Prostatic Smooth Muscle Research Articles

Altered prostate epithelial development and IGF‐1 signal in mice lacking the androgen receptor in stromal smooth muscle cells

Androgens and the androgen receptor (AR) play critical roles in the prostate development via mesenchymal-epithelial interactions. Smooth muscle cells (SMC), differentiated from mesenchyme, are one of the basic components of the prostate stroma. However, the roles of smooth muscle AR in prostate development are still obscure. We established the smooth muscle selective AR knockout (SM-ARKO) mouse model using the Cre-loxP system, and confirmed the ARKO efficiency at RNA, DNA and protein levels. Then, we observed the prostate morphology changes, and determined the epithelial proliferation, apoptosis, and differentiation. We also knocked down the AR in a prostate smooth muscle cell line (PS-1) to confirm the in vivo findings and to probe the mechanism. The AR was selectively and efficiently knocked out in the anterior prostates of SM-ARKO mouse. The SM-ARKO prostates have defects with loss of infolding structures, and decrease of epithelial proliferation, but with little change of apoptosis and differentiation. The mechanism studies showed that IGF-1 expression level decreased in the SM-ARKO prostates and AR-knockdown PS-1 cells. The decreased IGF-1 expression might contribute to the defective development of SM-ARKO prostates. The AR in SMCs plays important roles in the prostate development via the regulation of IGF-1 signal.

The Residual Nonadrenergic Contractile Response to Nerve Stimulation of the Mouse Prostate Is Mediated by Acetylcholine but Not ATP in a Comparison with the Mouse Vas Deferens

Neuronal release of noradrenaline is primarily responsible for the contraction of prostatic smooth muscle in all species, and this forms the basis for the use of α(1)-adrenoceptor antagonists as pharmacotherapies for benign prostatic hyperplasia. Previous studies in mice have demonstrated that a residual nonadrenergic component to nerve stimulation remains after α(1)-adrenoceptor antagonism. In the guinea pig and rat prostate and the vas deferens of guinea pigs, rats, and mice, ATP is the mediator of this residual contraction. This study investigates the mediator of residual contraction in the mouse prostate. Whole prostates from wild-type, α(1A)-adrenoceptor, and P2X1-purinoceptor knockout mice were mounted in organ baths, and the isometric force that tissues developed in response to electrical field stimulation or exogenously applied agonists was recorded. Deletion of the P2X1 purinoceptor did not affect nerve-mediated contraction. Furthermore, the P2-purinoceptor antagonist suramin (30 μM) failed to attenuate nerve-mediated contractions in wild-type, α(1A)-adrenoceptor, or P2X1-purinoceptor knockout mice. Atropine (1 μM) attenuated contraction in prostates taken from wild-type mice. In the presence of prazosin (0.3 μM) or guanethidine (10 μM), or in prostates taken from α(1A)-adrenoceptor knockout mice, residual nerve-mediated contraction was abolished by atropine (1 μM), but not suramin (30 μM). Exogenously administered acetylcholine elicited reproducible concentration-dependent contractions of the mouse prostate that were atropine-sensitive (1 μM), but not prazosin-sensitive (0.3 μM). Acetylcholine, but not ATP, mediates the nonadrenergic component of contraction in the mouse prostate. This cholinergic component of prostatic contraction is mediated by activation of muscarinic receptors.

Isoflavones isolated from red clover (Trifolium pratense) inhibit smooth muscle contraction of the isolated rat prostate gland

This study investigated whether red clover contains any bioactive constituents which may affect contractility of rat prostatic smooth muscle in an attempt to determine whether its medicinal use in the treatment of benign prostatic hyperplasia is supported by pharmacological effects. A commercially available red clover extract was chemically fractionated and various isoflavones (genistein, formononetin and biochanin A) were isolated from these fractions and their effects on contractility were examined on preparations of the isolated rat prostate gland. Contractile effects of the isolated fractions were compared with commercially available isoflavones (genistein, formononetin and biochanin A). Pharmacological tools were used to investigate the mechanism of action modifying smooth muscle contraction. Crude red clover extract (Trinovin) inhibited electrical field stimulation induced contractions of the rat prostate across a range of frequencies with an IC(50) of approximately 68 microg/ml. Contractions of the rat prostate elicited by exogenous administration of acetylcholine, noradrenaline or adenosine 5'-triphosphate (ATP) were also inhibited. Chromatographic separation, and final purification by high performance liquid chromatography (HPLC) permitted the isolation of the isoflavones: daidzein, calycosin, formononetin, prunetin, pratensin, biochanin A and genistein. Genistein, formononetin and biochanin A (100 microM) from either commercial sources or isolated from red clover extract inhibited electrical field stimulation induced contractions of the isolated rat prostate. It is concluded that isoflavones contained in red clover are able to inhibit prostatic smooth muscle contractions in addition to their antiproliferative effects. However, the high concentrations required to observe these smooth muscle relaxant effects mean that a therapeutic benefit from this mechanism is unlikely at doses used clinically.

