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Biomedical subjects

C C Harris

Publications and source records attributed to C C Harris.

At least 271 records · Page 15Linked to original sources
PubMed1987

Cell and species differences in metabolic activation of chemical carcinogens.

Metabolic activation and DNA binding of aflatoxin B1 (AFB1), N-nitrosodimethylamine (DMN) and benzo[a]pyrene (B[a]P) were compared in human, rat and mouse hepatocytes and human pulmonary alveolar macrophages (PAM). The degree of carcinogen activation by hepatocytes and PAM was measured by cell-mediated mutagenesis assays in which co-cultivated Chinese hamster V79 cells were used to monitor mutagenic metabolites. Hepatocytes from human, mouse and rat metabolized DMN and released the active metabolites to induce either ouabain- or 6-thioguanine-resistant mutation. The mutation frequencies mediated by hepatocytes of the 3 animal species were approximately 3-9 mutants/10(5) survivors at a concentration of 0.2 mM DMN. The variations of radioactivity bound to liver cell DNA were relatively small in cultured mouse, rat, and human hepatocytes exposed to 14C label DMN (0.5 mM) and the binding values were in a range of 6-12 X 10(3) pmoles/mg DNA. However, rat hepatocytes were at least 10-fold more effective than either human or mouse hepatocytes in generating mutagenic metabolites of AFB1 and also had a much higher AFB1 metabolite DNA-binding value. The AFB1 DNA-binding levels were 4.1, 12-27 (range), 120 pmoles/mg DNA respectively in mouse, human, and rat liver cells following AFB1 (3.3 microM) exposure for 20 h. Hepatocytes from the 3 animal species were unable to mediate mutation in the presence of 4 microM B[a]P; PAM activated B[a]P and effectively mediated mutation in the co-cultivated V79 cells. In contrast to results with hepatocytes, PAM failed to generate enough mutagenic metabolites of AFB1 (3.3 microM) and the mediation of mutations was seen only at very high concentration of DMN (80 mM). The genotoxic effects of the 3 carcinogens on hepatocytes from different species in vitro were in agreement with the in vivo animal experiments in that mice are relatively resistant to AFB1 carcinogenesis whereas rats are sensitive; B[a]P is not effective as a complete liver carcinogen in adult rat and mouse whereas DMN induces liver cancer.

Aflatoxin B1↗
PubMed1987

Chemical carcinogen-induced decreases in genomic 5-methyldeoxycytidine content of normal human bronchial epithelial cells.

The genomic content of DNA 5-methyldeoxycytidine (m5dC) was measured in dividing normal human bronchial epithelial cells treated with a broad range of chemical carcinogens. At noncytotoxic concentrations, all of the carcinogenic agents tested significantly reduced cellular DNA m5dC content whereas the weakly carcinogenic and noncarcinogenic agents, benzo[e]pyrene and phenanthrene (respectively), did not. These reductions varied from 8% to 31% depending on the agent and the donor cells. The reductions in genomic m5dC levels were concentration dependent for the carcinogenic polycyclic aromatic hydrocarbon benzo[a]pyrene. We speculate that carcinogen-induced perturbation of DNA m5dC patterns may lead to heritable changes in gene expression and contribute to the molecular alterations involved in the initiation and the subsequent steps of the carcinogenesis process.

Carcinogens↗
PubMed1987

Distribution and modulation of the cellular receptor for transforming growth factor-beta.

Scatchard analyses of the binding of transforming growth factor-beta (TGF-beta) to a wide variety of different cell types in culture revealed the universal presence of high affinity (Kd = 1-60 pM) receptors for TGF-beta on every cell type assayed, indicating a wide potential target range for TGF-beta action. There was a strong (r = +0.85) inverse relationship between the receptor affinity and the number of receptors expressed per cell, such that at low TGF-beta concentrations, essentially all cells bound a similar number of TGF-beta molecules per cell. The binding of TGF-beta to various cell types was not altered by many agents that affect the cellular response to TGF-beta, suggesting that modulation of TGF-beta binding to its receptor may not be a primary control mechanism in TGF-beta action. Similarly, in vitro transformation resulted in only relatively small changes in the cellular binding of TGF-beta, and for those cell types that exhibited ligand-induced down-regulation of the receptor, down-regulation was not extensive. Thus the strong conservation of binding observed between cell types is also seen within a given cell type under a variety of conditions, and receptor expression appears to be essentially constitutive. Finally, the biologically inactive form of TGF-beta, which constitutes greater than 98% of autocrine TGF-beta secreted by all of the twelve different cell types assayed, was shown to be unable to bind to the receptor without prior activation in vitro. It is proposed that this may prevent premature interaction of autocrine ligand and receptor in the Golgi apparatus.

