Influence of serum cholesterol and albumin on partitioning of PCB congeners between human serum and adipose tissue

The influence of serum lipids and proteins on partitioning of individual polychlorinated biphenyl (PCB) congener peaks between human serum and adipose tissue lipid was assessed using regression analysis. Subjects were 55 repair workers who were either currently or previously exposed, and 56 comparison workers without occupational exposure to PCBs. Seven congeners (2,4,5,4'-tetra CB, 2,3,5,2',3',6'-hexa CB, 2,3,4,2',3',6'-hexa CB, 2,3,4,2',4',5'-hexa CB, 3,4,5,3',4'-penta CB, 2,3,4,5,2',3',4'-hepta CB, and 2,3,4,5,2',3',4',5'-octa CB) which had been quantified in both serum and adipose tissue in at least one-third of the total study population were selected for evaluation. Initially the crude correlations between the serum PCB and certain candidate variables were assessed; more than one congener was correlated with serum cholesterol, albumin, triglycerides, low-density lipoproteins, very low-density lipoproteins, age, body fat content, and average servings of fish per day. Stepwise regression of log serum congener concentration on log adipose congener concentration and these variables was performed. Only cholesterol, albumin, and average servings of fish per day were significant for at least one congener peak. Congeners behaved in two groups, depending on their order of chromatographic elution. For 2,4,5,4'-tetra CB, 2,3,5,2',3',6'-hexa CB, and 2,3,4,2',3',6'-hexa CB, log serum concentration was not significantly correlated with log adipose congener concentration, and there was no consistent pattern for significant candidate variables. For congeners 2,3,4,2',4',5'-hexa CB, 3,4,5,3',4'-penta CB, 2,3,4,5,2',3',4'-hepta CB, and 2,3,4,5,2',3',4',5'-octa CB, log serum congener concentration was consistently significantly correlated with log adipose congener concentration and serum cholesterol positively, and with serum albumin negatively. For these four congeners the explanations of variation (R2) in serum congener concentration using adipose congener concentration alone were 3, 23, 20, and 30%, respectively, and after adding cholesterol and albumin were 13, 56, 43, and 46%. Thus we conclude that serum cholesterol and albumin can influence the distribution or partition of PCBs between serum and adipose tissue.     

Toxicity and Microsomal Enzyme Induction Effects of Several Polybrominated Biphenyls of Firemaster

Toxicity and Microsomal Enzyme Induction Effects of SeveralPolybrominated Biphenyls of Firemaster. Dannan, G.A., Sleight,S.D. and Aust, S.D. (1982). Fundam. Appl. Toxicol. 2:313-321.Some toxicological and pharmacological effects of 2,4,5,2',5'-penta-(congener 1), 2,3,4,2',4',5'-hexa- (congener 5), 2,4,5,3',4',5'-hexa-(congener 6), 2,3,4,5,3',4',-hexa- (congener 7), and 2,3,4,5,2',3',4'-hepta-bromobiphenyl(congener 9) were evaluated in male rats given a single 90 mg/kgip injection and killed seven days later. Only congener 7 depressedbody weight gain, spleen and thymus weights, and caused severehistopathological changes in the thymus. Congener 7 caused thelargest increase in liver weight and the most changes in liverpathology while congener 1 failed to enlarge this organ andcaused the mildest ultrastructural changes. Liver microsomeswere isolated and evaluated for enzyme induction from all treatedrats except those administered congener 6, which was previouslyidentified as a mixed-type enzyme inducer (Dannan et al. 1978b).All congeners increased the liver microsomal cytochrome P-450content, but only congener 7 shifted the carbon monoxide differencespectrum absorption maximum to 448.0 nm. The microsomal ethylisocyanide difference spectrum 455/430 nm ratio was increasedthe most by congener 7 (3 fold). All congeners increased cytochromeP-450 reductase and microsomal epoxide hydrase activities bynearly 1.5–3 fold. Congener 7 failed to induce amino-pyrine-N-demethylaseactivity but the remaining congeners increased it by 2 fold.Congener 7 was the most effective inducer of benzo[a]pyrenehydroxylase and p-nitrophenol UDP-glucuronyl transferase. Theseresults add to the suggestion that the presence of an orthohalogen on a polyhalo-genated biphenyl does not completely abolishtoxicity or 3-methylcholanthrene-type microsomal enzyme induction.     

