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660 Effect of ethanol on prolactin release in female rats: role of estrogens
Poster Session P34. Endocrine disrupters 658
EFFECTS OF FLAME RETARDANTS PBDE 99 AND PCB 0N MRNA LEVELS OF ESTROGEN TARGET GENES AFTER PRENATAL EXPOSURE.
R. Ceccatelli, O. Faass, I. Fleischmann, M. Conscience, M. Schlumpf, W. Lichtensteiger. Institut of Pharmakology and Toxicology, University of Zurich 190 Wintherthurerstrasse 8057 Zurich Polybrominated diphenyl ethers (PBDE) are used in large quantities as additive flame retardants in plastics and textile materials. PBDEs are persistent compounds and have been detected in wildlife and in human adipose tissue and plasma samples. Certain PCBs, a structurally related group of substances, show endocrine disrupting action in mammals. We are investigating the effects of PBDE 99 (2,2‘,4,4‘,5-PentaBDE) and a PCB mixture, Aroclor 1254, on the mRNA levels of estrogen target genes in reproductive organs of Long Evans rat by Real Time PCR, with cyclophilin as reference gene. PBDE 99 (1 or 10 mg/kg/day) or Aroclor 1254 (10mg/kg/day) was injected subcutaneosly to time-pregnant rats from gestational day 10 to 18 (vehicle control: olive oil). Prenatal exposure to PBDE 99 or Aroclor 1254 affected the development of reproductive organs as indicated by changes in organ weight of adult offspring (effects on ventral and dorsal prostate, epididymis and ovary weight with PBDE 99). The effect patterns differed between PBDE 99 and PCB mixture. These effects were accompanied by changes in mRNA levels in tissue of adult offspring, i.a., progesterone receptor mRNA (uterus) and mRNA encoding for androgen receptor and estrogen receptor α and β (ventral and dorsal prostate). Our data indicate that PBDE 99 can interfere with the development of reproductive organs and induce long term changes in gene expression patterns. 659
POSSIBLE EFFECTS OF METHOXYCHLOR ON ACTH SECRETION THROUGH DOPAMINE
A. Lafuente 1 , T. Martínez-Rivas 1 , González Carracedo 1 , A.I. Esquifino 2 . 1 Laboratorio de Toxicología, Facultad de Ciencias, Universidad de Vigo, Campus de Orense, Las Lagunas, 32004-Orense, Spain. 2 Dpt. de Bioquímica y Biología Molecular, Facultad de Medicina, Universidad Complutense, Madrid, Spain Methoxychlor (MTX) is an organochlorated insecticide, currently used as a substitute for DDT. This organochloride insecticide, a oestrogen like substance, may modify the feedback mechanisms of steroids on the hypothalamus and pituitary. This work was undertaken to explore the possible effects of MTX on adrenocorticotropin hormone (ACTH) secretion and to analyze whether these effects are mediated by dopamine. Adult male Sprague-Dawley rats were administered with 25 mg/kg/day of MTX dissolved in sesame oil, for 30 days. The control group animals received sc 0.3 mL of sesame oil. The plasma ACTH levels were measured as well as dopamine (DA) content in median eminence and in anterior, mediobasal and posterior hypothalamus and in striatum. Dopamine was analyzed by high performance liquid chromatography, using electrochemical detection (HPLC.EQ) and plasma ACTH levels were analyzed by specific double antibody radioimmunoassay. The exposure to the insecticide diminished the plasma levels of ACTH (P<0.001 vs. control group). In the groups treated with MTX there were an augmentation of DA content in the anterior and posterior hypothalamus (P<0.05 vs control group) and a decrease in the median eminence (P<0.001 vs. control group), without modifying its levels in the mediobasal hypothalamus and striatum. The increase in the DA content in the anterior hypothalamus could explain the decrease of this catecholamine found in the median eminence after MTX exposure. These data suggest that MTX could inhibit the ACTH release through an increase of the DA content in the anterior hypothalamus.
