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Stephen Safe - One of the best experts on this subject based on the ideXlab platform.
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characterization of the h4iie rat Hepatoma Cell bioassay for evaluation of environmental samples containing polynuclear aromatic hydrocarbons pahs
Archives of Environmental Contamination and Toxicology, 1997Co-Authors: Kristine L Willett, Piero R Gardinali, Jose L Sericano, Terry L Wade, Stephen SafeAbstract:The H4IIE rat Hepatoma Cell bioassay has been extensively used to assess the toxic equivalents (TEQs) of complex mixtures of halogenated aromatic hydrocarbons in environmental samples. However, there is often a discrepancy between bioassay induction results and toxic equivalents calculated from chemical analysis of samples; the former generally yield higher bioassay-TEQs. Polynuclear aromatic hydrocarbons (PAHs) are a class of chemicals which can significantly contribute to induction-TEQs. Benzo(a)pyrene (BAP), dibenz(a, h)anthracene (DBA), benz(a)anthracene (BA), benzo(k)fluoranthene (BkF), benzo(b)fluoranthene (BbF), chrysene (Chr), and indeno(1,2,3-c,d) pyrene (IdP) are carcinogenic PAHs found in environmental samples, including oysters collected from Galveston Bay. The induction potency of these PAHs relative to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was determined individually in rat Hepatoma H4IIE Cells seeded in 6-well plates, and the induction-derived equivalency factors (EFs) relative to TCDD were 0.000354, 0.00203, 0.000025, 0.00478, 0.00253, 0.00020, 0.0011 for BAP, DBA, BA, BkF, BbF, Chr, and IdP, respectively. Dilutions of a reconstituted PAH mixture containing 23 PAHs (744 to 4466 ng/g total PAHs) with constant percentages of BAP (4.5%), DBA (3.5%), BA (2.4%), BkF (3.7%), BbF (3.5%), Chr (4.7%), and IdP (4.2%) yielded bioassay-derived induction-EQs that ranged from 0.52 to 1.44 ng/g. Oysters exposed in the laboratory to the same PAH mixture for 30 days differentially accumulated the PAHs with time. Bioassay-EQs for these oyster extracts ranged from 0.94 to 5.79 ng/g. These results were similar to the chemically calculated EQs which varied from 0.81 to 3.13 ng/g.
John A Timbrell - One of the best experts on this subject based on the ideXlab platform.
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in vitro cytotoxicity assays comparison of ldh neutral red mtt and protein assay in Hepatoma Cell lines following exposure to cadmium chloride
Toxicology Letters, 2006Co-Authors: George Fotakis, John A TimbrellAbstract:Abstract The aim of this study was to compare four in vitro cytotoxicity assays and determine their ability to detect early cytotoxic events. Two Hepatoma Cell lines, namely HTC and HepG2 Cells, were exposed to cadmium chloride (0–300 μM) for 3, 5 and 8 h. Following exposure to the toxic metal cytotoxicity was determined with the lactate dehydrogenase leakage assay (LDH), a protein assay, the neutral red assay and the methyl tetrazolium (MTT) assay. In HTC Cells no toxicity was observed for any incubation period when the LDH leakage, the MTT and the protein assay were employed whereas the neutral red assay revealed early cytotoxicity starting after incubation of HTC Cells with CdCl 2 for 3 h. In the case of HepG2 Cells the MTT assay reveals cytotoxicity due to CdCl 2 exposure after 3 h whereas no such effect is seen with the other three assays. Following 5 h exposure of HepG2 Cells to CdCl 2 , toxicity is observed with the MTT assay at lower concentrations compared to the ones required for detection of toxicity with the LDH leakage and the neutral red assay. In conclusion different sensitivity was observed for each assay with the neutral red and the MTT assay being the most sensitive in detecting cytotoxic events compared to the LDH leakage and the protein assay.
