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G C Clark

Publications and source records attributed to G C Clark.

At least 37 records · Page 2Linked to original sources
PubMed1994

Tumor necrosis factor involvement in the toxicity of TCDD: the role of endotoxin in the response.

We have previously demonstrated that tumor necrosis factor is involved in the acute toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), since therapies designed to attenuate the effects of tumor necrosis factor resulted in reduced mortality and toxicity in mice exposed to an LD75 dose of TCDD. The current study addresses whether endotoxin may be a contributing factor in the cachexia and mortality resulting from TCDD exposure. Endotoxin-nonresponsive C3H/HeJ mice and endotoxin-responsive C57BL/6 mice were treated with 350 micrograms/kg TCDD and body weight and mortality were recorded. C3H/HeJ mice showed no trend in body weight loss (p = 0.554), while C57BL/6 mice demonstrated a statistically significant (p < 0.01) linear decline in body weight of -0.23 g/day, resulting in a net loss of 3.5 g over 15 days preceding mortality. Mortality was observed in the C57BL/6 mice beginning on day 16 with 100% of the mice dying by the 23rd day while no mortality was observed in C3H/HeJ mice until the 24th day of the study with only 22% mortality observed. These data further demonstrate that endotoxin is a contributing factor to the cachexia and lethality of TCDD.

Animals↗
PubMed1993

A mechanistic model of effects of dioxin on gene expression in the rat liver.

Improved methods for estimating the shape of the response curve for effects of exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) are needed in order to evaluate possible adverse health effects of TCDD. A mathematical model has been constructed to describe TCDD-mediated alterations in hepatic proteins in the rat. In this model it was assumed that TCDD mediates increases in the liver concentration of transforming growth factor-alpha (TGF-alpha) by a mechanism which requires the aryl hydrocarbon (Ah) receptor. TGF-alpha subsequently binds to the epidermal growth factor (EGF) receptor, a process which is known to cause internalization of this receptor in hepatocytes. This action is thought to be an early event in the generation of a mitogenic signal. Because TCDD decreases binding of EGF in the livers of intact female rats but not in ovariectomized rats, this effect was further assumed to be dependent on estrogen action. The model postulates Ah receptor-dependent effects on the concentration of cytochrome P450 1A2 (CYP1A2), which is involved in the metabolism of estradiol, and on the concentration of the estrogen receptor. The model also incorporates information on induction of cytochrome P450 1A1 (CYP1A1) by TCDD. The biochemical response curves for all these proteins were hyperbolic (Hill exponents in the equations for their expression were found to be 1), indicating a proportional relationship between target tissue dose and protein concentration at low administered doses of TCDD. The model successfully reproduced the observed tissue distribution of TCDD, the concentrations of CYP1A1 and CYP1A2, and the effects of TCDD on the Ah, estrogen, and EGF receptors over a wide dose range.

Animals↗
PubMed1993

CYP1A1 mRNA levels as a human exposure biomarker: use of quantitative polymerase chain reaction to measure CYP1A1 expression in human peripheral blood lymphocytes.

Accurate human risk assessment requires sensitive methods to evaluate dose-response relationships, especially following low level exposures. We have developed a reverse transcriptase polymerase chain reaction (RT-PCR) method to quantitative cytochrome P450-1A1 (CYP1A1) mRNA levels in human blood lymphocytes. Many polycyclic aromatic hydrocarbons (PAH) such as benzo[a]pyrene, and chlorinated PAH such as polychlorinated dibenzodioxins, dibenzofurans and biphenyls induce CYP1A1 expression through activation of an endogenous protein, the Ah receptor. Using a quantitative competitive RT-PCR method that included a synthetic internal standard we determined copy numbers of CYP1A1 mRNA in resting as well as mitogen-stimulated human blood lymphocytes. In mitogen-stimulated human blood lymphocytes assay variation was approximately 10% for measurement of this low expression gene and mRNA levels correlated well with ethoxyresorufin-O-deethylase (EROD) activity. The expression of mRNA was induced 20-fold upon culturing human lymphocytes with 10 nM TCDD. In nonstimulated, uninduced lymphocytes CYP1A1 levels are extremely low (1000 copies mRNA/10(4) cells) and cannot be measured by EROD activity. Studies of CYP1A1 mRNA expression in chemically-exposed populations are in progress.