Metabolism of polychlorinated biphenyls by Gunn rats: identification and serum retention of catechol metabolites

The tissue distributions of persistent metabolites of polychlorinated biphenyls (PCBs) in Wistar rats and homozygous uridine diphosphate glucuronosyltransferase (UGT) deficient Gunn rats exposed to 2,4,5,2',5'-pentachlorobiphenyl (CB101) and the commercial PCB mixture, Kanechlor-500 (KC500), were investigated. After exposure to CB101, four hydroxy and two methylsulfonyl (MeSO2) metabolites were detected in liver, lung, kidney, blood, and adipose tissues. One was identified as 3',4'-(OH)2-2,4,5,2',5'-pentaCB, which was retained selectively in the serum of Gunn rats. Comparative analysis of the metabolite profiles in both rat strains after exposure to KC500 showed higher formation ratios of several dihydroxy PCB metabolites in the liver of Gunn rats; major metabolites are the catechols from 2,5,3',4'-tetraCB, CB101, 2,3,6,3',4'-pentaCB, and 2,3,6,2',4',5'-pentaCB. Thus, Gunn rats effectively metabolized PCBs with 2,5- or 2,5,6-chlorine substitution to the 3,4-catechol, but less formed MeSO2 metabolites in the liver. Although both rat strains retained 4-OH-2,3,5,3',4'-pentaCB in serum, Gunn rats also retained the catechol PCBs, accounting for about 52% of the total phenolic PCBs. These results suggest that a lack of UGTs markedly alters the formation ratios and retention profiles of catechols and MeSO2 metabolites of PCBs.     

Metabolism of 2,2',3,3',6,6'-hexachlorobiphenyl and 2,2',4,4',5,5'-hexachlorobiphenyl by human hepatic microsomes

Since the metabolism of polychlorinated biphenyls (PCBs) is the critical factor that determines whether or not they accumulate in adipose tissue, we have studied the metabolism of two hexachlorobiphenyls (HCBs), 2,2'3,3',6,6'-hexachlorobiphenyl (236-HCB) and 2,2'4,4',5,5'-hexachlorobiphenyl (245-HCB), by human hepatic microsomes. Human microsomes were isolated from patients undergoing liver resection and were found to have cytochrome P-450 levels (0.28 nmoles/mg microsomal protein) and cytochrome P-450-dependent enzymatic activities similar to those reported by other workers. 245-HCB was not metabolized by human microsomes under various conditions, while 236-HCB was metabolized with an apparent Km of 8.8 microM and a Vmax of 5.1 pmoles/mg microsomal protein/min. Two major metabolites were formed and identified by gas chromatography-mass spectrometry as 2,2',3,3',6,6'-hexachloro-4-biphenylol and 2,2',3,3'6,6'-hexachloro-5-biphenylol. [14C]236-HCB equivalents were found to covalently bind to microsomal protein. Addition of 1 or 5 mM reduced glutathione decreased the degree of covalent binding. These data suggest that HCBs are metabolized through an arene oxide. The fact that 245-HCB was not metabolized explains why it is the predominant PCB found in human adipose tissue.     

Species differences in the tissue distribution of catechol and methylsulphonyl metabolites of 2,4,5,2',5'-penta- and 2,3,4,2',3',6'-hexachlorobiphenyls in rats, mice, hamsters and guinea pigs

Polychlorinated biphenyls (PCBs) are metabolized to phenolic or methylsulphonyl PCBs (MeSO(2)-CBs) in animal species. The study determined the species differences in the tissue distribution of persistent PCB metabolites in rats, mice, hamsters and guinea pigs 4 days after exposure to 2,4,5,2('),5(')-pentachlorobiphenyl (CB101) or 2,3,4,2('),3('),6(')-hexachlorobiphenyl (CB132). For CB101 metabolism, the hydroxylation in rats, mice and hamsters occurred primarily at the 3(')-position in the 2('),5(')-dichlorinated phenyl ring, whereas the hydroxylation in guinea pigs occurred preferentially at the 3-position. Metabolite profiles in tissues of hamsters were dominated by 3('),4(')-catechol-CB101, whereas metabolite profiles in rats and mice were dominated by 3(')- or 4(')-MeSO(2)-CBs. For CB132 metabolism, rats and mice produced 4(')- and 5(')-MeSO(2)-CBs at similar concentration ratios, whereas guinea pigs produced MeSO(2)-CBs at higher levels and selectively retained 5(')-MeSO(2)-CB in liver. In contrast, hamsters preferentially produced 4('),5(')-catechol-CB132 that was retained in serum. Consequently, hamsters produced catechols, whereas guinea pigs produced meta-substituted MeSO(2)-CBs, preferentially from CB132. These findings indicate that PCBs with 2,3,6-chlorine substitution are preferred substrates for the formation of catechols or MeSO(2)-CBs and the differences in metabolite profiles are related to species-dependent metabolic capacities.     

