Androgen signaling is essential for prostate development, morphogenesis, and regeneration. Emerging evidence indicates that Wnt/β-catenin signaling also contributes to prostate development specifically through regulation of cell fate determination. Prostatic Axin2-expressing cells are able to respond to Wnt signals and possess the progenitor properties to regenerate prostatic epithelium. Despite critical roles of both signaling pathways, the biological significance of androgen receptor (AR) in Axin2-expressing/Wnt-responsive cells remains largely unexplored. In this study, we investigated this important question using a series of newly generated mouse models. Deletion of Ar in embryonic Axin2-expressing cells impaired early prostate development in both ex vivo and tissue implantation experiments. When Ar expression was deleted in prostatic Axin2-expressing cells at pre-puberty stages, it results in smaller and underdeveloped prostates. A subpopulation of Axin2 expressing cells in prostate epithelium is resistant to castration and, following androgen supplementation, is capable to expand to prostatic luminal cells. Deletion of Ar in these Axin2-expressing cells reduces their regenerative ability. These lines of evidence demonstrate an indispensable role for the Ar in Wnt-responsive cells during the course of prostate development, morphogenesis, and regeneration, which also imply an underlying interaction between the androgen and Wnt signaling pathways in the mouse prostate. Stem Cells 2018;36:891-902.

The precise role of Wnt/β-catenin signaling during prostatic development and tumorigenesis is unclear. Axin2 is a direct transcriptional target of β-catenin. Recent studies have shown that Axin2-expressing cells have stem/progenitor cell properties in a variety of mouse tissues. Here, we genetically labeled Axin2-expressing cells at various time points and tracked their cellular behavior at different developmental and mature stages. We found that prostatic Axin2-expressing cells mainly express luminal epithelial cell markers and are able to expand luminal cell lineages during prostatic development and maturation. They can also survive androgen withdrawal and regenerate prostatic luminal epithelial cells following androgen replacement. Deletion of β-catenin or expression of stabilized β-catenin in these Axin2-expressing cells results in abnormal development or oncogenic transformation, respectively. Our study uncovers a critical role of Wnt/β-catenin-responsive cells in prostatic development and regeneration, and that dysregulation of Wnt/β-catenin signaling in these cells contributes to prostatic developmental defects and tumorigenesis.

Alzheimer's disease (AD) is the world's most common form of dementia, in which aggregation of amyloid-β (Aβ) is the hallmark. Unfortunately, few medicines have succeeded to completely cure AD. Yangxue Qingnao (YXQN) is a Chinese traditional medicine, and its pharmacological effect is improving cerebral blood flow. In this study, we firstly demonstrated that YXQN reduced AD-like pathology and cognitive impairment in APPswePS1dE9 (APP/PS1) mice with 2 months administration. Our data showed that YXQN substantially ameliorated behavioral defects in 10-month old APP/PS1 mice using Morris Water Maze and Y-maze tests, in which the cognitive ability of YXQN high-dose group approaches to wild type mice. Next, we focused on the brain pathological alterations in the YXQN group by three experiments, including thioflavin-S, congo-red, and Aβ-immunohistochemistry staining. The results demonstrated that the high-dose of YXQN dramatically suppressed amyloid plaques in the hippocampus and cortex of APP/PS1 mice, which showed a 47-72% reduction in plaque deposits, relative to the vehicle group. In addition, our data verified that YXQN decreased the cerebral amyloid load by attenuating β-secretase BACE1 and γ-secretase PS1 in the pathological processing of APP, and promoting the level of α-secretase ADAM10 in the physiological processing of APP to generate more sAPPα, which combats amyloidosis formation, and also carries out neurotropic and neuroprotective effect. Taken together, our results strongly suggest that YXQN could be a potential medicine for AD, and provide new evidence for further AD drug research and development.

Emerging evidence has shown that both prostatic basal and luminal cells are able to initiate oncogenic transformation. However, despite the diversity of tumor-initiating cells, most prostate cancer cells express the androgen receptor (AR) and depend on androgens for their growth and expansion, implicating an essential role of androgen signaling in prostate tumorigenesis. Prostatic basal cells express p63 and are able to differentiate into luminal, neuroendocrine, and basal cells. Here, we directly assessed the essential role of androgen signaling in prostatic p63-expressing cell initiated oncogenic transformation and tumor formation. Using novel and relevant mouse models, we demonstrated that, with stabilized β-catenin expression, prostatic p63-expressing cells possess the ability to initiate oncogenic transformation and, in the presence of androgens, they further transdifferentiate into luminal-like tumor cells and develop adenocarcinomas. Castration prior to activating stabilized β-catenin sensitizes p63-expressing cells and increases their sensitivity to androgens, resulting in aggressive and fast growing tumor phenotypes. These findings are consistent with what have been observed in human prostate cancers, demonstrating an essential role for androgen signaling in prostate cancer initiation and progression. This study also provides fresh insight into developing new therapeutic strategies for better treating prostate cancer patients.