Base Sequence↗
PubMed1993

TCDD-mediated changes in hepatic epidermal growth factor receptor may be a critical event in the hepatocarcinogenic action of TCDD.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent liver tumor promoter in rats, with females being more sensitive than males. The epidermal growth factor receptor (EGFR) pathway has been implicated in altered cell growth patterns induced by tumor promoters. We investigated hepatic EGFR levels in a two-stage initiation promotion model. The TCDD doses were chosen to encompass the dose range administered in a previous chronic bioassay currently used to determine the cancer potency commonly used for human health risk assessments. TCDD was administered biweekly by oral gavage to female Sprague-Dawley rats for 30 weeks following initiation by a single dose of diethylnitrosamine (DEN). TCDD-mediated decreased EGF receptor levels were demonstrated in intact but not ovariectomized animals, consistent with previous tumor data. Likewise, previous studies have shown that TCDD induces cell proliferation in intact rats but not ovariectomized rats. We report a significant dose-dependent decrease in plasma membrane EGF receptor maximum binding capacity in both initiated and non-initiated intact rats at TCDD doses equivalent to 3.5, 10.7, 35.7 and 125 ng/kg/day. There was a significant correlation between EGF receptor effects and liver TCDD concentration. The decrease in plasma membrane EGFR determined by equilibrium binding was confirmed quantitatively by EGF stimulation of EGFR autophosphorylation as well as qualitatively by immunohistochemical detection in control and treated rats. These results demonstrate that the observed down modulation of the EGFR by TCDD is ovarian-dependent and is a sensitive effect induced at dose levels associated with TCDD hepatocarcinogenicity in rodent bioassays.

Analysis of Variance↗
PubMed1992

Dose-response relationships for chronic exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin in a rat tumor promotion model: quantification and immunolocalization of CYP1A1 and CYP1A2 in the liver.

The mechanisms responsible for the braod spectrum of effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) are not entirely clear but seem to involve an initial interaction with the Ah receptor. A major uncertainty in risk assessment for TCDD is the lack of adequate dose-response relationships following chronic exposure to TCDD. Induction of cytochrome P-450 enzymes (CYP1A1 and CYP1A2) is one of the most sensitive responses to TCDD and its structural analogues. We have used a two-stage model for hepatocarcinogenesis in female Sprague-Dawley rats to evaluate dose-response relationships for induction of CYP1A1 and CYP1A2 in diethylnitrosamine-initiated as well as in noninitiated rats. After initiation with a single dose of diethylnitrosamine, TCDD was administered biweekly by p.o. gavage at doses equivalent to 3.5, 10.7, 35.7, and 125 ng/kg/day for 30 weeks. CYP1A1 and CYP1A2 concentrations were quantified in hepatic microsomes by radioimmunoassay and localized in hepatic tissue slices by immunohistochemical techniques. Radioimmunoassay data revealed a maximum induction of 200-fold for CYP1A1 and 10-fold for CYP1A2 and there were no statistically significant differences between initiated and noninitiated rats. Induction at the lowest dose (3.5 ng/kg/day) was 20-fold for CYP1A1 and 3-fold for CYP1A2. Mathematical analysis indicates that the best fit of the induction data are inconsistent with a threshold for this response. There was a linear relationship between administered dose and TCDD liver concentration over the entire dose range of the study. This indicates that induction of CYP1A2 does not significantly alter the distribution of TCDD in our chronic dosing regimen. Immunolocalization of CYP1A1 and CYP1A2 revealed the same localization and induction pattern for both isozymes in the cytoplasm of hepatocytes. However, the hepatic distribution pattern was not uniform with the most intense staining observed around central veins. These studies help to clarify dose-response relationships for dioxin-mediated effects and demonstrate different sensitivity of hepatocytes to the effects of TCDD.