660
s177
EFFECT OF ETHANOL ON PROLACTIN RELEASE IN FEMALE RATS: ROLE OF ESTROGENS
T. Shumkova, N. Bojadjieva. Department of Pharmacology and Toxicology, Medical University of Sofia, Sofia, Bulgaria Alcohol drinking is known to cause hyperprolactinemia in both in humans and laboratory animals. In order to better understand the mechanisms of alcohol’s effects on pituitary, we investigated the effects of estrogens on the alcohol stimulated hyperprolactinemia. Female rats were ovariectomized (OVX) and then treated with alcohol for 3 weeks. Group of rats were treated with ethanol and 17-β-estradiol. The plasma prolactin (PRL) levels and pituitary total protein levels were determined. The result demonstrated that ethanol treatment increased the levels of PRL in OVX rats. The 17-β-estradiol treated OVX rats did not show the significant changes in the plasma PRL levels. When ethanol was combined with 17-β-estradiol, the OVX rats showed higher levels of plasma prolactin, compared with ethanol treated rats. These results suggest that estrogen plays a role in ethanol stimulated hyperprolactinemia. 661
DETERMINATION OF ORGANOCHLORINE COMPOUNDS IN ADIPOSE BREAST TISSUE FROM WOMEN WITH BREAST CANCER
A.P.M. dos Santos 1 , J.P. Franco 1 , M.E. Vaz Pereira 2 , M.C.C. Batoréu 1 . 1 Laboratory of Toxicology, Faculty of Pharmacy, University of Lisbon, Av. Prof Gama Pinto, Lisbon, Portugal; 2 Service of Surgery, District Hospital of Setubal, Setubal, Portugal A recent epidemiologic study concluded that 73% of breast cancers are attributable to environmental factors. Evidence is growing that there is a connection between certain chemicals in the environment and the rising incidence of breast cancer. Several studies have shown that breast epithelial cells proliferate in response to estrogens, including environmental estrogens, and breast cancer has been correlated with cellular exposure to xenoestrogens. Xenoestrogens include several lipophilic, persistent compounds to which humans and wildlife have been exposed. Among these are a number of chlorinated organics, such as the insecticides dieldrin, DDT and its metabolites, endosulfan, chlordecone, and industrial pollutants such as some polychlorinated biphenyl (PCB) congeners and dioxins. Several of these chemicals have been shown to have estrogenic activity in vitro and/or in vivo assays. The objective of this study is to evaluate the chronic exposure to organochlorine compounds through the identification and quantification of those compounds accumulated in the adipose breast tissue from women with breast cancer. The adipose tissue samples were collected from the Service of Surgery of the Hospital of Setubal, which follows and treats women from an intensive agricultural area. Methodology: It was developed a method to extract and determine the organochlorine compounds in adipose breast tissue. A solid-liquid procedure was used for the extraction of organochlorine compounds from fat tissue: a pyrex glass column filled with Alumine Merck 90 was used for the extraction of 0.2 g of breast fat tissue dissolved in n-hexane. The eluate obtained was concentrated and dissolved again in n-hexane. After the addition of an internal standard, the extract was injected into a gas chromatographer Hewlett Packard 5890, with electron capture detector. The column installed was a HP-5. Our preliminary results indicate that the main organochlorine compounds detected in breast fat tissue are DDE (metabolite of DDT), dieldrin, endosulfan, and four PCBs congeners. Dieldrin was found in 40% of the samples analysed (0.313 – 0.429 µg/g) and was the compound with the higher concentration in tissue from women with breast cancer. In control samples, some organochlorine compounds were found nevertheless dieldrin was not detected.
EFFECTS OF FLAME RETARDANTS PBDE 99 AND PCB 0N MRNA LEVELS OF ESTROGEN TARGET GENES AFTER PRENATAL EXPOSURE.
R. Ceccatelli, O. Faass, I. Fleischmann, M. Conscience, M. Schlumpf, W. Lichtensteiger. Institut of Pharmakology and Toxicology, University of Zurich 190 Wintherthurerstrasse 8057 Zurich Polybrominated diphenyl ethers (PBDE) are used in large quantities as additive flame retardants in plastics and textile materials. PBDEs are persistent compounds and have been detected in wildlife and in human adipose tissue and plasma samples. Certain PCBs, a structurally related group of substances, show endocrine disrupting action in mammals. We are investigating the effects of PBDE 99 (2,2‘,4,4‘,5-PentaBDE) and a PCB mixture, Aroclor 1254, on the mRNA levels of estrogen target genes in reproductive organs of Long Evans rat by Real Time PCR, with cyclophilin as reference gene. PBDE 99 (1 or 10 mg/kg/day) or Aroclor 1254 (10mg/kg/day) was injected subcutaneosly to time-pregnant rats from gestational day 10 to 18 (vehicle control: olive oil). Prenatal exposure to PBDE 99 or Aroclor 1254 affected the development of reproductive organs as indicated by changes in organ weight of adult offspring (effects on ventral and dorsal prostate, epididymis and ovary weight with PBDE 99). The effect patterns differed between PBDE 99 and PCB mixture. These effects were accompanied by changes in mRNA levels in tissue of adult offspring, i.a., progesterone receptor mRNA (uterus) and mRNA encoding for androgen receptor and estrogen receptor α and β (ventral and dorsal prostate). Our data indicate that PBDE 99 can interfere with the development of reproductive organs and induce long term changes in gene expression patterns. 659
POSSIBLE EFFECTS OF METHOXYCHLOR ON ACTH SECRETION THROUGH DOPAMINE
A. Lafuente 1 , T. Martínez-Rivas 1 , González Carracedo 1 , A.I. Esquifino 2 . 1 Laboratorio de Toxicología, Facultad de Ciencias, Universidad de Vigo, Campus de Orense, Las Lagunas, 32004-Orense, Spain. 2 Dpt. de Bioquímica y Biología Molecular, Facultad de Medicina, Universidad Complutense, Madrid, Spain Methoxychlor (MTX) is an organochlorated insecticide, currently used as a substitute for DDT. This organochloride insecticide, a oestrogen like substance, may modify the feedback mechanisms of steroids on the hypothalamus and pituitary. This work was undertaken to explore the possible effects of MTX on adrenocorticotropin hormone (ACTH) secretion and to analyze whether these effects are mediated by dopamine. Adult male Sprague-Dawley rats were administered with 25 mg/kg/day of MTX dissolved in sesame oil, for 30 days. The control group animals received sc 0.3 mL of sesame oil. The plasma ACTH levels were measured as well as dopamine (DA) content in median eminence and in anterior, mediobasal and posterior hypothalamus and in striatum. Dopamine was analyzed by high performance liquid chromatography, using electrochemical detection (HPLC.EQ) and plasma ACTH levels were analyzed by specific double antibody radioimmunoassay. The exposure to the insecticide diminished the plasma levels of ACTH (P<0.001 vs. control group). In the groups treated with MTX there were an augmentation of DA content in the anterior and posterior hypothalamus (P<0.05 vs control group) and a decrease in the median eminence (P<0.001 vs. control group), without modifying its levels in the mediobasal hypothalamus and striatum. The increase in the DA content in the anterior hypothalamus could explain the decrease of this catecholamine found in the median eminence after MTX exposure. These data suggest that MTX could inhibit the ACTH release through an increase of the DA content in the anterior hypothalamus.
