Search PubMed⌕ Search

Biomedical subjects

R Montesano

Publications and source records attributed to R Montesano.

At least 199 records · Page 11Linked to original sources
PubMed1985

DNA ethylation in target and non-target organs of hamsters and rats treated with diethylnitrosamine.

Kinetics of ethylation of target and non-target organ DNA in vivo by diethylnitrosamine (DEN) was compared in rats and Syrian golden hamsters, since published reports indicate a single dose of DEN induces both kidney and liver tumors in rats and almost exclusively respiratory tract tumors in hamsters. Following treatment with 200 mg DEN/kg, 7-ethylguanine (7-etG) was lost more rapidly from hamster than from rat liver DNA, while O6-ethylguanine (O6-etG) persisted longer in hamster than in rat liver DNA. DNA ethylation was not detected in rat lung (non-target organ), while both 7-etG and O6-etG were quantitated in hamster lung (target organ) following DEN treatment. DNA ethylation in rat kidney DNA was approximately 1/10 of that in liver by 200 mg DEN/kg, and the persistence of 7-etG and O6-etG differed only slightly in these tissues. Ethylation of hamster liver DNA by DEN at doses between 20 and 200 mg/kg, as measured by 7-etG and O6-etG was proportional to the dose of carcinogen up to 160 mg/kg; at larger doses DNA ethylation sharply increased. Differences in the persistence of O6-etG between DEN-treated rats and hamsters cannot solely account for species differences in the organotropism of DEN carcinogenesis.

Alkylation↗
PubMed1985

Repair of DNA alkylation adducts in mammalian cells.

Carcinogenic alkylating agents, including nitrosamines, are able to alkylate DNA at various sites. This review presents evidence of the high degree of specificity in the type of DNA damage induced by various N-nitroso compounds and in the DNA repair processes among tissues or cells of different species. The O6-alkylguanine DNA alkyltransferase activity in various human and rodent tissues is discussed as well as the detection of O6-methylguanine in human DNA, using monoclonal antibodies and radioimmunoassay. The relevance of these findings to the mechanisms of cancer induction by nitrosamines is discussed.

Alkylating Agents↗
PubMed1985

O6-Alkylguanine DNA alkyltransferase activity in monkey, human and rat liver.

Previous studies have shown that in in vitro assays using methylated DNA substrates, human and rat liver fractions were able to catalyse the repair of the promutagenic lesion O6-methylguanine (O6-meG). These studies have been extended to include monkey liver fractions and the repair of O6-ethylguanine (O6-etG). Human and monkey liver fractions have similar capacities for the repair of O6-meG and are 8-10 times more active than rat liver fractions. In all species examined, the liver extracts showed a lower capacity to repair O6-etG and the rate of repair was dependent on the degree of modification of the DNA substrate and the protein concentration used. The in vitro metabolism of dimethylnitrosamine (DMN), an alkylating carcinogen, by liver slices, has been previously demonstrated in these three species, but the activity in monkey liver was considerably lower compared with human and rat liver. The lack of evidence of a carcinogenic effect of DMN in the monkey liver could be correlated to this low capacity to metabolise DMN and the high O6-alkylguanine-DNA alkyltransferase levels in this organ.

Animals↗
PubMed1985

Repair of O4-methyldeoxythymidine residues in DNA by mammalian liver extracts.

The enzymatic repair of O4-methyldeoxythymidine (O4-MedT) by mammalian liver extracts was investigated in vitro using double-stranded poly[d(A-T) X d(A-T)] alkylated with N-[methyl-3H]-N-nitrosourea as a substrate. The removal of O4-MedT by monkey liver extracts was proportional to the protein concentration of the reaction mixture up to approximately 4 mg/ml, while kinetic analysis revealed the repair reaction reached a plateau by 2 h at 37 degrees C. The removal of O4-MedT could not be accounted for by contaminating nuclease activity, for greater than 90% of the 3-methyldeoxythymidine (3-MedT) was always recovered, whereas O4-MedT declined to as little as 25% of control values. A DNA-glycoslyase is not involved in the repair of O4-MedT, for O4-methylthymine, the free base, was not detected in supernatants from monkey liver reaction mixtures following incubation with substrate and precipitation of macromolecules with ethanol, even though analysis of nucleic acid digests of these assays revealed O4-MedT was lost from the substrate. The order of O4MedT repair activity in human, monkey, partially hepatectomized rat and methylguanine DNA-methyltransferase activity, with human and monkey most active, partially hepatectomized rat intermediate in activity and control rat least active. Our findings suggest the same, or a similarly regulated mammalian liver DNA-methyltransferase repairs O6-methylguanine and O4-MedT residues in DNA, yet the affinity of the enzyme(s) differs for these promutagenic adducts, as we estimate 30-50 O6-methylguanines would be repaired for every one O4-MedT repaired.