Animals↗
PubMed1992

Inhibition of acute TCDD toxicity by treatment with anti-tumor necrosis factor antibody or dexamethasone.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) acute toxicity is characterized in part by a wasting syndrome with depletion of adipose tissue. Tumor necrosis factor (TNF) induces a similar response during chronic infection. The similarities of these toxic effects led to a hypothesis that TNF plays a role in TCDD acute toxicity. To test this hypothesis pharmacologic doses of an antibody specific for murine TNF and the potent anti-inflammatory agent Dexamethasone (DEX) were used to inhibit TCDD toxicity in mice. TNF antibody treatment resulted in a 54% reduction in TCDD-mediated mortality while DEX treatment, a glucocorticoid agonist that inhibits transcription of TNF, reduced mortality by 92%. Cyp 1A1 induction, the most commonly measured TCDD-mediated response, was not blocked by DEX, demonstrating separation of this biochemical effect from acute toxic responses to TCDD. These data suggest that TCDD-mediated changes in the TNF pathway may be an important mechanism for acute TCDD toxicity.

Animals↗
PubMed1992

Two dimensional gel electrophoresis of cellular and secreted proteins from rat alveolar macrophages after lipopolysaccharide treatment.

Two dimensional gel electrophoresis (2-D PAGE) and automated image analysis were used to study the effects of bacterial lipopolysaccharide (LPS) activation on secreted and cellular rat alveolar macrophage proteins. Primary alveolar macrophages were cultured and exposed to LPS in the presence of [35S]-methionine for 24 h. Image analysis of 2-D PAGE revealed that LPS treatment primarily modulated the proportions of several alveolar macrophage cellular proteins versus control in addition to limited protein induction and repression. The differential effect of LPS was more pronounced on secreted proteins where qualitative and quantitative differences from control cells were found. Immunoblots of secreted proteins with anti-tumor necrosis factor alpha (TNF alpha) and anti-interleukin-1 alpha(IL-1 alpha) antibodies identified these monokines from protein fluorographic patterns. IL-1 alpha was detected as a single polypeptide of 17 kD at pI = 5. Use of recombinant TNF alpha and monoclonal and polyclonal antibodies for immunodetection revealed the 17 kD form of TNF alpha as well as higher molecular weight species at 18 kD and 22 kD. Thus, analysis of radiolabeled rat macrophages treated with LPS reveals quantitative modulation of several cellular proteins as well as a distinctive pattern of secreted proteins which contain multiple monokine forms resolvable by 2-D PAGE.

Animals↗
PubMed1991

Tumor necrosis factor involvement in 2,3,7,8-tetrachlorodibenzo-p-dioxin-mediated endotoxin hypersensitivity in C57BL/6J mice congenic at the Ah locus.

An experimental model of endotoxin-induced release of tumor necrosis factor-alpha (TNF) into the serum of C57BL/6J mice congenic at the Ah locus was used to investigate the effects of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) on TNF production. TCDD exposure of Ah-responsive mice (Ahbb) resulted in a dose-dependent increase in the concentration of TNF in the serum of endotoxin-exposed mice, with a significant increase observed at a dose of 10 micrograms/kg TCDD. At a dose of 500 micrograms/kg TCDD, Ahbb mice demonstrated a 46-fold increase in serum TNF levels compared to control. In contrast, congenic Ah-receptor deficient mice (Ahdd) did not show a significant increase in serum TNF levels until exposed to 150 micrograms/kg TCDD, and the maximum response was an 8-fold increase over control. These data suggest that increased TNF production may be responsible for endotoxin hypersensitivity in TCDD-treated mice and that the Ah locus mediates this response.

Alleles↗
PubMed1991

2,3,7,8-Tetrachlorodibenzo-p-dioxin stimulation of tyrosine phosphorylation in B lymphocytes: potential role in immunosuppression.

