Prostatic Smooth Muscle Research Articles

Implication of Rho-kinase and soluble guanylyl cyclase enzymes in prostate smooth muscle dysfunction in middle-aged rats.

Aging is highly associated with benign prostate hyperplasia (BPH). We investigated here the alterations of the contractile and relaxant machinery in prostates of middle-aged rats, focusing on the Rho-kinase, nitric oxide (NO)-soluble guanylyl cyclase (sGC), α1- and β-adrenoceptor pathways. Male Wistar young (3.5-month old) and middle-aged rats (10-month old) were used. Quantitative image analysis of prostates and functional assays evaluating the prostate contractions and relaxations were employed. Measurement of [3 H]-noradrenaline efflux, western blotting for α1 and β1 sGC subunits, and cyclic nucleotide levels were carried out. Prostates of middle-aged rats showed significant increases in lumen and smooth muscle cells, but no alterations in the relative prostate weight were observed. In vivo, noradrenaline (10-7 -10-4 g/kg) produced greater prostatic contractions in middle-aged compared with control rats. Likewise, the in vitro contractions to phenylephrine (1 nM-100 μM) and α,β-methylene ATP (1-10 μM) were greater in middle-aged rats. Electrical-field stimulation (EFS, 1-32 Hz) promoted higher [3 H]-noradrenaline efflux and prostate contractions in middle-aged rats. Reduced expressions of α1 and β1 sGC subunits and diminished NO-mediated prostate relaxations in middle-age were observed. Isoproterenol-induced relaxations and cAMP levels were reduced in prostates of middle-aged rats. The Rho-kinase inhibitor fasudil (50 mg/kg, 2 weeks) normalized the prostate hypercontractility in middle-age rats. Prostate hypercontractility in middle-aging is associated with increased release of noradrenaline and Rho-kinase pathway, as well as with impairments of NO-sGC and β-adrenoceptor pathways. Middle-aged rats are suitable to explore the enhanced prostatic tone in the absence of prostate overgrowth. Neurourol. Urodynam. 36:589-596, 2017. © 2016 Wiley Periodicals, Inc.

Electrical properties of purinergic transmission in smooth muscle of the guinea-pig prostate.

Prostatic smooth muscle develops spontaneous myogenic tone which is modulated by autonomic neuromuscular transmission. This study aimed to investigate the role of purinergic transmission in regulating electrical activity of prostate smooth muscle and whether its contribution may be altered with age. Intracellular recordings were simultaneously made with isometric tension recordings in smooth muscle preparations of the guinea-pig prostate. Immunostaining for P2X1 receptors on whole mount preparations was also performed. In prostate preparations which generated spontaneous slow waves, electrical field stimulation (EFS)-evoked excitatory junction potentials (EJPs) which were abolished by guanethidine (10 μM), α-β-methylene ATP (10 μM) or pyridoxal phosphate-6-azophenyl-2,4-disulfonic acid (PPADS, 10 μM) but not phentolamine (1 μM). Consistently, immunostaining revealed the expression of P2X1 receptors on prostatic smooth muscle. EJPs themselves did not cause contractions, but EJPs could sum to trigger a slow wave and associated contraction. Yohimbine (1 μM) and 3,7-dimethyl-1-propargylxanthine (DMPX, 10 μM) but not propranolol (1 μM) potentiated EJPs. Although properties of EJPs were not different between young and aging guinea-pig prostates, ectoATPase inhibitor ARL 67156 (100 μM) augmented EJP amplitudes by 64.2 ± 29.6% in aging animals, compared to 22.1 ± 19.9% in young animals. These results suggest that ATP released from sympathetic nerves acts on P2X1 purinoceptors located on prostate smooth muscle to evoke EJPs, while pre-junctional α2-adrenergic and adenosine A2 receptors may play a role in preventing excessive transmitter release. Age-related up-regulation of enzymatic ATP breakdown may be a compensatory mechanism for the enhanced purinergic transmission which would cause hypercontractility arising from increased ATP release in older animals.

Prostatic Artery Embolization (PAE) for Symptomatic Benign Prostatic Hyperplasia (BPH): Part 2, Insights into the Technical Rationale.

Rationale of prostatic artery embolization (PAE) in the treatment of symptomatic benign prostatic hyperplasia is conventionally believed to include two parts: shrinkage of the enlarged prostate gland as a result of PAE-induced ischemic infarction and potential effects to relax the increased prostatic smooth muscle tone by reducing the number and density of α1-adrenergic receptor in the prostate stroma. This review describes new insights into the likely mechanisms behind PAE, such as ischemia-induced apoptosis, apoptosis enhanced by blockage of androgens circulation to the embolized prostate, secondary denervation following PAE, and potential effect of nitric oxide pathway immediately after embolization. Studies on therapeutic mechanisms in PAE may shed light on potentially new treatment strategies and development of novel techniques.

The expression of β3-adrenoceptors and their function in the human prostate.