Animals↗
PubMed1985

Modulation of repair of O6-methylguanine in parenchymal and nonparenchymal liver cells of rats treated with dimethylnitrosamine.

Previous studies have shown that chronic treatment of rats with dimethylnitrosamine (DMN) (2 mg/kg for 3 weeks) results in increased repair of O6-methylguanine (O6-mGua) in liver DNA. The experiments reported here try to determine if this increased repair is confined to one or more cell populations of the liver. Liver cells [parenchymal (PC) and nonparenchymal (NPC)] were separated by elutriation centrifugation at various times after the last administration of DMN. The in vivo alkylation studies show that at any time after a dose of [14C]DMN (2 mg/kg) the level of O6-mGua in PC cells of DMN pretreated rats was much lower than in the same cell population from control rats receiving only a single dose of DMN. In contrast, the pretreatment schedule resulted in no change in the levels of this DNA adduct in NPC cells. These results were confirmed by the determination of the levels of O6-methyldeoxyguanosine by radioimmunoassay in DNA from PC or NPC cells of rats similarly either pretreated for 3 weeks with DMN or receiving a single dose of DMN (2 mg/kg). The in vitro measurements of O6-mGua DNA alkyltransferase activity, using alkylated DNA as substrate, also show a higher activity of this repair enzyme in PC cells. The DMN pretreatment resulted in a 25-fold difference in O6-mGua DNA alkyltransferase activity between the two cell populations of the liver.

Alkylation↗
PubMed1984

Lack of mutagenicity of synthetic pyrethroids in Salmonella typhimurium strains and in V79 Chinese hamster cells.

Seven pyrethroids, i.e., cypermethrin, permethrin, deltamethrin, bioresmethrin, resmethrin, cismethrin and fenvalerate, were not found to be mutagenic in (a) Salmonella typhimurium strains TA100 or TA98 in the presence or absence of a rat liver activation system using the plate incorporation assay and fluctuation tests, or (b) V79 Chinese hamster cells in the presence or absence of hepatocytes.

Animals↗
PubMed1984

Leukocyte interleukins induce cultured endothelial cells to produce a highly organized, glycosaminoglycan-rich pericellular matrix.

We report here that interleukins have a dramatic effect on extracellular matrix production by cultured endothelial cells. Human umbilical vein endothelial cells incubated with growth media conditioned by lectin-activated human peripheral blood mononuclear leukocytes undergo marked changes in cell shape and elaborate a highly organized extracellular material that is not detectable in untreated cultures. This material has the following characteristics: (a) it is not recognizable by electron microscopy unless the cationic dye, Alcian blue, is added to the fixative; (b) it is visualized as a network of branching and anastomosing fibrils of various thickness that can be resolved into bundles of fine filaments; (c) it is associated with the cell surface, extends between contiguous cells, and coats the culture substrate; (d) it is removed by digestion with glycosaminoglycan-degrading enzymes, such as crude heparinase and chondroitinase ABC. These results demonstrate that soluble factors released by activated peripheral blood mononuclear leukocytes (interleukins) stimulate cultured human umbilical vein endothelial cells to produce a highly structured pericellular matrix containing glycosaminoglycans (probably chondroitin sulfate and/or hyaluronic acid) as a major constituent. We speculate that this phenomenon corresponds to an early step of angiogenesis as observed in vivo as a consequence of interleukin release.

Alcian Blue↗
PubMed1984

Identification of chemicals carcinogenic to man.

Although both the epidemiologic and experimental studies have led to the identification of chemical carcinogens, the limitations in epidemiologic approaches and the need for primary prevention of cancer require a greater reliance on experimental studies. Long-term carcinogenicity studies in experimental animals have been instrumental in identifying chemicals with carcinogenic activity, and, in some cases, the experimental evidence has preceded the epidemiologic evidence (for 4-aminobiphenyl, aflatoxin B1, diethylstilbestrol, melphalan, mustard gas, and vinyl chloride). A better understanding of the multistage process of carcinogenesis and the findings from various short-term tests available more recently may provide a more solid basis for extrapolating experimental findings to man.

Animals↗
PubMed1984

An enzyme-linked immunosorbent procedure for assaying aflatoxin B1.

