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Biomedical subjects

D L Busbee

Publications and source records attributed to D L Busbee.

At least 37 records · Page 2Linked to original sources
PubMed1986

In vitro cytotoxicity of chrysotile asbestos to human pulmonary alveolar macrophages is decreased by organosilane coating and surfactant.

Human pulmonary alveolar macrophages were used to quantitate the cytotoxic effect of surface-altered chrysotile asbestos. Little difference was observed in mortality between chrysotile asbestos that was surface-treated to a 42% extent by a hydrophobic organosilane or untreated chrysotile. Little or no effect on mortality was observed when human pulmonary alveolar macrophages were cultured with untreated chrysotile or acid-leached asbestos in the presence of 10 mM dipalmitoyl lecithin. However, when human pulmonary alveolar macrophages were cultured with a hydrophobically-treated (to a 42% or 95% extent) chrysotile asbestos in the presence of 10 mM dipalmitoyl lecithin, a statistically significant decrease in mortality was observed compared to untreated chrysotile. No mutagenic activity was observed when V79 cells were cultured with acid-leached, or 42% hydrophobically-treated chrysotile asbestos, even when human pulmonary alveolar macrophages were included as an activation source. The 95% hydrophobically-treated and acid-leached chrysotile also exhibited decreased binding of benzo[a]pyrene compared to untreated chrysotile asbestos.

Adult↗
PubMed1986

Inhibition of DNA polymerase activity by methyl methanesulfonate.

Methyl methanesulfonate (MMS) inhibits both thymidine incorporation into DNA in mitogen-activated human lymphocytes and deoxythymidine triphosphate incorporation into template DNA by DNA polymerase-alpha in a cell-free system. When MMS-modified DNA was used as the template for DNA synthesis utilizing unmodified DNA polymerase-alpha, nucleotide incorporation into template DNA was not inhibited. When unmodified DNA was used as the template for DNA synthesis utilizing MMS-modified DNA polymerase-alpha, nucleotide incorporation was differentially inhibited dependent on the MMS concentration. An analysis of the kinetics of DNA polymerase-alpha inhibition showed that incorporation of all 4 deoxynucleoside triphosphates into DNA template was noncompetitively inhibited by MMS, which is consistent with nonspecific MMS modification of the enzyme. These data indicate that MMS modification of DNA polymerase-alpha alone is sufficient to inhibit the incorporation of deoxynucleoside triphosphates into template DNA in vitro. The data further indicate that alkylation of both DNA polymerase-alpha and DNA template synergistically increases inhibition of DNA synthesis.

Alkylation↗
PubMed1985

DNA polymerase activity in a repair-deficient human cell line.

A human low-density-lipoprotein (LDL) receptor-deficient diploid fibroblast cell line (GM1915) was determined to be short patch competent (DNA polymerase-beta) and long patch deficient (DNA polymerase-alpha) for DNA excision repair. Analysis of DNA from GM1915 cells or from WI38 control cells, following treatment with a mutagen known to initiate long patch excision repair, showed that GM1915 cells exhibited decreased resynthesis of oligonucleotide segments excised during repair. When cells deficient in DNA polymerase-alpha activity were permeabilized to permit LDL entry, repair synthesis immediately increased. These data suggest that DNA polymerase-alpha is not activated by mutagen treatment in GM1915 cells and that introduction of LDL into the cells results in activation of the enzyme.

Cell Line↗
PubMed1985

Alkylation of deoxyguanosine by the sesquiterpene lactone hymenoxon.

Hymenoxon, a toxic sesquiterpene lactone found in the ruminant forage plant Hymenoxys odorata, binds deoxyguanosine in a cell-free system, and forms adducted guanine residues in sheep lymphocyte DNA. Mitogen-stimulated DNA synthesis in lymphocytes was inhibited by hymenoxon at concentrations greater than 100 microM. Unscheduled DNA synthesis in lymphocytes was initiated by hymenoxon concentrations exceeding 50 microM, and inhibited by concentrations above 100 microM. We describe an HPLC method which separates unmodified hymenoxon and deoxyguanosine from the hymenoxon-deoxyguanosine adduct, and allows the quantitation of adducts in hymenoxon-treated cells.

