Dataset Information

Ventral prostate and mammary gland phenotype in mice with complete deletion of the ER? gene.

ABSTRACT: Disagreements about the phenotype of estrogen receptor ? (ER?) knockout mouse, created by removing the DNA-binding domain of the ER? gene or interruption of the gene with a neocassette (Oliver Smithies ER? knockout mice [ER?OS-/-]), prompted us to create an ER? knockout mouse by deleting the ER? gene with the use of CRISPR/Cas9 technology. We confirmed that the ER? gene was eliminated from the mouse genome and that no ER? mRNA or protein was detectable in tissues of this mouse. Overall the phenotype of the ventral prostate (VP) and mammary gland (MG) in ER?crispr-/- mice was similar to, but more severe than, that in the ER?OS-/-mice. In the VP of 6-mo-old ER?crispr-/- mice there was epithelial hyperplasia, fibroplasia, inflammation, stromal overgrowth, and intraductal cancer-like lesions. This was accompanied by an increase in Ki67 and P63 and loss in DACH1 and PUR?, two androgen receptor (AR) repressors. In the MG there was overexpression of estrogen receptor ? and progesterone receptor, loss of collagen, increase in proliferation and expression of metalloproteases, and invasive epithelium. Surprisingly, by 18 mo of age, the number of hyperplastic foci was reduced, the ducts of the VP and MG became atrophic, and, in the VP, there was massive immune infiltration and massive desquamation of the luminal epithelial cells. These changes were coincident with reduced levels of androgens in males and estrogens in females. We conclude that ER? is a tumor suppressor gene in the VP and MG where its loss increases the activity AR and ER?, respectively.

SUBMITTER: Warner M

PROVIDER: S-EPMC7060692 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Ventral prostate and mammary gland phenotype in mice with complete deletion of the ERβ gene.

Warner Margaret M Wu Wan-Fu WF Montanholi Leticia L Nalvarte Ivan I Antonson Per P Gustafsson Jan-Ake JA

Proceedings of the National Academy of Sciences of the United States of America 20200219 9

Disagreements about the phenotype of estrogen receptor β (ERβ) knockout mouse, created by removing the DNA-binding domain of the ERβ gene or interruption of the gene with a neocassette (Oliver Smithies ERβ knockout mice [ERβ<sup>OS-/-</sup>]), prompted us to create an ERβ knockout mouse by deleting the ERβ gene with the use of CRISPR/Cas9 technology. We confirmed that the ERβ gene was eliminated from the mouse genome and that no ERβ mRNA or protein was detectable in tissues of this mouse. Overal ...[more]

PMID: 32075916

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Project description:Androgens are required for prostate development, growth and physiology, by activating the androgen receptor (AR) upon activation by testosterone and dihydrotestosterone (DHT), the AR undergoes conformational changes, dimerizes and translocates to the cell nucleus regulation important genes releted to cell survival. Understanding the mechanisms of androgen regulation in the prostate gland is important, because the prostate is affected by several different diseases, in particular prostate cancer (PCa). Several ways exist to treat prostate cancer and promote epithelial cell death. Treatments involving androgen manipulation include surgical castration (bilateral orchiectomy), antiandrogens (usually AR antagonists), or substances that inhibit androgen synthesis (5 alpha-reductase inhibitors, gonadotrophin-releasing hormone blockers). 17 beta-estradiol exerts anti-androgen effects by blocking the hypothalamic production of gonadotropin-releasing hormone and thereby inhibiting the production of testosterone by the testes , but also acts locally via interactions with either of the estrogen receptors found in the gland. It is known that the kinetics of apoptosis are different in the rat ventral prostate (VP) of castrated rats (Cas group) and in rats subjected to 17 beta-estradiol high dose (group E2) or their combination (group Cas+E2), with an evident additive effect in the latter situation (Garcia-Florez et al, 2005). The microarray approach was done to figure out what genes are expressed and how the cells of ventral prostate gland responses when the androgen is not available comparing three diferent androgen deprivation methods (sirurgical castration, high dose of 17-beta estradiol and both treatment combined). Forty-eight 21-day-old male Wistar rats were obtained from the Multidisciplinary Center for Biological Research (CEMIB), University of Campinas. The animals were kept under normal light conditions (12-h light:dark cycle) and received filtered tap water and Purina rodent chow ad libitum. On the 90th day after birth, the rats were divided in four groups (n=3) and assigned to different treatment groups. To cause androgen deprivation, we utilized three different procedures with different effects on epithelial cell apoptosis. Animals in the first group were castrated (Cas) by orchiectomy via scrotal incision under ketamine (150 mg/Kg body weight) and xylazin (10 mg/kg body weight) anesthesia. Animals in the second group received a 25 mg/Kg body weight dose of 17β-estradiol diluted in corn oil (E2 group). The third group received a combination of both treatments (Cas+E2 group) (combined orchiectomy and 17β-estradiol). In the control group (Ct; normal androgen and estrogen), the animals received only the vehicle. Three days after the treatments, the rats were killed by anesthetic overdose, and the ventral prostate was dissected out for the microarray and immunohistochemistry analyses.

Project description:The E2F transcription factors control key elements of development, including mammary gland branching morphogenesis, with several E2Fs playing essential roles. Additional prior data has demonstrated that loss of individual E2Fs can be compensated by other E2F family members, but this has not been tested in a mammary gland developmental context. Here we have explored the role of the E2Fs and their ability to functionally compensate for each other during mammary gland development. Using gene expression from terminal end buds and chromatin immunoprecipitation data for E2F1, E2F2 and E2F3, we noted both overlapping and unique mammary development genes regulated by each of the E2Fs. Based on our computational findings and the fact that E2Fs share a common binding motif, we hypothesized that E2F transcription factors would compensate for each other during mammary development and function. To test this hypothesis, we generated RNA from E2F1-/-, E2F2-/- and E2F3+/- mouse mammary glands. QRT-PCR on mammary glands during pregnancy demonstrated increases in E2F2 and E2F3a in the E2F1-/- mice and an increase in E2F2 levels in E2F3+/- mice. During lactation we noted that E2F3b transcript levels were increased in the E2F2-/- mice. Given that E2Fs have previously been noted to have the most striking effects on development during puberty, we hypothesized that loss of individual E2Fs would be compensated for at that time. Double mutant mice were generated and compared with the single knockouts. Loss of both E2F1 and E2F2 revealed a more striking phenotype than either knockout alone, indicating that E2F2 was compensating for E2F1 loss. Interestingly, while E2F2 was not able to functionally compensate for E2F3+/- during mammary outgrowth, increased E2F2 expression was observed in E2F3+/- mammary glands during pregnancy day 14.5 and lactation day 5. Together, these findings illustrate the specificity of E2F family members to compensate during development of the mammary gland.

Dataset Information

Ventral prostate and mammary gland phenotype in mice with complete deletion of the ER? gene.

Publications

Ventral prostate and mammary gland phenotype in mice with complete deletion of the ERβ gene.

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