Expression of cAMP-dependent Protein Kinase Isoforms in the Human Prostate: Functional Significance and Relation to PDE4

To investigate the expression of isoforms of the cyclic AMP (cAMP)-dependent protein kinase (cAK) in the transition zone of the human prostate and the functional significance of the enzyme in the control of prostate smooth muscle. Using Western blot analysis and immunohistochemistry, the expression and distribution in the prostate of cAKIalpha, cAKIbeta, cAKIIalpha, and cAKIIbeta in relation to alpha-actin and the phosphodiesterase PDE4 (types A and B) were investigated. The effects of the cAK inhibitor Rp-8-CPT-cAMPS on the reversion of the adrenergic tension of isolated prostate tissue induced by forskolin, rolipram, sodium nitroprusside (SNP), and tadalafil were examined by means of the organ bath technique. Immunosignals specific for cAKIalpha, cAKIIalpha, and cAKIIbeta were observed in the smooth musculature and glandular structures of the prostate. Double stainings revealed the colocalization of alpha-actin and PDE4 with the cAK isoforms. The expression of the cAK isoforms was confirmed by Western blot analysis. The relaxation of the tension induced by norepinephrine brought about by forskolin, rolipram, SNP, and tadalafil was significantly attenuated by Rp-8-CPT-cAMPS. The colocalization of smooth muscle alpha-actin and PDE4 with cAK, as well as the results from the organ bath experiments, provide further evidence for a pivotal role of the cAMP-dependent signaling in the regulation of prostate smooth muscle contractility. Compounds interacting with the cAMP/cAK pathway might represent a new therapeutic avenue to treat symptoms of benign prostatic hyperplasia and lower urinary tract symptomatology.

Abstract 1012: A liquid miscibility model of tumor-stromal interaction in prostate cancer

Abstract Preventing the spread of prostate cancer would significantly reduce the morbidity and mortality associated with this disease. Impeding the interaction between tumor and normal cells is a possible strategy for therapeutic intervention. The prostate gland contains two distinct compartments, epithelial and stromal. In aggressive prostate cancer the boundary between these compartments disappears, allowing tumor cells to intermix with stromal cells. Intermixing facilitates access of tumor cells to blood and lymphatic vessels, facilitating invasion and subsequent metastasis. Understanding the molecular and physical forces driving intermixing is a crucial first step in preventing the spread of prostate cancer. We present a model in which prostate cancer cell-stromal cell interaction can be viewed as a liquid miscibility/immiscibility phenomenon, rooted in the same thermodynamic principles that govern the interactions between simple liquids. The premise is that whether cancer and stromal cells remain segregated or intermix depends on their relative strength of self- and cross-adhesion. We apply an analogy of the high energy interaction between ethanol and water resulting in miscibility (intermixing) of the two phases, or the lower energy of interaction between oil and water resulting in immiscibility (segregation), to test the hypothesis that the model can differentiate between such interactions in mixtures of prostate cancer and stromal cells. We show that when differentially-labeled non invasive AT-2 or invasive MLL cells are mixed with rat prostatic fibroblasts (RPF), AT-2 cells segregate from RPF cells, while MLL cells remain intermixed, suggesting that the MLL-RPF interaction is of higher energy than that of AT-2 and RPF. A switch in the composition of the prostatic stroma from smooth muscle (SMCs) to fibroblastic is a hallmark of malignant transformation in prostate cancer. We mixed MLL cells either with rat prostatic SMCs or RPF cells and showed that MLL cells segregate from SMCs but intermix with fibroblasts. This indicates that MLL cells have higher affinity for prostatic fibroblasts than for SMCs and is consistent with the observation that the stroma of more aggressive prostate cancers is predominantly fibroblastic. Invasive CaP cells lack the ability to assemble a fibronectin matrix. We have previously shown that restoring this ability gives rise to increased cohesion between cells in 3D spheroids. We treated MLL cells with MEK inhibitors (MEKi) and showed that treatment restored the ability of MLL cells to assemble fibronectin into a fibrillar matrix, leading to increased cell-cell cohesion. When mixed with RPF cells MEKi-treated MLL segregated, suggesting that altering the cohesive relationship between tumor cells is sufficient to reduce their affinity for stromal cells. Our model provides a molecular and physical basis for exploring epithelial-stromal interaction in an easily-manipulated in vitro system. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1012.

