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Biomedical subjects

E Huberman

Publications and source records attributed to E Huberman.

At least 109 records · Page 6Linked to original sources
PubMed1982

Induction of differentiation of human promyelocytic leukemia (HL-60) cells by teleocidin and phorbol-12-myristate-13-acetate.

Human promyelocytic leukemia cells (HL-60) were induced to differentiate into macrophage-like cells in a dose-dependent manner by the tumor promoters phorbol-12-myristate-13-acetate and teleocidin, a non-phorbol ester promoter. An HL-60 cell variant, designated as R-59, which is resistant to differentiation induction by phorbol-12-myristate-13-acetate was also resistant to differentiation induction by teleocidin. Differentiation was determined by increases in the percent of morphologically mature cells and in lysozyme and nonspecific esterase activities. Both compounds inhibited the growth of HL-60 cells by blocking them from entering the synthesis phase of the cell cycle with an accumulation of cells after 48 h in G1 phase. No such effects were observed in the R-59 cells. They were, however, as susceptible as the parent HL-60 cells, to inducers which are not considered to be tumor promoters such as dimethylsulfoxide and retinoic acid. However, these inducers cause the HL-60 and R-59 cells to differentiate into granulocyte-like cells. These results indicate that teleocidin produces in both the HL-60 and R-59 cells effects which are similar to those cause by phorbol-12-myristate-13-acetate. The possibility that agents producing such effects in these two cell types may represent potential tumor promoters is discussed.

Alkaloids↗
PubMed1982

Cell-mediated mutagenesis and tumor-initiating activity of the ubiquitous polycyclic hydrocarbon, cyclopenta[c,d]pyrene.

The ubiquitous polycyclic aromatic hydrocarbon (PAH), cyclopenta[c,d]pyrene (CPP), was tested to determine its mutagenicity for 6-thioguanine and ouabain resistance in Chinese hamster V79 cells and its tumor-initiating activity in the skin of the tumor susceptible Sencar mice. The potent carcinogen/mutagen, benzo[a]pyrene (BP), was included for comparison. Inasmuch as V79 cells do not metabolize PAHs, mutagenesis was tested both in the presence and in the absence of X-irradiated golden hamster embryo fibroblasts capable of metabolizing PAH. Neither CPP nor BP showed mutagenicity for V79 cells in the absence of the embryo cells. In the presence of these cells (in the cell-mediated assay) both PAHs elicited, in a dose dependent manner, a cytotoxic and mutagenic response in V79 cells. CPP was however less active than BP in inducing both of these responses. At the optimal expression time and at the dose range of 0.1-1 microgram/ml, CPP induced 2-8 6-thioguanine resistant mutants per 10(5) colony forming cells compared to 9-50 mutants induced by BP. Similarly, these doses of CPP induced 1-9 ouabain resistant mutants per 10(6) colony forming cells compared to 7-75 mutants induced by BP. CPP was also active in initiating skin tumors in approximately 60% of the mice at 200 micrograms, the highest dose tested. BP was more efficient in tumor initiation and yielded a similar response with 10 micrograms. These results indicate that CPP and BP elicit, in the cell mediated assay, a mutagenic response similar to the activity of these PAH in the skin of Sencar mice.

Animals↗
PubMed1982

The control of phospholipid methylation by phorbol diesters in differentiating human myeloid HL-60 leukemia cells.

Treatment of human HL-60 promyelocytic leukemia cells with phorbol-12-myristate-13-acetate (PMA), a tumor promoter and inducer of differentiation, stimulated the incorporation of label from L-[methyl-3H]methionine into the cellular phospholipids. Such a stimulation of phospholipid methylation was not observed in an HL-60 cell variant that is resistant to phorbol ester-induced differentiation. Enhanced methylation of phospholipids was detected 6 h after treatment and reached a maximum level of about twice the control level at 24-48 h. The degree of phospholipid methylation was dependent on the phorbol ester dose. The stimulation in phospholipid methylation by PMA was confirmed by measuring the activity of phosphatidylethanolamine methyltransferase in cellular lysates. After 24 or 48 h of exposure, the enzyme activity was elevated in the HL-60 cell lysates but not in the resistant cells. Phospholipid methylation was also stimulated after treatment of the HL-60 cells with the phorbol diester phorbol-12,13-dibutyrate or teleocidin, which is not a phorbol ester compound. These two chemicals and PMA are tumor promoters and inducers of cell differentiation in the HL-60 cells. Phorbol-12,13-diacetate and 4-O-methyl PMA, which are not tumor promoters or inducers of cell differentiation in the HL-60 cells, did not stimulate phospholipid methylation. The possible role of enhanced phospholipid methylation in cell differentiation of the HL-60 by these chemicals is discussed.