Relative potencies of PAHs and PCBs based on the response of human cells

P450 reporter gene system (RGS) utilizes a human hepatoma cell line stably transfected with a plasmid containing firefly luciferase linked to human CYP1A1 promoter sequences. Luciferase expression in these cells is used to detect and quantify compounds that activate the Ah receptor (AhR) and induce CYP1A1. In this study, concentration-response curves were used to determine the relative potencies and calculate induction equivalency factors (IEFs) for non-ortho and mono-ortho coplanar PCBs and high-molecular weight PAHs. Relative potencies of PCBs were: 3,4,4',5-TetraCB (81)>3,3',4,4',5-PentaCB (126)>3,3',4,4'-TetraCB (77)≈2,3,4,4',5-PentaCB (114)>2,3',4,4',5-PentaCB (118)≈2',3,4,4',5-PentaCB (123)>3,3',4,4',5,5'-HexaCB (169). In addition, two other mono-ortho congeners, 2,3,3',4,4'-PentaCB (105) and 2,3,3',4,4',5-HexaCB (156), did not induce luciferase in these cells. Relative potencies of the PAHs were: benzo[k]fluoranthene>dibenz[a,h]anthracene>benzo[b]fluoranthene≈indeno[1,2,3-cd]pyrene>benzo[a]pyrene>chrysene≈benzo[a]anthracene>benzo[g,h,i]perylene. Relative potencies of PAHs are similar to those of PCBs.     

Disposition of polychlorinated biphenyl congeners in occupationally exposed persons

Relative concentrations of 37 polychlorinated biphenyl (PCB) congeners and the adipose-plasma partition of 28 PCB congeners were investigated in 26 persons occupationally exposed to various PCBs (20 to 54% chlorine). Concentrations of PCBs in adipose and plasma were related to duration and intensity of exposure in the workplace. PCB concentration in adipose tissue was proportional to that in plasma, with a partition for total PCBs of approximately 190:1 indicated from regression analysis. PCB congeners with chlorines in both 4-positions of the biphenyl ring were the major components in plasma and adipose tissue. Congeners with unsubstituted 3,4-positions (e.g., 2,5-chlorine substitution) on at least one of the biphenyl rings were observed at lower concentrations and had lower adipose-plasma partition than other congeners. In contrast, those compounds with substituents at the 2,4- and/or 3,4-positions on both rings were present in much higher proportions in blood or adipose than in the PCB mixtures in use. These components also had higher adipose-plasma partition than those with unsubstituted 3,4-positions, regardless of the degree of chlorination. PCBs with 2,4-substitution patterns on both rings, including 2,4,4′-tri- and 2,4,5,4′-tetra-CBs, had somewhat higher adipose-plasma partition than congeners with 3,4-substitution on at least one of the biphenyl rings (e.g., 2,4,3′,4′-tetra-CB).     

Comparison of 2,2'4,4',5,5'-Hexachloro[14C]biphenyl Levels in Different Adipose Tissues of Dogs and Monkeys

Comparison of 2,2',4,4',5,5'-Hexachloro[¹⁴C]biphenyl Levelsin Different Adipose Tissues of Dogs and Monkeys. RYERSON, B.A., CARTER, D. E., AND SIPES, 1. G. (1984). Fundam. Appl. Toxicol.4, 120–124. The polychlorinated biphenyl isomer, 2,2',4,4',5,5'-hexachloro[¹⁴C]biphenyl(2,4,5-HCB) was administered as a single iv dose at 0.6 mg/kgto dogs and monkeys. Adipose tissue, which included omentum,pericardial perirenal, peritesticular, and subcutaneous fat,and blood were collected at various termination times and analyzedfor total ¹⁴C and the parent hexachlorobiphenyl (HCB). Significantdifferences (p < 0.0005) in the total hexachlorobiphenylconcentration as measured by total radioactivity (¹⁴C equivalents)were noted in the various adipose tissues and in the same adiposetissues with time. Peritesticular fat was consistently lowerin the concentration of ¹⁴C equivalents than the other adiposetissues, which were nearly equal. The concentrations in subcutaneousfat samples were inconsistent. Total ¹⁴C equivalent concentrationsin the adipose tissues either peaked or reached a maximum atDay 1 and Day 4 for dog and monkey, respectively. However, parentHCB fat/blood ratios continually increased over the time courseof the experiment, because concentrations in blood decreasedmore rapidly than those in adipose tissue.     