Animals↗
PubMed1987

Experimentally measured scatter fractions and energy spectra as a test of Monte Carlo simulations.

A method for the validation of Monte Carlo photon transport calculations is presented, with particular emphasis on the scatter component of such calculations. The method is based on a quantitative comparison of calculated and experimental scatter fractions. In addition, the method includes a qualitative comparison of point spread functions and energy spectra. An application of the method is demonstrated by comparing the results of an existing Monte Carlo code with experimental results obtained with a gamma camera viewing a point source of 99Tcm (140 keV gamma rays) centred within a water-filled cylinder. The results of the comparisons show good agreement between experiment and calculation. These results allow the code to be used with increased confidence in a variety of situations, and they define more precisely the region of applicability of the code. In addition, the determination of scatter fractions and energy spectra is useful for other applications. For example, scatter fractions can be a useful parameter for evaluating possible techniques for scatter compensation.

Computer Simulation↗
PubMed1987

Localization of aflatoxin B1--nucleic acid adducts in mitochondria and nuclei.

The reaction of chemical carcinogens with cellular macromolecules, particularly DNA, is considered to be an important event during chemical carcinogenesis. Until recently, most studies have dealt primarily with nuclear DNA (nDNA). Mitochondrial DNA (mtDNA) is increasingly being investigated for its possible role during chemical carcinogenesis. Using monoclonal antibodies against aflatoxin B1 (AFB1)-modified guanosine and ultrastructural immunocytochemistry, we have localized the AFB1-guanosine adducts predominantly in mitochondria and nuclei. Morphometric analysis of the electron micrographs demonstrates that localization of the AFB1-guanosine adducts is several-fold greater in mitochondria than in the nuclei. However, biochemical analysis demonstrates that 77% of the covalent binding was detected in nucleic acids of the mitochondria relative to the nuclear fractions. In RNA-free preparations, the extent of covalent modification of mt circular DNA was less than 50% of that of nuclear DNA. Our results indicate that 67% of AFB1 binding to mitochondrial nucleic acids is associated primarily with mtRNA.

Aflatoxin B1↗
PubMed1987

Genotoxicity of fecapentaene-12 in bacterial and mammalian cell assay systems.

Fecapentaene-12 (Fec-12), a compound thought to be responsible for much of the mutagenicity in fecal extracts from groups at high risk for colon cancer, was assayed for genotoxic potential in a battery of bacterial and mammalian cell short-term assays. This compound demonstrated significant mutagenic activity with Salmonella typhimurium tester strains TA104, TA100 and TA98, inducing approximately 1400, 700 and 100 revertants/micrograms, respectively. Fec-12 caused dose-dependent increases in unscheduled DNA synthesis in both rat hepatocytes and human fibroblasts, indicating its potential genotoxicity to mammalian as well as bacterial cells. Finally, Fec-12 had the ability to induce neoplastic transformation in mouse BALB/c 3T3 cells in the absence of exogenous metabolic activation.

Animals↗
PubMed1987

Strontium phosphate transfection of human cells in primary culture: stable expression of the simian virus 40 large-T-antigen gene in primary human bronchial epithelial cells.

Strontium ion formed DNA-phosphate precipitates analogous to those formed by calcium but lacking the lethal and differentiation-inducing effects of calcium on many epithelial cell types in primary culture. Human primary bronchial epithelial cells were transiently and stably transfected by using strontium phosphate; the frequency of stable transformation with a plasmid carrying the simian virus 40 large-T-antigen gene was greater than 10(-4).

Antigens, Viral, Tumor↗
PubMed1987

Biochemical and molecular epidemiology of human cancer: indicators of carcinogen exposure, DNA damage, and genetic predisposition.