The prototype halogenated aromatic hydrocarbon 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is carcinogenic and toxic in experimental animals. At the cellular level, TCDD toxicity is often expressed as an inhibition or alteration in normal cell maturation. In this respect, we and others have demonstrated that exposure of experimental animals to TCDD causes immunosuppression, including inhibition of B lymphocyte maturation and antibody synthesis. Although the immunological effects of TCDD are well described, little is known about its mechanism of action. In the present studies, it was found that TCDD increases membrane protein phosphorylation, which is, in part, associated with tyrosine-specific phosphorylation in B lymphocytes. This increase in phosphorylation occurred within minutes following TCDD treatment and was not associated with protein kinase C. The increase in tyrosine kinase by TCDD appears to be primarily due to de novo synthesis of new protein, because the protein synthesis inhibitors puromycin and cycloheximide, as well as the transcriptional inhibitor actinomycin D, partially inhibited the effect, although increased activity of preexisting protein cannot be fully dismissed. The dose response for increased phosphorylation by TCDD was identical to that we previously reported for inhibition of antibody synthesis, suggesting that immunosuppression by TCDD may be expressed through alterations in regulatory processes controlled by tyrosine kinases. These studies are discussed in terms of the potential role of TCDD-induced tyrosine phosphorylation in immunosuppression.

Animals↗
PubMed1991

Cervicothoracic vascular injuries. Presentation, management, and outcome.

One hundred consecutive patients with cervicothoracic vascular trauma were analyzed. The injury severity score, mechanism of injury, age, initial findings, management, and results were tabulated. There were 48 arterial and 61 venous injuries in the stable Group A patients, 11 arterial and 12 venous injuries in the unstable Group B patients, and three arterial and five venous injuries in the morbid Group C patients. Treatment included primary repair, resection with end-to-end anastomosis, or ligation. Twenty-three patients developed postoperative complications, the most common being respiratory in nature. The overall mortality rate was six per cent. Five patients died during or immediately after operation of exsanguination, and one died of ischemic brain death on the seventh postinjury day. The usefulness of preoperative angiograms, especially in the detection of arteriovenous fistulas, is important in planning the surgical approach. The overall outcome was favorable.

Adolescent↗
PubMed1990

Comparison of the inhalation toxicity of kretek (clove cigarette) smoke with that of American cigarette smoke. II. Fourteen days, exposure.

The comparative repeat dose toxicity of American cigarettes and kreteks (Indonesian cigarettes containing approximately 60% tobacco and 40% shredded clove buds) was assessed by exposure of groups of five male and five female rats to equivalent (approximately 2 mg/l in terms of total particulate matter) concentrations of smoke from each type of cigarette over 15 consecutive days. The smoke was delivered "nose only" using an HRC Rodent Smoking Machine (Mark IV). For each type of cigarette, three doses were used. These were achieved by regulating the daily total duration of exposure to smoke. The different doses used were 2 x, 4 x and 6 x, 15-min exposures, presented daily over a period of approximately 6 h. Inter-group comparisons were made between American and kretek groups which received the same daily durations of smoke exposure. Higher doses of smoke resulted in reduced bodyweight gains and food consumption in male groups; the response of female groups was not as clear. At the highest dose, male rats exposed to kretek smoke gained significantly more weight by comparison with males exposed to American smoke. Higher doses of smoke tended to increase water consumption in both sexes of groups exposed to American smoke; kretek smoke produced no obvious effect. Smoke exposures produced the expected responses in certain haematological and blood biochemical parameters attributed to exposure to CO and the irritants present in cigarette smoke. Such responses were, however, confined largely to the groups exposed to American smoke. Macroscopic pathological findings attributed to smoke inhalation were confined to the lungs, and consisted of minimal to moderate discolouration and incomplete collapse of the lung.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Inhalation↗
PubMed1990

Pertussis toxin inhibition of anti-immunoglobulin-stimulated proliferation and inositol phosphate formation.