PCB reduction and clinical improvement by detoxification: an unexploited approach?

1. A detoxification trial was administered to a female worker from a capacitor factory who had been exposed to polychlorinated biphenyls (PCBs) and other lipophilic industrial chemicals. 2. The patient presented with severe abdominal complaints, chloracne, liver abnormalities, and a spontaneous nipple discharge of approximately 50 ml d-1. 3. PCB levels were high in adipose tissue (102 mg kg-1), serum, (512 micrograms l-1), skin lipids (66.3 mg kg-1), and in the nipple discharge (712 micrograms l-1). 4. The patient's history, the medical evaluation and prior unsuccessful symptomatic treatments were indicative of consequences elicited by occupational exposure to chemicals. 5. Detoxification treatment reduced the PCB levels in adipose tissue to 37.4 mg kg-1 and in serum to 261 micrograms l-1, a 63% and 49% reduction, respectively. 6. The nipple discharge ceased and the symptoms improved. 7. Excretion of intact PCBs in sebum was appreciable before treatment and was enhanced by up to five-fold during detoxification. 8. This therapeutic approach appears promising for cases involving occupational exposure to lipophilic chemicals.     

Developmental exposure to low-dose PBDE-99: tissue distribution and thyroid hormone levels

Thyroid hormone concentrations, hepatic enzyme activities and tissue concentrations of 2,2',4,4',5-pentabromodiphenyl ether (PBDE-99) were evaluated in Wistar rats (dams and offspring) after treatment by gavage on gestation day (GD) 6 with a single low dose of either 60 or 300 microg PBDE-99/kg body weight (bw), respectively. Tissue concentration analysis confirmed that PBDE-99 is persistent in rodents as significant amounts of the parent compound were detected in adipose tissue 37 days after exposure. The dose of 300 microg PBDE-99/kg bw reduced thyroxin (T4) concentration in dams at the beginning of lactation (post-gestational day [PGD] 1), and caused a slight reduction in T4 on PGD 22, although not statistically significant. In offspring, reduced T4 was observed only at PND 22, probably due to cumulative effects of PBDE-99 during lactation. PBDEs have been shown to reduce T4 concentrations in several studies, but this is the first study demonstrating endocrine disruption at low doses. The adipose tissue concentration of PBDE-99 measured in this study was close to those reported for PBDE-99 in non-occupationally exposed humans. In addition, we have previously reported permanent changes in the reproductive systems and locomotor activity of male and female offspring using these same dosages.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the distribution and biliary excretion of polychlorinated biphenyls in rats.

Plasma disappearance, biliary excretion, and tissue distribution of two polychlorinated biphenyls (PCBs), 2,5,2′,5′-[³H]tetrachlorobiphenyl (4-CB) and 2,4,5,2′,4′,5′-[³H]hexachlorobiphenyl (6-CB), was determined in rats 10 days after oral administration of a single 10 or 25 μg/kg dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Plasma disappearance of both PCBs was not altered by TCDD treatment, but biliary excretion was depressed. Associated with the depressed excretion was a reduction in bile flow and concentration of 4-CB- and 6-CB-derived ³H in bile. Treatment with TCDD resulted in less PCB being distributed to the skin and a greater percentage of the dose being accumulated in the liver. The content of PCB-derived ³H in skeletal muscle, adipose tissue, and urine was similar in control and TCDD-treated rats. Extraction of bile with hexane showed that the majority of biliary radioactivity was in the form of polar metabolites and that the proportion of parent PCB (hexane-extractable radioactivity) to polar metabolites was not altered by TCDD treatment. In rats given 4-CB and sacrificed 1 hr later, the majority of radioactivity in the liver was hexane extractable, and the smaller amount of hepatic radioactivity due to polar 4-CB metabolites was greater in the TCDD treatment group than in the control group. When biliary metabolites of 4-CB were administered, biliary excretion of the metabolites was depressed in TCDD-treated animals. Thus, TCDD treatment impairs the initial and main excretory pathway for PCB elimination in the rat—biliary excretion—and alters the distribution of PCBs to the skin and liver.     