Alkaloids↗
PubMed1982

Cell-mediated mutagenicity in Chinese hamster V79 cells of dibenzopyrenes and their bay-region fluorine-substituted derivatives.

The polycyclic aromatic hydrocarbons dibenzo(a,i)pyrene and dibenzo(a,h)pyrene, each of which possesses two bay regions, and their bay-region difluorinated derivatives were tested for mutagenicity for ouabain and 6-thioguanine resistance in Chinese hamster V79 cells. Since V79 cells do not metabolize polycyclic aromatic hydrocarbons, mutagenesis was tested in both the presence and the absence of golden hamster embryo cells capable of metabolizing polycyclic aromatic hydrocarbons. Neither of the dibenzopyrenes nor their fluorinated derivatives were mutagenic in the absence of the golden hamster embryo cells. In the presence of these cells (cell-mediated assay), both dibenzopyrenes were mutagenic, whereas the difluorinated derivatives, 2,10-difluorodibenzo-(a,i)pyrene and 3,10-difluorodibenzo(a,h)pyrene, either were inactive or exhibited (on a dose basis) a weak response. However, the mutagenicity of the dibenzopyrenes was eliminated when they were coincubated with 7,8-benzoflavone, a mixed-function oxidase inhibitor. The results suggest that metabolic oxidation of these polycyclic aromatic hydrocarbons at the bay region (presumably to diol-epoxides) is required for a mutagenic response in the cell-mediated assay.

Animals↗
PubMed1981

Mutagenesis by chemical agents in V79 chinese hamster cells: a review and analysis of the literature. A report of the Gene-Tox Program.

The report reviews and evaluates the current literature (about 125 primary publications) on chemically induced specific locus mutations in the V79 Chinese hamster lung cell line. The V79 cell is convenient to use for mutagenesis studies since it has a rapid growth rate, high plating efficiency, and a stable karyotype. Mutation can be easily measured at either the hypoxanthine-guanine phosphoribosyl transferase or the Na+/K+ ATPase locus, both of which have been well characterized. Other less-studied markers are also described. We discuss the protocols for quantitative mutation studies including measurements of cytotoxicity, mutant expression times, mutant selection agents, cell densities during selection, and the stability and verification of mutant phenotypes. Mutations in the V79 cells by chemicals that require activation can be tested after their metabolism by cell homogenates or by intact cells, and the results with each type of activation are compared. For purposes of analysis, we classified a compound as mutagenic if it induced a mutation frequency that is at least 3 times higher than the spontaneous mutant frequency reported for that specific experiment. By this criterion two-thirds of the chemicals analyzed were mutagenic--; 11% with and 55% without metabolic activation. Of the 191 chemicals examined; 119 were polycyclic aromatic hydrocarbons; 25 were nitro or nitroso compounds, 9 were alkyl halides; 7 were purine or pyrimidine derivatives and the remaining 31 were from other chemical classes. We also defined mutagenic potency as the concentration of a compound that increases the mutant frequency by 10 times the spontaneous frequency. Mutagenic potencies of the compounds examined varied over a range of 5 X 10(6). We have also found large interlaboratory variations in the mutagenic potencies. Such variation in potency could be reduced by normalizing the results to a standard mutagen such as N-methyl-N'-nitro-N-nitrosoguanidine. The role of the V79 assay in mutagenicity and carcinogenicity testing is discussed and recommendations are suggested for future investigation.