Alkylation↗
PubMed1985

Polychlorinated biphenyl uptake and transport by lymph and plasma components.

The uptake and vascular transport of ingested Aroclor 1242, an isomeric mixture of polychlorinated biphenyls (PCB), was investigated in experimental animals. High concentrations of ingested PCB were found in the chylomicron fraction of thoracic duct lymph. When the lymph flow was exteriorized PCB were not subsequently found in the vascular circulation. When lymph was not exteriorized plasma PCB concentrations reached maximal levels 6 hr after ingestion. Less than 1% of total plasma PCB was detected in cellular fractions of blood over a 10-hr period following ingestion. Chylomicrons contained 31% of total plasma PCB 30 min after ingestion, decreasing to less than 6% at 4 hr. A maximum of 10% of plasma PCB at 1 hr, and less than 5% at 6 hr, after ingestion was associated with very low density lipoproteins (VLDL) or low density lipoproteins (LDL). Although PCB enter the vascular circulation with the chylomicron fractions of lymph, delipoproteinated plasma contained 52% of the total PCB in blood collected 30 min after ingestion. This level increased to 78% after 2 hr, and remained constant at about 80% for an additional 8-hr period. High performance liquid chromatographic (HPLC) examinations of delipoproteinated plasma from blood taken 6 hr after PCB ingestion showed elution of greater than 95% of plasma PCB to coincide with the albumin peak. Electrophoretic examinations of delipoproteinated plasma showed the association of PCB with albumin to be noncovalent. The results suggest that apolar PCB are absorbed into intestinal epithelial cells from which they are secreted into the lymphatic drainage sequestered within the apolar core of chylomicrons, that these PCB transit the thoracic duct and enter the vascular circulation within chylomicrons and are metabolized or otherwise released from chylomicrons during hepatic chylomicron clearance, and that resulting PCB or PCB derivatives circulate in association with plasma albumins.

Animals↗
PubMed1984

Human lymphocytes treated with r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene require low-density lipoproteins for DNA excision repair.

Human lymphocytes which were non-mitogen-stimulated, and which were depleted of lipoproteins, were found to be deficient in DNA excision repair typically initiated in these cells in response to treatment with a direct-acting polynuclear aromatic hydrocarbon carcinogen. Lymphocytes either depleted of lipoproteins or supplemented with human low-density lipoproteins formed DNA-carcinogen adducts which were not chromatographically distinguishable. The state of lipoprotein depletion did not alter lymphocyte uptake of thymidine from the medium. Lymphocytes which were depleted of lipoproteins, treated with carcinogen, and subsequently supplemented with low-density lipoproteins, regained the ability to engage in DNA excision repair. The data suggest that either low-density lipoprotein(s), or a component(s) of low-density lipoprotein(s), is required by human lymphocytes in order to initiate excision repair of carcinogen-damaged DNA.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1984

Inhibition of DNA synthesis by an electrophilic metabolite of benzo[a]pyrene.