A variety of receptor agonists activate cells by stimulating polyphosphoinositide hydrolysis. Increasing evidence supports the concept that receptor-stimulated phosphoinositide hydrolysis is mediated by a guanosine triphosphate binding protein, which in some cell systems is inhibited by pertussis toxin through ADP-ribosylation. The cross-linking of membrane immunoglobulin by antigen or anti-Ig stimulates phosphoinositide hydrolysis resulting in the formation of inositol phosphate and diacylglycerol which act as second messengers in initiating B lymphocyte activation. In this report, we demonstrate that anti-Ig-stimulated inositol phosphate formation is enhanced by the nonhydrolyzable guanosine triphosphate analogue, GppNHp, in permeabilized B lymphocytes and also inhibited by pretreatment of intact cells with pertussis toxin. This latter effect is associated with the pertussis toxin-catalyzed ADP-ribosylation of a 41-kDa membrane protein which is of the same molecular weight as the guanosine triphosphate binding protein reported to mediate receptor-stimulated phosphoinositide hydrolysis in other cellular receptor systems. B lymphocyte proliferation induced by agents such as lipopolysaccharide and PMA plus calcium ionophore, which activate cellular proliferation without stimulating phosphoinositide breakdown, is not inhibited by pertussis toxin. We conclude that anti-Ig activation of B lymphocytes contains pertussis toxin- and guanosine triphosphate-sensitive components which are involved in regulating phosphoinositide breakdown and initiating cellular activation.

Animals↗
PubMed1989

Comparison of the inhalation toxicity of kretek (clove cigarette) smoke with that of American cigarette smoke. I. One day exposure.

The comparative acute toxicity of a branded American cigarette and kreteks (Indonesian cigarettes containing approximately 60% tobacco and 40% ground clove buds) was assessed by exposure of groups of ten male and ten female rats to three different but equivalent (in terms of total particulate matter) concentrations of smoke from each type of cigarette. The smoke was delivered "nose only" using an HRC Rodent Smoking Machine (Mark IV) within a single 1-h period, with a total delivery of 30 min smoke and a 15 min air-breathing period between the two smoke exposures. Comparison of the immediate response to smoke exposure was made by monitoring respiration during exposure and by observation of the animals immediately following exposure. At this level, the only differences observed were more severe signs of smoke intoxication in the American smoke exposed animals which, at least in part, was attributed to the higher concentrations of carbon monoxide (CO) to which these animals were exposed; CO concentrations in the American smoke atmospheres were 2-2.5 times higher than the corresponding kretek smoke atmospheres. Comparison of any delayed response was made by observation and measurement of body weight, food and water consumption for a sub-population maintained for 14 days following exposures. This comparison revealed no differences between the groups which could be attributed to the smoke exposures. Comparison of any lung changes induced were made at two intervals, 24 h and 14 days following smoke exposures. These intervals were selected to provide information on any damage to the lung attributable to the smoke exposures and any subsequent development or repair.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Comparative inhalation toxicity of technical chlordane in rats and monkeys.

Technical chlordane (1,2,4,5,6,7,8,8-octachloro-3a,4,7,7-tetrahydro-4,7-methanoinda ne) is used extensively for control of certain wood-boring insects. The present study was conducted to evaluate the inhalation toxicity of technical chlordane in rats and monkeys. Range-finding (28-d) and subchronic (90-d) inhalation studies with Wistar rats, and subchronic (90-d) inhalation studies with cynomolgus monkeys were conducted. In the range-finding study in rats, the threshold of toxicity for technical chlordane was approximately 5.8 micrograms/l. Among the observations made during the course of the 90-d study, in which technical chlordane was administered by inhalation to rats and monkeys at concentrations close to 0.1, 1.0, and 10 micrograms/l, the most significant were associated with alterations in the liver and were confined to rats only. However, in the rat, the effects on the liver were largely reversible during 90 d following cessation of administration of technical chlordane. The no-effect level of chlordane inhalation in rats appears to be between 0.1 and 1.0 microgram/l, while in monkeys the no-effect level is in excess of 10 micrograms/l. This study demonstrated that the monkey, a species closely related to humans, can tolerate relatively high chlordane concentrations without any adverse effects.

Administration, Inhalation↗
PubMed1988

Acute inhalation toxicity of eugenol in rats.