1405 EVALUATING THE SIGNIFICANCE OF THE CYCLIC AMP-MEDIATED SIGNALLING IN THE CONTROL OF SMOOTH MUSCLE OF THE HUMAN PROSTATE – A FUNCTIONAL AND BIOCHEMICAL STUDY

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Basic Research1 Apr 20101405 EVALUATING THE SIGNIFICANCE OF THE CYCLIC AMP-MEDIATED SIGNALLING IN THE CONTROL OF SMOOTH MUSCLE OF THE HUMAN PROSTATE – A FUNCTIONAL AND BIOCHEMICAL STUDY George Kedia, Dietmar Schönfeld, Michael Seidler, Markus Kuczyk, and Stefan Ückert George KediaGeorge Kedia More articles by this author , Dietmar SchönfeldDietmar Schönfeld More articles by this author , Michael SeidlerMichael Seidler More articles by this author , Markus KuczykMarkus Kuczyk More articles by this author , and Stefan ÜckertStefan Ückert More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2010.02.1097AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Basic and clinical research on the use of phosphodiesterase (PDE) inhibitors to treat lower urinary tract symptomatology (LUTS) suggestive of benign prostatic hyperplasia (BPH) has been focused mainly on the nitric oxide (NO)/cyclic GMP pathway and the cyclic GMP-specific PDE5. Up until today, only a very few studies have addressed the potential significance of the cyclic AMP-dependent signal transduction and related proteins (for example the PDE4) in the control of the function of the human prostate. This prompted us to evaluate further the significance of the cyclic AMP signalling in the mechanism of prostate smooth muscle relaxation. METHODS Human prostate tissue from the transition zone was obtained from male patients who had undergone pelvic surgery. Using the organ bath technique, the effects of the PDE4 inhibitors rolipram, Ro 20-1724 and ZK 803616 on the tension induced by norepinephrine (NE, 40 μM) of isolated prostate tissue were investigated in comparison to the PDE5 inhibitors sildenafil citrate, DASPP and MSPP, and BAY 13-1197, a NO-independent activator of the soluble guanylyl cyclase (1 nM - 10 μM each). By means of chromatographic techniques, total PDE activity was isolated from the microsomal fraction prepared from rough homogenates of specimens taken from the transition zone and subjected to biochemical analysis. RESULTS In the organ bath experiments, rolipram, Ro 20-1724 and ZK 803616 dose-dependently reversed the tension induced by NE of the isolated prostate tissue by 32%, 59%, and 53%, respectively. Thus, the PDE4 inhibitors were as effective as MSPP (54%) and more effective than BAY 13-1197 (18%), sildenafil (17%), and DASPP (14%). Only a single peak of PDE activity was isolated from the microsomal fraction. This activity was suggestive of representing PDE4 since it was sensitive to increasing concentrations (0.01 μM - 100 μM) of papaverine (non-specific PDE inhibitor), and the PDE4 inhibitors rolipram and Ro 20-1724. CONCLUSIONS Our results present evidence for a significance of the cyclic AMP signalling in controlling prostate smooth muscle function. These findings might be of importance for the identification of new drug options to treat of symptoms of the benign prostatic syndrome (BPS). Hannover, Germany© 2010 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 183Issue 4SApril 2010Page: e542 Peer Review Report Advertisement Copyright & Permissions© 2010 by American Urological Association Education and Research, Inc.MetricsAuthor Information George Kedia More articles by this author Dietmar Schönfeld More articles by this author Michael Seidler More articles by this author Markus Kuczyk More articles by this author Stefan Ückert More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

