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Endpoint:
additional toxicological information
Type of information:
experimental study
Adequacy of study:
key study
Study period:
no information
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study well documented, meets generally accepted scientific principles, acceptable for assessment
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
publication
Title:
Cadmium mimics the in vivo effects of estrogen in the uterus and mammary gland
Author:
Johnson MD, Kenney N, Stoica A, Hilakivi-Clarke L, Singh B, Chepko G, Clarke R, Sholler PF, Lirio AA, Foss C, Reiter R, Trock B, Paik S and Martin MB
Year:
2003
Bibliographic source:
Nat. Med. 9(8):1081-1084

Materials and methods

Type of study / information:
Type: other: estrogen-like activity
Principles of method if other than guideline:
Study was conducted to analyse the estrogenic activity of cadmium.
GLP compliance:
not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Cadmium chloride
- Procured from Sigma

Results and discussion

Any other information on results incl. tables

- 3.8-fold increase in uterine wet weight in the estradiol-treated animals.

- In the cadmium-treated animals, there was a 1.9-fold increase in uterine wet weight that was blocked by the antiestrogen.

- A similar increase in uterine wet weight was also observed when the animals were given a single intraperitoneal 10 µg/kg dose of cadmium (1.7-fold increase) or when the animals were ovariectomized on Day 40 (1.8-fold increase).

-Histological examination showed that the cadmium-induced increase in uterine wet weight was due to a mitogenic response and not due to toxicity (Fig. 1).

- In the ovariectomized control animals, the endometrial lining was flat to cuboidal. No vacuolation or stromal inflation were observed in either the endometrial or stromal cells.

- In the estradiol-treated animals, the endometrial lining showed epithelial hyperplasia and hypertrophy. The endometrial cells were taller and had abundant granular cytoplasm. The surrounding stroma was hypercellular and was infiltrated by numerous eosinophils. Cadmium-treated animals also showed hyperplasia and hypertrophy. In addition, the cells showed abundant subnuclear and supranuclear vacuolation. The stroma was more cellular in the cadmium-treated animals than in the ovariectomized animals and was less cellular than in the animals treated with estradiol. No stromal inflammatory infiltrate was noted in the cadmium-treated animals. Both estradiol- and cadmium-treated animals showed rare mitoses in the endometrial cells.