The acute toxicity of inhaled eugenol was assessed by exposure of three groups of five male and five female rats to a submicron aerosol of eugenol for 4 h followed by a 14-day observation period. A fourth group, also of five male and five female rats and exposed to air only under similar conditions, served as a control group for comparison. The three concentrations of eugenol to which the different groups were exposed were 2.58, 1.37 and 0.77 mg/l. The mass median aerodynamic diameters and geometric standard deviations of the aerosols were, respectively, 0.82 micron (sigma g 2.26), 0.88 micron (sigma g 2.05) and 0.9 micron (sigma g 1.87). Clinical signs observed during exposure consisted principally of moderately increased salivation and restlessness (indicative of irritation) and abnormal breathing patterns. The signs were graded, being less marked in animals exposed to the lower concentrations of eugenol. All three groups, exposed to high, medium and low levels of eugenol, lost weight overnight following exposure. Associated with the weight loss were marked reductions in food and water intake. The responses appeared to be largely independent of the concentration of eugenol inhaled, although there was some evidence of a graded effect on water intake. There was rapid recovery, with food and water consumption data comparable with control values throughout most of the remainder of the 14-day observation period. Also, by the end of the observation period, group mean body weights were comparable. Upon sacrifice and macroscopic examination of the animals, abnormalities were detected in the lungs only of a few animals: 3/10 control, 2/10 eugenol 2.58 mg/l, and 2/10 eugenol 0.77 mg/l.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Inhalation↗
PubMed1988

Biochemical events associated with inhibition of B-cell proliferation by phorbol diesters.

Phorbol myristate acelate (PMA), a potent tumor promoter, has a variety of effects on cells of the immune system resulting in altered patterns of cell proliferation and differentiation. Although PMA is mitogenic or co-mitogenic for human lymphocytes and murine T-cells, it inhibits proliferation of murine B-cells stimulated by LPS or anti-Ig. PMA, however, does not inhibit the ability of LPS or anti-Ig to activate B-cells, as evidenced by increased Ia antigen expression and RNA synthesis. In the present studies it was shown that inhibition of DNA synthesis by PMA coincided with qualitative and quantitative changes in phosphorylated proteins. In particular, PMA treatment resulted in a unique profile of phosphoproteins independent of LPS or anti-Ig treatment. Inhibition of DNA synthesis occurred over a wide range of PMA concentrations. At concentrations up to 10(-9) M, inhibition of proliferation correlated with decreased phosphatidylinositol turnover and decreased intracellular Ca2+ levels, suggesting that PMA affects the phosphoinositide signal transduction pathway. However, at PMA concentrations less than 10(-10) M, inhibition of anti-Ig- and LPS-mediated proliferation occurred without inhibition of the phosphoinositide transduction signal. At these concentrations, PMA-induced inhibition of DNA synthesis was highly sensitive to recombinant IL-2. These data suggest that the antiproliferative effects of PMA on B-cells stimulated by LPS or anti-Ig may be mediated by two mechanisms. At high concentrations, PMA causes a feedback regulation of the phosphoinositide-dependent messenger system, while at lower concentrations, PMA alters the response to specific growth factors. Since PMA induces unique phosphoproteins and both of these events can be regulated by protein phosphorylation, it is possible that these unique phosphoproteins are responsible for the antiproliferative effects of PMA.

Animals↗
PubMed1988

Variables affecting outcome in blunt chest trauma: flail chest vs. pulmonary contusion.

We reviewed 144 consecutive patients with flail chest and/or pulmonary contusion between 1979 and 1984. The purpose was to analyze the factors adversely affecting morbidity and mortality. There were 97 males and 47 females, with an average age of 40 years +/- 18 S.D. (range, 2-83). Seventy-five per cent of the injuries were caused by motor vehicle accidents, with the remainder due to falls (17%), cardiopulmonary resuscitation (4%), altercations (2%), or falling objects (2%). The Injury Severity Score (ISS) averaged 32 +/- 14 S.D. in all survivors versus 60 +/- 14 S.D. in those who died. Eighty-three patients (58%) required mechanical ventilation. Thirty-six patients died (25%). Isolated pulmonary contusion or flail chest had a mortality of 16% each. However, the mortality more than doubled when there was a combined pulmonary contusion and flail chest (42%). More than half of all deaths were directly attributed to central nervous system injuries with another third due to massive hemorrhage. Factors that were associated with a higher morbidity and mortality included severe associated thoracic injuries, a high ISS, the presence of shock, falls from heights, and the combination of pulmonary contusion and flail chest.

Adolescent↗