Meng-Yin Tsai

Abnormal Mammary Gland Development and Growth Retardation in Female Mice and MCF7 Breast Cancer Cells Lacking Androgen Receptor

Phenotype analysis of female mice lacking androgen receptor (AR) deficient (AR −/−) indicates that the development of mammary glands is retarded with reduced ductal branching in the prepubertal stages, and fewer Cap cells in the terminal end buds, as well as decreased lobuloalveolar development in adult females, and fewer milk-producing alveoli in the lactating glands. The defective development of AR −/− mammary glands involves the defects of insulin-like growth factor I–insulin-like growth factor I receptor and mitogen-activated protein kinase (MAPK) signals as well as estrogen receptor (ER) activity. Similar growth retardation and defects in growth factor–mediated Ras/Raf/MAPK cascade and ER signaling are also found in AR −/− MCF7 breast cancer cells. The restoration assays show that AR NH2-terminal/DNA-binding domain, but not the ligand-binding domain, is essential for normal MAPK function in MCF7 cells, and an AR mutant (R608K), found in male breast cancer, is associated with the excessive activation of MAPK. Together, our data provide the first in vivo evidence showing that AR-mediated MAPK and ER activation may play important roles for mammary gland development and MCF7 breast cancer cell proliferation.

Activin A Enhances Prostate Cancer Cell Migration Through Activation of Androgen Receptor and Is Overexpressed in Metastatic Prostate Cancer

Bone metastasis is the major cause of mortality associated with prostate cancer. Whereas activin A is known to inhibit prostate cancer cell growth and promote apoptosis, the correlation of elevated activin A with increasing serum prostate‐specific antigen (PSA) levels in bone metastatic stages of prostate cancer is well documented. The molecular mechanisms explaining these paradoxical effects of activin A and how activin A influences the progression of prostate cancer with bone metastasis remain unclear. By comparing expression profiles of primary prostate cancer biopsies, with and without bone metastasis, we discovered that the expression of activin A is increased in cases with bone metastatic propensity and correlates with increased androgen receptor (AR), PSA expression, and Gleason scores. Activin A promotes migration of prostate cancer cells to osteoblasts, elevates the AR gene transcription through Smads through binding to AR promoter, and induces nuclear translocation of AR to interact with Smad3. Knockdown of Smad3 by siRNA decreases activin A–promoted AR expression and cancer cell migration. Overexpression of AR reversed Smad3‐siRNA suppression on activin A–mediated cell migration to osteoblasts. These data suggest that activation of the AR through Smads is required for activin A–promoted prostate cancer cell migration to bone matrix, thereby promoting the bone metastatic phenotype, and the activin A–Smad–AR axis may be considered a therapeutic target in bone metastatic diseases.

Androgen receptor regulates expression of skeletal muscle-specific proteins and muscle cell types.

C2C12 myoblasts expressing the androgen receptor (AR) were used to analyze the role of androgen-AR signaling pathway in skeletal muscle development. Marked up-regulation of AR expression was observed in differentiated myotubes. A nuclear run-on transcription assay demonstrated that transcription of the AR gene is increased during skeletal muscle cell differentiation. Regulation of skeletal muscle-specific protein expression by the androgen-AR signaling pathway was further analyzed using quadriceps skeletal muscle from wild-type (WT) and AR knock-out (ARKO) male mice. A histological analysis of quadriceps skeletal muscle indicates no morphological differences between ARKO and WT mice. However, the androgen-AR signaling pathway increases expression of slow-twitch-specific skeletal muscle proteins and downregulates fast-twitch-specific skeletal muscle proteins, resulting in an increase of slow-twitch muscle fiber type cells in quadriceps muscle.

Tumor Suppressor PAX6 Functions as Androgen Receptor Co-Repressor to Inhibit Prostate Cancer Growth

PAX6, a transcription factor, has currently been suggested to function as a tumor suppressor in glioblastoma and to act as an early differentiation marker for neuroendocrine cells. The androgen receptor (AR) plays a pivotal role in prostate cancer development and progression due to its transcriptional activity in regulating genes involved in cell growth, differentiation, and apoptosis. To determine the role of PAX6 in prostate cancer, we investigated whether PAX6 interacts with AR to affect prostate cancer development.

Roles of testicular orphan nuclear receptors 2 and 4 in early embryonic development and embryonic stem cells.

The testicular orphan nuclear receptors (TRs) 2 and 4 act as either transcriptional activators or regulatory proteins of other nuclear receptor superfamily members. With no identified cognate ligands, their physiological roles remain unclear. Here we showed the phenotypes of TR2(-/-):TR4(-/-) mutant embryos, which reveal that the loss of TR2 and TR4 causes early embryonic lethality and increased cell death. We also found that TR2 and TR4 are expressed in blastocysts and embryonic stem (ES) cells, and can act as transcriptional activators in ES cells. The results on further investigating the roles of TR2 and TR4 in ES cells showed that TR2 and TR4 were differentially expressed when ES cells were induced into different specialized cell types, and their expression is regulated by retinoic acid. Knocking down TR2 and TR4 mRNAs decreased the expression of Oct-3/4 and Nanog genes. Mechanism dissection suggests that TR2 and TR4 may affect the Oct-3/4 gene by binding to a direct repeat-1 element located in its promoter region, which is influenced by retinoic acid. Together, our findings highlight possible roles for TR2 and TR4 in early embryonic development by regulating key genes involved in stem cell self-renewal, commitment, and differentiation.