Differential binding affinities of PCBs, HO-PCBs, and aroclors with recombinant human, rainbow trout (Onchorhynkiss mykiss), and green anole (Anolis carolinensis) estrogen receptors, using a semi-high throughput competitive binding assay.

A comparative study was undertaken to assess the ability of 44 polychlorinated biphenyls (PCBs), 9 hydroxylated PCBs (HO-PCBs), and 8 aroclors at concentrations ranging from 1 nM to 10 microM to compete with [3H]17beta-estradiol (E2) for binding to bacterially expressed fusion proteins using a semi-high throughput competitive-binding assay. The fusion proteins consisted of the D, E, and F domains of human (alpha), cloned reptilian (Anolis carolinensis) and recloned rainbow trout (Onchorhynkiss mykiss) estrogen receptors (ER) linked to the glutathione S-transferase (GST) protein. GST-hERalphadef (human), GST-aERdef (reptile) and GST-rtERdef (rainbow trout) fusion proteins exhibited high affinity for E2 with dissociation constants (Kd) of 0.4+/-0.1 nM, 0.7+/-0.2 nM, and 0.6+/-0.1 nM, respectively. Of the 44 PCBs examined, only PCBs 104, 184, and 188 effectively competed with [3H]E2 for binding to the GST-rtERdef protein with IC50 values ranging from 0.4-1.3 microM. In contrast, these same congeners only caused a 30% displacement of [3H]E2 in GST-hERalphadef and GST-aERdef proteins. Several additional congeners were found to bind to the GST-rtERdef fusion protein, although the degree of interaction varied among congeners. Among the HO-PCBs, 2',3',4',5'-tetrachloro-4-biphenylol and 2,6,2',6'-tetrachloro-4-biphenylol bound to all three fusion proteins with IC50 values ranging from 0.1-0.3 microM. Dimethyl sulphoxide (DMSO) concentrations of 20% significantly increased the ability of PCBs 104, 184, and 188 to compete with [3H]E2 for binding to the GST-ERdef fusion proteins, whereas at 20% DMSO, a significant reduction in saturable binding was observed. These results demonstrate that ERs from different species exhibit differential ligand preferences and relative binding affinities for PCBs, which can be dramatically affected by DMSO concentration.     

Reduced levels of 1,25-dihydroxyvitamin D(3) in rat dams and offspring after exposure to a reconstituted PCB mixture.

Previous studies revealed effects of polychlorinated biphenyls (PCBs) and other polyhalogenated hydrocarbons on steroid hormone levels and hormone-dependent functions including behavior. In the present study serum concentrations of the vitamin D(3) metabolites 25-hydroxycholecalciferol (25-D) and 1,25-dihydroxycholecalciferol (1,25-D) were determined in rat dams and offspring after exposure to a PCB mixture that was reconstituted according to the congener pattern found in human breast milk. Unmated females were exposed to diets adulterated with 0; 5; 20; or 40 mg PCBs/kg diet. Exposure started 50 days prior to mating and was terminated at birth. Gestational exposure reduced serum concentrations of 1,25-D in dams in a dose-dependent manner. Concentration of 25-D was also decreased at the time of delivery, but not at weaning. Determination of 1,25-D in offspring at weaning revealed reductions in both high-exposure groups. Levels of 25-D were diminished only at the highest exposure level. Internal PCB concentrations in adipose tissue and brains exhibited a linear relation to dosages in diet. Concentrations of PCBs in brains were similar in dams and offspring at birth, but decreased at the end of lactation in dams. In offspring, values increased during this period because of continued exposure via the milk. In the adipose tissue, PCB levels were much lower in offspring than in dams. To our knowledge, this is the first report of PCB-induced effects on vitamin D(3) metabolites. In dams, reductions were seen even at the lowest exposure level used. Further studies are needed to evaluate the biological significance of these reductions in pregnant dams and possible consequences for the developing offspring.     

