Report of the WHO working group on the assessment of health risks for human infants from exposure to PCDDs, PCDFs and PCBs.

On Nov. 20-22, 1995, a World Health Organization working group consisting of 12 scientific representatives from 6 different countries met to reassess the health risks to infants associated with perinatal exposure to polyhalogenated aromatic hydrocarbons (PHAHs). Following a review of previous WHO/EURO consultations, as part of their comprehensive programme on PCDDs, PCDFs and PCBs, current exposure information and recent experimental and epidemiologic data were discussed. Exposure assessments within the past decade have revealed that in the case of breast milk samples concentrations of PCDDs/DFs and PCBs have shown a continual decline, in certain countries by up to 50%. New experimental data has revealed that a variety of structural, functional and behaviourial alterations can be induced in rodent species following exposure to PHAHs while a Dutch collaborative PCB/dioxin study has illustrated subtle clinical, endocrine and mental/psychomotor development effects can occur in breast fed infants. The provisional conclusions of the working group were: 1) current evidence does not warrant altering the previous WHO recommendation for promotion/support of breast feeding and 2) based on new clinical data which supports the biological plausibility of certain observed experimental observations, continued and enhanced effort should be directed towards identifying and controlling sources of environmental input for these contaminants.

Consumption of fatty fish from the Baltic Sea and PCB in whole venous blood, plasma and cord blood from delivering women in the Aland/Turku archipelago.

The present study aimed to assess the role of fish consumption for the body burden of polychlorinated biphenyls (PCBs) in mothers living in the Aland and Turku archipelago in Finland. The overall objective was to investigate whether there exists an appropriate population for a full-scale prospective study on PCB-related developmental effects in infants. Concentrations of the four major PCBs were determined in whole venous blood and cord blood from 30 delivering mothers, of which 20 subjects consumed fatty fish from the Baltic Sea (2.5-12.5 meals per month) and the remaining 10 mothers did not. The concentrations of CB-118, CB-138, CB-153, and CB-180 in cord blood were generally two- to threefold lower than in whole blood from the mothers, but strong correlations were observed between PCBs in the two matrices (r = .67-.80). Neither the venous blood nor cord blood concentrations of PCBs, however, were correlated with stated fish intake. Moreover, the concentration of CB-153 in plasma was only weakly associated with fish intake, and the level of organic mercury in erythrocytes was not correlated with fish intake at all. The present results of CB-153 concentrations in women's blood are lower than those reported in other recent investigations. A reasonable contributing explanation is the rapid decline during the last decades of PCB in Baltic Sea fish, which has resulted in less impact of fish intake on the body burdens of PCB in relatively young women (median 30 yr in the present study) as compared with older females. The relatively low PCB levels in blood taken together with the low number of yearly deliveries in the archipelago population makes it an inappropriate study base for a prospective study of PCB-related health effects in infants.

Subchronic toxicity of 2,2',3,3',4,4'-hexachlorobiphenyl in rats.

The subchronic toxicity of 2,2',3,3',4,4'-hexachlorobiphenyl (PCB 128) was investigated in rats following dietary exposure at 0, 0.05, 0.5, 5, or 50 ppm for 13 wk. The growth rate was not affected by treatment and no apparent clinical signs of toxicity were observed. There was a significant increase in liver weight in the 50 ppm females. The liver ethoxyresorufin deethylase (EROD) activity was increased by five- and fourfold in the highest dose males and females, respectively, while aminopyrine demethylase (ADPM) activity was significantly increased only in the highest dose females. Liver vitamin A was significantly reduced in the highest dose females. No other biochemical or hematological effects were observed. Treatment-related histopathological changes were seen in the thyroid and liver, and to a lesser extent in the bone marrow and thymus. Residue data showed a dose-dependent accumulation of PCB 128 in the following tissues: fat, liver, kidney, brain, spleen, and serum, with the highest concentration being found in fat followed by liver and kidney. Based on these data, the no-observable-adverse-effect level of PCB 128 was judged to be 0.5 ppm in diet or 42 micrograms/kg body weight.

Promotion of altered hepatic foci by 2,3',4,4',5-pentachlorobiphenyl in Sprague-Dawley female rats.