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cadmium chloride induced dna and lysosomal damage in a Hepatoma Cell line
Toxicology in Vitro, 2005Co-Authors: George Fotakis, Eduardo Cemeli, D Anderson, John A TimbrellAbstract:Abstract Cadmium is a toxic metal and no uniform mechanism of toxicity has so far been proposed. The aim of this study was to investigate the biochemical effects of cadmium chloride in a rat Hepatoma Cell line (HTC Cells) and the Cellular events mediating DNA damage. HTC Cells were exposed to various concentrations of cadmium chloride for 5 and 8 h and lysosomal damage was assessed with the neutral red assay (NR) and fluorescence microscopy. Mitochondrial integrity was assessed from ATP levels and DNA damage determined with the single Cell gel electrophoresis/comet assay. The formation of reactive oxygen species (ROS) was also determined under the same experimental conditions with the dichlorofluorescein assay. Cytotoxicity was assessed with the LDH leakage assay and the levels of glutathione were measured and correlated with the other effects. The results indicate that lysosomal damage occurs at a lower concentration of cadmium chloride (20 μM) than DNA damage (500 μM) in HTC Cells. The latter effect was accompanied by an increase of reactive oxygen species without any significant LDH leakage whereas lysosomal damage was significant as determined by the neutral red assay and confirmed with fluorescence microscopy. The effect of CdCl2 on mitochondria and glutathione levels were observed at concentrations or incubation times higher than the ones required to induce lysosomal damage. The data suggest that DNA damage may be due to the formation of reactive oxygen species. It is possible that cadmium induced lysosomal damage is an earlier event than DNA damage and can mediate other Cellular events that lead to Cell death.
Irving Kushner - One of the best experts on this subject based on the ideXlab platform.
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effects of cytokine combinations on acute phase protein production in two human Hepatoma Cell lines
Journal of Immunology, 1991Co-Authors: Andrzej Mackiewicz, T Speroff, M K Ganapathi, Irving KushnerAbstract:We evaluated the effects of binary combinations of four cytokines on production of the positive acute phase proteins alpha-1 antichymotrypsin, haptoglobin and fibrinogen, and the negative acute phase proteins albumin and alpha-fetoprotein (AFP) in two human Hepatoma Cell lines. The effects of the cytokine combinations on the five proteins varied; each protein exhibited a unique and specific pattern of response to the cytokine combinations. In Hep G2 Cells, antichymotrypsin was induced by all four cytokines, IL-6, IL-1, TNF-alpha, and transforming growth factor beta 1 alone, and their effects in binary combinations could be attributed to additive or minimally synergistic interactions. Fibrinogen was induced only by IL-6 and this induction was inhibited by IL-1 alpha, TNF-alpha or transforming growth factor beta 1. Haptoglobin was also induced only by IL-6, but TNF-alpha was the only cytokine that inhibited this induction at all concentrations of IL-6. Each of the four cytokines alone down regulated production of AFP and albumin. However, binary combinations of the four cytokines were simply additive, for the most part, in inhibiting AFP production, whereas the inhibitory effects of combinations of cytokines on albumin production differed significantly from simple additive effects. These observations, taken together with studies of effects of cytokine combinations on other acute phase proteins, indicate that the various acute phase proteins respond differently to different combinations of cytokines and that the potential exists for highly specific regulation of synthesis of individual plasma proteins by cytokine interactions. These findings imply that the acute phase response in vivo represents the integrated sum of multiple, separately regulated changes in gene expression.
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Transforming growth factor Β1 influences glycosylation of α1-protease inhibitor in human Hepatoma Cell lines
Inflammation, 1990Co-Authors: Andrzej Mackiewicz, Irving KushnerAbstract:We have previously shown that changes in acute-phase protein glycosylation result from alterations occurring within hepatocytes as a result of regulation by cytokines, that the glycosylation patterns of proteins secreted by Hep 3B and Hep G2 Cells respond differently to the crude mixtures of cytokines found in conditioned medium from LPS-stimulated monocytes, and that interleukin-6 (IL-6) causes increased concanavalin A (Con A) binding of α l protease inhibitor in Hep 3B Cells and decreased Con A binding of this protein in Hep G2 Cells. In the present study we found that transforming growth factor Β l (TGF- Β ), like IL-6, led to secretion of forms of α l-protease inhibitor with increased Con A binding in Hep 3B Cells, and that IL-6 and TGF- Β in combination were additive. In contrast, in Hep G2 Cells, TGF- Β had an effect opposite to that produced by IL-6, leading to secretion of forms of α l-protease inhibitor with increased Con A binding. When employed in combination with IL-6, TGF- Β abolished the effect of that cytokine. These studies indicate that TGF- Β influences glycosylation of al-protease inhibitor in two human Hepatoma Cell lines in a manner that can be differentiated from that of IL-6. The identification of TGF- Β as a second defined cytokine capable of influencing glycoprotein glycosylation and the demonstration that the effect of one cytokine can be modulated by another cytokine support the view that changes in glycosylation of plasma proteins are mediated by combinations of cytokines.