Species differences in the tissue distribution of catechol and methylsulphonyl metabolites of 2,4,5,2',5'-penta- and 2,3,4,2',3',6'-hexachlorobiphenyls in rats,mice, hamsters and guinea pigs

Polychlorinated biphenyls (PCBs) are metabolized to phenolic or methylsulphonyl PCBs (MeSO₂-CBs) in animal species. The study determined the species differences in the tissue distribution of persistent PCB metabolites in rats, mice, hamsters and guinea pigs 4 days after exposure to 2,4,5,2',5'-pentachlorobiphenyl (CB101) or 2,3,4,2',3',6'-hexachlorobiphenyl (CB132). For CB101 metabolism, the hydroxylation in rats, mice and hamsters occurred primarily at the 3'-position in the 2',5'-dichlorinated phenyl ring, whereas the hydroxylation in guinea pigs occurred preferentially at the 3-position. Metabolite profiles in tissues of hamsters were dominated by 3',4'-catechol-CB101, whereas metabolite profiles in rats and mice were dominated by 3'- or 4'-MeSO₂-CBs. For CB132 metabolism, rats and mice produced 4'- and 5'-MeSO₂-CBs at similar concentration ratios, whereas guinea pigs produced MeSO₂-CBs at higher levels and selectively retained 5'-MeSO₂-CB in liver. In contrast, hamsters preferentially produced 4',5'-catechol-CB132 that was retained in serum. Consequently, hamsters produced catechols, whereas guinea pigs produced meta-substituted MeSO₂-CBs, preferentially from CB132. These findings indicate that PCBs with 2,3,6-chlorine substitution are preferred substrates for the formation of catechols or MeSO₂-CBs and the differences in metabolite profiles are related to species-dependent metabolic capacities.     

Role of the Ah Locus in Suppression of Cytotoxic T Lymphocyte Activity by Halogenated Aromatic Hydrocarbons (PCBs and TCDD): Structure-Activity Relationships and Effects in C57BI/6 Mice Congenic at the Ah Locus

Role of the Ah Locus in Suppression of Cytotoxic T LymphocyteActivity by Halogenated Aromatic Hydrocarbons (PCBs and TCDD):Structure-Activity Relationships and Effects in C57B1/6 MiceCongenic at the Ah Locus. KERKVLIET, N. I., BAECHER-STEPPAN,L., SMITH, B. B., YOUNGBERG, J. A., HENDERSON, M. C, AND BUHLER,D. R. (1990). Fundam. Appl. Toxicol. 14, 532–541. Previousstudies have shown that the generation of cytotoxic T lymphocytes(CTL) following allogeneic tumor challenge is suppressed inAh-responsive C57B1/6 mice treated with a single oral dose ofthe toxic, Ah receptor-binding 3,4,5,3',4',5'-hexachlorobiphenyl(HxCB). The present studies have examined the specific roleof the Ah receptor in this immuno-toxic response by utilizingHxCB isomers of known, varied affinity for the Ah receptor aswell as by comparing effects of high-affinity Ah receptor ligands(3,4,5,3',4',5'-HxCB and 2,3,7,8-tetrachlorodibenzo-p-dioxin[TCDD]) on the CTL response of mice that differ only at theAh locus, that is, Ah-responsive (Ah1^*) and Ah-nonresponsive(Ah^dd) congenic C57B1/6 mice. Correlative changes in thymicweight, serum corticosterone (CS) levels, and spleen cellularitywere also measured. The potency of HxCB congeners (3,4,5,3',4',5'-;2,3,4,5,3',4'-; 2,4,5,2',4',5'-) and 2,3,7,8-TCDD to suppressthe CTL response, to reduce spleen cellularity, to cause thymicatrophy, and to elevate serum CS levels was directly correlatedwith the binding affinity of the congener for the Ah receptor.Furthermore, these parameters of immunotoxicity in Ah^*1 C57B1/6mice were significantly more resistant to alterations inducedby either 3,4,5,3',4',5'-HxCB or 2,3,7,8-TCDD as compared toAh1^* C57B1/6 mice. These results strongly support an Ah receptor-dependentimmunotoxic mechanism in suppression of the CTL response followingacute exposure to halogenated aromatic hydrocarbons.     

Effects of 2,2',4,4'-tetrachlorobiphenyl on granulocytic HL-60 cell function and expression of cyclooxygenase-2.

