemistry, was comparable between the groups. Mutations in the H-ras gene were observed in one case each of the BBN and BBN+MNU groups, and both cases showed a G:C to A:T transition at codon 12. The present study thus suggested that while an additional single treatment with MNU of rats bearing BBN-induced bladder neoplasias is associated with significant, possibly mutation-dependent tumor progression, H-ras mutations are not necessary events.

Administration, Oral↗
PubMed1994

Absence of p53 mutation at codon 249 in duck hepatocellular carcinomas from the high incidence area of Qidong (China).

Dietary aflatoxin and hepatitis B virus infection may play a role in generating the p53 tumor suppressor gene codon 249 hotspot mutation found in human hepatocellular carcinomas (HCCs) from Qidong (China) and southern Africa. No data are available on the HCC site-specific mutation of the p53 gene in hepadnavirus-infected animals exposed to AFB1. We have searched for the presence of p53 gene codon 249 mutations in both duck hepatitis B virus (DHBV) positive and negative HCCs of domestic ducks from Qidong, where the human p53 hotspot is so prevalent, as well as in duck HCCs experimentally induced by AFB1. Direct sequencing of DNA amplification products encompassing p53 codon 249 did not reveal any mutations in 11 HCCs from Qidong ducks, regardless of the status of DHBV infection. In addition no mutation was detected in four HCCs from AFB1-treated ducks. This contrasts with the human data; however, in humans, the mutation and the preferential binding of AFB1 to codon 249 occurs at the third nucleotide G, while in duck, the codon 249 lacks this G residue. The DNA sequence of adjacent codons is also different in the two species even though the amino acid sequence is identical. This may explain the low frequency of mutation we have observed. In addition, species differences in metabolism and DNA repair could influence the occurrence of codon 249 mutations.

Aflatoxin B1↗
PubMed1994

TGF-beta 1 influences the relative development of the exocrine and endocrine pancreas in vitro.

Pancreatic rudiments from E12.5 mouse embryos undergo extensive development and differentiation when cultured in three-dimensional gels of extracellular matrix proteins for up to 12 days. Whereas collagen gels promote the formation of numerous exocrine acini and relatively small clusters of endocrine cells, in basement membrane (EHS) matrices the development of endocrine cells is dramatically favoured over that of acinar tissue. Buds embedded in a collagen gel contiguous to an EHS gel also fail to develop acini, suggesting the involvement of diffusible factor(s). Addition of cytokines to cultures of pancreatic buds in collagen gels modifies the relative proportions of the epithelial components of the gland. In the presence of EGF the proportion of the tissue occupied by ducts overrides that of acinar structures, whereas the endocrine portion of the tissue is not significantly modified. TGF-beta 1 partially mimicks the effect of EHS matrix in inhibiting the development of acinar tissue without decreasing the amount of ducts and mesenchyme; TGF-beta 1 also promotes the development of endocrine cells, in particular of insulin-containing beta cells and of cells expressing genes of the PP-fold family. These results show that cytokines can modulate the development of the pancreas and suggest a role for TGF-beta 1 in regulating the balance between the acinar and endocrine portions of the gland in vivo. More generally, they are compatible with the notion that, during organogenesis, cytokines act as paracrine factors responsible for the development and maintenance of appropriate proportions of different tissue constituents.

Animals↗
PubMed1994

Database of p53 gene somatic mutations in human tumors and cell lines.

A data base is described in which over 2,500 mutations in the p53 gene of human tumors and tumor cell lines are compiled from a systematic search of reports published before 1 January 1994. Data from 1994 are being added intermittently, with a systematic search and update scheduled for December, 1994. The compilation has been deposited with the EMBL Data Library and is available in electronic form free of charge. This report contains a rationale for the compilation, a brief summary of the major findings and a description of the data base.

DNA Mutational Analysis↗