Animals↗
PubMed1981

Alterations in polyamine levels induced by phorbol diesters and other agents that promote differentiation in human promyelocytic leukemia cells.

Polyamine levels were evaluated in human HL-60 promyelocytic leukemia cells after treatment with inducers of terminal differentiation. Differentiation in these cells was determined by increases in the percentage of morphologically mature cells and in lysozyme activity. Treatment of the HL-60 cells with phorbol 12-myristate-13-acetate (PMA), phorbol 12,13-didecanoate or other inducers of terminal differentiation such as dimethylsulfoxide and retinoic acid resulted in increased levels of putrescine. However, no increase in putrescine could be detected after PMA treatment of a HL-60 cell variant that exhibited a decreased susceptibility to PMA-induced terminal differentiation. Similarly, no increase in putrescine was observed with two non-tumor-promoters (phorbol 12,13-diacetate and 4-O-methyl-PMA) or with anthralin, a non-phorbol tumor promoter. In addition to enhancing putrescine levels, PMA also increased the amount of spermidine and decreased the amount of spermine. The increase in putrescine and spermidine preceded the expression of the various differentiation markers. Unlike the changes observed in the polyamine levels after PMA treatment, the activities of ornithine and S-adenosylmethionine decarboxylases, which are polyamine biosynthetic enzymes, did not significantly change. alpha-Methylornithine and alpha-difluoromethylornithine and methylglyoxal bis(guanylhydrazone), which are inhibitors of the polyamine biosynthetic enzymes, did not affect differentiation in control or PMA-treated cells. Because of these observations, we suggest that the change in polyamine levels involve biochemical pathways other than the known biosynthetic ones. By-products of these pathways may perhaps be the controlling factors involved in the induction of terminal differentiation in the HL-60 and other cell types as well.

Cell Differentiation↗
PubMed1981

Down regulation of specific binding of [20-3H]phorbol 12,13-dibutyrate and phorbol ester-induced differentiation of human promyelocytic leukemia cells.

Binding of [20-3H]phorbol 12,13-dibutyrate ([3H]PDB) to intact human promyelocytic leukemia cells susceptible (HL-60) or resistant (R-35) to phorbol ester-induced differentiation was characterized. Specific binding of [3H]PDB to both HL-60 and R-35 cells at 37 degrees C reached a maximum within 15-20 min. Maximal specific [3H]PDB binding to HL-60 cells was followed by a decline (down regulation) of radioactivity. This down regulation was temperature dependent, because no loss of radiolabel occurred by 1 hr at 4 degrees C. The down regulation of bound [3H]PDB seen in HL-60 cells at 37 degrees C was not observed with R-35 cells. Prior exposure of the HL-60 cells but not of R-35 cells to 1 microM phorbol 12-myristate 13-acetate for 90 min at 37 degrees C caused a marked reduction in the specific binding of [3H]PDB. When [3H]PDB binding was carried out at 4 degrees C, [3H]PDB bound to both cell types in a rapid, specific, and reversible manner. At equilibrium, HL-60 and R-35 cells were found to contain almost the same number of binding sites, which had dissociation constants of about 50 nM, indicating that the failure of R-35 cells to undergo PDB-induced differentiation was not associated with any change in the affinity or in the number of [3H]PDB binding sites. These results indicate that the down regulation of specific [3H]PDB binding may be a crucial early event in the control of phorbol ester-induced terminal differentiation in HL-60 cells. Furthermore, we suggest that such down regulation may be involved in other cellular and biochemical effects of phorbol diester tumor promoters.

Binding Sites↗
PubMed1981

Mutagen-induced resistance to mycophenolic acid in hamster cells can be associated with increased inosine 5'-phosphate dehydrogenase activity.

