Inhibition of gap junctional intercellular communication by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in rat hepatocytes

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent rodenthepatic tumor promoter. Unlike observations with the majorityof tumor promoting chemicals studied to date, most investigationshave failed to demonstrate down- regulation of gap junctionalintercellular communication (GJIC) in cultured cells by TCDD.The present study examined the effect of TCDD on GJIC in rathepatocytes in primary culture. At non-cytolethal doses TCDDinhibited GJIC In a time- (1, 4, 24 and 48 h) and concentration(1x10^–8–1x10^–14M)-dependent manner. This inhibitionoccurred within 4 h of treatment at doses of 1x10^–81x10–^–12MTCDD and persisted for up to 48 h, despite removal of TCDD.Treatment of rat hepatocytes with TCDD resulted in a decreasein hepatocyte connexin 32 mRNA, but had no apparent effect onconnexin 26 mRNA. Co-incubation of rat hepatocytes with TCDDand     

Growth modulation of hepatocytes and rat liver epithelial cells (WB-F344) by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)

Modulation of DNA synthesis by 2,3,7,8-tetrachlorodi-benzo-p-dioxin(TCDD) was studied in primary cultures of hepatocytes and inrat liver epithelial cells (WB-F344) to develop models for studieson the interactions between the activated Ah receptor and cellulargrowth control. In hepatocytes TCDD either positively or negativelymodulated EGF-stimulated DNA synthesis. In the presence of ethlnylestradiol10^–12 M TCDD moderately increased EGF-stimulated DNA synthyesis(     

Induction of hepatic DNA single strand breaks in rats by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)

Previous studies have demonstrated that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces lipid peroxidation in hepatic and extrahepatic tissues. DNA single strand breaks as well as other forms of DNA damage are believed to occur in conjunction with lipid peroxidation. We have therefore examined the effect of TCDD on hepatic DNA single strand breaks. Ten days after the administration of 100 micrograms TCDD/kg to female rats, a 7.5-fold increase in the DNA elution constant (single strand breaks) occurred. Similar changes were observed in the content of thiobarbituric acid reactive substances (TBARS) in the nuclei as well as the NADPH-dependent production of TBARS. The accumulation of TBARS appeared to precede the accumulation of DNA single strand breaks. The tumor promoting effects of TCDD may be associated with the enhanced formation of DNA single strand breaks.     

Inhibition of high-density growth arrest in human squamous carcinoma cells by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)

The highly toxic environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a potent carcinogen and tumor promoter, and affects cellular proliferation and differentiation both in vivo and in vitro. This report presents data showing that TCDD enhances the proliferation of two human squamous carcinoma cell lines in monolayer culture by inhibiting growth arrest at high cell density. SCC-15G and SCC-25 cells were treated with 0-100 nM TCDD in culture medium, and examined for changes in proliferation and differentiation. TCDD stimulated increases in cell number and DNA synthesis of both cell lines, and inhibited differentiation of SCC-15G cells in a dose-dependent manner. The minimum effective concentrations for increases in proliferation were 0.1 nM in SCC-15G cells and 1 nM in SCC-25G cells. The saturation density of SCC-15G cells grown in 10 nM TCDD was approximately double that of untreated controls, while the saturation density of SCC-25 cells was 50% above controls. TCDD-induced increases in proliferation were detectable only in cells exposed at subconfluent density, then assayed after control cultures had reached high-density growth arrest. There was no difference in cell number or DNA synthesis between control and TCDD-treated cultures when cells were both treated and assayed during the logarithmic phase of growth, nor in cultures treated after the cells had reached high density growth arrest. Therefore, TCDD-induced proliferation resulted from failure of treated cells to undergo normal density-dependent growth arrest rather than from direct mitogenic stimulation of the cells. Differentiation (envelope competence and keratin staining) of SCC-15G cells was inhibited by TCDD, despite the fact that in these cultures cell density was twice that of the controls. The sensitivity of SCC-15G cells to modulation of growth and differentiation by TCDD provides the basis for a model to examine the biological mechanisms of TCDD-induced alterations in proliferation and differentiation of epidermal cells.     