- The ability of the antiestrogen to block the effects of cadmium suggests that the effects of the metal are mediated by the estrogen receptor. There was no evidence of toxicity in the liver or kidney, organs sensitive to the toxic effects of the metal (data not shown), and there was no effect on whole body weight. To determine whether cadmium mimics the effects of estrogens in the mammary gland, epithelial density was measured on days 4 and 14 after treatment. In the ovariectomized control animals, the mammary gland consisted of a simple ductal network with low epithelial density. After exposure to estradiol, there was a 50% increase in epithelial density on Day 4 and 14, the result of an increase in mammary ducts and secretory lobuloalveolar structures. In animals exposed to cadmium, there was also a 50% increase in epithelial density by Day 4 and a 30% increase by Day 14. The cadmium-induced augmentation was due to a rise in quaternary branching of the ducts as well as an increase in lobuloalveolar structures. In rats treated with the antiestrogen, the epithelial density was not significantly different from control rats but there were fewer secretory structures in the glands of antiestrogen-treated rats on day 4, with a more pronounced effect observed on day 14. The antiestrogen also blocked the effects of cadmium on epithelial density and secretory structures, suggesting that the response of the mammary gland to cadmium is mediated by the estrogen receptor. To assess whether cadmium induced a secretory differentiation of the gland, the expression of casein and whey acidic protein were also examined on day 14. In control rats, the mammary gland was devoid of casein, whereas animals treated with estradiol for 14 d synthesized significant amounts of the protein. Casein was found in the ductal lumen, alveolar cells and alveolus. Rats exposed to a single dose of cadmium also synthesized significant amounts of casein. The protein was localized in lumenal cells, alveolar cells, alveolus and ductal lumen, with most of the casein immunolocalized in the cytoplasm. Expression of whey acidic protein was also detected but was not as abundant. The ability of cadmium to induce the synthesis of both casein and whey acidic protein indicates that similar to estradiol, the metal induces milk protein synthesis in the mammary gland. To determine whether cadmium also mimics the effects of estradiol on gene expression, we measured the amounts of PgR and complement protein C3 mRNA. In the uterus, estradiol treatment resulted in a 3-fold increase in PgR mRNA and a 124-fold increase in C3 mRNA. Similarly, treatment with cadmium induced a 2-fold increase in PgR mRNA and a 12-fold increase in complement C3 mRNA (P < 0.001). In the mammary gland, estradiol induced a 42- and 416-fold increase in PgR mRNA and C3 mRNA, respectively, and cadmium induced a 9-fold increase in PgR mRNA and a 16-fold increase in C3 mRNA (P< 0.001). The increase in PgR and C3 mRNA expression in both organs was blocked by the antiestrogen, providing additional evidence that the effects of cadmium are mediated by the estrogen receptor. The amount of cadmium in the uterus and mammary gland was also determined using anodic stripping voltammetry, an electrochemical technique that offers detection limits in the subparts-per-billion range. Cadmium was not detected in the organs of control animals but was detectable in most organs of the metal-treated animals. However, the amount of cadmium was too low to accurately quantitate. When detectable in the uterus or mammary gland, the amount of cadmium was approximately 10-2 pg per g of tissue (10-5 parts per billion). To assess the estrogenic effects of cadmium after in utero exposure, pregnant rats were given two injections of cadmium intraperitoneally at a dose of either 0.5 or 5 µg per kg body weight on days 12 and 17 of gestation. Cadmium did not alter pregnancy weight gain, number of pups per litter or birth weights. On postnatal day 35, however, female offspring exposed to the lower dose of cadmium had significantly increased body weights (135.8 ± 2.4 g, mean ± s.e.m.) compared with control offspring. This temprorary increase in weight is consistent with in utero exposure to estrogenic compounds. Also consistent with in utero exposure to low doses of estrogens, there was no difference in uterine wet weight (either crude or adjusted for body weight) between control animals and animals exposed to cadmium. In utero exposure to cadmium also induced an earlier onset of vaginal opening. Vaginal opening occurred on average on day 30.6 ± 0.6 in control animals, and on day 27.2 ± 1.1 (P <0.05) and day 26.7 ± 1.1 (P < 0.05) in animals exposed to cadmium doses of 0.5 and 5 µg/kg, respectively. The effects of in utero exposure to cadmium on mammary gland development were assessed on postnatal day 35 during the rapid growth phase of the gland. As with perinatal exposure to estrogenic compounds, in utero exposure to cadmium increased the parenchymal area of the mammary gland and the number of terminal end buds and decreased the number of alveolar buds. The mammary epithelial area was significantly larger, at 70.7 ± 5.2 mm2 and 66.5 ± 7.7 mm2 (mean ± s.e.m.) in rats exposed to cadmium doses of 0.5 and 5 µg/kg, respectively, compared with 45.5 ± 4.2 mm2 in control rats. The mammary glands of rats exposed to the lower dose of cadmium also contained significantly more terminal end buds (12.5 ±1.0) than control rats (9.4 ± 0.2). Both doses of cadmium reduced the number of alveolar buds in the mammary gland from 15.0 ± 3.9 in control rats to ab. 7.5 ± 1.5 in rats exposed to the metal. The ability of cadmium to mimic the in utero effects of estrogens provides additional support that environmentally relevant doses of the metal have potent estrogen-like activities. To date, most animal studies that have examined the toxic and carcinogenic effects of cadmium have used doses in the range of 1-5 mg/kg (ab 5-25 µmol/kg). To mimic human exposure, we used 0.1 % of this dose of cadmium, similar to dietary exposures according to the World Health Organization recommended Provisional Tolerable Weekly Intake of 7 µg per kg body weight per week.

Applicant's summary and conclusion

Conclusions:
The data presented in this study provide strong evidence that cadmium is a potent nonsteroidal estrogen in vivo.
Executive summary:

A study was conducted to confirm the estrogen-like activity of Cadmium.

 

Cadmium chloride (Sigma) was dissolved in sterile PBS and administered as a single intraperitoneal injection at a dose of 5 µg per kg body weight (ab.27 nmol/kg). An estradiol 30-d release pellet (Innovative Research of America) was implanted subcutaneously. The antiestrogen ICI-182,780 (Tocris) was dissolved in peanut oil and given intraperitoneally at a dose of 500 µg per kg per d. Animals were killed either 4 or 14 d later and the effects on histology and gene expression were examined.           

 

Exposure to cadmium increased uterine wet weight, promoted growth and development of the mammary glands and induced hormone-regulated genes in ovariectomized animals. In the uterus, the increase in wet weight was accompanied by proliferation of the endometrium and induction of progesterone receptor (PgR) and complement component C3. In the mammary gland, cadmium promoted an increase in the formation of side branches and alveolar buds and the induction of casein, whey acidic protein, PgR and C3. In utero exposure to the metal also mimicked the effects of estrogens. Female offspring experienced an earlier onset of puberty and an increase in the epithelial area and the number of terminal end buds in the mammary gland.

 

The results of the present study show that cadmium also has potent estrogen-like activity in vivo.