Polychlorinated biphenyls (PCBs) are persistent environmental contaminants that affect a number of cellular systems, including neutrophils. It has been demonstrated that noncoplanar PCBs (i.e., ortho- substituted PCBs) alter function of primary rat neutrophils. The objectives of these experiments were to determine if responses in a human, neutrophil-like cell line exposed to PCBs were similar to those reported for rat neutrophils and to explore further PCB-mediated alterations in neutrophil function. The human promyelocytic leukemia cell line (HL-60) was differentiated with DMSO to a neutrophil-like phenotype. Treatment of differentiated HL-60 cells with 2,2',4,4'-tetrachlorobiphenyl, a noncoplanar, ortho-substituted PCB congener, caused an increase in f-Met-Leu-Phe-induced degranulation, as measured by release of myeloperoxidase (MPO). Treatment with the coplanar, non-ortho-substituted congener 3,3',4,4'-tetrachlorobiphenyl had no effect on MPO release. 2,2',4,4'-Tetrachlorobiphenyl caused a time- and dose-dependent release of [3H]-arachidonic acid (3H-AA). A significant increase in 3H-AA release was observed after 60 min of exposure, and concentrations of 10 microM or larger increased 3H-AA release. In contrast, 3,3',4,4'-tetrachlorobiphenyl had no effect on 3H-AA release. The effect of PCBs on mRNA levels for cyclooxygenase-2 (COX-2) was examined using semiquantitative RT-PCR. COX-2 mRNA was significantly elevated in response to 2,2',4,4'-tetrachlorobiphenyl in a concentration-dependent manner. COX-2 expression was maximal by 30 min of exposure to 2,2',4,4'-tetrachlorobiphenyl. COX-2 protein and activity were also increased after exposure to 2,2',4,4'-tetrachlorobiphenyl; COX-1 protein and activity were unaffected. 3,3',4,4'-Tetrachlorobiphenyl did not increase COX-2 mRNA levels. These results demonstrate that a noncoplanar PCB alters the functional status of granulocytic HL-60 cells, causing enhanced degranulation and upregulation of COX-2, whereas a coplanar PCB lacks this activity. These data suggest that noncoplanar PCBs alter HL-60 cell function and COX-2 expression via an Ah-receptor-independent mechanism.     

The role of type I and type II 5' deiodinases on hexachlorobenzene-induced alteration of the hormonal thyroid status

Treatment of male Wistar rats with hexachlorobenzene (HCB) (1000 mg/kg b.w.) for 3-30 days decreases circulating levels of thyroxine (T4) but does not affect triiodothyronine (T3). Time courses were determined for 5' deiodinase type I (5' D-I) activity in thyroid, liver, and kidney and 5' deiodinase type II (5' D-II) activity in brown adipose tissue (BAT) to test the possibility that increased deiodinase activity might contribute to the maintenance of the serum T3 level. Specific 5' D-I activity was increased in the thyroid at 21 days and thereafter. No significant changes were observed in the liver, however, total 5' D-I activity in this tissue was increased at 30 days of treatment as a consequence of liver weight enhancement. HCB decreased kidney 5' D-I activity after 15 days, and BAT 5' D-II activity after 21 days of treatment. Total body 5' D-I activity was significantly increased by 30 days of HCB-treatment. HCB increased the activity of hepatic T4 uridine diphosphoglucuronosyl transferase (UDPGT) in a time-dependent manner, without changes in T3 UDPGT. We propose that increased T4 to T3 conversion in the thyroid and in the greatly enlarged liver may account for the maintenance of serum T3 concentration in hypothyroxinemic HCB-treated rats.     

Induction of DNA-repair synthesis (UDS) in rat pleural mesothelial cells by urine of subjects exposed to genotoxic agents.

Unscheduled DNA synthesis was determined in confluent rat pleural mesothelial cells arrested in G0/G1 with hydroxyurea by the measurement of [3H]thymidine incorporation into DNA. Cells were treated with concentrated urine or serum from subjects exposed to certain genotoxic agents, i.e. eight cancer patients treated with radiotherapy and/or chemotherapy (r/c cancer patients) and six chromium workers. Two additional groups consisted of six nonoccupationally exposed healthy smokers and five control volunteers who were nonsmokers and nonexposed. [3H]thymidine incorporated into DNA of all samples was measured by liquid scintillation counting and of urine samples from r/c cancer patients by autoradiography. Compared to the level observed in untreated cells, a statistically significant increased [3H]thymidine incorporation was found in cells treated with urine from 7 of 8 r/c cancer patients and from 5 of 6 chromium workers. In contrast, urine from control volunteers had no effect on the unscheduled DNA synthesis response and urine from only one smoker significantly enhanced [3H]thymidine incorporation into DNA. No clear-cut difference between groups was obtained with serum. These results suggest that urine could be useful to monitor subjects exposed to genotoxic agents.     