Prevention of apoptosis by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the MCF-10A cell line: correlation with increased transforming growth factor alpha production

We have recently reported that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) inhibits epidermal growth factor (EGF) withdrawal-induced apoptosis in the human mammary epithelial cell line MCF-10A. We hypothesized that TCDD-mediated inhibition of apoptosis was due to its ability to stimulate the EGF receptor (EGFR) pathway. Indeed, in the present studies, the EGFR inhibitor AG1478 was able to prevent TCDD-, EGF-, and transforming growth factor alpha (TGF-alpha)-dependent cell recovery and inhibition of apoptosis. These effects were specific for an EGFR-mediated pathway because cotreatment with AG825, an erbB2 inhibitor, had little effect on apoptosis. In addition, TCDD was able to mimic the EGF and TGF-alpha signaling as demonstrated by increasing Akt and extracellular signal-regulated kinase 1,2 phosphorylation. These effects were dependent on EGFR activity because AG1478, but not AG825, was able to prevent EGF-, TGF-alpha, or TCDD-mediated Akt and extracellular signal-regulated kinase 1,2 phosphorylation. The ability of TCDD to stimulate the EGFR pathway and inhibit apoptosis may be due to the ability of TCDD to increase expression of TGF-alpha, a ligand for EGFR. Treatment with 10 nM TCDD increased TGF-alpha mRNA at 2 h and TGF-alpha protein at 6 h. These data suggest a mechanism whereby TCDD is able to inhibit apoptosis in human mammary epithelial cells by stimulating TGF-alpha production, resulting in an autocrine effect.     

Effects of cycloheximide on the induction of CYP1A1 gene expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in three human breast cancer cell lines

Treatment of MCF-7, MDA-MB-231 and Hs578-T human breast cancercell lines with 10_–9 M 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD) induces CYP1A1 gene expression in the MCF-7 but not inthe MDA-MB-231 or Hs578-T cells. Pretreatment of the cells with10_–5 M cycloheximide results in significantly increasedP4501A1 mRNA levels in all three cells lines. However, in cellsco-treated with 10_–5 M cycloheximide plus 10_–9 MTCDD, an induced response by TCDD was observed in the MCF-7and MDA-MB-231 but not in Hs578-T cells. Gel-retardation assaysof nuclear extracts from the three cell lines complexed witha ³²P-labeled dioxin-responsive element (DRE) gave a TCDD-inducibleretarded band only in the MCF-7 and MDA-MB-231 cells. A retardedband with a similar mobility was observed in nuclear extractsfrom Hs578-T cells treated with either 10_–9 M TCDD orDMSO (solvent control). These results suggest that aryl hydrocarbonnon-responsive MDA-MB-231 and Hs578-T human breast cancer celllines contain the CYP1A1 gene and treatment with cycloheximideincreases both constitutive and TCDD-induced CYP1A1 gene expression.     

Interactions of rutaecarpine alkaloids with specific binding sites for 2,3,7,8-tetrachlorodibenzo-p-dioxin in rat liver

Rutaecarpine alkaloids have the capacity to inhibit specific2,3,7,8-[1,6-³H]tetrachlorodibenzo-p-dioxin (TCDD) binding inrat liver cytosol, as analysed by electrofocusing in polyacrylamidegel. The IC₅₀ value for binding of 7,8-dehydrorutaecarpine wasestimated to 7 nM indicating a high-affinity interaction, whereasrutaecarpine appeared less active (IC₅₀ 110 nM). These findingsare of interest in view of the fact that analogues to thesecompounds may be formed following UV-irradiation of tryptophanand that such photo-products have been suggested to constitute(the) endogenous ligand(s) for the TCDD receptor. As furthersupport of this notion, the rutaecarpine alkaloids investigatedcould be fitted into a rectangle of 6.8x13.7 A, a characteristiccommon for most high affinity ligands of the TCDD receptor hithertostudied. In view of their structural similarity to dehydrorutaecarpineand the agreement of their mol. wt with that of the photoproductwith the highest affinity for the TCDD receptor, we suggestdeaza-analogues of dehydrorutaecarpine to represent possiblecandidates for the endogenous TCDD receptor ligand.     

Ontogeny of the rat hepatic receptor for 2,3,7,8-tetrachlorodibenzo-p-dioxin and its endocrine indepence.