The tumor promotion potential of 2,3',4,4',5-pentachlorobiphenyl (PCB-118) was studied in a two-stage initiation/promotion bioassay in female Sprague-Dawley rats. The animals were initiated by intraperitoneal administration of N-nitrosodiethylamine after partial hepatectomy. After 5 weeks of recovery, the promotion period commenced by once-weekly subcutaneous administrations of PCB-118 at six dose levels (10, 40, 160, 640, 2500, and 10,000 microg/kg body weight/week) for 20 weeks. In addition, three of these dose levels (40, 640, and 10,000 microg/kg body weight/week) were administered for 52 weeks. Evaluation of hepatic foci positive for glutathione S-transferase P demonstrated that the mono-ortho chlorine substituted congener PCB-118 significantly increased the number of foci/cm3 of liver in the two highest dose groups after 20 weeks, but did not significantly increase the percentage of the liver occupied by foci. After 52 weeks of treatment, both the percentage and the number of foci/cm3 were significantly increased in the highest dose group. A toxic equivalency factor based on foci development during 20 weeks of treatment would be less than 0.00002. Altered relative liver and thymus weights were observed after treatment with both substances as well as an induction of methyl cholanthrene- and phenobarbital-inducible isoenzymes of cytochrome P450 monooxygenase. These results show that PCB-118 has a potency to enhance foci growth in rat liver, although the potency is low compared to that of structurally related compounds.

Toxicity of 2,4,4'-trichlorobiphenyl in rats following 90-day dietary exposure.

The toxicity of 2,4,4'-trichlorobiphenyl (PCB 28) was investigated in rats after a 90-d dietary exposure. Groups of 10 male and 10 female weanling Sprague-Dawley rats were administered PCB 28 in the diet at 0, 0.05, 0.50, 5.0, or 50.0 ppm for 13 wk. Growth rate and food consumption were not affected by treatment, and no clinical signs of toxicity were observed. Mottled liver was noted in both control and PCB-treated males, but was found with increased incidence in the highest treatment group. Increased urinary ascorbic acid and hepatic microsomal ethoxyresorufin O-deethylase activity were observed in the 50.0 ppm group of both sexes. The vitamin A content in liver, lung, and kidney was not significantly affected by treatment. Analysis of brain biogenic amines showed a decreased dopamine concentration in the substantia nigra region of female rats receiving 0.5 ppm PCB 28 and higher doses. Female rats appeared to be more sensitive than males to the neurochemical effects of PCB 28. Dose-dependent histologic changes were observed in the thyroid and liver, with biologically significant changes occurring at 5.0 ppm and above. Based on these data, the no observable-adverse-effect level (NOAEL) for this PCB congener was considered to be 0.5 ppm in diet or 36 micrograms/kg body weight/d.

Toxicity of 2,2',4,4',5,5'-hexachlorobiphenyl in rats: effects following 90-day oral exposure.

The subchronic toxicity of 2,2',4,4',5,5'-hexachlorobiphenyl (PCB 153) was investigated in rats after 13 weeks of dietary exposure. Groups of 10 male and 10 female rats were administered PCB 153 in their diet at levels of 0.05, 0.50, 5.0 or 50 ppm for 13 weeks. The control groups received the diet containing 4% corn oil. Growth rate and dietary consumption were not affected by treatment. Clinical signs of toxicity were not observed. Enlarged, fatty liver was observed in treated animals at necropsy, but most were confined to the two highest dose groups. Increased hepatic microsomal ethoxyresorufin-O-deethylase, aminopyrine-N-demethylase and aniline hydroxylase activities occurred in high-dose groups of both sexes, with increased ethoxyresorufin-O-deethylase activity being observed starting at 0.05 ppm in females and at 0.5 ppm in males. Treatment-related reduction in hepatic and pulmonary vitamin A was seen in the highest dose group of both sexes. Changes in brain biogenic amines and intermediate products were observed mainly in females; these included decreased dopamine and 5-hydroxytryptamine concentrations in the frontal cortex region, and dihydroxyphenylacetic acid in the caudate nucleus region at 5.0 and 50 ppm. Female rats appeared to be more sensitive to the neurotoxic effects of PCB 153 than males. Dose-dependent histological changes were observed in the thyroid and liver of rats of both sexes and significant changes occurred at 5.0 and 50 ppm. Based on these data, the no-observable-adverse-effect level (NOAEL) of PCB 153 was judged to be 0.5 ppm in the diet or 34 micrograms kg-1 body wt. day-1.

Mechanistical studies of the inhibition of intercellular communication by organochlorine compounds.