Differential effects of PCBs on the induction of apoptosis machinery and PKCalpha translocation in rat renal tubular cell cultures

We have demonstrated previously [Pérez-Reyes, P.L., Sánchez-Alonso, J.A., López-Aparicio, P., Recio, M.N., Pérez-Albarsanz, M.A., 2001. Different molecular capacity in the induction of apoptosis by polychlorinated biphenyl congeners in rat renal tubular cell cultures. Biosci. Rep. 6, 765-778] that the polychlorinated biphenyls (PCBs) cause loss of cell viability and accelerate apoptosis in cell kidney cultures. Further investigations are necessary to elucidate the mechanism of apoptosis induction. In this way, we have analyzed in the present work the effects of PCBs on protein kinase C (PKC, a protein family intimately involved in the regulation of cell survival) and the expression of two proapoptotic (caspase-3 and Bax) and one antiapoptotic (Bcl-2) proteins. Aroclor 1248 (a commercial PCB mixture with 48% chlorine by weight), PCB 153 (2,2',4,4',5,5'-hexachlorobiphenyl, a di-ortho-substituted nonplanar congener) and PCB 77 (3,3',4,4'-tetrachlorobiphenyl, a non-ortho-substituted planar congener), significantly increased PKCalpha activity compared to control cells in the cytosolic and particulate cell fractions, and increased the PKCalpha protein content in the particulate fraction. The nonplanar PCB 153 showed stronger effects than the coplanar congener PCB 77. In addition, Aroclor 1248 decreased both, procaspase-3 levels and the Bcl-2/Bax protein ratio. These findings indicate that PCBs, particularly nonplanar congeners, can induce apoptosis in primary renal tubular cells through the PKCalpha, caspase-3 and Bcl-2/Bax pathway.     

Polychlorinated and polybrominated biphenyl congeners and retinoid levels in rat tissues: structure-activity relationships.

The purpose of this study was to examine the structural requirements of polychlorinated and polybrominated biphenyls (PCBs and PBBs) for altering tissue levels of retinoids. Seven congeneric PCBs and PBBs were studied: 3,3',4,4'-tetrachlorobiphenyl (TCB), 2',3,3',4,5- and 3,3',4,4',5-pentachlorobiphenyls (-PeCBs), 3,3',4,4'- and 3,3',5,5'-tetrabromobiphenyls (-TBBs), 2,2',3,3',5,5'-hexachlorobiphenyl (-HCB), and 3,3',4,4',5,5'-hexabromobiphenyl (-HBB). Male Sprague-Dawley rats were fed a vitamin A-adequate diet (1.3 mg/kg) for 30 days before being given a single IP injection of one of seven polyhalogenated biphenyls (150 mumol/kg) in corn oil (10 ml/kg) or vehicle alone. Rats were killed 1 week later. Except for 3,3',4,4',5,5'-HBB, all PCBs and PBBs studied significantly decreased serum retinol levels and, except for 3,3',4,4',5,5'-HBB and 2,2',3,3',5,5'-HCB, all PCBs and PBBs also lowered the serum retinol-binding-protein (RBP) content. The activity of hepatic retinyl ester hydrolase (REH) was reduced by the treatment of 3,3',4,4',5-PeCB, 3,3',4,4'-TBB, and 3,3',4,4',5,5'-HBB. The levels of hepatic retinol were decreased by 2,2',3,3',5,5'-HCB, 2',3,3',4,5-PeCB, and 3,3',4,4',5-PeCB, while levels of hepatic retinyl palmitate were decreased by 2',3,3',4,5-PeCB, 3,3',4,4',5-PeCB, 3,3',4,4'-TCB, 3,3',4,4'-TBB, and 3,3',4,4',5,5'-HBB. The substantial decreases in hepatic retinyl palmitate levels could not be explained solely on the basis of hepatomegaly caused by acutely toxic PCBs and PBBs. All halogenated biphenyls which caused a decrease in hepatic retinyl palmitate also caused an increase in renal retinyl palmitate except 3,3',4,4',5-PeCB. In summary, the acutely toxic (nonortho substituted) congeners had pronounced effects on hepatic, renal, and serum retinoids whereas other biphenyls only decreased serum retinol levels. The effects of these seven compounds on REH activity were not correlated with the effects on serum retinol or RBP levels. Therefore, this study shows that the structure-activity relationships for altering hepatic retinoids differ from those for serum retinol, implying the involvement of multiple mechanisms.