The ontogeny of the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) receptor was studied in Sprague-Dawley rats by quantitation of the receptor in liver cytosol using isoelectric focusing in polyacrylamide gel. No differences by sex in the receptor concentration were seen at any of the ages studied. Newborn, 21-day-old, and 42-day-old rats contained significantly more receptor in the liver cytosol than did 56-day-old rats. There was no significant difference in the receptor concentration in liver cytosol from 7-day-old rats compared to that from 56-day-old rats. The maximum receptor concentration was found in cytosol from 21-day-old rats [36.1 +/- 24.0 (S.D.) fmol/mg protein]. Adult rats (56 days old) contained the lowest concentration of receptor (13.3 +/- 6.3 fmol/mg protein). The level of TCDD receptor in liver cytosol from adult rats was not significantly changed by orchiectomy, ovariectomy, adrenalectomy, or hypophysectomy. The maximum for TCDD receptor concentration at puberty corresponds to the reported maximum for the induction of aryl hydrocarbon hydroxylase activity. However, no further conclusion can as yet be drawn concerning the regulation of the TCDD receptor.     

Metabolism of benzo[a]pyrene in rat prostate glands following 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure

The present study was undertaken to determine, using h.p.l.c.,both the rate and pattern of benzo[a]pyrene (BP) metabolismin monooxygenase(s)-induced and non-induced prostate glandsof adult Sprague Dawley rats (CD-strain). H.p.l.c. analysisshowed that for non-induced prostate glands, the initial syntheticrates of dihydrodiols, quinones and phenols in the organic extractablephase were 0.16, 0.18 and 0.16 pmol/min/mg protein, while forthe total BP metabolites in the aqueous phase it was 5.4 pmol/min/mgprotein. Therefore, BP metabolism in the non-induced prostategland is extremely low, with the total of BP metabolites inthe aqueous phase being approximately 11 times greater thanthe total of BP metabolites in the organic phase. In contrast,the rates of phenol, quinone and dihydrodiol synthesis in 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD)-induced prostate glands were 45.9, 34.5 and 6.0 pmol/min/mgprotein, respectively, and are thus significantly increasedby 290, 192 and 38 times, respectively over control rates. Atthe highest TCDD dose (10 µg/kg body weight), the syntheticrates for phenols and quinones increased by 200–300-foldand for the diols about 35-fold over controls. The rate of 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrenesynthesis was greater than 4,5-dihydroxy-4,5-dihydrobenzo[a]pyreneor 9,10-dihydroxy-9,10-dihydrobenzo[a]pyrene synthesis. Furthermore,the ratios of the total BP metabolites in the organic phaseto those in the aqueous phase of control and TCDD-induced were0.09 and 3.3, respectively. Thus, it is clear that one of themajor effects of TCDD in rat prostate glands is to significantlyincrease organic-soluble BP metabolites over water-soluble BPmetabolites. These experiments also demonstrated that the specificaryl hydrocarbon hydroxylase (AHH), epoxide hydrotase (EH) andglutathione transferase (GSH-T) activities at the maximum substrateconcentrations of BP and 4,5-benzo[a]pyrene 4,5-oxide in non-inducedprostate glands of CD-rats are 0.25, 50, 12,500 pmol/min/mgprotein, respectively. Thus, it is clearly shown that specificprostatic AHH activity is extremely low when compared to EHor GSH-T activity, which are 200 or 50,000 times greater thanAHH activity, respectively. However, TCDD treatment significantlyincreased the specific prostalic AHH activity (200 times), aswell as the cytochrome P-446 species (7 times), while specificEH and GSH-T activities were not altered.     

Carcinogenicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin in the Syrian golden hamster

The carcinogenic potential of 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD) was examined in Syrian golden hamsters, at differentdose levels. Twenty-one percent of the hamsters that receiveda total of 600 µg/kg body weight of TCDD either by thes.c. or i.p. route developed squamous cell carcinomas of theskin of facial region within 12-13 months from the beginningof the experiment. Neoplasms were not observed in any otherorgans. These studies suggest that TCDD may be a complete carcinogenin hamsters, the species most resistant to the toxic effectof this compound.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) does not inhibit intercellular communication in Chinese hamster V79 cells

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a ubiquitous environmentalpollutant which has been shown to be both a potent teratogenand carcinogen and also to have tumorpromoting activity. Wehave compared TCDD with the prototype phorbol ester tumor promoter,12-O-tetradecanoylphor-bol-13-acetate (TPA) using the metaboliccooperation assay as a measure of tumor promotional competence.Unlike TPA, TCDD was found to be ineffective in inhibiting metaboliccooperation at concentrations which elicit many of TCDD's biologicalresponses. These results suggest that TPA and TCDD elicit tumorpromotion by different pathways. We discuss these results inthe context of cell-type-specific and TCDD receptor-mediatedbiological responses to tumor promoters.     

