Mouse Prostate Epithelium Research Articles

Pten null prostate tumorigenesis and AKT activation are blocked by targeted knockout of ER chaperone GRP78/BiP in prostate epithelium

GRP78/BiP has recently emerged as a novel biomarker for aggressive prostate cancer. Here, we report that homozygous deletion of Grp78 specifically in mouse prostate epithelium suppresses prostate tumorigenesis without affecting postnatal prostate development and growth. Mouse prostates with double conditional knockout of Grp78 and Pten exhibit normal histology and cytology, in contrast to the invasive adenocarcinoma in mouse prostates with Pten inactivation. AKT activation in Pten null prostate epithelium is inhibited by Grp78 homozygous deletion, corresponding with suppression of AKT phosphorylation by GRP78 knockdown in prostate cancer cell line. Thus, inactivation of GRP78 may represent a previously undescribed approach to stop prostate cancer and potentially other cancers resulting from the loss of PTEN tumor suppression and/or activation of the oncogenic AKT.

Expression and sub‐cellular localization of the CCAAT/enhancer binding protein α in relation to postnatal development and malignancy of the prostate

C/EBPalpha is a critical mediator of terminal differentiation and a tumor suppressor through its strong antiproliferative actions on cell cycle regulatory proteins. C/EBPalpha also appears to regulate androgen receptor (AR) AR signaling. There, is a paucity of information on the expression and sub-cellular localization of C/EBPalpha in normal mouse and human prostate and in prostate cancer. Immunohistochemistry of tissues including tissue arrays, quantitative polymerase chain reaction and mRNA expression database mining. In the mouse prostate epithelium, C/EBPalpha was present at 1 week postnatal localized in the cytosol, began to show nuclear localization at 8 weeks and continued to show prominent nuclear expression at 10 weeks and beyond; C/EBPalpha mRNA was expressed at all ages. In humans, C/EBPalpha showed prominent nuclear localization from peripubescence up to middle age but was sequestered in the cytosol in older individuals; the mRNA level for C/EBPalpha remained essentially unchanged. Most prostate adenocarcinomas expressed a range of levels of C/EBPalpha mRNA and protein that were relatively high in metastatic tumors in a manner that correlated with AR expression; however, most cells showed C/EBPalpha sequestered in the cytosol. Temporal changes in sub-cellular localization of C/EBPalpha are consistent with a role in prostate differentiation and as a prostate tumor suppressor; the cytoplasmic sequestration of C/EBPalpha, unique to older human prostates, is arguably a permissive condition for the greater frequency of proliferative disorders of the prostate. In malignant prostate C/EBPalpha may be available to regulate AR signaling through transient changes in its sub-cellular localization.

Temporally controlled ablation of PTEN in adult mouse prostate epithelium generates a model of invasive prostatic adenocarcinoma

Studies of prostate cancer pathogenesis and development of new therapies have been hampered by a lack of appropriate mouse models. We have generated PSA-Cre-ER(T2) mice that express the tamoxifen-dependent Cre-ER(T2) recombinase selectively in prostatic epithelium, thus allowing us to target floxed genes selectively in epithelial cells of fully differentiated prostate of adult mice and to modulate the number of genetically altered cells. Our present mouse model, in which prostate carcinogenesis is initiated through Cre-ER(T2)-mediated somatic biallelic ablation of the tumor suppressor gene PTEN after puberty, closely mimics the course of human cancer formation. Indeed, mutant mice developed prostate epithelium hyperplasia within 4 weeks after PTEN ablation and prostatic intraepithelial neoplasia (PIN) in all lobes within 2-3 months, with the highest incidence in the dorsolateral lobe, which is considered to be the most similar to the peripheral zone of the human prostate, in which adenocarcinoma is preferentially localized. Eight to 10 months after PTEN ablation some PINs of the dorsolateral lobe had progressed to adenocarcinoma, but no distant metastases were found up to 20 months after PTEN ablation, indicating that progression to metastasis requires an additional mutation or mutations. Interestingly, monoallelic Cre-ER(T2)-mediated PTEN ablation in epithelial cells of adult prostate also generated focal hyperplasia and PINs, but exclusively in the dorsolateral lobe, and in much lower number and after a longer latency. However, no progression to adenocarcinoma was observed. Because PTEN expression was undetectable in epithelial cells from these PINs, loss of PTEN function appears to act as a permissive event for uncontrolled cell proliferation.