The primary goal of biochemical and molecular epidemiology is to identify individuals at high cancer risk by obtaining evidence of high exposure to carcinogens, leading to pathobiological lesions in target cells, and/or increased oncogenic susceptibility due to either inherited or acquired host factors. This emerging and multidisciplinary area of cancer research combines epidemiological and laboratory approaches. Because DNA is considered to be an important target for modification by mutagens and carcinogens, damage to DNA can be used as an internal, molecular dosimeter of carcinogen exposure. The reactive species of these carcinogens may directly bind to DNA to form adducts and may indirectly cause secondary DNA lesions, e.g., via induction of free radicals and aldehydes. Highly sensitive and specific methods have been developed to measure the minute amounts of DNA lesions and DNA repair products found in biological specimens from humans exposed to carcinogens. For example, DNA adducts have been measured in cells and tissues from people occupationally exposed to carcinogenic polycyclic aromatic hydrocarbons. Antibodies recognizing carcinogen-DNA adducts have also been detected in human sera. Inherited predisposition to cancer has been revealed by recent advances in molecular genetics, including restriction-fragment-length polymorphism. For example, the hypothesis that rare alleles of the Ha-ras proto-oncogene are associated with an increased risk of lung cancer is currently being tested. These approaches afford the potential of biochemical and molecular epidemiology to predict disease risk for individual persons, instead of for populations, and before the onset of clinically evident disease.

Animals↗
PubMed1987

Biochemical and molecular effects of N-nitroso compounds in human cultured cells: an overview.

In-vitro models using human tissues and cells provide a bridge between studies of humans and of laboratory animals. Cultured human cells activate N-nitrosamines to DNA-damaging metabolites, including alkyldiazonium ions and aldehydes; these metabolites can also inactivate DNA repair processes, such as O6-alkylguanine DNA-alkyltransferase (AAT) activity, and are mutagenic in human cells. Both human epithelial and mesenchymal cells have been transformed in vitro by N-nitroso compounds.

Cell Transformation, Neoplastic↗
PubMed1987

Effects of tumor promoters and cocarcinogens on growth and differentiation of cultured human esophageal epithelial cells.

The acute effects of 12-O-tetradecanoylphorbol-13-acetate [(TPA) CAS: 56937-68-9], T-2 toxin (CAS: 21259-20-1), capsaicin (CAS: 404-86-4), cigarette smoke condensate (CSC), and ethanol (CAS: 3807-77-0) were examined in secondary cultured human esophageal epithelial cells in serum-free LHC-8 medium. Effects were evaluated by morphology and measurement of clonal growth rate (population doublings per day), cross-linked envelope (CLE) formation, and the enzymatic activities of ornithine decarboxylase (ODC) and plasminogen activator (PA). All compounds tested were inhibitory to clonal growth; concentrations causing 50% growth inhibition were estimated as 10 nM TPA, 6 nM T-2 toxin, 40 microM capsaicin, 8 micrograms CSC/ml, 540 mM ethanol, and 0.8 microgram CSC/ml with 220 mM ethanol. None of the compounds tested induced CLE formation, although calcium ionophore (A23187) could induce CLE in at least 60% of the cells. TPA (10 and 100 nM) decreased the ODC activity of cells, and capsaicin (100 microM) induced ODC by 220%. TPA (1-100 nM) and capsaicin (100 microM) also induced PA activity. Slight increases in ODC activity by CSC (10 micrograms/ml), CSC (1 microgram/ml) with ethanol, and T-2 toxin (1 nM) were observed, but PA activity was not affected by these compounds. The results indicated that the response of human esophageal epithelial cells to TPA is both similar to and different from that reported for human epidermal and bronchial cells in vitro. Enhancement of PA activity and decrease in ODC by TPA are found in all three human epithelial cell types. However, these changes are not associated in esophageal cells with increased CLE formation as reported in studies with the use of bronchial and epidermal epithelial cells. The results from these acute studies provide the basis for designing in vitro carcinogenesis investigations using these agents.

Capsaicin↗
PubMed1987

Pathobiological effects of aldehydes in cultured human bronchial cells.

Effects of the tobacco smoke-related aldehydes, i.e., acetaldehyde, formaldehyde and acrolein, have been investigated in cultured human bronchial epithelial cells and fibroblasts. As determined from loss of colony-forming efficiency of epithelial cells, acrolein is 200- and 5000-fold more toxic than formaldehyde and acetaldehyde, respectively. The aldehydes differ markedly in their potencies to induce terminal differentiation, as indicated by cessation of growth and enhanced formation of cross-linked envelopes. The cellular content of glutathione is markedly decreased by acrolein, whereas formaldehyde and acetaldehyde only slightly decrease glutathione levels. All three aldehydes produce DNA damage, as indicated by the formation of DNA single-strand breaks and DNA protein cross-links. Both formaldehyde and acrolein are weakly mutagenic in fibroblasts. In-vitro assays of O6-methylguanine-DNA methyltransferase (MMT) activity indicate that it is markedly inhibited by acrolein, and to a lesser extent by formaldehyde. However, formaldehyde significantly inhibits removal of O6-methylguanine (O6-meG) in N-methyl-N-nitrosourea (MNU)-exposed cells. Thus, the many biological effects induced by aldehydes include: inhibition of proliferation, enhanced cellular differentiation, thiol depletion, DNA damage, mutation and inhibition of DNA repair in human cells. Furthermore, we speculate that exogenous or metabolically generated aldehydes may increase the genotoxicity of N-nitroso compounds by the dual action of causing DNA damage and inhibiting the repair of promutagenic O6-meG DNA lesions in human cells.