Characterization of the promotion of altered hepatic foci by 2,3,7,8-tetrachlorodibenzo-p-dioxin in the female rat.

Previous studies have suggested that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) acts as a promoting agent in various organ systems including the rat liver. Since a major characteristic of the stage of tumor promotion is its operational reversibility, we have assessed whether TCDD-induced promotion is reversible in a two-stage model of hepatocarcinogenesis. In this model, female Fischer F344 rats were administered a single, intragastric dose of the initiating agent, diethylnitrosamine (DEN, 10 mg/kg), at the peak of proliferation induced by a partial hepatectomy. TCDD was then administered biweekly (0.14 micrograms/kg, s.c.) for 1, 3 or 5 months. One group of animals was killed at each of these time points, while a second group was maintained for each time point for an additional 6 months in the absence of further TCDD. Four serial frozen sections of liver were each stained with a different enzyme marker of altered hepatic foci (AHF). The AHF were identified and the number and volume fraction determined by quantitative stereology. Exposure to TCDD resulted in an increase in the number and size of AHF in the initiated relative to the uninitiated rats. Increasing the duration of promotion with TCDD led to an increase in the number of AHF per liver, the volume fraction of the liver occupied by AHF and the number of markers expressed aberrantly by a single AHF. Discontinuation of TCDD administration for 6 months before killing the animals resulted in a decrease in the total number of AHF observed, but those AHF that remained increased in size with an overall increase in volume fraction of AHF. Analysis of the size class distribution for AHF for each of the periods of TCDD promotion revealed an increase in the larger AHF but a decrease in the smaller, thereby resulting in an overall decrease in number of AHF with an increase in the volume fraction of AHF. Increasing the duration of the TCDD exposure prior to its withdrawal led to an increased AHF size, phenotypic complexity and number of AHF remaining after cessation of TCDD administration. Although the levels of TCDD in livers of rats 6 months after cessation of TCDD administration were still greater than background, they were markedly reduced compared to immediately after administration. Thus, cessation of exposure to TCDD after a brief duration led to a reversal of its promotional effects on the majority of AHF, while prolonged exposure led to maintained promotion of a minority of AHF.     

Suppression of estrogen-regulated extracellular tissue plasminogen activator activity of MCF-7 cells by 2,3,7,8-tetrachlorodibenzo-p-dioxin

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) suppresses the estrogen enhancement of tissue plasminogen activator (t-PA) by MCF-7 breast cancer cells. 17 beta-estradiol treatment of MCF-7 cells was previously shown to enhance t-PA secretion in a receptor-mediated process dependent on RNA and protein synthesis. The current studies demonstrate that treatment with TCDD, at a concentration as low as 10(-11) M, reduces the 17 beta-estradiol-induced enhancement of t-PA secretion in these cells. Treatment of MCF-7 cells with TCDD alone does not alter t-PA activity nor was inhibition of t-PA activity observed when TCDD was added directly to the enzyme assay. Kinetic studies and the lack of inhibition following in vitro mixing of conditioned media from TCDD-treated and control 17 beta-estradiol stimulated MCF-7 cells argue against TCDD induction of a plasminogen activator inhibitor. The related polychlorinated dibenzofuran, 2,3,7,8,-tetrachlorodibenzofuran, while also active, is less potent that TCDD. Other polychlorinated dibenzodioxins, polychlorinated dibenzofurans, and polychlorinated biphenyls do not suppress 17 beta-estradiol induction of t-PA over the concentrations tested. These results are in agreement with the structure-activity relationships established using these compounds in other assay systems. Treatment with TCDD does not alter the number or affinity of 17 beta-estradiol receptors of MCF-7 cells. TCDD treatment does not suppress constitutive t-PA activity in the estrogen independent breast cancer line MDA-MB-231 nor the t-PA induced by 12-O-tetradecanoylphorbol-13-acetate in HeLa cells. These effects suggest that TCDD is not acting directly on expression of the t-PA genome. Induction of aryl hydrocarbon hydroxylase by TCDD, a cytochrome P-450 regulated metabolic enzyme for which TCDD is the most potent known inducer, was observed in MCF-7 cells but not in MDA-MB-231 or HeLa cells. A plausible mechanism for the antiestrogenic activity of TCDD is based on the metabolic conversion of 17 beta-estradiol to less active derivatives by TCDD induced cytochrome P-450 metabolic enzymes.     