Activation of Nonreceptor Tyrosine Kinase Bmx/Etk Mediated by Phosphoinositide 3-Kinase, Epidermal Growth Factor Receptor, and ErbB3 in Prostate Cancer Cells

Pathways activated downstream of constitutively active phosphatidylinositol (PI) 3-kinase in PTEN-deficient prostate cancer (PCa) cells are possible therapeutic targets. We found that the nonreceptor Tec family tyrosine kinase Bmx/Etk was activated by tyrosine phosphorylation downstream of Src and PI 3-kinase in PTEN-deficient LNCaP and PC3 PCa cells and that Bmx down-regulation by short interfering RNA markedly inhibited LNCaP cell growth. Bmx also associated with ErbB3 in LNCaP cells, and heregulin-beta1 enhanced this interaction and further stimulated Bmx activity. Epidermal growth factor (EGF) similarly stimulated an interaction between Bmx and EGF receptor and rapidly increased Bmx kinase activity. Bmx stimulation in response to heregulin-beta1 and EGF was Src-dependent, and heregulin-beta1 stimulation of Bmx was also PI 3-kinase-dependent. In contrast, the rapid tyrosine phosphorylation and activation of Bmx in response to EGF was PI 3-kinase-independent. Taken together, these results demonstrate that Bmx is a critical downstream target of the constitutively active PI 3-kinase in PTEN-deficient PCa cells and further show that Bmx is recruited by the EGF receptor and ErbB3 and activated in response to their respective ligands. Therefore, Bmx may be a valuable therapeutic target in PCa and other epithelial malignancies in which PI 3-kinase or EGF receptor family pathways are activated.

Bisphenol A induces permanent squamous change in mouse prostatic epithelium

Bisphenol A (BPA) is a monomer of plastic products widely used in daily life, and has weak estrogenic activity. In this study, male BALB/c mice were treated with BPA and diethylstilbestrol (DES) in adult and fetal periods to investigate whether BPA could affect prostatic epithelial differentiation. Eight-to 9-week-old mice treated for 3 weeks with subcutaneous implants of 0.2–200 mg BPA pellets induced the expression of cytokeratin 10 (CK10) in prostatic basal epithelial cells in a dose-dependent manner. Utilizing organ culture of adult prostate, 1 nM and 1 μM BPA also induced CK10 expression and squamous metaplasia with multilayering of basal epithelial cells, respectively. Fetal exposure to low-dose BPA (20 μg/kg/day) from gestation day (GD) 13 to GD18 induced permanent CK10 expression in basal cells of the adult prostate similar to DES (0.2 μg/kg/day). These results indicate that in mouse, BPA can directly elicit CK10 expression in prostatic epithelium, and that this change can be elicited by doses as low as 20 μg/kg/day. We speculate that low-dose BPA during fetal life may also induce permanent squamous change in human prostate.

Castration‐induced epithelial cell death in human prostate tissue is related to locally reduced IGF‐1 levels

Castration rapidly reduces stroma insulin-like growth factor (IGF)-1 synthesis and action in mouse prostate epithelium. We explore if similar changes are of importance for castration-induced prostate regression in humans. Epithelial and surrounding stroma cells were micro-dissected from patient biopsies obtained before and shortly after castration. IGF-1 mRNA levels were quantified by RT-PCR and related to epithelial apoptosis and IGF-1, IGF-1 receptor, and androgen receptor (AR) immunoreactivity. IGF-1 mRNA was principally produced in the stroma and IGF-R1 in the epithelium. Stroma IGF-1 mRNA levels were significantly decreased after castration in non-malignant but not malignant tissue. Lack of stroma IGF-1 reduction after castration was associated with low stroma AR expression before therapy. Reduction of IGF-1 mRNA levels in the tumor stroma and/or epithelium was associated with epithelial apoptosis after therapy. Low AR expression and maintained stroma IGF-1 synthesis may result in limited tumor cell death after castration therapy.