Aldehydes↗
PubMed1986

Genomic 5-methylcytosine determination by 32P-postlabeling analysis.

A simple and sensitive method for the quantitation of 5-methyldeoxycytidine in DNA has been developed by the adaptation of the Randerath 32P-postlabeling technique. Nucleic acids were digested to 3'-monophosphate nucleotides, which were converted to 32P-labeled 3',5'-bisphosphate nucleotides, the 3'-phosphate was cleaved by the action of nuclease P1, and the resultant 5'-[32P]-monophosphate nucleotides were separated by two-dimensional thin-layer chromatography. Less than 1 microgram of DNA was required for the precise quantitation of 5-methyldeoxycytidine content to a detectable limit of 0.01% of the total cytidine residues methylated. The genomic 5-methyldeoxycytidine content may thus be quantitated in tissue samples, small or selective cell populations, senescing or terminally differentiating cells, or DNA from any source. We report here, for the first time, the genomic 5-methyldeoxycytidine content of normal human bronchial epithelial and normal human pulmonary mesothelial cells. The chromatographic separation of all of the normal and some of the rare monophosphate deoxyribonucleotides and ribonucleotides has been characterized. Thus, 5-bromodeoxyuridine and the RNA contamination of DNA or the DNA contamination of RNA can also be quantitated during the same analysis.

5-Methylcytosine↗
PubMed1986

Increased proto-oncogene expression in peripheral blood lymphocytes from patients with systemic lupus erythematosus and other autoimmune diseases.

Using RNA hybridization techniques, we examined the expression of proto-oncogenes associated with lymphocyte activation in vitro in patients with systemic lupus erythematosus and other autoimmune diseases. T and B lymphocytes from these patients were found to have significantly increased expression of c-myc, c-myb, and c-raf RNA when compared with those of normal individuals. Among the mononuclear cell subpopulations, B lymphocytes expressed higher levels of RNA for these proto-oncogenes compared with the T lymphocytes. Since prompt expression of these and other proto-oncogenes occurs in fibroblasts and lymphocytes following mitogenic stimulation, we propose that the present findings reflect the pathologically activated state of various lymphocytic subpopulations which is observed in systemic lupus erythematosus and in other autoimmune diseases. Endogenous and exogenous factors which lead to the expression of autoimmunity might share the induction of proto-oncogene expression as a common pathogenetic step.

Autoimmune Diseases↗
PubMed1986

Inter-individual variations in carcinogen metabolism.

Explant cultures of human tissues metabolized chemical carcinogens into the ultimate carcinogenic form as measured by binding to cellular DNA. A wide inter-individual variation in the binding level of many chemical carcinogens to target cell DNA was observed, whereas lesser inter-individual variation was observed in primary epithelial cell cultures derived from the explants. The binding levels in the explants showed a unimodal distribution without any sub-populations being present. The binding level of benzo(a)pyrene to human bronchial DNA has been analyzed with respect to genetic and environmental factors such as family history of cancer and smoking history.

Benzo(a)pyrene↗
PubMed1986

Sulphoxidation of S-carboxymethyl-L-cysteine in the rhesus monkey (Macaca mulatta), cynomologus monkey (Macaca fascicularis), African green monkey (Cercopithecus aethiops) and the marmoset (Callithrix jacchus).

The metabolic pathways giving rise to the urinary metabolites of S-carboxymethyl-L-cysteine have been identified for the rhesus, cynomologus, African green and marmoset species of monkey. The formation of a sulphoxide metabolite from the sulphide precursor is a reaction important in these species. The metabolic profile displayed by the marmoset was distinct from the three Old World species, with the rhesus and cynomologus being similar to man.

Animals↗