Many hydrocarbons are environmental pollutants that, due to their lipophilicity and chemical stability, accumulate in biological systems including milk and body fat. A number of investigations have demonstrated that many organochlorine compounds can act as tumour promoters in vivo and inhibit gap junctional intercellular communication between cells in culture. In the present study we have investigated the dioxin 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), different polychlorinated biphenyls, chlorinated paraffins and the pesticide endosulfan. Using techniques of scrape loading dye/transfer and Western blot analysis the function, expression and phosphorylation of different connexins in vitro and in vivo were studied. The results show a good correlation between the ability to act as a tumour promoter and to interfere with gap junctional intercellular communication. All tested compounds inhibited the intercellular communication in a liver derived cell line (IAR 20). However, the results show that the time to inhibition varies between the different agents. Endosulfan and chlorinated paraffins inhibit the communication within one hour, whereas dioxin like substances need to expose the cells for 48 hours before the communication is affected.

Interaction of 3,4,5,3',4'-pentachlorobiphenyl and 2,4,5,2',4',5'-hexachlorobiphenyl in promotion of altered hepatic foci in rats.

This study was undertaken to investigate tumour promoting interactions of 2,4,5,2',4',5'-hexachlorobiphenyl (PCB 153) and 3,4,5,3',4'-pentachlorobiphenyl (PCB 126) in female Sprague-Dawley rats. Five weeks before the promotion treatment, the rats were partially hepatectomized and initiated with nitrosodiethylamine. The test substances were administered by weekly, subcutaneous injections for 20 weeks. The results from this study suggest that treatment with a combination of these two congeners causes a more than additive effect on the formation of gamma-glutamyltranspeptidase-positive hepatic foci. Co-exposure to PCB 126 and PCB 153 caused a dose-dependent reduction of the PCB 153-induced CYP2B1/B2-activity in these livers.

Toxicity of PCB 77 (3,3',4,4'-tetrachlorobiphenyl) and PCB 118 (2,3',4,4'5-pentachlorobiphenyl) in the rat following subchronic dietary exposure.

The toxicity of 3,3',4,4'-tetrachlorobiphenyl (PCB 77) and 2,3',4,4',5-pentachlorobiphenyl (PCB 118) was investigated in rats following subchronic dietary exposure. Groups of 10 male and 10 female weanling Sprague-Dawley rats were administered PCB 77 in the diet at 0, 10, 100, 1000, or 10,000 ppb for 13 weeks. PCB 118 was administered to males in the diet at 0, 10, 100, 1000, and 10,000 ppb, while the female groups received 0, 2, 20, 200, or 2000 ppb of the congener for 13 weeks. Growth rate and food consumption were not affected by treatment. No clinical signs of toxicity were observed. Increased spleen weight occurred in male rats fed 1000 or 10,000 ppb PCB 77. Male rats receiving 10,000 ppb PCB 118 had increased liver weight and hepatic ethoxyresorufin O-deethylase (EROD) activity. Increased hepatic EROD activity but not liver weight was observed in female rats given the 2000-ppb PCB 118 diet. Increased EROD activity was also noted in male rats given 10,000 ppb and in female groups receiving 1000 or 10,000 ppb PCB 77. Male rats exposed to 10,000 ppb PCB 77 had decreased vitamin A in the liver and lung and elevated levels in the kidney. Liver vitamin A of both 1000- and 10,000-ppb PCB 77 female groups was decreased. PCB 118 had no effects on tissue vitamin A at the levels studied. No hematological changes or serum biochemical changes were seen in any of PCB 118- and PCB 77-treated groups, nor were liver uroporphyrin levels altered. A reduction in dopamine and homovanillinic acid in substantia nigra region of the brain was observed in female rats fed 2000 ppb PCB 118, while 10,000 ppb PCB 77 was associated with an elevation in 3,4-dihydroxyphenylacetic acid in the nucleus accumbens region of male rat brains. Mild to moderate changes were observed in the liver and thyroid of rats given PCB 77 or PCB 118. PCB 118 accumulated in a dose-dependent manner in fat and to a much lesser extent in liver. In contrast, very low levels of PCB 77 residue were found in the tissues examined. Based on the above data it was concluded that NOAEL of PCB 77 is 100 ppb in diet or 8.7 micrograms/kg and that of PCB 118 is 200 ppb in diet or 17 micrograms/kg body wt/day.

Functional aspects of developmental toxicity of polyhalogenated aromatic hydrocarbons in experimental animals and human infants.