Tyrosine Kinase Etk/BMX Is Up-regulated in Human Prostate Cancer and Its Overexpression Induces Prostate Intraepithelial Neoplasia in Mouse

The nonreceptor tyrosine kinase Etk/BMX was originally identified from the human prostate xenograft CWR22. Here, we report that Etk is up-regulated in human prostate tumor specimens surveyed. Knocking down Etk expression by a specific small interfering RNA (siRNA) in prostate cancer cells attenuates cell proliferation, suggesting an essential role of Etk for prostate cancer cell survival and growth. Targeted expression of Etk in mouse prostate epithelium results in pathologic changes resembling human prostatic intraepithelial neoplasia, indicating that up-regulation of Etk may contribute to prostate cancer development. A marked increase of luminal epithelial cell proliferation was observed in the Etk transgenic prostate, which may be attributed in part to the elevated activity of Akt and signal transducers and activators of transcription 3 (STAT3). More interestingly, the expression level of acetyltransferase cyclic AMP-responsive element binding protein-binding protein (CBP) is also increased in the Etk transgenic prostate as well as in a prostate cancer cell line overexpressing Etk, concomitant with elevated histone 3 acetylation at lysine 18 (H3K18Ac). Down-modulation of Etk expression by a specific siRNA leads to a decrease of H3 acetylation in prostate cancer cell lines. Our data suggest that Etk may also modulate chromatin remodeling by regulating the activity of acetyltransferases, such as CBP. Given that Etk may exert its effects in prostate through modulation of multiple signaling pathways altered in human prostate cancer, the Etk transgenic mouse model may be a useful tool for studying the functions of Etk and identification of new molecular markers and drug targets relevant to human diseases.

NKX3.1 stabilizes p53, inhibits AKT activation, and blocks prostate cancer initiation caused by PTEN loss

We demonstrate that PTEN loss causes reduced NKX3.1 expression in both murine and human prostate cancers. Restoration of Nkx3.1 expression in vivo in Pten null epithelium leads to decreased cell proliferation, increased cell death, and prevention of tumor initiation. Whereas androgen receptor (AR) positively regulates NKX3.1 expression, NKX3.1 negatively modulates AR transcription and consequently the AR-associated signaling events. Consistent with its tumor suppressor functions, NKX3.1 engages cell cycle and cell death machinery via association with HDAC1, leading to increased p53 acetylation and half-life through MDM2-dependent mechanisms. Importantly, overexpression of Nkx3.1 has little effect on Pten wild-type epithelium, suggesting that PTEN plays a predominant role in PTEN-NKX3.1 interplay. Manipulating NKX3.1 expression may serve as a therapeutic strategy for treating PTEN-deficient prostate cancers.

Characterization of an antibody that can detect the Kai1/CD82 murine metastasis suppressor

Kai1, also known as CD82, is a member of the tetraspanin family (TM4SF). The human homolog, KAI1, is an activation antigen of T-cells and is a metastasis suppressor for prostate and other cancers. Little is known about the mouse protein because of the lack of antibody reagents. Peptide immunized rabbits were used to generate polyclonal antibody to Kai1. The antibody was analyzed using immunoblotting, flow cytometry, and immunohistochemistry. This antibody specifically recognizes murine Kai1 protein, crossreacts with rat Kai1 but not with human KAI1. The normal tissue distribution of this protein in mice is shown to be similar to that of the human homolog. Interestingly, mouse prostatic epithelium showed differential expression within the lobes. This antibody, the first described that can specifically detect murine Kai1/CD82, should be very useful in addressing the mechanism of action of Kai1 in metastatic suppression.