1415 RHO KINASE-SIGNALING IN THE HUMAN PROSTATE DOES NOT INVOLVE EZRIN/RADIXIN/MOESIN, ADDUCIN, OR LIM KINASE

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Basic Research1 Apr 20101415 RHO KINASE-SIGNALING IN THE HUMAN PROSTATE DOES NOT INVOLVE EZRIN/RADIXIN/MOESIN, ADDUCIN, OR LIM KINASE Frank Strittmatter, Christian Gratzke, Boris Schlenker, Philipp Weinhold, Christian G. Stief, and Martin Hennenberg Frank StrittmatterFrank Strittmatter More articles by this author , Christian GratzkeChristian Gratzke More articles by this author , Boris SchlenkerBoris Schlenker More articles by this author , Philipp WeinholdPhilipp Weinhold More articles by this author , Christian G. StiefChristian G. Stief More articles by this author , and Martin HennenbergMartin Hennenberg More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2010.02.1107AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Rho kinase is an important mediator of contraction and proliferation in prostatic smooth muscle cells. Here, we investigated, whether ezrin/radixin/moesin (“ERM proteins”), adducin or LIM kinase are substrates and suitable markers of Rho kinase in the human prostate. METHODS Human prostate tissue was obtained from 5 patients underoing radical prostatectomies. The effect of the Rho kinase inhibitor Y27632 on prostate tone was tested in myographic measurements (n=5). The effect of Y27632 on the phosphorylation state of putative Rho kinase substrates was investigated by Western blot analysis with phospho-specific antibodies after incubation (45 minutes) of prostate tissue with Y27632 (10 or 30 μM) and solvent (n=5). RESULTS In myographic measurements, the resting tone of prostate strips was reduced by Y27632 (from 0.503+/-0.004 g to 0.315+/-0.061 g, p<0.02), indicating the presence and constitutive activity of Rho kinase, the effectiveness of Y27632, and substantial regulation of prostate tone by Rho kinase. The phosphorlyation state of the ERM proteins ezrin (threonine 567), radixin (threonine 564), and moesin (threonine 558) remained unaltered by Y27632 (phospho-ERM was 157+/-32% after 10 μM Y27632 and 262+/-91% after 30 μM Y27632 compared to solvent, phospho-moesin was 100+/-22% after 10 μM Y27632 and 119+/-29% after 30 μM Y27632 compared to solvent). Likewise, Y27632 did not change the state of adducin phosphorylation at threonine 448, or of LIM kinase-1/2 phosphorylation at threonines 508/505 (phospho-adducin was 184+/-23% after 10 μM Y27632 and 181+/-31% after 30 μM Y27632 compared to solvent, phospho-LIM kinase-1/2 was 118+/-56% after 10 μM Y27632 and 116+/-28% after 30 μM Y27632 compared to solvent). CONCLUSIONS Rho kinase is an important regulator of human prostatic smooth muscle, raising the need for suitable markers of prostatic Rho kinase activity. Futher studies are required to evaluate the prostate-specific regulation of Rho kinase, which differs from Rho kinase regulation in other smooth muscle types. Munich, Germany© 2010 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 183Issue 4SApril 2010Page: e546 Peer Review Report Advertisement Copyright & Permissions© 2010 by American Urological Association Education and Research, Inc.MetricsAuthor Information Frank Strittmatter More articles by this author Christian Gratzke More articles by this author Boris Schlenker More articles by this author Philipp Weinhold More articles by this author Christian G. Stief More articles by this author Martin Hennenberg More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

The Forefront for Novel Therapeutic Agents Based on the Pathophysiology of Lower Urinary Tract Dysfunction: Pathophysiology of Voiding Dysfunction and Pharmacological Therapy

Normal lower urinary tract function consists of voiding and storage. During voiding, the pontine micturition reflex center orders the sacral parasympathetic nucleus to increase parasympathetic activity, resulting in urinary bladder detrusor contraction via activation of post-synaptic muscarinic receptors (M2/3) and in the relaxation of both urethral and prostatic smooth muscle by nitric oxide (NO). In addition, the rhabdosphincter relaxes by inhibition of the pudendal nucleus at the sacral portion. During the storage phase, increase in sympathetic activity relaxes the urinary bladder via activation of post-synaptic β3-receptors and in the contraction of both urethral and prostatic smooth muscles via α1-adrenoceptor. Many factors influence voiding function, including lower urinary tract disorders (benign prostatic hyperplasia in males, urethral stricture) and neurological disorders (central and peripheral). Theories of pharmacotherapy for voiding dysfunction are 1) increase detrusor contractility and 2) decrease urethral resistance. The former includes agonists for muscarinic receptors and cholinesterase inhibitor; and the latter includes α1-adrenoceptor antagonists, NO donors, benzodiazepines, baclofen, dantrolene, and boturinum toxin.

Isolation and pharmacological characterization of AdTx1, a natural peptide displaying specific insurmountable antagonism of the α1A‐adrenoceptor

Venoms are a rich source of ligands for ion channels, but very little is known about their capacity to modulate G-protein coupled receptor (GPCR) activity. We developed a strategy to identify novel toxins targeting GPCRs. We studied the interactions of mamba venom fractions with alpha(1)-adrenoceptors in binding experiments with (3)H-prazosin. The active peptide (AdTx1) was sequenced by Edman degradation and mass spectrometry fragmentation. Its synthetic homologue was pharmacologically characterized by binding experiments using cloned receptors and by functional experiments on rabbit isolated prostatic smooth muscle. AdTx1, a 65 amino-acid peptide stabilized by four disulphide bridges, belongs to the three-finger-fold peptide family. It has subnanomolar affinity (K(i)= 0.35 nM) and high specificity for the human alpha(1A)-adrenoceptor subtype. We showed high selectivity and affinity (K(d)= 0.6 nM) of radio-labelled AdTx1 in direct binding experiments and revealed a slow association constant (k(on)= 6 x 10(6).M(-1).min(-1)) with an unusually stable alpha(1A)-adrenoceptor/AdTx1 complex (t(1/2diss)= 3.6 h). AdTx1 displayed potent insurmountable antagonism of phenylephrine's actions in vitro (rabbit isolated prostatic muscle) at concentrations of 10 to 100 nM. AdTx1 is the most specific and selective peptide inhibitor for the alpha(1A)-adrenoceptor identified to date. It displays insurmountable antagonism, acting as a potent relaxant of smooth muscle. Its peptidic nature can be exploited to develop new tools, as a radio-labelled-AdTx1 or a fluoro-labelled-AdTx1. Identification of AdTx1 thus offers new perspectives for developing new drugs for treating benign prostatic hyperplasia.