Mitogen-stimulated scheduled DNA synthesis and DNA excision repair in human lymphocytes, as well as DNA polymerase a activity in a cell-free system, were inhibited by an electrophilic metabolite of benzo[a]pyrene. This metabolite, (+/-)-anti-(7r,8t)-dihydroxy-(9,10t)-epoxy-7,8,9,10-tetrahyd robenzo[a]pyrene (BPDE), covalently binds to cellular macromolecules and is mutagenic, carcinogenic, and cytotoxic. Human lymphocytes treated with BPDE at concentrations greater than 500-800 ng/ml showed decreases in both mitogen-stimulated DNA synthesis and excision repair of damaged DNA but did not exhibit overt cytotoxicity (excluded trypan blue and maintained an adenylate charge of greater than 0.7). Formation of, and total concentration of, BPDE-DNA adducts was not correlated with inhibition of DNA synthesis. DNA polymerase alpha studies using a cell-free system showed that enzymatic activity was not diminished when purified polymerase was treated with BPDE prior to the addition of template DNA. When the template DNA concentration was varied, BPDE inhibition of enzyme activity was uncompetitive. BPDE inhibition of enzyme activity was found to be noncompetitive when concentrations of dATP, dCTP, or dTTP were varied and competitive when the concentration of dGTP was varied. The data indicate that BPDE competitively inhibits interaction of dGTP with the template-DNA polymerase alpha complex.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1984

Benzo[a]pyrene uptake by lymph: a possible transport mode for immunosuppressive chemicals.

Benzo[a]pyrene, a lipophilic promutagen, reached maximal concentrations in the thoracic duct lymphatic circulation within 2 h after gastric instillation. Benzo[a]pyrene in lymph obtained by thoracic duct cannulation decreased to approximately control levels within 4 h after treatment. When lymph was not allowed to enter the blood vascular circulation, serum levels of benzo[a]pyrene increased very slowly, suggesting minimal mesenteric blood vascular absorption of the lipophilic hydrocarbon. Benzo[a]pyrene partitions into lymph lipoproteins as a function of the lipoprotein concentration. Data suggest that low-density lipoproteins may take up benzo[a]pyrene more efficiently than do very low-density or high-density lipoproteins, and that lymph components other than lipoproteins do not take up and transport benzo[a]pyrene. We propose that lipophilic xenobiotic compounds interact with cells of the immune system via lymphatic lipoprotein transport of potentially mutagenic, carcinogenic, or immunosuppressive agents.

Absorption↗
PubMed1984

Benzo[a]pyrene uptake by serum lipids: correlation with triglyceride concentration.

An in vitro study of the relationship between benzo[a]pyrene (BaP) association with serum lipoproteins (LP) and LP composition was conducted using human subjects. BaP partitioning into different serum LP ranged from 53 to 71% of available BaP. Efficiency of BaP partitioning was examined for the relationship with lipid components of different sera. The data indicate that triglyceride (TG) concentrations were more directly correlated with BaP uptake than were concentrations of other LP components. Adjusting sera to a uniform TG concentration (96.5 mg/dl) resulted in the same BaP uptake for each serum type, while adjusting sera to contain a uniform cholesteryl ester concentration (104.6 mg/dl) did not result in similar BaP uptake among serum types. Analysis of serum LP composition suggested that marked differences in both BaP uptake and serum TG concentrations among the subjects were due mainly to differences in serum very low density lipoprotein (VLDL) concentrations. A correlation study using 14 human subjects showed that serum TG concentration was the best predictor (r = 0.973, P less than 0.001) for BaP uptake by serum, followed by phospholipid (r = 0.658, P less than 0.01) and total cholesterol (r = 0.514, P less than 0.05) concentrations. The results indicate that serum TG concentration (typically VLDL-TG) may be the primary factor affecting BaP uptake by serum LP, and suggest that a small change in serum TG concentration could cause a significant increase in BaP uptake by serum LP, contributing to an increased level of circulating carcinogen.

Adult↗
PubMed1983

High-density lipoproteins decrease both binding of a polynuclear aromatic hydrocarbon carcinogen to DNA and carcinogen-initiated cell transformation.