Little is known about β3-adrenoceptor (AR) expression and function in human prostate. We examined the expression and distribution of β-AR subtypes in normal prostate and benign prostatic hyperplasia (BPH) tissues, and investigated which selective β-AR subtype agonist was most involved in the relaxation of isolated human prostate strips. Messenger RNA (mRNA) expression for β1-, β2-, and β3 -ARs was investigated using reverse transcriptase-polymerase chain reactions (RT-PCR). Quantitative analysis of mRNA expression of β-AR subtypes between normal prostate and BPH tissues was performed using quantitative RT-PCR (qPCR). Distributions were examined by immunohistochemistry (IHC). Strips of human normal prostate or BPH were suspended in organ baths and exposed to isoproterenol, dobutamine, procaterol, and TRK-380 to investigate their relaxant effects on KCl-induced contractions, and their inhibitory effects on electrical field stimulation (EFS)-induced contractions. We confirmed the presence of mRNA for β1-, β2-, and β3-ARs both in normal prostate and in BPH tissues. For β3-AR, mRNA expression in BPH tissues was significantly higher than in normal prostate tissues, but there was no significant difference in β1- and β2-AR expression between normal and BPH tissues. IHC revealed differences in staining intensity between smooth muscle cells and glandular cells, with different proportions for different β-AR subtypes. Staining of β3-AR was particularly intense in smooth muscle cells as opposed to glandular cells. Isoproterenol and TRK-380 significantly decreased the tone of KCl-induced contractions of the normal prostate strips. The rank order of relaxant effects was isoproterenol > TRK-380 > procaterol > dobutamine. All selective β-AR agonists significantly decreased the amplitude of EFS-induced contractions of the normal prostate strips. The rank order of inhibitory effects was isoproterenol > dobutamine >TRK-380 > procaterol. In BPH strips, all selective β-AR agonists showed no significant relaxant or inhibitory effects on KCl- or EFS-induced contractions. β3 -AR is abundant in human prostate smooth muscle, whose relaxation is mediated by β1- and β3-AR stimulation. β3-AR agonists may have clinical use in the treatment of male non-BPH patients or neurogenic bladder patients with voiding dysfunction.

Inhibition of prostate smooth muscle contraction and prostate stromal cell growth by the inhibitors of Rac, NSC23766 and EHT1864.

Medical therapy of lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) targets smooth muscle contraction in the prostate, or prostate growth. However, current therapeutic options are insufficient. Here, we investigated the role of Rac in the control of smooth muscle tone in human prostates and growth of prostate stromal cells. Experiments were performed using human prostate tissues from radical prostatectomy and cultured stromal cells (WPMY-1). Expression of Rac was examined by Western blot and fluorescence staining. Effects of Rac inhibitors (NSC23766 and EHT1864) on contractility were assessed in the organ bath. The effects of Rac inhibitors were assessed by pull-down, cytotoxicity using a cell counting kit, cytoskeletal organization by phalloidin staining and cell growth using an 5-ethynyl-2'-deoxyuridine assay. Expression of Rac1-3 was observed in prostate samples from each patient. Immunoreactivity for Rac1-3 was observed in the stroma, where it colocalized with the smooth muscle marker, calponin. NSC23766 and EHT1864 significantly reduced contractions of prostate strips induced by noradrenaline, phenylephrine or electrical field stimulation. NSC23766 and EHT1864 inhibited Rac activity in WPMY-1 cells. Survival of WPMY-1 cells ranged between 64 and 81% after incubation with NSC23766 (50 or 100 μM) or EHT1864 (25 μM) for 24 h. NSC23766 and EHT1864 induced cytoskeletal disorganization in WPMY-1 cells. Both inhibitors impaired the growth of WPMY-1 cells. Rac may be a link connecting the control of prostate smooth muscle tone with proliferation of smooth muscle cells. Improvements in LUTS suggestive of BPH by Rac inhibitors appears possible.

