Generierung eines genetischen Mausmodells zur Analyse der FoxP3-abhängigen Regulation der ErbB2/HER2-Expression während der Brustdrüsenentwicklung und der Tumorgenese des Mammakarzinoms
Final Report Abstract
The transcription factor Foxp3 is a master regulator of T-cell fate, and its constitutive expression in a subset of CD4+ T cells is crucial for the generation and maintenance of socalled regulatory T cells (Treg). Treg cells play an important role in sustaining self-tolerance by controlling the activity of CD4+ and CD8+ effector T cells. While Foxp3 is highly expressed in Treg cells, low levels of Foxp3 expression were previously reported for normal mammary gland and other epithelial tissues. Foxp3 has been shown to act as a tumor suppressor in the mammary gland controlling expression of cellular proto-oncogenes such as ErbB2 (HER2). Scurfy mice carry an inactivating mutation in the X-chromosomally encoded Foxp3 gene (Foxp3sf). Homozygous and hemizygous scurfy mice develop fatal autoimmune disease after birth due to the loss of functional Treg cells. For heterozygous Foxp3sf/+ mice the development of mammary carcinomas and other tumors with increased frequency was reported, which was attributed to random silencing of the remaining functional Foxp3 allele by X chromosome inactivation. Mammary carcinomas in these mice were characterized by elevated ErbB2 levels. To investigate whether Foxp3 also plays a role during normal mammary gland development, and to study in more detail the contribution of a loss of Foxp3 to malignant transformation, we developed a genetic mouse model characterized by targeted deletion of Foxp3 in the mammary gland. We generated Foxp3fl/fl;Cre mice, which carry a conditional allele of Foxp3 flanked by loxP sites, and an MMTV-LTR-driven Cre transgene predominantly expressed in the mammary tissue. Analysis of sorted cell populations revealed successful knock-out of Foxp3 in luminal, myoepithelial, and stem-cell-like cells of the mammary gland. Histological analysis of mammary tissues of virgin and lactating Foxp3fl/fl;Cre mice showed no differences to those of control mice, and uni- and multiparous animals displayed no defects in lactation. These results demonstrate that Foxp3 is not critical for normal mammary gland development. In aged Foxp3fl/fl;Cre females we observed ductal and alveolar hyperplasia. Nevertheless, this phenotype became apparent later than reported for Foxp3sf/+ mice, and in contrast to the latter we did not see the appearance of malignant breast cancers. We also observed a mild and latent autoimmune phenotype in female Foxp3fl/fl;Cre and male Foxp3fl/y;Cre animals, likely due to activation of the MMTV-Cre transgene in lymphocytes. This resulted in markedly reduced numbers of functional Treg cells. Hence, our genetic mouse model with attenuated Treg numbers may also be useful for immunological studies. We noted the presence of few, but in comparison to controls increased numbers of lymphocytes in mammary tissues and ducts of some Foxp3fl/fl;Cre mice, possibly associated with the reduced Treg cell numbers. This raises the question whether hyperplastic growth of mammary epithelial cells in aged Foxp3fl/fl;Cre mice is a cell-intrinsic function, or indirectly caused by inflammatory reactions and possible chronic inflammation in mammary tissue. Ongoing work focuses on investigating the expression of estrogen and progesterone receptors, the proliferation marker Ki-67, and the Foxp3 target genes ErbB2 and c-Myc in Foxp3fl/fl;Cre mice, which may serve as early markers for disregulated growth of Foxp3-negative mammary epithelial cells.