A polyclonal rabbit antibody preparation against aflatoxin B1 (AFB1), produced by immunization with a bovine serum albumin-AFB1 conjugate, has been used to develop an enzyme-linked immunosorbent assay (ELISA). AFB1-ovalbumin, obtained by reacting either AFB1-8,9-dichloride or -8,9-dibromide with ovalbumin, was used to coat each well of a polyvinyl microtitre dish. Rabbit antibody, diluted 1:100 000, was added to each well and left to attach for 90 min, then the excess was washed off with phosphate-buffered saline/Tween. Anti-rabbit IgG coupled to peroxidase was then added, left for 90 min and the excess washed away. Residual peroxidase activity was assayed using tetramethylbenzidine as substrate; the reaction was quenched at the end of the 15-min incubation period with 2 N sulfuric acid. Inhibitor studies to assay AFB1 and related compounds in urine involved prior incubation of the diluted anti-AFB1 antibody with inhibitor for 60 min at 37 degrees C, prior to dispensing into the multi-well plates. Inhibitor studies with AFB1 showed inhibition over a concentration range of 10(-1) to 10(-5) micrograms/ml. The minimum detectable concentration was approximately 10(-5) micrograms/ml (0.032 pmol/ml). Anti-AFB1 antibody was also inhibited by iro-AFB1-DNA, one of the forms of AFB1-DNA, as well as by AFB1-guanine. Undiluted urine from normal subjects inhibited antibody binding to plates; this inhibition could be prevented by either dilution or extraction procedures.(ABSTRACT TRUNCATED AT 250 WORDS)

Aflatoxin B1↗
PubMed1984

Enhanced repair of O6-alkylguanine in mammalian tissues.

N-nitroso compounds and related alkylating agents react with several nucleophilic sites in DNA. One of the products, O6-alkylguanine, may be responsible for the carcinogenic effects of such agents as it can mispair with thymine during DNA replication, thereby affecting the integrity of the genome and consequently the repair of this lesion could be an important determinant in carcinogenesis. The use of both in vivo and in vitro procedures for the assay of the activity of this repair system have shown that it can be enhanced in rat liver by treatment with a variety of agents while in other species, the system is not only less effective but does not appear to respond to such treatments. Differences in the capacity and inducibility of this repair system may be related to the susceptibility of different species to tumour induction by chronic administration of nitrosamines.

2-Acetylaminofluorene↗
PubMed1983

Specialization of the macrophage plasma membrane at sites of interaction with opsonized erythrocytes.

We incubated mouse peritoneal macrophages for 3-8 min at 37 degrees C with antibody-coated sheep erythrocytes and examined regions of close interaction between the two cell types by electron microscopy. At sites of focal macrophage-erythrocyte contact we observed a distinctive specialization of the macrophage plasma membrane consisting of a prominent subplasmalemmal band of electron-dense material, approximately 25-35 nm in thickness. In many instances, this band showed a periodic substructure similar to that seen in clathrin coats. Moreover, many slender erythrocyte processes penetrated into invaginations of the macrophage surface which were bristle-coated at their blind extremity. As previously shown for clathrin-coated pits, the segments of the macrophage plasma membrane beneath which the defense material was found were selectively resistant to the membrane-perturbing effect of the antibiotic, filipin. This structural specialization of the macrophage plasma membrane at sites of ligand-receptor interaction during immune phagocytosis of antibody-coated erythrocytes may represent the morphological counterpart of the zipper mechanism of phagocytosis previously demonstrated by functional studies.

Animals↗
PubMed1983

Collagen matrix promotes reorganization of pancreatic endocrine cell monolayers into islet-like organoids.

To evaluate the capacity of pancreatic endocrine cells to reassociate in vitro according to the characteristic topographical pattern observed in the islets of Langerhans in situ, we cultured cells dissociated from neonatal rat pancreas within a three-dimensional collagen matrix. Cell monolayers grown on the surface of collagen gels were covered with a second layer of collagen. This induced the monolayers of endocrine cells to reorganize into smooth-contoured, three-dimensional aggregates, in which non-B cells (identified by electron microscopy and immunofluorescence) had a preferential distribution at the periphery, whereas B cells were concentrated in a central position. These results show that cultured pancreatic endocrine cells have the capacity to reassociate into islet-like organoids in vitro, and that collagen matrices may have a permissive effect on the expression of this potential.

Animals↗
PubMed1983

In vitro rapid organization of endothelial cells into capillary-like networks is promoted by collagen matrices.

We have studied the behavior of cloned capillary endothelial cells grown inside a three dimensional collagen matrix. Cell monolayers established on the surface of collagen gels were covered with a second layer of collagen. This induced the monolayers of endothelial cells to reorganize into a network of branching and anastomosing capillary-like tubes. As seen by electron microscopy, the tubes were formed by at least two cells (in transverse sections) delimiting a narrow lumen. In addition, distinct basal lamina material was present between the abluminal face of the endothelial cells and the collagen matrix. These results showed that capillary endothelial cells have the capacity to form vessel-like structures with well-oriented cell polarity in vitro. They also suggest that an appropriate topological relationship of endothelial cells with collagen matrices, similar to that occurring in vivo, has an inducive role on the expression of this potential. This culture system provides a simple in vitro model for studying the factors involved in the formation of new blood vessels (angiogenesis).

Animals↗