The Forefront for Novel Therapeutic Agents Based on the Pathophysiology of Lower Urinary Tract Dysfunction: α-Blockers in the Treatment of Male Voiding Dysfunction — How Do They Work and Why Do They Differ in Tolerability?

α1-Adrenoceptor antagonists are the mainstay of medical treatment of male voiding dysfunction which typically is attributed to benign prostatic hyperplasia. While original concepts have assumed that they relieve voiding dysfunction by relaxing prostatic smooth muscle, newer data indicate that their therapeutic effects at least partly occur independent of prostatic relaxation, perhaps involving direct effects on blood vessels, urothelium, afferent nerves, and/or smooth muscle of the urinary bladder. The adverse event profiles differ among α1-adrenoceptor antagonists, with tamsulosin having a particularly good cardiovascular tolerability. While this was originally attributed to its selectivity for α1A-adrenoceptors, it appears that alfuzosin which lacks subtype-selectivity, has a very similar tolerability. In contrast, doxazosin and terazosin, which are chemically and pharmacologically more closely related to alfuzosin than to tamsulosin, appear to have more side effects attributable to the cardiovascular system. More recent data indicate that tolerability differences between α1-adrenoceptor antagonists may at least partly relate to pharmacokinetic rather than to pharmacodynamic differences. Taken together, these data emphasize the idea that concepts about drug efficacy and tolerability despite being highly plausible may not necessarily be true and always require thorough experimental testing.

Translational Pharmacology in Aging Men with Benign Prostatic Hyperplasia: Molecular and Clinical Approaches to Alpha1-Adrenoceptors

Benign prostatic hyperplasia (BPH) occurs in over 50% of men over 60 years of age, most of whom have lower urinary tract symptoms (LUTS). The incidence of BPH appears to be increasing due to the increased longevity of men. LUTS associated with BPH (LUTS/BPH) greatly affects the patient's quality of life. Physicians should be aware of other issues associated with advancing age, such as cardiovascular diseases, sexual dysfunction and cataract, that may complicate the treatment of aging men with LUTS/BPH; therefore, management of LUTS/BPH requires careful selection of the most appropriate treatment for each patient. Alpha1-adrenoceptor (alpha(1)-AR) stimulation plays an important role in the regulation of prostate smooth muscle contraction. Recent remarkable advances in molecular biology offer the possibility of new findings about the role of alpha(1)-AR subtypes and new therapeutic options for BPH; however, even though alpha(1)-AR antagonists are used as the first-line medical treatment for patients affected by LUTS/BPH, responses to alpha(1)-AR antagonists differ among patients. The risks of unexpected adverse events, acute urinary retention and the need for invasive therapy also differ among patients. Translational pharmacology is the collaboration between researchers and clinicians to discover more effective medical therapies and to identify new drugs for various diseases. This review summarizes the recent molecular and physiological findings of alpha(1)-AR subtypes and discusses potential new strategies for BPH medical treatment. In the future, the promise of genetic-based prescriptions and therapeutic plans as a useful strategy to improve clinical outcomes of BPH medical therapy may become credible and warrants further investigation.

Icaritin induces growth inhibition and apoptosis of human prostatic smooth muscle cells in an estrogen receptor-independent manner

Icaritin has selective estrogen receptor (ER) modulating activity. ERs are expressed in the prostate stroma, and estrogens have an important role in the pathology of benign prostatic hyperplasia (BPH). However, the impact of icaritin on BPH was not studied. Human prostatic smooth muscle cells (PSMCs) were treated with 0-100 microM icaritin, also using 10 microM ICI182780 as a specific ER antagonist. The effects on cell growth and apoptosis were determined by cell counting and sandwich-enzyme-immunoassay. Western blotting was employed to illustrate the possible mechanisms. Cell growth was strongly inhibited by icaritin, and this was accompanied by an augmented apoptosis. Few changes in icaritin-induced growth inhibition and apoptosis were observed after pretreatment in the presence of ICI182780. Consistent with growth inhibition and apoptosis induction, icaritin decreased cyclin D1 and CDK4 expression and increased Bax/Bcl-2 ratio in human PSMCs. Furthermore, icaritin induced sustained phosphorylation of extracellular signal-regulated kinase (ERK) in human PSMCs. PD98059, a specific ERK inhibitor, blocked the activation of ERK by icaritin and abolished the icaritin-induced growth inhibition and apoptosis. The results indicate that icaritin reduces growth and induces apoptosis in human PSMCs via ERK signaling pathway without involvement of ERs.