Kristine L Willett - One of the best experts on this subject based on the ideXlab platform.
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characterization of the h4iie rat Hepatoma Cell bioassay for evaluation of environmental samples containing polynuclear aromatic hydrocarbons pahs
Archives of Environmental Contamination and Toxicology, 1997Co-Authors: Kristine L Willett, Piero R Gardinali, Jose L Sericano, Terry L Wade, Stephen SafeAbstract:The H4IIE rat Hepatoma Cell bioassay has been extensively used to assess the toxic equivalents (TEQs) of complex mixtures of halogenated aromatic hydrocarbons in environmental samples. However, there is often a discrepancy between bioassay induction results and toxic equivalents calculated from chemical analysis of samples; the former generally yield higher bioassay-TEQs. Polynuclear aromatic hydrocarbons (PAHs) are a class of chemicals which can significantly contribute to induction-TEQs. Benzo(a)pyrene (BAP), dibenz(a, h)anthracene (DBA), benz(a)anthracene (BA), benzo(k)fluoranthene (BkF), benzo(b)fluoranthene (BbF), chrysene (Chr), and indeno(1,2,3-c,d) pyrene (IdP) are carcinogenic PAHs found in environmental samples, including oysters collected from Galveston Bay. The induction potency of these PAHs relative to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was determined individually in rat Hepatoma H4IIE Cells seeded in 6-well plates, and the induction-derived equivalency factors (EFs) relative to TCDD were 0.000354, 0.00203, 0.000025, 0.00478, 0.00253, 0.00020, 0.0011 for BAP, DBA, BA, BkF, BbF, Chr, and IdP, respectively. Dilutions of a reconstituted PAH mixture containing 23 PAHs (744 to 4466 ng/g total PAHs) with constant percentages of BAP (4.5%), DBA (3.5%), BA (2.4%), BkF (3.7%), BbF (3.5%), Chr (4.7%), and IdP (4.2%) yielded bioassay-derived induction-EQs that ranged from 0.52 to 1.44 ng/g. Oysters exposed in the laboratory to the same PAH mixture for 30 days differentially accumulated the PAHs with time. Bioassay-EQs for these oyster extracts ranged from 0.94 to 5.79 ng/g. These results were similar to the chemically calculated EQs which varied from 0.81 to 3.13 ng/g.
Young Min Park - One of the best experts on this subject based on the ideXlab platform.
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expression of cyclooxygenase 2 cox 2 in hepatoCellular carcinoma and growth inhibition of Hepatoma Cell lines by a cox 2 inhibitor ns 398
Clinical Cancer Research, 2001Co-Authors: Eun Sun Jung, Young Min ParkAbstract:Cyclooxygenase-2 (COX-2) has been suggested to be associated with carcinogenesis. In hepatoCellular carcinoma (HCC), the expression pattern of COX-2 protein has been well correlated with the differentiation grade, suggesting that abnormal COX-2 expression plays an important role in hepatocarcinogenesis. We investigated the expression pattern and clinical significance of COX-2 in HCC tissues. In addition, we evaluated the efficacy of a selective COX-2 inhibitor, NS-398, in three Hepatoma Cell lines. Thirty-six HCC tissues, 15 Hepatoma Cell lines, 1 colorectal Cell line (HT-29), and 1 fibroblast Cell line (SV80) were included in the study. We evaluated serological tests and histological and radiological evaluations of HCC tissues. Immunohistochemical staining for COX-2 was performed on 36 HCC tissues and 17 cancer Cell lines. A Cell viability assay for growth inhibition of NS-398 in five Cell lines was performed. Immunohistochemically, all six well-differentiated HCCs were positive, whereas 83% (10 of 12) of the poorly differentiated HCCs were negative. There was no significant relationship between the intensity of COX-2 expression and the level of α-fetoprotein, tumor size, presence of portal vein thrombosis, tumor capsule and metastasis, Tumor-Node-Metastasis staging, and growth types ( P > 0.05). According to the Cell viability assay, NS-398 suppressed the growth of all Cell lines, independent of the degree of COX-2 expression. The inhibitory effect on each Cell line was identified in 10 μm NS-398 and was significantly strong in 100 μm NS-398. All Cell lines exhibited apoptosis, which was identified by 4′-6diamidino-2-phenylindole staining. In conclusion, COX-2 may be a determinant of the differentiation grade of HCC, and the inhibition of COX-2 can induce growth suppression of Hepatoma Cell lines via induction of apoptosis.