Levels and membrane localization of the c-K-ras p21 protein in lungs of mice of different genetic strains and effects of 2,3,7,8- tetrachlorodibenzo-p-dioxin (TCDD) and Aroclor 1254

Mutational activation of the K-ras oncogene often occurs in human and mouse lung adenocarcinomas. Since K-ras p21 functions in trans-membrane signaling, we have investigated whether the amount of this protein in lung cell membranes is a variable that could influence lung tumorigenesis, either due to genetic differences or in response to tumor promoters. The six mouse strains assessed showed little difference in the total lung K-ras p21 after immunoprecipitation and immunoblotting. However, amount of ras p21 in the membrane fraction showed significant differences, with C57BL/6 and BALB/c having 3-5-fold more than NIH Swiss, AKR and DBA mice. Interestingly, a congenic AKR strain having the Ahr(b-1) Ah receptor allele from C57BL/6 mice (designated AKR.B6Ah) had high lung membrane K-ras p21 similar to that of C57BL/6. To test for possible changes related to lung tumor promotion, mice were treated with a promotional dose of TCDD (5 nmol/kg). After 48 h C57BL/6 lungs showed an increase in p21 in both total and membrane fractions. BALB/c, DBA and Swiss mice showed an increase only in membranes. There was no change in the AKR and AKR.B6Ah. Aroclor 1254 (250 mg/kg) caused an increase in membrane/cytosol ratio in Swiss mice. Thus the membrane:cytosol K-ras p21 ratio may be influenced by the Ahr phenotype, and TCDD and PCBs can induce p21 or increase its membrane level in certain strains, but these properties are not fully dependent on Ahr receptor type. In confirmation of the relevance of these findings for the tumor target cell type, the immortalized alveolar type 2 E10 cell line presented K- ras p21 in membrane, and this was increased 4-fold by treatment with 10 nM TCDD.      

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on protein kinase C and inositol phosphate metabolism in primary cultures of rat hepatocytes

Adult rat hepatocytes, after maintenance for 24 h in serum-freeculture, were treated with the tumor promoters, 12-O-tetradecanoylphorbol-13-acetate(TPA) or 2,3,7,8-tetrachloro-dibenzo-p-dioxin (TCDD). Short-termtreatment (15 min) with TPA, 1 µM, increased protein kinaseC (PKC) activity in the particulate fraction of hepatocytesand, concomitantly, decreased the vasopressin (100 nM)-stimulatedsynthesis of inositol phosphates. The latter effect of TPA couldbe prevented by prior addition of the PKC inhibitor, H7 (100µM). After short-term treatment (15 min) with TCDD, 1pM, no effects on PKC or inositol phosphate metabolism wereobserved. However, after prolonged exposure to TCDD (3–48h), the particulate PKC was significantly activated (1.5-fold).In contrast to the effect of TPA (24 h), no down-regulationwas found. Moreover, long-term treatment with TCDD significantlyenhanced vasopressin-stimulated inositol 1,3,4,5-tetrakisphosphatesynthesis, while TPA treatment (24 h) stimulated the synthesisof inositol trisphosphates and inositol 1,3,4,5-tetrakisphosphate.The results suggest that the tumor promoters, TPA and TCDD,act differently on the signal transduction pathways in hepatocytes.Thus, the effects of TCDD on PKC and inositol phosphate metabolismmight be mediated by a yet unknown mechanism rather than bydirect activation of PKC as seen with TPA.     