Extracts of bark from the traditional Chinese herb Phellodendron amurense inhibit contractility of the isolated rat prostate gland

Aim of the study To assess the effectiveness of the traditional Chinese herb Phellodendron amurense in treating urological disorders. Materials and methods Prostate smooth muscle relaxant activity of an extract from the bark of Phellodendron amurense was tested on contractions of isolated rat prostate gland induced by electrical nerve stimulation and direct muscle stimulation. Results Electrical field stimulation (0.5 ms, 60 V, 1–20 Hz) induced nerve mediated contractions of isolated rat prostate were inhibited by Phellodendron amurense extract dissolved in either dimethylsulfoxide (DMSO), acetic acid or water ( P ≤ 0.022, n = 6 for each) but not boiling water ( P = 0.619, n = 6). Phellodendron amurense extract also inhibited contractions of isolated rat prostates elicited by exogenous administration of noradrenaline (10 nM to 100 μM), acetylcholine (10 nM to 100 μM) or adenosine 5′-triphosphate (ATP, 100 nM to 100 μM) ( P ≤ 0.004, n = 6–8 for each). Conclusion Phellodendron amurense is able to inhibit prostatic contractility suggesting that it may be useful in the treatment of urological disorders caused by prostatic urethral obstruction such as in the case of benign prostatic hyperplasia (BPH).

Combination of Alfuzosin and Tadalafil Exerts an Additive Relaxant Effect on Human Detrusor and Prostatic Tissues In Vitro

BackgroundLower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) and erectile dysfunction (ED) are highly prevalent in aging men and are strongly linked. Alpha1-blockers such as alfuzosin are effective monotherapies for LUTS. Phosphodiesterase type 5 (PDE5) inhibitors such as tadalafil are the first-line treatment for ED. Both drugs act by two different mechanisms of action on common urogenital target organs and, thus, may have additive effects. ObjectivesWe evaluated in vitro the effects of alfuzosin, tadalafil, and the combination of both on human detrusor and prostatic smooth muscle. Design, setting, and participantsProstatic and bladder tissue were obtained from patients (n=20 and n=17, respectively) undergoing cystoprostatectomy for bladder cancer. MeasurementsIn organ baths, isolated prostatic strips and isolated bladder strips were incubated with vehicle, tadalafil (10−6 M and 10−5 M), alfuzosin (3×10−8 M or 10−6 M and 10−5 M) or a combination. Concentration-response curves (CRCs) to norepinephrine were generated on prostatic strips and detrusor strips precontracted with carbachol. Strips were also submitted to electrical field stimulation (EFS). Results and limitationsWhen alfuzosin and tadalafil were combined, the maximal relaxation to norepinephrine on carbachol-precontracted detrusor strips was significantly increased compared with tadalafil alone, and EFS-induced detrusor contractions were better inhibited compared with each compound alone. Tadalafil significantly inhibited norepinephrine-induced prostatic strip contractions by reducing the maximal effect, whereas alfuzosin shifted the CRC of norepinephrine to the right. Combining both tadalafil and alfuzosin resulted in a greater relaxant effect. Likewise, the combination was more effective at reducing EFS-induced contractions compared with each compound alone. ConclusionsThe combination of alfuzosin and tadalafil exerts an additive effect of inhibiting adrenergic smooth muscle tone of prostatic tissue and EFS-induced detrusor contractions and conversely, of enhancing adrenergic relaxation of detrusor precontracted with carbachol. These experiments provide experimental support for the clinical investigation of the combination of α1-blockers and PDE5 inhibitors in the treatment of LUTS.

Inositol trisphosphate‐dependent Ca2+ stores and mitochondria modulate slow wave activity arising from the smooth muscle cells of the guinea pig prostate gland