MP21-01 HYDROGEN SULFIDE IS A POTENT RELAXING FACTOR IN THE RAT BLADDER AND PROSTATE

You have accessJournal of UrologyBladder and Urethra: Anatomy, Physiology and Pharmacology II1 Apr 2015MP21-01 HYDROGEN SULFIDE IS A POTENT RELAXING FACTOR IN THE RAT BLADDER AND PROSTATE Takahiro Shimizu, Shogo Shimizu, Kumiko Nakamura, Masashi Honda, Keiji Inoue, and Motoaki Saito Takahiro ShimizuTakahiro Shimizu More articles by this author , Shogo ShimizuShogo Shimizu More articles by this author , Kumiko NakamuraKumiko Nakamura More articles by this author , Masashi HondaMasashi Honda More articles by this author , Keiji InoueKeiji Inoue More articles by this author , and Motoaki SaitoMotoaki Saito More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.953AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Hydrogen sulfide (H2S) is a third endogenous gasotransmitter besides NO and CO, and induces smooth muscle relaxation. In this study, we investigated pharmacological profiles of exogenous H2S-induced relaxation of bladder and prostate smooth muscle and expression levels of H2S synthesis enzymes, cystathionine β-synthase (CBS) and 3-mercaptopyruvate sulftransferase (MPST), in these tissues of the rat. METHODS Bladder dome and trigone (BL-D and BL-T), and ventral and dorsolateral prostate (PR-V and PR-D) were prepared from male Wistar rats (350–400 g). Relaxation effects of NaHS, an H2S donor, were evaluated by organ bath studies on acetylcholine (1x10−5 M)-induced contractions of bladder and on noradrenaline (1x10−5 M)- induced ones of prostate. Protein levels of CBS and MPST in each part were also investigated by Western blotting. RESULTS NaHS dose-dependently elicited relaxation on BL-D and BL-T pre-contracted by acetylcholine and on PR-V and PR-D pre-contracted by noradrenaline. There were no significant differences in values of EC50 and relaxation rate against the pre-contractions among these tissues. CBS was detected in PR-V and PR-D, and the expression levels in PR-D were higher than those in PR-V. In BL-D and BL-T, CBS was not detected. MPST was detected in all these tissues. Expression levels of MPST in PR-V were the highest and those in prostate were higher than those in bladder. CONCLUSIONS H2S induces relaxation of bladder and prostate smooth muscle. H2S synthesis enzymes are expressed in both bladder and prostate. These results suggest that H2S could function as an endogenous relaxation factor in both tissues. Endogenous H2S might be a new therapeutic target for lower urinary tract dysfunction such as overactive bladder and benign prostatic hyperplasia. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e232 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Takahiro Shimizu More articles by this author Shogo Shimizu More articles by this author Kumiko Nakamura More articles by this author Masashi Honda More articles by this author Keiji Inoue More articles by this author Motoaki Saito More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

MP31-12 TAMSULOSIN IMPROVES MEMORY FUNCTION BY ACTIVATING NMDA RECEPTORS IN RAT HIPPOCAMPUS

INTRODUCTION AND OBJECTIVES: Prostate smooth muscle contraction may be critical for pathophysiology and therapy of lower urinary tract symptoms (LUTS) suggestive of benign prostate hyperplasia. Although application of a1-blockers represents the goldstandard for medical treatment, their effects are limited to max. 50%. The reasons are unknown, while only little is known about contributions of nonadrenergic mediators to prostate contraction. In vivo, different mediators probably do not occur separately, but rather act simultaneously on smooth muscle cells. Here, we compared different contractile mediators with norepinephrine, and adressed their cooperative effects in contraction of human prostate smooth muscle. METHODS: Prostate tissues were obtained from radical prostatectomy (n1⁄472 patients). Contractility of prostate strips was assessed in an organ bath. Receptor expression was detected by Western blot analysis and fluorescence staining. RESULTS: Maximum endothelin-1 (ET-1)(0.1-3 mM), endothelin-2 (ET-2)(0.1-3 mM), and norepinephrine (NE)(0.1-100 mM) induced contractions were of similar magnitude (ET-1 116 23% of KCl-induced contraction; ET-2 86 17%; NE 117 18%). Maximum contractions by the thromboxane analogue, U46619 (10 mM) amounted on average around 50% of NE-induced contraction (63 9% of KCl). ET-1or ET-2-induced contractions were not affected by the a1-blocker tamsulosin (300 nM), while NE-induced contraction was virtually completely abolished and U46619-induced contraction was slightly inhibited. Dopamine (0.1-300 mM) and serotonin (0.1-300 mM) induced only very slight contractions (22 4% and 9 2% of KCl) of prostate strips, which may be of minor or local relevance. Carbachol and uridine adenosine tetraphosphate did not induce contractions. After maximumNE-induced precontraction, ET-1 did not further increase tension. In contrast, contractile effects of ET-1 (3 mM) and U46619 (10 mM) were additive, so that tension after combined application of ET-1 and U46619 (172 25% of KCl) was higher than maximumNE(100 mM) induced tension (p<0.05). Endothelin receptor A and B were detectable by Western blot in all samples. Double fluorescence staining suggests expression in stromal smooth muscle cells. CONCLUSIONS: NE-induced contraction of human prostate smooth muscle can be exceeded by cooperative actions of endothelins and thromboxane. These contractions may even occur under treatment with a1-blockers. Such contributions of non-adrenergic mediators might explain the limited efficacy of a1-blockers.