Lipophilic carcinogens partition into the three major classes of lipoproteins potentially present in serum used as a medium supplement for cell culture. Different serum lots sequester differing quantities of the procarcinogen benzo[a]pyrene, dependent on the serum lipoprotein concentrations. In the presence of high-density lipoproteins a mutagenic benzo[a]pyrene metabolite was bound to cellular DNA at decreased levels when compared to cells exposed to the mutagen in the absence of high-density lipoproteins. Fetal calf serum with low levels of lipoproteins, specifically, high-density lipoproteins, is associated with efficient methylcholanthrene-initiated transformation of C3H/10T1/2 cells, while calf serum with a significant concentration of high-density lipoproteins requires up to a 500% increase in methylcholanthrene concentration to achieve similar levels of transformation in this mouse embryo cell line. When concentrated serum lipoproteins or purified HDL were added to fetal calf serum containing MCA at 1 microgram/ml, the C3H/10T1/2 transformation frequency was decreased compared to the transformation frequency achieved in the presence of 1 microgram/ml of MCA in fetal calf serum without supplementation. The results suggest that high-density lipoprotein partitioning of lipophilic polynuclear aromatic hydrocarbon mutagens from the cell culture medium may effectively reduce the concentration of carcinogen available for interaction with cellular DNA in vitro, which, in turn, may be associated with decreased carcinogen-induced transformation of cells.

Animals↗
PubMed1982

Binding of benzo[a]pyrene and intracellular transport of a bound electrophilic benzo[a]pyrene metabolite by lipoproteins.

Human serum lipoproteins were isolated by means of size exclusion h.p.l.c. Non-covalent uptake of [3H]benzo[a]pyrene was quantitated for fractions collected from the effluent of a liquid chromatographic separation of human serum, and was found to directly correlate with the lipoprotein concentration. An electrophilic benzo[a]pyrene metabolite, [3H]trans 7,8-dihydrodiol-9,10-epoxybenzo[a]pyrene, non-covalently associated with low density lipoproteins was transferred to human lymphocytes in vitro and bound acid-precipitable nucleic acids of the lymphocytes as a function of time. Benzo[a]pyrene metabolite binding to lymphocyte DNA was demonstrated by means of CsCl density gradient analysis. Non-mitogen-stimulated lymphocytes exposed to very low concentrations of carcinogen in the presence of low density lipoprotein demonstrated [3H]thymidine incorporation; without the concomitant addition of low density lipoprotein the low concentrations of carcinogen did not stimulate [3H]thymidine incorporation.

Benzo(a)pyrene↗
PubMed1981

Reassessment of the relationship between aryl hydrocarbon hydroxylase and lung cancer.

Aryl hydrocarbon hydroxylase was measured in cultured human lymphocytes induced with benzathracene and in pulmonary alveolar macrophages induced in situ in cigarette smokers. Considered separately, neither lymphocyte AHH nor macrophage AHH levels were distinctly different in either noncancer or lung cancer patients. Considered simultaneously, lymphocyte and macrophage AHH levels are quite different in noncancer and lung cancer patients. The lung cancer patient group was seen to contain a significantly higher percentage of persons with high levels of AHH than did an age-matched group of noncancer patients, (P less than 0.001), when more than one tissue was assayed to determine the individual's enzyme levels.

Aryl Hydrocarbon Hydroxylases↗
PubMed1981

Separation and detection of lipoproteins in human serum by use of size-exclusion liquid chromatography: a preliminary report.

Components of human serum can be separated on the basis of differences in relative molecular mass by using size-exclusion "high-performance" liquid chromatography. Lipoproteins in fractions of the eluate can be quantitated by conventional chemical and enzymatic methods. Alternatively, if lipoproteins in the serum are selectively prestained with diformazan dye, the column effluent can be monitored spectrophotometrically at 580 nm, so that only the lipoprotein components of serum are detected. Samples of purified low-density lipoproteins, so stained and analyzed, provide peak-area values that are proportional to their concentration as evaluated by chemical methods. With this technique, the various lipoprotein classes can be quickly separated and their concentration estimated. These techniques should prove useful in clinical and research laboratories.

Azo Compounds↗
PubMed1980

Correlation of carcinogen-induced unscheduled DNA synthesis and NAD reduction in fresh human lymphocytes.