Changes in smooth muscle tone of the prostate gland are involved in aetiology of symptomatic prostatic hyperplasia, however the control mechanisms of prostatic smooth muscle are not well understood. Here, we have examined the role of internal Ca(2+) compartments in regulating slow wave activity in the guinea pig prostate. Standard intracellular membrane potential recording techniques were used. The majority (89%) of impaled cells displayed 'slow wave' activity. Cyclopiazonic acid (10 micromol.L(-1)) transiently depolarized (3-9 mV) the membrane potential of the prostatic stroma and transiently increased slow wave frequency. Thereafter, slow wave frequency slowly decreased over 20-30 min. Ryanodine transiently increased slow wave frequency, although after 30 min exposure slow wave frequency and time course returned to near control values. Caffeine (1 mmol.L(-1)) reduced slow wave frequency, accompanied by membrane depolarization of about 8 mV. Blockade of inositol trisphosphate receptor (IP(3)R)-mediated Ca(2+) release with 2-aminoethoxy-diphenylborate (60 micromol.L(-1)) or Xestospongin C (3 micromol.L(-1)) or inhibiting phospholipase C and IP(3) formation using U73122 (5 micromol.L(-1)) or neomycin (1 and 4 mmol.L(-1)) reduced slow wave frequency, amplitude and duration. The mitochondrial uncouplers, p-trifluoromethoxy carbonyl cyanide phenyl hydrazone (1-10 micromol.L(-1)), carbonyl cyanide m-chlorophenylhydrazone (1-3 micromol.L(-1)) or rotenone (10 micromol.L(-1)), depolarized the membrane (8-10 mV) before abolishing electrical activity. These results suggest that slow wave activity was dependent on the cyclical release of Ca(2+) from IP(3)-controlled internal stores and mitochondria. This implies that intracellular compartments were essential in the initiation and/or maintenance of the regenerative contractile activity in the guinea pig prostate gland.

Effects of Phosphodiesterase Inhibitors on Contraction Induced by Endothelin-1 of Isolated Human Prostatic Tissue

To study the effects of selective phosphodiesterase (PDE) inhibitors on the contraction induced by endothelin-1 (ET-1) of isolated human prostatic tissue. Using the organ bath technique, the effects of the cumulative addition of the PDE1 inhibitor vinpocetine, PDE2 inhibitor erythro-9-(2-hydroxy-3-nonyl)adenine hydrochloride (EHNA), PDE4 inhibitor rolipram, and PDE5 inhibitors sildenafil, vardenafil, and tadalafil (1 nM to 10 microM) were investigated on the contraction brought about by the peptide ET-1 (1 microM) on normal human prostatic tissue isolated from the transition zone. In the present study, the nitric oxide donor drug sodium nitroprusside and adenylyl cyclase activator forskolin were used as reference compounds. ET-1 induced stable and long-lasting contraction of the isolated prostatic tissue. The tension induced by ET-1 was dose-dependently reversed by the drugs with the following rank order of efficacy: rolipram, forskolin, tadalafil, sodium nitroprusside, sildenafil, vinpocetine, vardenafil, EHNA. The maximal reversion of tension ranged from 67% (rolipram) to 20% (EHNA). Our results have provide additional evidence that the function of prostatic smooth muscle is under the control of peptidergic and cyclic nucleotide (cyclic guanosine monophosphate, cyclic adenosine monophosphate)-mediated pathways. This might give a rationale for the use of drugs interacting with cyclic guanosine monophosphate or cyclic adenosine monophosphate signaling, such as PDE inhibitors or nitric oxide donors, in the pharmacotherapy of the benign prostatic syndrome.

１．排尿障害の病態と新しい治療

Normal lower urinary tract function consists of voiding and storage. During voiding, the pontine micturition reflex center orders the sacral parasympathetic nucleus to increase parasympathetic activity, resulting in urinary bladder detrusor contraction via activation of post-synaptic muscarinic receptors (M2/3), and the relaxation of both urethral and prostatic smooth muscle by NO. In addition, the rhabdosphincter relaxes by inhibition of pudendal nucleus at the sacral portion. During the storage phase, an increase in sympathetic activity relaxes urinary bladder via activation of post-synaptic β3 receptors, and both the urethral and prostatic smooth muscles are contracted via α1-adrenoceptor. Many factors influence voiding function, including lower urinary tract disorders (benign prostatic hyperplasia in males, urethral stricture), and neurological disorders (central and peripheral). Theories of action of pharmacotherapy for voiding dysfunction are: 1) increase detrusor contractility, and 2) decrease urethral resistance. The former includes agonists for muscarinic receptors, and choline-esterase inhibitor, and the latter includes α1-adrenoceptor antagonists, nitric oxide donors, Benzodiazepines, Baclofen, Dantrolene, and Boturinum toxin.