MP31-10 ADDITIVE CONTRACTIONS BY THROMBOXANE A2 AND ENDOTHELIN-1 ARE STRONGER THAN NOREPINEPHRINE-INDUCED CONTRACTIONS IN HUMAN PROSTATE SMOOTH MUSCLE

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Basic Research1 Apr 2015MP31-10 ADDITIVE CONTRACTIONS BY THROMBOXANE A2 AND ENDOTHELIN-1 ARE STRONGER THAN NOREPINEPHRINE-INDUCED CONTRACTIONS IN HUMAN PROSTATE SMOOTH MUSCLE Martin Hennenberg, Alice C. Acevedo, Alexander Tamalunas, Yiming Wang, Beata Rutz, Frank Strittmatter, Raphaela Waidelich, Christian G. Stief, and Christian Gratzke Martin HennenbergMartin Hennenberg More articles by this author , Alice C. AcevedoAlice C. Acevedo More articles by this author , Alexander TamalunasAlexander Tamalunas More articles by this author , Yiming WangYiming Wang More articles by this author , Beata RutzBeata Rutz More articles by this author , Frank StrittmatterFrank Strittmatter More articles by this author , Raphaela WaidelichRaphaela Waidelich More articles by this author , Christian G. StiefChristian G. Stief More articles by this author , and Christian GratzkeChristian Gratzke More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.1365AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Prostate smooth muscle contraction may be critical for pathophysiology and therapy of lower urinary tract symptoms (LUTS) suggestive of benign prostate hyperplasia. Although application of α1-blockers represents the goldstandard for medical treatment, their effects are limited to max. 50%. The reasons are unknown, while only little is known about contributions of non-adrenergic mediators to prostate contraction. In vivo, different mediators probably do not occur separately, but rather act simultaneously on smooth muscle cells. Here, we compared different contractile mediators with norepinephrine, and adressed their cooperative effects in contraction of human prostate smooth muscle. METHODS Prostate tissues were obtained from radical prostatectomy (n=72 patients). Contractility of prostate strips was assessed in an organ bath. Receptor expression was detected by Western blot analysis and fluorescence staining. RESULTS Maximum endothelin-1 (ET-1)- (0.1-3 μM), endothelin-2 (ET-2)- (0.1-3 μM), and norepinephrine (NE)- (0.1-100 μM) induced contractions were of similar magnitude (ET-1 116±23% of KCl-induced contraction; ET-2 86±17%; NE 117±18%). Maximum contractions by the thromboxane analogue, U46619 (10 μM) amounted on average around 50% of NE-induced contraction (63±9% of KCl). ET-1- or ET-2-induced contractions were not affected by the α1-blocker tamsulosin (300 nM), while NE-induced contraction was virtually completely abolished and U46619-induced contraction was slightly inhibited. Dopamine (0.1-300 μM) and serotonin (0.1-300 μM) induced only very slight contractions (22±4% and 9±2% of KCl) of prostate strips, which may be of minor or local relevance. Carbachol and uridine adenosine tetraphosphate did not induce contractions. After maximum NE-induced precontraction, ET-1 did not further increase tension. In contrast, contractile effects of ET-1 (3 μM) and U46619 (10 μM) were additive, so that tension after combined application of ET-1 and U46619 (172±25% of KCl) was higher than maximum NE- (100 μM) induced tension (p<0.05). Endothelin receptor A and B were detectable by Western blot in all samples. Double fluorescence staining suggests expression in stromal smooth muscle cells. CONCLUSIONS NE-induced contraction of human prostate smooth muscle can be exceeded by cooperative actions of endothelins and thromboxane. These contractions may even occur under treatment with α1-blockers. Such contributions of non-adrenergic mediators might explain the limited efficacy of α1-blockers. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e359 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Martin Hennenberg More articles by this author Alice C. Acevedo More articles by this author Alexander Tamalunas More articles by this author Yiming Wang More articles by this author Beata Rutz More articles by this author Frank Strittmatter More articles by this author Raphaela Waidelich More articles by this author Christian G. Stief More articles by this author Christian Gratzke More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

MP61-18 ELK-1 PROMOTES PROSTATE CANCER PROGRESSION AND SILODOSIN THAT INACTIVATES ELK-1 IN PROSTATE CANCER CELLS INCREASES SENSITIVITY TO GEMCITABINE