660
s177
EFFECT OF ETHANOL ON PROLACTIN RELEASE IN FEMALE RATS: ROLE OF ESTROGENS
T. Shumkova, N. Bojadjieva. Department of Pharmacology and Toxicology, Medical University of Sofia, Sofia, Bulgaria Alcohol drinking is known to cause hyperprolactinemia in both in humans and laboratory animals. In order to better understand the mechanisms of alcohol’s effects on pituitary, we investigated the effects of estrogens on the alcohol stimulated hyperprolactinemia. Female rats were ovariectomized (OVX) and then treated with alcohol for 3 weeks. Group of rats were treated with ethanol and 17-β-estradiol. The plasma prolactin (PRL) levels and pituitary total protein levels were determined. The result demonstrated that ethanol treatment increased the levels of PRL in OVX rats. The 17-β-estradiol treated OVX rats did not show the significant changes in the plasma PRL levels. When ethanol was combined with 17-β-estradiol, the OVX rats showed higher levels of plasma prolactin, compared with ethanol treated rats. These results suggest that estrogen plays a role in ethanol stimulated hyperprolactinemia. 661
DETERMINATION OF ORGANOCHLORINE COMPOUNDS IN ADIPOSE BREAST TISSUE FROM WOMEN WITH BREAST CANCER
A.P.M. dos Santos 1 , J.P. Franco 1 , M.E. Vaz Pereira 2 , M.C.C. Batoréu 1 . 1 Laboratory of Toxicology, Faculty of Pharmacy, University of Lisbon, Av. Prof Gama Pinto, Lisbon, Portugal; 2 Service of Surgery, District Hospital of Setubal, Setubal, Portugal A recent epidemiologic study concluded that 73% of breast cancers are attributable to environmental factors. Evidence is growing that there is a connection between certain chemicals in the environment and the rising incidence of breast cancer. Several studies have shown that breast epithelial cells proliferate in response to estrogens, including environmental estrogens, and breast cancer has been correlated with cellular exposure to xenoestrogens. Xenoestrogens include several lipophilic, persistent compounds to which humans and wildlife have been exposed. Among these are a number of chlorinated organics, such as the insecticides dieldrin, DDT and its metabolites, endosulfan, chlordecone, and industrial pollutants such as some polychlorinated biphenyl (PCB) congeners and dioxins. Several of these chemicals have been shown to have estrogenic activity in vitro and/or in vivo assays. The objective of this study is to evaluate the chronic exposure to organochlorine compounds through the identification and quantification of those compounds accumulated in the adipose breast tissue from women with breast cancer. The adipose tissue samples were collected from the Service of Surgery of the Hospital of Setubal, which follows and treats women from an intensive agricultural area. Methodology: It was developed a method to extract and determine the organochlorine compounds in adipose breast tissue. A solid-liquid procedure was used for the extraction of organochlorine compounds from fat tissue: a pyrex glass column filled with Alumine Merck 90 was used for the extraction of 0.2 g of breast fat tissue dissolved in n-hexane. The eluate obtained was concentrated and dissolved again in n-hexane. After the addition of an internal standard, the extract was injected into a gas chromatographer Hewlett Packard 5890, with electron capture detector. The column installed was a HP-5. Our preliminary results indicate that the main organochlorine compounds detected in breast fat tissue are DDE (metabolite of DDT), dieldrin, endosulfan, and four PCBs congeners. Dieldrin was found in 40% of the samples analysed (0.313 – 0.429 µg/g) and was the compound with the higher concentration in tissue from women with breast cancer. In control samples, some organochlorine compounds were found nevertheless dieldrin was not detected.