Incorporation of [3H]thymidine into the DNA of fresh human lymphocytes, treated with various chemical mutagens, was measured and correlated with cellular NAD levels before and after treatment. NAD levels in lymphocytes were significantly reduced following treatment with mutagenic chemicals. Reduction of cellular NAD pools was directly correlated with [3H]thymidine incorporation. As NAD levels decreased, [3H]thymidine incorporation increased. Theophylline, a known inhibitor of poly(ADP-ribose)polymerase, inhibited both the NAD reduction in cells treated with DNA damaging agents and the incorporation of [3H]thymidine into DNA. The inhibitory effect of theophylline on NAD depletion and on [3H]thymidine incorporation was dose and cell number dependent. Near normal responses to carcinogen exposure could be restored to theophylline-treated cells following the removal of theophylline. These data suggest that conversion of NAD to poly(ADP-ribose) may be necessary, or at least closely associated with, DNA repair in human lymphocytes.

Benz(a)Anthracenes↗
PubMed1980

Reduction of nicotinamide adenine dinucleotide levels by ultimate carcinogens in human lymphocytes.

The effect of several classes of DNA-damaging chemicals and closely related compounds on cellular nicotinamide adenine dinucleotide (NAD) levels was studied in freshly isolated peripheral human lymphocytes. Of the 21 compounds examined, 7 were direct DNA-damaging agents and 14 were non-DNA-damaging compounds or required metabolic activation to casue DNA damage. Rapid lowering of cellular NAD levels was caused by each of the direct DNA-damaging chemicals examined in this study including N-methyl-N'-nitr-N-nitrosoguanidine, methyl methanesulfonate, N-acetoxy-2-acetylaminofluorene, 7-bromomethylbenz(a)anthracene, and the benzo(a)pyrene derivatives, r-7,t-8-dihydroxy-9, 10-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene and benzo(a)pyrene-4,5-epoxide. The indirect-acting carcinogen 2-acetylaminofluorene, 13 polycyclic aromatic hydrocarbons, and derivatives that were non-DNA-damaging did not cause lowering of NAD. The results suggest a general correlation between DNA damage and acute lowering of cellular NAD pools.

Carcinogens↗
PubMed1979

Aryl hydrocarbon hydroxylase activity in pulmonary alveolar macrophages and lymphocytes from lung cancer and noncancer patients: a correlation with family histories of cancer.

Aryl hydrocarbon hydroxylase (AHH) activity was measured in pulmonary alveolar macrophages (PAMs) and peripheral blood lymphocytes from cigarette smokers with and without primary lung cancer. Frequency distribution analysis of AHH induction ratios for the two groups revealed an increased number of individuals in the lung cancer patient group with high lymphocyte induction values (P less than 0.05). A similar increase was not shown for high-PAM AHH values in lung cancer patients (P greater than 0.2). When individual PAM and lymphocyte AHH values were compared between noncancer and lung cancer patients, a positive correlation was observed for noncancer patients (r=0.195, P less than 0.001), but no correlation of these values was noted for lung cancer patients. The lung cancer patients were divided into three subgroups of patients showing (I) high PAM and low lymphocyte AHH levels, (II) low PAM and low lymphocyte AHH levels, and (III) low PAM and high lymphocyte AHH levels. When the incidence of family history of cancer was compared for these subgroups, no family cancer history was recorded for persons in subgroup II; however, individuals in subgroups I and III presented family cancer history incidence of 9.5% and 39.3%, respectively. Patients in group III averaged 6 years younger than those in group I. These data suggest that familial factors may be identified among lung cancer patients and that these factors appear to associate as either a cause of an effect with the capacity of pulmonary alveolar macrophages and lymphocytes to be induced for AHH. The data support the hypothesis that high AHH values may be characteristic of lung cancer patients but show that enzyme values determined from a single tissue, either PAMs or lymphocytes, may not be appropriate for showing whether high AHH inducibility is correlated with lung cancer.

Aryl Hydrocarbon Hydroxylases↗