INTRODUCTION AND OBJECTIVES: LK-1 is a transcriptional factor whose phosphorylation is necessary for c-fos proto-oncogene activation. However, little is known about biological significance of ELK1 in prostate cancer. In addition, silodosin, a selective a1-adrenergic blocker used for the symptomatic treatment of benign prostatic hyperplasia, has been shown to decrease ELK-1 expression in human prostate smooth muscle cells. In this study, we aim to investigate the functions of ELK-1 in prostate cancer growth and the efficacy of silodosin, in combination with chemotherapeutic drugs, in cell proliferation. METHODS: We first immunohistochemically determined the levels of phospho-ELK-1 (pELK-1) expression in radical prostatectomy specimens. We then assessed the effects of ELK-1 knockdown via short hairpin RNA on cell proliferation, migration, and invasion as well as those of silodosin on ELK-1 expression/activity in prostate cancer lines and their viability. RESULTS: Immunohistochemistry showed that the levels of pELK-1 expression were significantly higher in carcinoma (positivity & score, P<0.001) or high-grade prostatic intraepithelial neoplasia (HGPIN) (positivity & score, P<0.001) than in benign as well as in carcinoma than in HGPIN (positivity, P1⁄40.330; score, P1⁄40.002). Although none of the 7 patients with pELK-1-negative carcinoma had biochemical recurrence after radical prostatectomy, Kaplan-Meier and log-rank tests revealed no significant correlations of pELK-1 levels with patient outcomes. In PC3 and DU145 cells expressing ELK-1 (mRNA, protein), ELK-1 silencing resulted in decreases in the expression of cfos, a downstream target of ELK-1 signals, as well as in cell viability/ colony formation, cell migration/invasion, and the expression of matrix metalloproteinase (MMP)-2 and MMP-9. In these cells, silodosin reduced the expression of a1-adrenergic receptor, ELK-1, and c-fos as well as the transcriptional activity of ELK-1. However, silodosin alone (up to 10 mM) only marginally inhibited PC3/DU145 cell growth. Interestingly, silodosin treatment in the prostate cancer lines increased sensitivity to gemcitabine, but not to cisplatin or docetaxel. CONCLUSIONS: ELK-1 is likely activated in prostate cancer cells. Our results also suggest that ELK-1 plays an important role in prostate cancer progression. Thus, ELK-1 inhibition has the potential of being a therapeutic approach for prostate cancer. Furthermore, silodosin that inactivates ELK-1 in prostate cancer cells enhances the cytotoxic activity of gemcitabine.

MP61-19 A NOVEL STRATEGY TO OPTIMISE TREATMENT OF DOCETAXEL-RESISTANT PROSTATE CANCER

INTRODUCTION AND OBJECTIVES: LK-1 is a transcriptional factor whose phosphorylation is necessary for c-fos proto-oncogene activation. However, little is known about biological significance of ELK1 in prostate cancer. In addition, silodosin, a selective a1-adrenergic blocker used for the symptomatic treatment of benign prostatic hyperplasia, has been shown to decrease ELK-1 expression in human prostate smooth muscle cells. In this study, we aim to investigate the functions of ELK-1 in prostate cancer growth and the efficacy of silodosin, in combination with chemotherapeutic drugs, in cell proliferation. METHODS: We first immunohistochemically determined the levels of phospho-ELK-1 (pELK-1) expression in radical prostatectomy specimens. We then assessed the effects of ELK-1 knockdown via short hairpin RNA on cell proliferation, migration, and invasion as well as those of silodosin on ELK-1 expression/activity in prostate cancer lines and their viability. RESULTS: Immunohistochemistry showed that the levels of pELK-1 expression were significantly higher in carcinoma (positivity & score, P<0.001) or high-grade prostatic intraepithelial neoplasia (HGPIN) (positivity & score, P<0.001) than in benign as well as in carcinoma than in HGPIN (positivity, P1⁄40.330; score, P1⁄40.002). Although none of the 7 patients with pELK-1-negative carcinoma had biochemical recurrence after radical prostatectomy, Kaplan-Meier and log-rank tests revealed no significant correlations of pELK-1 levels with patient outcomes. In PC3 and DU145 cells expressing ELK-1 (mRNA, protein), ELK-1 silencing resulted in decreases in the expression of cfos, a downstream target of ELK-1 signals, as well as in cell viability/ colony formation, cell migration/invasion, and the expression of matrix metalloproteinase (MMP)-2 and MMP-9. In these cells, silodosin reduced the expression of a1-adrenergic receptor, ELK-1, and c-fos as well as the transcriptional activity of ELK-1. However, silodosin alone (up to 10 mM) only marginally inhibited PC3/DU145 cell growth. Interestingly, silodosin treatment in the prostate cancer lines increased sensitivity to gemcitabine, but not to cisplatin or docetaxel. CONCLUSIONS: ELK-1 is likely activated in prostate cancer cells. Our results also suggest that ELK-1 plays an important role in prostate cancer progression. Thus, ELK-1 inhibition has the potential of being a therapeutic approach for prostate cancer. Furthermore, silodosin that inactivates ELK-1 in prostate cancer cells enhances the cytotoxic activity of gemcitabine.

Increased Rho-kinase-mediated prostate contractions associated with impairment of β-adrenergic-cAMP-signaling pathway by chronic nitric oxide deficiency

