Transforming growth factor-B (TGF-B) isoforms are growth factors that function physiologically to regulate development, cellular proliferation, and immune responses. The role of TGF-B signaling in mammary tumorigenesis is complex, as TGF-B has been reported to function as both a tumor suppressor and tumor promoter. To elucidate the role of TGF-B signaling in mammary gland development, tumorigenesis, and metastasis, the gene encoding type II TGF-B receptor, Tgfbr2, was conditionally deleted in the mammary epithelium (Tgfbr2

TGF-β is an important paracrine factor in tumorigenesis. Ligand binding of the type I and II TGF-β receptors initiate downstream signaling. The role of stromal TGF-β signaling in prostate cancer progression is unknown. In mice the conditional stromal knockout of the TGF-β receptor type II expression (Tgfbr2fspKO) resulted in the development of prostatic intraepithelial neoplasia and progression to adenocarcinoma within seven months. Clinically, we observed a loss of TGF-β receptor type II expression in 69% of human prostate cancer-associated stroma compared to 15% of stroma associated with benign tissues (n =140, p value < 0.0001). To investigate the mechanism of paracrine TGF-β signaling in prostate cancer progression, we compared the effect of the prostatic stromal cells from Tgfbr2fspKO and Tgfbr2floxE2/floxE2 mice on LNCaP human prostate cancer cells in vitro and tissue recombination xenografts. Induction of LNCaP cell proliferation and tumorigenesis was observed by Tgfbr2fspKO prostate stroma as a result of elevated Wnt3a expression. Neutralizing antibodies to Wnt3a reversed LNCaP tumorigenesis. The TGF-β inhibition of Wnt3a expression was in part through the suppression of Stat3 activity on the Wnt3a promoter. In conclusion, the frequent loss of stromal TβRII expression in human prostate cancer can relieve the paracrine suppression of Wnt3a expression.

Mechanisms of androgen dependence of the prostate are critical to understanding prostate cancer progression to androgen independence associated with disease mortality. Transient elevation of transforming growth factor-B (TGF-B) occurs after androgen ablation. To determine the role of TGF-B on prostate response to androgen ablation, conditional TGF-B type II receptor knockout mouse models of the epithelia (Tgfbr2 NKX3.1KO ) and stromal fibroblasts (Tgfbr2 fspKO ) were used. After castration, the prostates of Tgfbr2 NKX3.1KO mice had apoptosis levels similar to those expected for control Tgfbr2 floxE2/floxE2 mice. Prostates of Tgfbr2 fspKO mice, however, had reduced regression and high levels of proliferation associated with canonical Wnt activity throughout the glandular epithelia regardless of androgen status. In contrast, Tgfbr2 floxE2/floxE2 prostates had epithelial canonical Wnt activity only in the surviving proximal ducts after castration. In vitro studies showed that androgen antagonist, bicalutamide, transiently elevated both Tgfbr2 floxE2/floxE2 and Tgfbr2 fspKO stromal expression of Wnt-2, Wnt-3a, and Wnt-5a. The neutralization of Wnt signaling by the expression of secreted frizzled related protein-2 (SFRP-2) resulted in decreased LNCaP prostate epithelial cell proliferation in stromal conditioned media transfer experiments. In vivo tissue recombination studies using Tgfbr2 fspKO prostatic stromal cells in combination with wild-type or SV40 large T antigen expressing epithelia resulted in prostates that were refractile to androgen ablation. The expression of SFRP-2 restored the Tgfbr2 fspKO -associated prostate responsiveness to androgen ablation. These studies reveal a novel TGF-B, androgen, and Wnt paracrine signaling axis that enables prostatic regression of the distal ducts after androgen ablation while supporting proximal duct survival.

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Manipulatable models of bladder development which interrogate specific pathways are badly needed. Such models will allow a systematic investigation of the multitude of pathologies which result from developmental defects of the urinary bladder. In the present communication, we describe a model in which mouse embryonic stem (ES) cells are directed to differentiate to form bladder tissue by specific interactions with fetal bladder mesenchyme. This model allows us to visualize the various stages in the differentiation of urothelium from ES cells, including the commitment to an endodermal cell lineage, with the temporal profile characterized by examining the induction of specific endodermal transcription factors (Foxa1 and Foxa2). In addition, final functional urothelial differentiation was characterized by examining uroplakin expression. It is well established that ES cells will spontaneously develop teratomas when grown within immunocompromised mouse hosts. We determined the specific mesenchymal to ES cell ratios necessary to dictate organ-specific differentiation while completely suppressing teratomatous growth. Embryonic mesenchyme is well established as an inductive tissue which dictates organ-specific programming of epithelial tissues. The present study demonstrates that embryonic bladder mesenchyme can also steer ES cells towards developing specific endodermal derived urothelium. These approaches allow us to capture specific stages of stem cell differentiation and to better define stem cell hierarchies.