Cell variants resistant to the cytotoxic effect of mycophenolic acid, an inhibitor of IMP dehydrogenase (IMP:NAD+ oxidoreductase, EC 1.2.1.14), were selected by a one-step procedure from Chinese hamster V79 cells. The frequency of these variants was increased in a dose-dependent manner after treatment with the mutagen N-methyl-N'-nitro-N-nitrosoguanidine and after an expression time of 8 days. The degree of resistance in five of the six isolated cell variants was associated with a comparable increase in the specific activity of IMP dehydrogenase, which was 3- to 6-fold higher than that of the parent V79 cells. The IMP dehydrogenase activity from both the variants and the V79 cells had a similar affinity for the substrate IMP with a Km of about 20 microM and a similar response to mycophenolic acid with a Ki of 12-16 nM. It is suggested that cell variants with an altered regulation of IMP dehydrogenase activity may be helpful in studying the control of nucleic acid biosynthesis, cell growth, and carcinogenesis. Mycophenolic acid resistance also may be useful as a marker in short-term assays for the identification of potential chemical carcinogens.

Animals↗
PubMed1981

The relationship between the carcinogenicity and mutagenicity of nitrosamines in a hepatocyte-mediated mutagenicity assay.

A quantitative relationship was established for 26 nitrosamines between their carcinogenic effectiveness in experimental animals and their mutagenic activity in a mammalian cell-mediated assay. Mutagenesis was measured in Chinese hamster V79 cells co-cultivated with primary rat hepatocytes, which are capable of activating nitrosamines. Resistance to ouabain and to 6-thioguanine served as the genetic markers.

Animals↗
PubMed1981

[Giant duodenal diverticula with heterotopic gastric mucosa. Its demonstration with technetium 99].

The duodenal diverticulum, except for occasional findings, are diagnosed by their complications, these are found in 10% of the cases and they are: 1) diverticulitis 2) haemorrhage 3) mechanic disturbances 4) perforation 5) obstruction 6) malabsorption syndrome. The case that motivated our communication appeared as a massive haemorrhage, that was medically treated considering the high mortality, rate of the surgical procedures. As the duodenal diverticuli can be cause of acute or chronic digestive haemorrhage, and this is frequently due to the erosion produced, by the heterotopic gastric mucosae, we consider that its demonstration with technetium 99 can be a valuable diagnostic method, as has already been demonstrated in other pathologies that present heterotopic mucosae, and as has been demonstrated in our case.

Diverticulitis↗
PubMed1980

Postreplication repair and the susceptibility of Chinese hamster cells to cytotoxic and mutagenic effects of alkylating agents.

A cell variant (VR-43) resistant to the cytotoxic effect of N-methyl-N'-nitro-N-nitrosoguanidine and N-methyl-N'-nitro-N-nitrosourea was isolated from Chinese hamster V79 cells for use in studies of the relationship among cell survival, mutagenesis, and DNA repair by alkylating agents. Resistance to ouabain or 6-thioguanine was used as the genetic marker. After treatment with N-methyl-N'-nitro-N-nitrosoguanidine, the VR-43 cells exhibited mutation frequencies that were lower, on a dose basis, than those of the wild-type V79 cells. However, when analyzed at equicytotoxic doses, the VR-43 cells were more mutable than the V79 cells. No difference in cell survival or mutagenicity could be observed after treatment with other mutagens such as N-ethyl-N'-nitro-N-nitrosoguanidine, ethyl methanesulfonate, or x-rays. Postreplication repair was analyzed by determination of the molecular weight of the newly synthesized DNA by alkaline sucrose gradients. After treatment with N-methyl-N'-nitro-N-nitrosoguanidine, the VR-43 cells exhibited an enhanced postreplication repair relative to the V79 cells. No such enhancement was found after N-ethyl-N'-nitro-N-nitrosoguanidine or ethyl methanesulfonate treatment. Based on these results we propose that, after treatment of these and presumably other mammalian cells with some methylating mutagens, postreplication repair can cope with DNA lesions responsible for cytotoxicity and, to a lesser degree, with lesions responsible for mutagenicity.

Alkylating Agents↗
PubMed1980

The use of liver cell cultures in mutagenesis studies.