Impairment of nitric oxide (NO) – cyclic GMP signaling pathway is likely to contribute to human begnin prostate hyperplasia (BPH). In the present study we have used a model of chronic NO synthesis inhibition to evaluate the functional alterations of prostate smooth muscle (PSM) machinery, and involvement of Rho-kinase pathway. Wistar rats were treated with the NO inhibitor Nω-nitro-l-arginine methyl ester (L-NAME, 20mg/kg/day; 4 weeks), after which contractile responses to phenylephrine (α1-adrenoceptor agonist; 1nM to 100µM), carbachol (muscarinic agonist; 1nM to 1mM) and α,β-methylene ATP (P2X receptor agonist; 1–10µM), as well as to electrical-field stimulation (EFS; 1–32Hz) were evaluated. PSM relaxations to isoproterenol (non-selective β-adrenoceptor agonist, 0.1nM to 10µM) and sodium nitroprusside (NO donor, 1nM to 10mM) were also evaluated. The ratio prostate weight/body weight was 22% greater (P<0.05) in L-NAME compared with control group. The PSM contractions to phenylephrine, carbachol and α,β-methylene ATP were higher in L-NAME (Emax: 3.85±0.25, 3.52±0.35 and 2.03±0.2mN, respectively) compared with control group (Emax: 3.08±0.17, 2.37±0.18 and 1.57±0.18mN, respectively). The PSM contractions induced by EFS were also significantly greater in L-NAME group. Prior incubation with the Rho-kinase inhibitor Y27632 (1µM) fully reversed the enhanced contractions to phenylephrine and carbachol. Isoproterenol-induced PSM relaxations were 34% lower in L-NAME group, which was associated with reduced levels of cAMP in prostate tissue. The relaxations to sodium nitroprusside remained unaltered in L-NAME group. In summary, chronic NO deficiency leads to increased Rho-kinase-mediated PSM contractile responses accompanied by impairment of β-adrenergic-cAMP-signaling pathway.

897 Translocation of nuclear factor-κB and expression of cyclooxygenase-2 is enhanced by ketamine-induced ulcerative cystitis in rat bladder

INTRODUCTION AND OBJECTIVES: Hydrogen sulfide (H2S) is a third endogenous gasotransmitter besides NO and CO, and induces smooth muscle relaxation. In this study, we investigated pharmacological profiles of exogenous H2S-induced relaxation of bladder and prostate smooth muscle and expression levels of H2S synthesis enzymes, cystathionine b-synthase (CBS) and 3-mercaptopyruvate sulftransferase (MPST), in these tissues of the rat. METHODS: Bladder dome and trigone (BL-D and BL-T), and ventral and dorsolateral prostate (PR-V and PR-D) were prepared from male Wistar rats (350e400 g). Relaxation effects of NaHS, an H2S donor, were evaluated by organ bath studies on acetylcholine (1x10 5 M)-induced contractions of bladder and on noradrenaline (1x10 5 M)induced ones of prostate. Protein levels of CBS and MPST in each part were also investigated by Western blotting. RESULTS: NaHS dose-dependently elicited relaxation on BL-D and BL-T pre-contracted by acetylcholine and on PR-V and PR-D precontracted by noradrenaline. There were no significant differences in values of EC50 and relaxation rate against the pre-contractions among these tissues. CBS was detected in PR-V and PR-D, and the expression levels in PR-D were higher than those in PR-V. In BL-D and BL-T, CBS was not detected. MPST was detected in all these tissues. Expression levels of MPST in PR-V were the highest and those in prostate were higher than those in bladder. CONCLUSIONS: H2S induces relaxation of bladder and prostate smooth muscle. H2S synthesis enzymes are expressed in both bladder and prostate. These results suggest that H2S could function as an endogenous relaxation factor in both tissues. Endogenous H2S might be a new therapeutic target for lower urinary tract dysfunction such as overactive bladder and benign prostatic hyperplasia. Source of Funding: none

891 Inhibition of prostate smooth muscle contraction and prostate stromal cell growth by NSC23766 and EHT1864, two novel inhibitors of the small GTPase Rac

891 Inhibition of prostate smooth muscle contraction and prostate stromal cell growth by NSC23766 and EHT1864, two novel inhibitors of the small GTPase Rac

791 Effects of two novel group I p21-activate kinases, FRAX486 and IPA3, on neuronal and agonist-induced contraction of hyperplastic human prostate smooth muscle

791 Effects of two novel group I p21-activate kinases, FRAX486 and IPA3, on neuronal and agonist-induced contraction of hyperplastic human prostate smooth muscle

896 Additive contractions by thromboxane A2 and endothelin-1 are stronger than norepinephrineinduced contractions in human prostate smooth muscle

896 Additive contractions by thromboxane A2 and endothelin-1 are stronger than norepinephrineinduced contractions in human prostate smooth muscle

Structural modifications of the prostate in hypoxia, oxidative stress, and chronic ischemia.