Critical Function for ADAM9 in Mouse Prostate Cancer

ADAM9 is a membrane-anchored metalloprotease that is markedly up-regulated in several human carcinomas. Here, we show that ADAM9 is similarly up-regulated in mouse models for prostate, breast, and intestinal carcinoma. To assess whether ADAM9 is critical for the pathogenesis of prostate carcinoma, one of the most common cancers in men, we evaluated how loss of ADAM9 affects tumorigenesis in W(10) mice, a mouse model for this disease. In the absence of ADAM9, most tumors in 50-week-old W(10) mice were well differentiated, whereas littermate controls expressing wild-type ADAM9 had predominantly poorly differentiated, and in some cases significantly larger, tumors. Moreover, gain-of-function experiments in which ADAM9 was overexpressed in mouse prostate epithelium resulted in significant abnormalities, including epithelial hyperplasia at 4 to 6 months of age, and prostatic intraepithelial neoplasia after 1 year. A potential underlying mechanism for the role of ADAM9 in prostate cancer emerged from cell-based assays: ADAM9 can cleave and release epidermal growth factor and FGFR2iiib from cells, both of which have pivotal functions in the pathogenesis of this disease. Taken together, these results suggest that ADAM9 contributes to the pathogenesis of prostate cancer and potentially also other carcinomas, raising the possibility that ADAM9 might be a good target for antitumor drugs.

Beta-catenin: a transforming actor on many stages.

Mutations and deletions that result in the stabilization of beta-catenin are frequently found in a number of tumors, including those of the colon, the liver and the ovary, but are less frequently found in breast cancer. To investigate and understand the molecular nature of cell-specific beta-catenin signaling, experimental mouse genetics has been employed extensively. Gain-of-function and loss-of-function mutations have provided evidence that beta-catenin plays essential roles in development and tumorigenesis. Specifically, the Wnt/beta-catenin signaling pathway controls cell fate decisions throughout development, and a unique role in differentiated epithelia has emerged. Not only beta-catenin, but also the activation of other components of this pathway in differentiated mammary epithelium and prostate epithelium of transgenic mice can induce neoplasias and transdifferentiation to squamous metaplasias. This suggests that the Wnt/beta-catenin pathway is dominant over existing differentiation programs and can impose an epidermal fate or neoplasias onto a variety of cell types. Although there is evidence for a contextual specificity of the Wnt signaling, the experimental systems and designs used in different studies probably influence the cellular responses.

The Metaplastic Effects of Estrogen on Mouse Prostate Epithelium: Proliferation of Cells with Basal Cell Phenotype

The Metaplastic Effects of Estrogen on Mouse Prostate Epithelium: Proliferation of Cells with Basal Cell Phenotype

The metaplastic effects of estrogen on mouse prostate epithelium: proliferation of cells with basal cell phenotype.

The exogenous administration of estrogens to male mice alters the hypothalamic-pituitary-gonadal axis and reduces androgen levels, leading to a regression of the prostatic epithelium. As well, a specific direct response to estrogens is the induction of epithelial squamous metaplasia. The aims of this study were to identify the process by which the prostatic epithelium is transformed in intact adult male mice using the synthetic estrogen, diethylstilbestrol. A comparison of the effects of diethylstilbestrol in the three lobes revealed a hierarchy of response, with the anterior lobe being the most responsive, the dorsolateral lobe less responsive, and the ventral lobe the least responsive. The effect of castration was used to distinguish between the epithelial responses to estrogen administration and androgen deprivation. The results demonstrate that transformation of the epithelium involved proliferation of cells with a basal cell phenotype, the onset of cytokeratin 10 expression, up-regulation of progesterone receptor expression, and loss of the cell cycle inhibitor, p27(Kip1) expression; none of these changes was observed after castration. Mice lacking functional estrogen receptor alpha failed to respond, demonstrating a requirement for estrogen receptor alpha in the epithelium and/or stroma to mediate the proliferative response to estrogen in the prostate gland.

Estrogenic effects on prostatic differentiation and carcinogenesis.