Carcinogenic effects of infantile and long-term 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment in the mouse

An infantile carcinogenesis assay was carried out with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) injections administered intraperitoneally at 0, 1, 30 and 60 micrograms/kg b.w. doses to (C57BL/6J X C3Hf)F1 (B6C3) and to (C57BL/6J X BALB/c)F1 (B6C) mice, starting from the 10th day of life, once weekly repeated 5 times. Animals were then observed until 78 weeks of age. The induction of thymic lymphomas was related to treatment at 60 micrograms/kg dose level in both sexes of both hybrids, and at 30 micrograms/kg dose level in both sexes of B6C mice and in male but not female B6C3 mice. The incidence of hepatocellular adenomas was increased by TCDD treatment at 60 micrograms/kg dose level in B6C3 of both sexes but not in B6C mice. Hepatocellular carcinomas were seen at increased incidence at 30 and 60 micrograms/kg doses in B6C3 males but not in B6C3 females or in B6C mice of both sexes. The incidence of other tumor types was not related to treatment in both hybrids. A long-term carcinogenesis bioassay with TCDD was carried out in B6C3 mice treated by gavage at 0, 2.5 and 5.0 micrograms/kg b.w. doses from 6 weeks of age, once weekly for 52 weeks. The animals were observed until 110 weeks of age. An increased incidence of hepatocellular adenomas and carcinomas was related to treatment, at both doses and in both sexes. The incidence of other tumor types was uniformly low in treated and control groups, without any association with treatment, in both sexes.     

Carcinogen-binding proteins in the rat ventral prostate: specific and nonspecific high-affinity binding sites for benzo(a)pyrene, 3-methylcholanthrene, and 2,3,7,8-tetrachlorodibenzo-p-dioxin

The polychlorinated dibenzodioxin [3H]-2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) and the carcinogens [3H]benzo(a)pyrene and [3H]-3-methylcholanthrene bound to saturable binding sites in cytosol from the rat ventral prostate. Analysis of equilibrium binding parameters in diluted cytosol preparations indicated an apparent Kd of approximately 2 nM and a binding capacity of approximately 1 nmol/mg cytosolic protein, corresponding to approximately 5% of the total protein content. However, gel permeation chromatography analysis as well as velocity sedimentation analysis on sucrose gradients of [3H]TCDD-labeled rat prostatic cytosol indicated binding of [3H]TCDD to two discrete species. These analyses indicated a sedimentation coefficient of 3.6-3.8S, a Stokes radius of 25-28 A, and a calculated relative molecular weight of 42,000-45,000 for the most abundant binding species. The other binding species sedimented at 4-5S under high ionic strength conditions and at 8-10S under low ionic strength conditions and had a Stokes radius of approximately 60 A, a relative molecular weight of approximately 100,000 and an estimated concentration of 5-20 fmol/mg cytosolic protein. Binding of [3H]TCDD to this species was displaceable by a 200-fold M excess of 2,3,7,8-tetrachlorodibenzofuran. Therefore, this species was tentatively identified as the TCDD receptor. The properties of the high-capacity binder of [3H]TCDD were found to be similar to the characteristics of a protein previously purified from the rat ventral prostate, prostatic secretory protein, which binds androgens as well as estramustine, a nitrogen mustard derivative of estradiol. The binding of estramustine to diluted prostatic cytosol was shown to be competitively inhibited by 2,3,7,8-tetrachlorodibenzofuran. Moreover, purified prostatic secretory protein bound [3H]TCDD, [3H]benzo(a)pyrene, as well as [3H]-3-methylcholanthrene. It is suggested that binding to this protein is responsible for the high-binding capacity of carcinogens in cytosol from the rat ventral prostate.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) promotes the transformation of C3H/10T1/2 cells

Continuous treatment of C3H/10T1/2 cells with low concentrations(>4pM) of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) enhancedfocus production in cultures pretreated with N-methyl-N'-nitro-N-nitrosoguanidine.Maximal enhancement occurred at 40 pM TCDD, a concentration10 000-fold lower than that required to produce an optimal responsewith 12-O-tetradecanoylphorboI-13-acetate. Single treatmentswith 0.06 nM-5 µM TCDD did not transform C3H/10T1/2 cellsor initiate the process of transformation in cultures subsequentlyexposed to the tumor promoter 12-Otetradecanoylphorbol-13-acetate.Promotion of transformation is thus the predominant effect ofTCDD in the C3H/101/2 cell transformation system.