PurposeClinical studies have reported a correlation between pelvic ischemia and voiding dysfunction in elderly men. The aim of this study was to identify and compare prostate structural modifications in cultured cells and in a rabbit model after exposure to hypoxia, oxidative stress, and chronic ischemia.Materials and MethodsCultured human prostate smooth muscle cells (SMCs), epithelial cells (ECs), and stromal cells (SCs) were incubated under normoxia, hypoxia, and oxidative stress conditions by use of a computerized oxycycler system. We developed a rabbit model of chronic prostate ischemia by creating aorto-iliac arterial atherosclerosis. Markers of oxidative stress were examined by using fluorometric analysis and enzyme immunoassay. Prostate structure was examined by using Masson's trichrome staining and transmission electron microscopy (TEM).ResultsLipid peroxidation was found in SMCs exposed to hypoxia and in all cell types exposed to oxidative stress. We identified protein oxidation in ECs exposed to hypoxia and in all cell types exposed to oxidative stress. Markers indicating oxidative damage were present in chronically ischemic rabbit prostate tissue. These reactions were associated with DNA damage. Prostate ischemia resulted in epithelial atrophy, loss of smooth muscle, and diffuse fibrosis. TEM showed swollen mitochondria with degraded cristae, loss of membrane, loss of Golgi bodies, degenerated nerves, and disrupted cell-to-cell junctions.ConclusionsHuman prostate cells exhibited differential reactions to hypoxia and oxidative stress with widespread DNA damage. Structural modifications in ischemic prostate tissue were similar to those in cells exposed to oxidative stress. Structural changes due to ischemia and oxidative stress may contribute to prostatic noncompliance in aging men.

The beta-3 adrenoceptor agonist, mirabegron relaxes isolated prostate from human and rabbit: new therapeutic indication?

Alpha1 (α1)-blockers, 5-alpha reductase and phosphodiesterase type-5 inhibitors are pharmacological classes currently available for benign prostatic hyperplasia (BPH) treatment. Mirabegron, a beta-3 adrenoceptor (β3-AR) agonist has been approved for the therapy of overactive bladder and may constitute a new therapeutic option for BPH treatment. This study is aimed to evaluate the in vitro effects of mirabegron in human and rabbit prostatic smooth muscle. In rabbit prostate, electrical field stimulation (EFS)-induced contraction and concentration-response curve (CRC) to mirabegron in phenylephrine pre-contracted tissues were carried out. The potency (pEC50 ) and maximal response (Emax ) values were determined. In human prostate, CRC to phenylephrine was carried out in the absence and presence of mirabegron. Immunohistochemistry analysis for β3-AR was also carried out. In human prostate, immunohistochemistry analysis revealed the presence of β3-AR on the transition zone and mirabegron reduced by 42% the phenylephrine-induced contractions. In rabbit prostate, mirabegron produced concentration-dependent relaxations (pEC50 : 6.01 ± 0.12; Emax : 106 ± 3%), which were fully resistant to the blockade of β1-AR and β2-AR. The β3-AR blocker L748,337 caused a six-fold rightward shift in mirabegron-induced relaxations. Mirabegron (10 μM) reduced by 63% the EFS-induced contractions. Inhibitors of nitric oxide (L-NAME) and of soluble guanylate cyclase (ODQ) along with a cocktail of K+ channel blockers (apamin, charybdotoxin, glibenclamide, tetraethylammonium) all failed to significantly affect the mirabegron-induced rabbit relaxations. Mirabegron relaxes prostatic smooth muscle, providing an experimental support for the clinical investigation of its combination with an α1-blockers or PDE5 inhibitors in the treatment of BPH. Prostate 75:440-447, 2015. © 2014 Wiley Periodicals, Inc.

Effects of Schisandra chinensis extract on the relaxation of isolated human prostate tissue and smooth muscle cell

Ethnopharmacological relevanceSchisandra chinensis has been commonly used as a traditional herbal medicine to treat various diseases including body weakness, dysentery, impotence, enuresis and frequent urination in many countries including Korea, China and Russia. Benign prostate hyperplasia is a common disease for the elderly men and it induces lower urinary tract symptoms which hinder general activity and quality of life. We evaluated the therapeutic potential of Schisandra chinensis extract (SCE) in benign prostate hyperplasia using human prostate tissue. Materials and methodsSchisandra chinensis fruit was collected and extracted with ethanol. Human prostate tissues were obtained from 14 prostate cancer patients. Macroscopically normal tissue was excised from the transition zone and the periurethral regions. Isolated prostate tissue strips were mounted in an organ-bath system, and the relaxation effect of SCE was evaluated by cumulative addition to prostate strips pre-contracted with 10−5M norepinephrine. The effect of tamsulosin was compared, and the additive effect was evaluated. Electrophysiological studies using cultured human prostate smooth muscle cells (HPrSMC) were conducted. ResultsCumulative dosing of SCE induced concentration-dependent relaxation in contracted prostate tissue (n=18, P<0.05). Simultaneous dosing of SCE and tamsulosin showed an additive relaxation effect. The relaxation effect of SCE was abolished by inhibition of K+ channels by pre-treatment with tetraethylammonium. In HPrSMC, extracellular application of 100μg/mL SCE significantly increased outward currents, and this effect was significantly attenuated by treatment with 100nM Iberiotoxin. ConclusionsSCE showed a dose dependent relaxation effect on human prostate tissue as well as an additive effect with tamsulosin. The relaxation effects of SCE on HPrSMC were, in part, due to the activation of K+ channels.

Honokiol, a constituent of Magnolia species, inhibits adrenergic contraction of human prostate strips and induces stromal cell death.