E-cadherin complexes with the actin cytoskeleton via cytoplasmic catenins and maintains the functional characteristics and integrity of the epithelia in normal epithelial tissues. Lost expression of E-cadherin disrupts this complex resulting in loss of cell polarity, epithelial denudation and increased epithelial permeability in a variety of tissues. Decreased expression of E-cadherin has also been observed in invasive and metastatic human tumors. In this study, we investigated the effect of E-cadherin loss in prostatic epithelium using newly developed genetically engineered mouse models. Deletion of E-cadherin in prostatic luminal epithelial cells with modified probasin promoter driven Cre (PB-Cre4) induced the development of mouse prostatic intraepithelial neoplasia (PIN). An increase in levels of cytoplasmic and nuclear β-catenin appeared in E-cadherin deleted atypical cells within PIN lesions. Using various experimental approaches, we further demonstrated that the knockdown of E-cadherin expression elevated free cytoplasmic and nuclear β-catenin and enhanced androgen-induced transcription and cell growth. Intriguingly, pathological changes representing prostatic epithelial cell denudation and increased apoptosis accompanied the above PIN lesions. The essential role of E-cadherin in maintaining prostatic epithelial integrity and organization was further demonstrated using organoid culture approaches. To directly assess the role of loss of E-cadherin in prostate tumor progression, we generated a new mouse model with bigenic Cdh1 and Pten deletion in prostate epithelium. Early onset, aggressive tumor phenotypes presented in the compound mice. Strikingly, goblet cell metaplasia was observed, intermixed within prostatic tumor lesions of the compound mice. This study provides multiple lines of novel evidence demonstrating a comprehensive role of E-cadherin in maintaining epithelial integrity during the course of prostate oncogenic transformation, tumor initiation and progression.

The tumor suppressor p16Ink4a, encoded by the INK4a gene, is an inhibitor of cyclin D-dependent kinases 4 and 6, CDK4 and CDK6. This inhibition prevents the phosphorylation of the retinoblastoma protein (pRb), resulting in cellular senescence through inhibition of E2F-mediated transcription of S phase genes required for cell proliferation. The p16Ink4a plays an important role in tumor suppression, whereby its deletion, mutation, or epigenetic silencing is a frequently observed genetic alteration in prostate cancer. To assess its roles and related molecular mechanisms in prostate cancer initiation and progression, we generated a mouse model with conditional deletion of p16Ink4a in prostatic luminal epithelium. The mice underwent oncogenic transformation and developed prostatic intraepithelial neoplasia (PIN) from eight months of age, but failed to develop prostatic tumors. Given the prevalence of aberrant androgen signaling pathways in prostate cancer initiation and progression, we then generated R26hARL/wt:p16L/L: PB-Cre4 compound mice, in which conditional expression of the human AR transgene and deletion of p16Ink4a co-occur in prostatic luminal epithelial cells. While R26hARL/wt:PB-Cre4 mice showed no visible pathological changes, R26hARL/wt:p16L/L: PB-Cre4 compound mice displayed an early onset of high-grade PIN (HGPIN), prostatic carcinoma, and metastatic lesions. Strikingly, we observed tumors resembling human sarcomatoid carcinoma with intermixed focal regions of signet ring cell carcinoma (SRCC) in the prostates of the compound mice. Further characterization of these tumors showed they were of luminal epithelial cell origin, and featured characteristics of epithelial to mesenchymal transition (EMT) with enhanced proliferative and invasive capabilities. Our results not only implicate a biological role for AR expression and p16Ink4a deletion in the pathogenesis of prostatic SRCC, but also provide a new and unique genetically engineered mouse (GEM) model for investigating the molecular mechanisms for SRCC development.