Suppression of androgen receptor enhances the self-renewal of mesenchymal stem cells through elevated expression of EGFR.

Bone marrow derived mesenchymal stem cells (BM-MSCs) have been widely applied in several clinical trials of diseases, such as myocardial infarction, liver cirrhosis, neurodegenerative disease, and osteogenesis imperfecta. Although most studies demonstrated that transplantation of BM-MSCs did exert a temporary relief and short-term therapeutic effects, eventually all symptoms recur, therefore it is essential to improve the therapeutic efficacy of transplantation by either elevating the self-renewal of BM-MSCs or enhancing their survival rate. Herein we demonstrated that the BM-MSCs and adipocyte derived mesenchymal stem cells (ADSCs) isolated from the androgen receptor (AR) knockout mice have higher self-renewal ability than those obtained from the wild-type mice. Knockdown of AR in MSC cell lines exhibited similar results. Mechanistic dissection studies showed that the depletion of AR resulted in activation of Erk and Akt signaling pathways through epidermal growth factor receptor (EGFR) activation or pathway to mediate higher self-renewal of BM-MSCs. Targeting AR signals using ASC-J9® (an AR degradation enhancer), hydroxyflutamide (antagonist of AR), and AR-siRNA all led to enhanced self-renewal of MSCs, suggesting the future possibility of using these anti-AR agents in therapeutic approaches.

Altered TNSALP Expression and Phosphate Regulation Contribute to Reduced Mineralization in Mice Lacking Androgen Receptor

ABSTRACT While androgen receptor (AR)-deficient mice developed osteopenia in endochondral bones due to the high bone turnover with increased bone resorption by osteoclasts, little is known about the mechanism of intramembranous bone loss contributed by AR in osteoblasts. Here, we discovered a dramatic decrease in the area of calcification, new bone, and the number of osteocytes in calvaria from AR-deficient mice related to a reduction in mineralization caused, in part, by the diminished activity of AR-deficient osteoblasts. Enforced AR expression in differentiated osteoblasts boosts mineralization while knockdown of AR expression prevents androgen-induced mineralization. We identified the tissue-nonspecific alkaline phosphatase (TNSALP) and several members of small integrin binding ligand N-linked glycoprotein (SIBLING) gene family as androgen target genes required for AR-mediated bone formation. We show that inorganic phosphate (Pi) levels and TNSALP activity increased in response to androgen/AR and Pi signals increase the expression and translocation of AR. The ectopic expression of TNSALP or Pi partially rescued the bone loss due to AR deficiency. Thus, androgen/AR signaling plays an essential role in bone formation by coordinating the expression of genes associated with phosphate regulation.

Susceptibility to Autoimmunity and B Cell Resistance to Apoptosis in Mice Lacking Androgen Receptor in B Cells

Departments of Pathology, Urology, Microbiology and Immunology, and the Cancer Center (S.A., K.-H.C., K.-P.L., J.-J.L., H.-Y.L., F.M.Y., A.B., M.-Y.T., W.-P.Z., H.-C.C., S.Y., C.C.), University of Rochester Medical Center, Rochester, New York 14642; Clinical Research Laboratory (S.A.), Saad Specialist Hospital, Al Khobar 31952, Saudi Arabia; Center for Menopause and Reproductive Medicine Research (M.-Y.T.)., Chang Gung University/Hospital, Kaohsiung 333, Taiwan; and Department of Urology (H.-C.C.), National Taiwan University/Hospital, Taipei 10603, Taiwan

Interaction of Cell Cycle Regulatory Proteins with the Androgen Receptor

Androgen signaling is a key determinant in the development and maintenance of the male reproductive system, and also plays a critical role in associated pathologies such as prostate cancer. The androgen receptor (AR), a member of the steroid receptor (SR) subgroup of the nuclear receptor (NR) superfamily, modulates gene expression in response to androgens in target tissues including the brain, liver, pituitary gland and prostate (Chang et al., 1995). SRs have a conserved domain structure that includes ligand-independent and ligand-dependent activation functions (AF-1 & AF-2) and a general mechanism of action (White et al., 1998). In response to ligands, SRs undergo a conformational change within the ligand-binding domain (LBD), dissociate from chaperone proteins and form homodimers (White et al., 1998). The activated receptor dimer then enhances target gene expression from response elements within the target gene promoter by modulating the activity of the general transcription machinery (White et al., 1998).