PurposeSmooth muscle contraction and prostate growth are important targets for medical therapy of lower urinary tract symptoms (LUTS) in patients with benign prostatic hyperplasia. Honokiol and Magnolol are lignan constituents of Magnolia species, which are used in traditional Asian medicine. Here, we examined effects of honokiol and magnolol on contraction of human prostate tissue and on growth of stromal cells.MethodsProstate tissues were obtained from radical prostatectomy. Contraction of prostate strips was examined in organ bath studies. Effects in stromal cells were assessed in cultured immortalized human prostate stromal cells (WPMY-1). Ki-67 mRNA was assessed by reverse transcription-polymerase chain reaction, and proliferation by a fluorescence 5-ethynyl-2′-deoxyuridine assay.ResultsHonokiol (100μM) reduced noradrenaline-induced contractions, which was significant at 10 to 100μM noradrenaline. Honokiol reduced phenylephrine-induced contractions, which was significant at 3 to 100μM phenylephrine. Honokiol reduced electric field stimulation-induced contractions very slightly. In WPMY-1 cells, honokiol (24 hours) induced cell death. Magnolol (100μM) was without effects on contraction, and cellular viability.ConclusionsHonokiol inhibits smooth muscle contraction in the human prostate, and induces cell death in cultured stromal cells. Because prostate smooth muscle tone and prostate growth may cause LUTS, it appears possible that honokiol improves voiding symptoms.

The role of alpha blockers prior to removal of urethral catheter for acute urinary retention in men.

Acute urinary retention is a urological emergency in men and requires urgent catheterisation. Any intervention which aims at improving urinary symptoms following an acute urinary retention episode could be potentially beneficial. Alpha blockers relax prostatic smooth muscle cells thereby decreasing the resistance to urinary flow and by doing so could improve urinary symptoms. To assess the effectiveness of alpha blockers on successful resumption of micturition following removal of a urethral urinary catheter after an episode of acute urinary retention in men. In the absence of internationally agreed outcome measures for the success of a trial without catheter, success was defined as the return to satisfactory voiding without need for re-catheterisation within 24 hours. We searched the Cochrane Incontinence Group Specialised Trials Register, which contains trials identified from the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, MEDLINE in process, and handsearching of journals and conference proceedings (searched 9 October 2013), CENTRAL (2013, Issue 5) (searched 5 June 2013), MEDLINE 1946 to May Week 4 2013, MEDLINE in Process (covering to 3 June 2013), EMBASE Classic and EMBASE 1947 to 2013 Week 22 (all searched 4 June 2013) and the reference lists of relevant articles. No language or other restrictions were imposed on the searches. Only randomised and quasi-randomised clinical trials of alpha blockers for trial without a urethral catheter following an episode of acute urinary retention in men were included. Two review authors independently examined all the citations and abstracts derived from the search strategy. Any disagreement about trial selection and inclusion was resolved by discussion. A third independent judgement was sought where disagreement persisted. Two review authors extracted independently, cross-checked and processed the data as described in the Cochrane Handbook for Systematic Reviews of Intervention. Quality of evidence of the critical outcomes was assessed by adopting the GRADE approach. Nine randomised clinical trials were included in this review. Eight trials compared alpha blockers versus placebo (five trials tested alfuzosin and two trials tested tamsulosin, one trial tested both alfuzosin and tamsulosin, one trial tested silodosin) and one trial compared an alpha blocker (doxazosin) versus no treatment. Trial without catheter was performed after treatment with the drug for one to three days in seven trials and for eight and 32 days in two other trials respectively. There was moderate quality evidence to suggest that the rate of successful trial without catheter favoured alpha blockers over placebo ( 366/608, 60.2%, of men using an alpha blocker were able to void spontaneously after catheter removal compared with 185/486, 38.1%, using placebo, risk ratio (RR) 1.55, 95% confidence interval (CI) 1.36 to 1.76). The incidence of recurrent acute urinary retention was lower in groups treated with an alpha blocker (RR 0.69, 95% CI 0.60 to 0.79). This evidence was of moderate quality and was statistically significant for alfuzosin, tamsulosin and silodosin, though not for doxazosin. Of the trials mentioning adverse effects (for example, postural hypotension, dizziness), there was not enough information to detect statistically significant differences between the groups (RR 1.19, 95% CI 0.75 to 1.89) and the evidence was of low quality. Overall, adverse effect rates were low for both placebo and alpha blockers and, for example, vasodilatation-related adverse effects did not often result in discontinuation. There was some evidence to suggest that alpha blockers increase the success rates of trial without catheter, and the incidence of adverse effects was low. There was some evidence of a decreased incidence of acute urinary retention. The need for further surgery, cost effectiveness and recommended duration of alpha blocker treatment after successful trial without catheter remain unknown as these were not reported by any trial. There is a lack of internationally agreed outcome measures for what constitutes successful trial without catheter. This makes meta-analysis difficult. Large, well-designed controlled trials, which use the recommendations set out in the CONSORT statement, and include clinically important outcome measures, are required.