~(14)C标记1,2,7,8-TCDD在鲤体内分布及代谢的初步研究

利用14C标记1,2,7,8-四氯代二苯并二噁英(1,2,7,8Tetrachloro[U—14C]dibenzodioxin,14C1,2,7,8TCDD)初步研究了其在鲤体内的分布和代谢规律。14C1,2,7,8-TCDD溶解于丙酮/植物油中,腹腔暴露。暴露1、2、4、8、12d后取样,肝脏、胆汁、腹腔脂肪等消化制样后用液闪仪测量放射性活度。肝脏和胆汁内的放射性活度同步变化,都是第8d达到峰值后下降。腹腔脂肪内1—2d放射性明显高于随后取样的样品。肝和腹腔脂肪的分布量之比呈现“S”型变化趋势。暴露4d

2,3,7,8-四氯代二苯并二英(TCDD)和苯并芘(B[a]P)对原代培养鲫鱼肝细胞中卵黄蛋白原诱导的影响

通过测定原代培养鲫鱼(Carassius auratus)肝细胞中雌激素受体所介导的卵黄蛋白原(Vtg)生成以及芳香烃受体所介导的CYP1A1基因转录水平的变化, 建立了一种类雌激素体外实验模型. 实验结果表明, Vtg和Vtg mRNA的表达与己烯雌酚(DES)之间均有很好的剂量-效应关系, Vtg和Vtg mRNA均可作为指示类雌激素毒性的生物标志物. TCDD, B[a]P可显著抑制鱼肝细胞中DES诱导的Vtg和Vtg mRNA的表达, 呈明显的抗雌激素效应, 并同时激活了CYP1A1 基因的表达;

2,3,7,8-TCDD对稀有鮈鲫幼体超微结构影响的观察

国家自然科学基金29607004

Effects of 2,3,7,8-TCDD and benzo[a]pyrene on modulating vitellogenin expression in primary culture of crucian carp (Carassius auratus) hepatocytes

Vitellogenin (Vtg) is the precursor of yolk protein. Its expression and secretion are estrogen-regulated and are crucial for oocyte maturation. An in vitro xenoestrogen screening model was established by measuring Vtg induction in cultured primary hepatocytes from crucian carp. Vtg production was detected by biotin-avidin sandwich ELISA method while Vtg and cytochrome P4501A1 (CYP1A1) mRNA induction were measured by semi- quantitative PCR-primer dropping technique. Vtg and Vtg mRNA were dose-dependently induced by diethylstilbestrol (DES, 0.2-200 ng/mL) in hepatocytes of crucian carp. Co-treatment of the DES-induced hepatocytes with either 2,3,7,8-TCDD (TCDD, 0.1-4 pg/mL) or benzo[a]pyrene (B[a]P, 5-1000 ng/mL) resulted in a reduction of Vtg production and an increment of CYP1A1 mRNA expression both in a dose dependent manner, indicating the anti-estrogenic effects of the compounds. However, at lower tested concentrations, TCDD (0.1, 0.2 pg/mL), B[a]P (5 ng/mL) seemed to have a potentiating effect on Vtg expression and secretion, although by their own these compounds had no observable estrogenic effect on Vtg induction. Tamoxifen (a selective estrogen receptor modulators, 1 nmol/L-1 mumol/L), and P-naphtho-flavone (beta-NF, an aryl hydrocarbon receptor inducing compounds, 2.5-1000 ng/mL) also were employed to study the possible interactions in DES-induced Vtg expression. In co-treatment of the DES-induced hepatocytes with beta-NF or tamoxifen, the decrease in Vtg production did parallel induction of CYP1A1 for beta-NF, but tamoxifen inhibited Vtg induction did not parallel induced CYP1A1 expression in all test concentrations. On the contrary, it was found that in co-treatment of the TCDD-induced hepatocytes with DES, TCDD induced CYP1A1 mRNA production was inhibited by DES also. These results implicated a possible cross talk between estrogen receptor- and aryl hydrocarbon receptor-mediated pathways in the hepatocytes.

Immunofluorescence detection and localization of B[a]P and TCDD in earthworm tissues

An immunohistochemical method using antibodies against polycyclic aromatic hydrocarbons (PAHs) and dioxins was developed on frozen tissue sections of the earthworm Eisenia andrei exposed to environmentally relevant concentrations of benzo[a]pyrene (B[a]P) (0.1, 10, 50 ppm) and 2,3,7,8-tetrachlorodibenzo-para-dioxin (TCDD) (0.01, 0.1, 2 ppb) in spiked standard soils. The concentrations of B[a]P and TCDD in E. andrei exposed to the same conditions were also measured using analytical chemical procedures. The results demonstrated that tissues of worms exposed to even minimal amount of B[a]P and TCDD reacted positively and specifically to anti-PAHs and -dioxins antibody. Immunofluorescence revealed a much more intense staining for the gut compared to the body wall; moreover, positively immunoreactive amoeboid coelomocytes were also observed, i.e. cells in which we have previously demonstrated the occurrence of genotoxic damage. The double immunolabelling with antibodies against B[a]P/TCDD and the lysosomal enzyme cathepsin D demonstrated the lysosomal accumulation of the organic xenobiotic compounds, in particular in the cells of the chloragogenous tissue as well as in coelomocytes, involved into detoxification and protection of animals against toxic chemicals. The method described is timesaving, not expensive and easily applicable.

Análise do imprinting e da expressão dos genes H19 e IGF2 em células de Sertoli humanas apos exposição in vitro ao 2,3,7,8-Tetraclorodibenzo-p-dioxina (TCDD)

Pós-graduação em Biologia Geral e Aplicada - IBB

Defining suitable reference genes for RT-qPCR analysis on human sertoli cells after 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Toxicologia reprodutiva de ratos adultos expostos ao 2,3,7,8-Tetraclorodibenzo-P-Dioxina (TCDD) in útero

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Tissue-specific induction of EROD activity and CYP1A protein in Sparus aurata exposed to B(a)P and TCDD

The purpose of this study was to compare xenobiotic CYP1A induction in liver, gills, and excretory kidney of gilthead seabream, Sparus aurata. Fishes were exposed via water for 20 days to different concentrations of benzo(a)pyrene (B(a)P) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). CYP1A was measured at the enzyme activity level as 7-ethoxyresorufin-O-deethylase (EROD) activity, and at the protein level by means of ELISA. The liver displayed the highest absolute levels of EROD activity, both under non-exposed and exposed conditions. Organ- or inducer-related differences in the time course of CYP1A induction were moderate; however, the magnitude of the induction response varied between the organs and between B(a)P and TCDD. In the case of TCDD, liver, and kidney yielded a comparable induction response, whereas in the case of B(a)P, the kidney showed a substantially higher maximum induction factor than the liver. In the gills, the two xenobiotics resulted in similar maximum induction factors. In B(a)P-exposed seabream, EROD activities and CYP1A protein levels showed a good correlation in all three organs, whereas with TCDD as inducer the correlation was poor, what was mainly due to a decrease of EROD activities at the higher concentrations of TCDD, while CYP1A protein levels showed no concomitant decline. Overall, the study revealed both similarities and differences in the time-, concentration-, and inducer-dependent CYP1A responses of the three target organs, liver, kidney, and gills. Although, the findings of this study principally confirm the notion of the liver as the major metabolic organ in fish, they also provide evidence for substantial metabolic potential in gills and particularly in the kidney.

Health assessment document for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related compounds.

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