Subtypes of α1-adrenoceptors in BPH: future prospects for personalized medicine

The alpha(1)-adrenoceptors (alpha(1)-ARs) are involved in regulation of prostatic smooth muscle tone, and are a critical mediator of lower urinary tract symptoms and pathophysiology in benign prostatic hyperplasia (BPH). As a result, alpha(1)-AR antagonists are now used as first-line medical treatment for BPH. Three alpha(1)-AR subtypes (alpha(1a)-AR, alpha(1b)-AR, alpha(1d)-AR) have been identified on the basis of results of pharmacological and molecular cloning studies; however, the precise physiological role of individual alpha(1)-AR subtypes remains elusive. The expression levels of alpha(1)-AR subtypes in the prostate differ between patients, and individual differences in the genetic background of patients with BPH might be associated with variation in responses to subtype-selective alpha(1)-AR antagonists. In addition, single nucleotide polymorphism and microarray-based gene expression profiling studies might provide an opportunity to identify markers that predict clinical response and therapeutic tolerance to alpha(1)-AR antagonists. Further genomic studies will refine our knowledge of the functions of alpha(1)-AR subtypes, lead to new strategies for the clinical management of BPH and, perhaps, enable personalized treatment of BPH in the future.

Effects of 4-hydroxytamoxifen on prostate smooth muscle cells: an in vitro experiment

To investigate the effects of 4- hydroxytamoxifen (OHT) on the proliferation and apoptosis of prostate smooth muscle cells and the expression of estrogen receptor (ER) and androgen receptor (AR). Prostate smooth muscle cells were isolated from the resected specimens of prostate glands of 10 patients with benign prostatic hypertrophy (BPH), cultured, and exposed to estradiol (E(2)), diethylstilbestrol (DES), and OHT of different concentrations (1 x 10(-8) - 1 x 10(-5) mol/L) or mixture of E(2) (1 x 10(-8) - 1 x 10(-6) mol/L) with OHT (1 x 10(-7) mol/L). Flow cytometry was used to test the proliferation and apoptosis of the cells, and immunocytochemistry was used to test the expression of estrogen and androgen receptors. E(2) and DES promoted the proliferation of the prostate smooth muscle cells in a certain concentration range, but not dose-dependently, and OHT at the concentration of 1 x 10(-8) mol/L slightly increased the G(2)-M peak rate of the prostate smooth muscle cells, but suppressed the G(2)-M peak rate dose-dependently when its concentration was >or= 1 x 10(-7) mol/L (P < 0.05) and this suppression effect was dose-dependently (r = -0.312, P = 0.011). E(2) at the concentration >or= 1 x 10(-5) mol/L and DES at the concentration >or= 1 x 10(-6) mol/L slightly promoted the apoptosis of the prostate smooth muscle cells, but not dose-dependently, and OHT at the concentrations from 1 x 10(-8) mol/L to 1 x 10(-5) mol/L promoted the apoptosis of the prostate smooth muscle cells dose-dependently (r = 0.363, P = 0.021) and this effect could not be reversed by administration of E(2) at the concentration 1 x 10(-8) - 1 x 10(-6) mol/L (P > 0.05). E(2), DES, and OHT of different concentrations all increased the ERalpha and AR positive staining rates of the prostate smooth muscle cells (all P < 0.05). OHT suppresses the proliferation and promotes the apoptosis of prostate smooth muscle cells, and these functions do not depend on the estrogen receptor pathway. Low blood OHT concentration after oral administration of TAM and up-regulation of estrogen receptors by OHT may be the caused of the inefficiency of TAM for treatment of BPH.

Haploinsufficiency of the Maspin tumor suppressor gene leads to hyperplastic lesions in the prostate: Shao LJ, Shi HY, Ayala G, Rowley D, Zhang M, Departments of Molecular and Cellular Biology and Pathology, Baylor College of Medicine, Houston, TX; Department of Molecular Pharmacology and Biological Chemistry, Robert H. Lurie Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, IL

Maspin is a key tumor suppressor gene in prostate and breast cancers with diverse biological functions. However, how maspin regulates prostate tumor progression is not fully understood. In this study, we have used maspin heterozygous knockout mice to determine the effect of maspin haploinsufficiency on prostate development and tumor progression. We report that loss of one copy of maspin gene in Mp(+/–) heterozygous knockout mice leads to the development of prostate hyperplastic lesions, and this effect was mediated through decreased level of cyclin-dependent kinase inhibitors p21 and p27. Prostate hyperplastic lesions in Mp(+/–) mice also induced stromal reaction, which occurred in both aged prostate tissues and in neonatal prostates during early ductal morphogenesis. We showed that maspin was also expressed in prostate smooth muscle cells (PSMC), and recombinant maspin increased PSMC cell adhesion but inhibited cell proliferation. We also observed a defective interaction between epithelial cells and basement membrane in the prostate of Mp(+/–) mice, which was accompanied with a changed pattern of matrix deposition and a loss of epithelial cell polarity. Therefore, we have identified a novel property of maspin, which involves the control of the proliferation in prostate epithelial and smooth muscle cells. This is the first report that a partial loss of maspin caused an early developmental defect of the prostate and prostate hyperplastic lesions in mouse.