A scientific evaluation was made of functional aspects of developmental toxicity of polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) in experimental animals and in human infants. Persistent neurobehavioral, reproductive and endocrine alterations were observed in experimental animals, following in utero and lactational exposure to PCBs, PCDDs and PCDFs. The lowest observable adverse effect levels (LOAELs) for developmental neurobehavioral and reproduction endpoints, based on body burden of TCDD-toxic equivalents (TEQs) in animals, are within the range of current background human body burdens. Relatively subtle adverse effects on neurobehavioral development and thyroid hormone alterations have also been observed in infants and children exposed to background levels. Exclusive use of the toxic equivalency factor (TEF) approach may underestimate the risk of neurodevelopmental effects, because both Ah receptor dependent and independent mechanisms may be involved in these effects. The use of marker congeners and/or bioassays based on Ah receptor mediated mechanisms are rapid, low cost pre-screening alternatives for expensive and time consuming gas chromatographic-mass spectrometric analysis.

Liver tumour promoting activity of 3,4,5,3',4'-pentachlorobiphenyl and its interaction with 2,3,7,8-tetrachlorodibenzo-p-dioxin.

This study was undertaken to compare the tumour promoting effects induced by 3,4,5,3',4'-pentachlorobiphenyl (PCB 126) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In addition, interactive effects in rats treated with combinations of PCB 126 and TCDD were studied. Partially hepatectomized female Sprague-Dawley rats were initiated with nitrosodiethylamin. After 5 weeks of recovery the promotion treatment started and continued for 20 weeks. The results from the present study demonstrate that PCB 126 elicit approximately 10% of TCDD's tumour promoting activity measured as enhancement of the development of gamma-glutamyl-transpeptidase-positive altered hepatic foci in the liver. The factor required for the PCB to match the response of TCDD was adopted as a toxic equivalency factor and was in this case 0.1, which is the same as the factor suggested by Ahlborg et al. (1994). In the groups treated with a mixture of PCB 126 and TCDD the tumour promoting effect indicated an additive response. This result suggests that PCB 126 and TCDD act by the same mechanistical pathway, which in turn, supports that the toxic equivalency factor-concept can be used for TCDD-like tumour promoters.

Organochlorine compounds in relation to breast cancer, endometrial cancer, and endometriosis: an assessment of the biological and epidemiological evidence.

There is an increasing public and scientific concern that certain chlorinated compounds, recognized as environmental pollutants, may cause estrogen-related neoplastic disease in humans. The main hypothesis has been that certain organochlorines, through their estrogenic actions, might cause breast cancer. From experimental studies, both in vitro and in vivo, there is evidence that certain organochlorine compounds may cause estrogenic effects, whereas others may cause antiestrogenic effects. In limited studies, some of these compounds in high doses have also been shown to increase and reduce the frequency of estrogen-related tumors in animals. The epidemiological findings regarding the association between organochlorines and breast cancer are inconclusive. However, the largest and best designed study has been interpreted as negative with respect to DDT and polychlorinated biphenyls (PCB) in relation to breast cancer. Associations between organochlorine exposure and endometrial cancer or endometriosis have even more limited empirical basis. The hypothesis that human exposure to environmental levels or organochlorines would favor an estrogenic overactivity leading to an increase in estrogen-dependent formation of mammary or endometrial tumors is not supported by the existing in vitro, animal and epidemiological evidence. It can, however, not be conclusively rejected on the basis of available data.

Effect on tissue vitamin A levels in the rat following subchronic exposure to four individual PCB congeners (IUPAC 77, 118, 126, and 153).

The present study was carried out to investigate the effect on tissue vitamin A levels in rats exposed to 3,3',4,4'-tetraCB (CB-77), 2,3',4,4',5-pentaCB (CB-118), 3,3',4,4',5-pentaCB (CB-126), and 2,2',4,4',5,5'-hexaCB (CB-153). The obtained results show that hepatic vitamin A levels are reduced both in male and female rats following dietary exposure to individual PCB congeners for 13 weeks. However, there are pronounced potency differences between congeners. Compared to TCDD, the hepatic vitamin A reducing potencies of CB-126, CB-77 and CB-153, are 0.05, 0.0001 and 0.00001, respectively, in male rats. Compared to male rats, female rats are equally sensitive to hepatic vitamin A reduction both by TCDD and dioxinlike CBs. Effects on renal and pulmonary vitamin A levels vary between CBs and between sexes.

Effect of 2,3,7,8-tetrachlorodibenzoy-p-dioxin on the hepatic 7-ethoxyresorufin O-deethylase activity in four rodent species.