Recent genome analysis of human prostate cancers demonstrated that both AR gene amplification and TP53 mutation are among the most frequently observed alterations in advanced prostate cancer. However, the biological role of these dual genetic alterations in prostate tumorigenesis is largely unknown. In addition, there are no biologically relevant models that can be used to assess the molecular mechanisms for these genetic abnormalities. Here, we report a novel mouse model, in which elevated transgenic AR expression and Trp53 deletion occur simultaneously in mouse prostatic epithelium to mimic human prostate cancer cells. These compound mice developed an earlier onset of high-grade prostatic intraepithelial neoplasia and accelerated prostate tumors in comparison with mice harboring only the AR transgene. Histological analysis showed prostatic sarcomatoid and basaloid carcinomas with massive squamous differentiation in the above compound mice. RNA-sequencing analyses identified a robust enrichment of the signature genes for human prostatic basal cell carcinomas in the above prostate tumors. Master regulator analysis revealed SOX2 as a transcriptional regulator in prostatic basal cell tumors. Elevated expression of SOX2 and its downstream target genes were detected in prostatic tumors of the compound mice. Chromatin immunoprecipitation analyses implicate a coregulatory role of AR and SOX2 in the expression of prostatic basal cell signature genes. Our data demonstrate a critical role of SOX2 in prostate tumorigenesis and provide mechanistic insight into prostate tumor aggressiveness and progression mediated by aberrant AR and p53 signaling pathways.

Androgen signaling is essential for prostate development, morphogenesis, and regeneration. Emerging evidence also indicates a regulatory role of Notch signaling in prostate development, differentiation, and growth. However, the collaborative regulatory mechanisms of androgen and Notch signaling during prostate development, growth, and regeneration are largely unknown. Hairy and Enhancer of Split 1 (Hes1) is a transcriptional regulator of Notch signaling pathways, and its expression is responsive to Notch signaling. Hes1-expressing cells have been shown to possess the regenerative capability to repopulate a variety of adult tissues. In this study, we developed new mouse models to directly assess the role of the androgen receptor in prostatic Hes1-expressing cells. Selective deletion of AR expression in embryonic Hes1-expressing cells impeded early prostate development both in vivo and in tissue xenograft experiments. Prepubescent deletion of AR expression in Hes1-expressing cells resulted in prostate glands containing abnormalities in cell morphology and gland architecture. A population of castration-resistant Hes1-expressing cells was revealed in the adult prostate, with the ability to repopulate prostate epithelium following androgen supplementation. Deletion of AR in Hes1-expressing cells diminishes their regenerative ability. These lines of evidence demonstrate a critical role for the AR in Notch-responsive cells during the course of prostate development, morphogenesis, and regeneration, and implicate a mechanism underlying interaction between the androgen and Notch signaling pathways in the mouse prostate.

Prostate embryonic development, pubertal and adult growth, maintenance, and regeneration are regulated through androgen signaling-mediated mesenchymal-epithelial interactions. Specifically, the essential role of mesenchymal androgen signaling in the development of prostate epithelium has been observed for over 30 years. However, the identity of the mesenchymal cells responsible for this paracrine regulation and related mechanisms are still unknown. Here, we provide the first demonstration of an indispensable role of the androgen receptor (AR) in sonic hedgehog (SHH) responsive Gli1-expressing cells, in regulating prostate development, growth, and regeneration. Selective deletion of AR expression in Gli1-expressing cells during embryogenesis disrupts prostatic budding and impairs prostate development and formation. Tissue recombination assays showed that urogenital mesenchyme (UGM) containing AR-deficient mesenchymal Gli1-expressing cells combined with wildtype urogenital epithelium (UGE) failed to develop normal prostate tissue in the presence of androgens, revealing the decisive role of AR in mesenchymal SHH responsive cells in prostate development. Prepubescent deletion of AR expression in Gli1-expressing cells resulted in severe impairment of androgen-induced prostate growth and regeneration. RNA-sequencing analysis showed significant alterations in signaling pathways related to prostate development, stem cells, and organ morphogenesis in AR-deficient Gli1-expressing cells. Among these altered pathways, the transforming growth factor β1 (TGFβ1) pathway was up-regulated in AR-deficient Gli1-expressing cells. We further demonstrated the activation of TGFβ1 signaling in AR-deleted prostatic Gli1-expressing cells, which inhibits prostate epithelium growth through paracrine regulation. These data demonstrate a novel role of the AR in the Gli1-expressing cellular niche for regulating prostatic cell fate, morphogenesis, and renewal, and elucidate the mechanism by which mesenchymal androgen-signaling through SHH-responsive cells elicits the growth and regeneration of prostate epithelium.