Estrogens, alone or in combination with androgens, can induce aberrant growth and/or malignancy of the prostate gland. Squamous metaplasia is an abnormal form of prostatic epithelial differentiation elicited by exogenous estrogen alone. Estrogens elicit their effects via estrogen receptors (ER) in the prostate. Experiments using ERalpha and ERbeta null mice demonstrated that ERalpha, but not ERbeta is essential in the induction of prostatic squamous metaplasia. To determine the respective roles of epithelial versus stromal ERalpha in this response, the following tissue recombinants were constructed with prostatic epithelium (PRE) and stroma (S) from wild-type (wt) and ERalpha knockout (alphaERKO) mice: wt-S + wt-PRE, alphaERKO-S + alphaERKO-PRE, wt-S + alphaERKO-PRE and alphaERKO-S + wt-PRE. A metaplastic response to diethylstilbestrol (DES) was only observed in wt-S + wt-PRE tissue recombinants. Tissue recombinants containing alphaERKO-PRE and/or alphaERKO-S (alphaERKO-S + alphaERKO-PRE, wt-S + alphaERKO-PRE and alphaERKO-S + wt-PRE) failed to respond to DES. Therefore, full and uniform epithelial squamous metaplasia requires ERalpha in both the epithelium and stroma. Estradiol (E2) in combination with testosterone (T) was shown to be effective in inducing prostatic carcinogenesis in a tissue recombinant model composed of rat urogenital sinus mesenchyme plus mouse prostatic epithelium. A particularly efficient model of prostatic carcinogenesis in mice involves T + E2 treatment of mice bearing grafts of wild-type rat urogenital mesenchyme (rUGM) plus retinoblastoma gene (Rb) knockout (Rb-KO) prostatic epithelium (rUGM + Rb-KO-PRE). Such rUGM + Rb-KO-PRE tissue recombinants developed hyperplasia, atypical hyperplasia and invasive prostatic carcinoma with high efficiency. During carcinogenesis in rUGM + Rb-KO-PRE tissue recombinants, epithelial E-cadherin almost totally disappeared and epithelial PCNA labeling was elevated. These epithelial changes were associated with almost total loss of smooth muscle cells in the stroma. The results of this study demonstrate that the absence of the Rb tumor suppressor gene predisposes prostatic epithelial cells to hormonal carcinogenesis.

Apoptosis during castration-induced regression of the prostate is Fos dependent.

Apoptotic cell death was shown to be accompanied or preceded by an elevated expression of the c-fos protooncogene and DNA binding activity of transcription factor AP-1. We used Fos-deficient mice to study the role of c-Fos during programmed cell death in the prostate. In normal mice apoptosis is induced in the prostate within 2-4 days after castration. Histological features of reduced secretory activity and morphological signs of programmed cell death become obvious. No apparent decrease in secretory activity and no epithelial cell death were observed in Fos-deficient animals after castration. Fragmentation of nuclear DNA was measured by in situ terminal transferase reaction. DNA fragmentation was observed in the prostate epithelium of control mice after castration whereas no similar fragmentation was found in Fos-deficient animals. After castration an AP-1 complex accumulated in the prostate of Fos deficient mice which mainly consists of FosB, Fra-2 and JunD whereas in control animals the AP-1 complex in addition contained c-Fos. Our data strongly suggest that c-Fos is required for programmed cell death of prostate epithelial cells.

Interaction of Retinoic Acid and 3-Methylcholanthrene on the Fine Structure of Mouse Prostate Epithelium in Vitro

The effects of 3-methylcholanthrene (MCA) and retinoic acid (RA) on the fine structure of AKR mouse prostate epithelium in organ culture were correlated with changes in cell proliferation. In intact glands before explantation, the epithelial cytoplasm showed concentric flat or globular cisternae of endoplasmic reticulum in both supranuclear and basal areas, a well-developed Golgi complex, secretory vesicles, and numerous microvilli at the luminal surface. After explantation, the cytoplasmic organelles, particularly the endoplasmic reticulum, regressed and tonofilaments appeared. The regression was largely prevented by RA. MCA induced considerable epithelial hyperplasia and squamous metaplasia. The fine structure of the newly formed cells revealed a complete loss of endoplasmic reticulum, Golgi apparatus, secretory vesicles, and microvilli, with the appearance of bundles of tonofilaments and a striking increase in the number of desmosomes. Administration of RA to explants pretreated with the carcinogen partially reversed the hyperplasia and squamous metaplasia. The tonofilaments disappeared and the number of desmosomes greatly decreased, whereas endoplasmic reticulum, Golgi complex, secretory vesicles, and microvilli were largely reestablished. Planimetric measurements of the alveolar epithelium showed that the squamous transformation and its partial reversal by RA coincide with the rise and decline of epithelial hyperplasia. The data suggest that the restoration of secretory differentiation by RA was responsible for the initial breakdown of the hyperplastic epithelium, whereas the lowering of DNA synthesis by RA prevented further hyperplasia and kept cell replication within normal limits.