A sensitive cell-mediated assay has been developed for testing the mutagensis of liver carcinogens. Mutagenesis was detected in Chinese hamster V79 cells that were cocultivated with hepatocytes isolated after collagenase/hyaluronidase digestion of rat liver slices. Mutations were characterized by resistance to ouabain and 6-thioguanine. Seven of the nitrosamines, which are potent liver carcinogens, exhibited a mutagenic response. Mutagensis with the carcinogens could be detected at micromolar doses. The polyaromatic hydrocarbon benzo(a)pyrene, which is not a liver carcinogen, but can cause fibrosarcomas, was not mutagenic in this assay, but was mutagenic in a fibroblast-mediated assay. The liver carcinogen, aflatoxin B1, which usually does not induce fibrosarcomas, exhibited an inverse situation; it was mutagenic for V79 cells in the presence of liver cells but not in the presence of fibroblasts. We suggest that the use of various cell types, including hepatocytes prepared by the slicing method for carcinogen metabolism, and mutable V79 cells offers a sensitive assay for determining the mutagenic potential of chemical carcinogens, and may also allow a study of their organ specificity.

Aflatoxin B1↗
PubMed1980

[Colonic angiodysplasia: report of a case and considerations].

An important etiology of gastrointestinal disease hemorrhage is the angiodisplasic in the last decades the recognition of this pathology has advanced due to the endoscopic and arteriographic techniques. Higher frequency is observed in patients older than 60 years old attributing its etiology to degenerative changes being associated between 20 and 25% with aortic stenosis. Expectant or surgical therapeutics depend on the importance of the hemorrhage and the general state of the patient, the patient that motivated this review presented previous low gastrointestinal hemorrhages. The use of selective arteriography on the last internation allowed us to make the diagnosis of colonic angio displasia.

Angiography↗
PubMed1980

The induction of mutation and differentiation in mammalian cells by chemicals which initiate or promote tumor formation.

A cell-mediated mutagenesis assay was developed to predict the potential carcinogenic hazard of some environmental chemicals. In this assay, cells with appropriate markers for mutagenesis, such as the Chinese hamster V79 cells, are co-cultivated with cells capable of metabolizing chemical carcinogens. Use of this assay made it possible to demonstrate a relationship between the degree of carcinogenicity and mutagenicity of a series of polycyclic hydrocarbons, nitrosamines and aflatoxins, and to establish means to study organ specificity of some chemical carcinogens. However, most shortterm in vitro assays are designed to detect mutagenic activity and therefore do not detect tumor promoting agents which are devoid of this activity. By analyzing various markers of terminal differentiation in cultured human melanoma and myeloid leukemia cells, we have established a relationship between the activity of a series of tumor promoting phorbol diesters in the mouse skin and their ability to induce terminal differentiation. We suggest that measuring alterations in the differentiation. We suggest that measuring alterations in the differentiation characteristics of some cultured cells may represent an approach by which environmental tumor promoting agents can be studied and detected.

Animals↗
PubMed1980

A sensitive hepatocyte-mediated assay for the metabolism of nitrosamines to mutagens for mammalian cells.

A sensitive cell-mediated assay has been developed for testing mutagenesis in Chinese hamster V79 cells by carcinogenic nitrosamines. Mutations were characterized by resistance to ouabain and 6-thioguanine. Since V79 cells do not metabolize nitrosamines, mutagenesis in the V79 cells was tested in the presence of primary hepatocytes capable of metabolizing nitrosamines. The hepatocytes were isolated after collagenase and hyaluronidase digestion of liver slices. All seven liver carcinogens of the nine tested nitrosamines exhibited a mutagenic response in this cell-mediated assay. The potent liver carcinogens nitrosodimethylamine, nitrosodiethylamine, nitrosoethylmethylamine, and nitrosodipropylamine could be detected with doses as low as 1 muM. The noncarcinogenic nitrosodiphenylamine was not mutagenic. Nitrosomethoxymethylamine was the only nitrosamine that exhibited mutagenic activity in the absence of hepatocytes, and this activity was diminished in the presence of hepatocytes. It is suggested that the use of hepatocytes prepared by the slicing method for carcinogen metabolism and mutable V79 cells offers a highly sensitive assay for determining the mutagenic potential of carcinogenic nitrosamines and probably of other classes of hazardous chemicals occurring in the environment.

Animals↗