Temporal and dose-related effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on hepatic 7-ethoxyresorufin O-deethylase (EROD) activity were investigated in young male Hartley guinea pigs, Sprague-Dawley rats, C57Bl/6 mice, DBA/2 mice and Golden Syrian hamsters. Animals were terminated 1, 7, 14, 28, 56 and 112 days after the administration of a single i.p. dose of TCDD. The maximal induction of EROD activity, at doses producing a similar toxic and limited lethal effect in all species/strains, was 42-, 18-, 7- and 3-fold in rats, DBA/2 mice, C57Bl/6 mice and guinea pigs, respectively. No treatment-related induction of EROD activity was observed in hamsters. Generally, maximal induction occurred 1-4 weeks after injection in all species. The guinea pig alone maintained the same magnitude of induction of EROD activity throughout the study. Observed toxic effects, i.e., lethality, loss of body weight gain, liver enlargement and thymic atrophy, did not correlate with the TCDD-induced hepatic EROD activity. The obtained results suggest that the fold induction of cytochrome P450 1A1 activity is not a critical event for the expression of the lethal effect of TCDD in rodents.

Subchronic toxicity of 3,3',4,4',5-pentachlorobiphenyl in the rat. I. Clinical, biochemical, hematological, and histopathological changes.

The systemic toxicity of 3,3',4,4',5-pentachlorobiphenyl (PCB 126) following subchronic dietary exposure was investigated in Sprague-Dawley rats. PCB 126 was administered to rats of both sexes at concentrations of 0.1, 1.0, 10, or 100 ppb in their diet for 13 weeks. Another group of rats received a loading dose of 5 micrograms PCB/kg body wt at the start of the feeding period followed by exposure to 10 ppb PCB diet for the same period of time as the other groups. Growth suppression and decreased food consumption were observed in the highest dose groups of both sexes. Increased organ/body weight ratios for the liver occurred in the 10 and 100 ppb groups of both sexes. Rats of both sexes exposed to the highest dose of the PCB also exhibited increased relative kidney, spleen, and brain weights. Hematological and most serum biochemical changes were confined to the 100 ppb groups. These included elevated alkaline phosphatase, bilirubin, cholesterol, and aspartate aminotransferase, and decreased serum glucose, hemoglobin, erythrocytes, hematocrit, and platelets. A dose-dependent increase in liver ethoxyresorufin-O-deethylase activity was observed in rats of both sexes starting at 0.1 ppb. A dose-dependent increase in liver uroporphyrin levels was observed in both sexes and significant changes occurred in the female rats at 1.0 ppb and higher dose groups. Decreased liver vitamin A was observed in the 10 ppb group and higher in both sexes. Kidney vitamin A was elevated in the 100 ppb group. No statistically significant changes were noted in concentrations of brain biogenic amines. PCB 126 residues were 10-fold higher in liver than in fat. Treatment-related histopathological changes were observed in the thymus, thyroid, bone marrow, and liver of rats exposed to the 10 ppb diet, but increased frequency of mild changes was observed in most of these tissues at the 1.0 ppb level. Based on the above data, the no adverse effect level was judged to be 0.1 ppb in the diet or 0.01 micrograms/kg body wt/day.

Relative tumour promoting activity of three polychlorinated biphenyls in rat liver.

The relative tumour promoting activity of three structurally and toxicologically diverse polychlorinated biphenyls (3,4,5,3',4'-penta- 2,3,4,3',4'-penta- and 2,4,5,2',4',5'-hexachlorobiphenyl) was measured in an initiation/promotion assay in nitrosodiethylamine-initiated female Sprague-Dawley rats. The congeners under study were administered by once-weekly subcutaneous injections for 20 weeks. Evaluation of the development of gamma-glutamyl transpeptidase (GGT)- and glutation transferase P (GST-P)-positive hepatic foci showed that all congeners promoted altered hepatic foci, although 3,4,5,3',4'-pentachlorobiphenyl was far more potent. The volume fraction of the liver occupied by GGT-positive tissue in the 3,4,5,3',4'-pentachlorobiphenyl-treated animals (100 micrograms/kg per week) was 23%, while the volume fractions of altered liver tissue in the rats treated with 2,3,4,3',4'-pentachlorobiphenyl (5000 micrograms/kg per week) and 2,4,5,2',4',5'-hexaCB (20,000 micrograms/kg per week) were 1.2 and 2.3, respectively. The enhancement of GGT- and GST-P-positive foci was accompanied by an increased incidence of histological changes in the livers.