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A need exists for local (ie, bladder-specific) interventions to treat overactive bladder (OAB) with low risk of unwanted postprocedural outcomes. Gene therapy targeted to leverage endogenous physiology in bladder cells may assist in restoring normal cell and organ function. Herein, we review the potential promise of gene therapy for treating OAB, focusing on gene transfer of URO-902, a non-viral naked plasmid DNA expressing the big potassium (BK) channel. We searched PubMed for articles concerning functional aspects of the BK channel and its potential use for gene transfer as local OAB treatment. Results from preclinical, phase 1, and phase 2 studies of URO-902 for erectile dysfunction and phase 1 studies of URO-902 for OAB are included. The BK channel has been extensively studied; however, URO-902 is the first gene therapy used in clinical trials directed toward treating OAB via the BK channel. In both URO-902 studies, there were no serious adverse events considered treatment related and no adverse events leading to early withdrawal. Both studies included secondary efficacy endpoints with promising results suggesting improvement in OAB symptoms, and quality of life, with use of URO-902 versus placebo. Gene therapy involving the BK channel, such as gene transfer with URO-902, has demonstrated promising safety and efficacy results in women with OAB. Findings warrant further investigation of the use of URO-902 for OAB treatment.

Aims Two phase 1 trials were performed in healthy women with the overactive bladder (OAB) syndrome and urodynamically demonstrated detrusor overactivity (DO), with the aim to demonstrate the safety and potential efficacy of URO‐902, which comprises a gene therapy plasmid vector expressing the human big potassium channel α subunit. Methods ION‐02 (intravesical instillation) and ION‐03 (direct injection) were double‐blind, placebo‐controlled, multicenter studies without overlap in enrollment between studies. Active doses were administered and evaluated sequentially (lowest dose first) for safety. ION‐02 participants received either 5000 µg or 10 000 µg URO‐902, or placebo. ION‐03 participants received either 16 000 or 24 000 µg URO‐902, or placebo, injected directly into the bladder wall using cystoscopy. Primary outcome variables were safety parameters occurring subsequent to URO‐902 administration; secondary efficacy variables also were evaluated. Results Among the safety outcomes, there were no dose‐limiting toxicities or significant adverse events (AEs) preventing dose escalation during either trial, and no participants withdrew due to AEs. For efficacy, in ION‐02 (N = 21), involuntary detrusor contractions on urodynamics at 24 weeks in patients receiving URO‐902 (P

OBJECTIVE The aim of this study was to evaluate the effectiveness of a genital vibratory stimulation device in improving sexual function in women with arousal and orgasm disorders. METHODS In this single-arm, prospective study, baseline and 1- and 3-month assessments were performed to evaluate women with sexual arousal and/or orgasmic disorders, who received therapy using a genital vibratory stimulation device. Sexual function, satisfaction, and distress were evaluated using the Female Sexual Function Index (FSFI), the Female Sexual Distress Scale, and the Female Intervention Efficacy Index questionnaires. Genital sensation was evaluated using quantitative sensory testing. RESULTS Seventy women, aged 19 to 64 years, were evaluated from October 2009 to August 2013. Forty-seven (67.1%) and 37 (52.9%) women completed 1- and 3-month follow-ups, respectively. The FSFI arousal and orgasm domain scores and total FSFI scores improved at 1 and 3 months (P < 0.001 for all outcomes). Mean (SD) total FSFI scores increased from 20.04 (4.65) (baseline) to 25.03 (5.21) (1 month) to 26.66 (5.42) (3 months; both Ps < 0.0001). Female Sexual Distress Scale scores reflected significantly decreased distress at 1 (P = 0.0006) and 3 (P < 0.0001) months compared with baseline and at 3 months compared with 1 month (P = 0.03). Neurological sensation was increased at all genital sites at 1 and 3 months (P < 0.0001 for all). After adjustment for age, there was a significant interaction between arousal domain scores and clitoral and right labial sensation. At 3 months, perceptions of increased vaginal lubrication, orgasm, and genital sensation were reported by 67.5%, 65.0%, and 82.5% of the participants. No major adverse events were noted. CONCLUSIONS Genital vibratory stimulation device use resulted in uniform improvements in sexual function, satisfaction, sexually related distress and genital sensation.

Objectives To investigate the effect of diabetes on urothelial modulation of bladder contractility. Methods Bladder strips (urothelium intact or denuded) were prepared from 8-week-old streptozotocin-induced diabetic (n = 19) and non-diabetic control rats (n = 10). The effect of modulators of MaxiK (iberiotoxin and tetraethylammonium) and Kv7 (XE991 and retigabine) potassium channel activity were investigated for their effects on both carbachol-induced force generation and spontaneous contractile activity. Results In bladder strips from non-diabetic animals, the presence of the urothelium resulted in marked sensitivity to carbachol-induced force generation by modulators of MaxiK and Kv7 channel activity, whereas in the diabetic animal urothelial sensitivity to these agents was significantly diminished. Urothelial-intact bladder strips from non-diabetic animals were more sensitive to modulators of Kv7 activity in reducing the amplitude of spontaneous phasic contractions than urothelial-denuded bladder strips, whereas in diabetic animals the presence or absence of the urothelium did not alter the sensitivity to modulators of Kv7 activity. Spontaneous activity in the presence of tetraethylammonium was not affected by the urothelium in bladder strips from either diabetic or non-diabetic animals. Conclusions The presence of the urothelium in bladders from non-diabetic animals modulates the activity of potassium blockers to affect bladder contractility, whereas in the diabetic bladder this effect is attenuated. These findings could help to explain the lack of success of pharmaceutical treatments targeting potassium channels to treat bladder pathology in patients with diseases imparing urothelial function.

Men with sickle cell disease (SCD) risk developing priapism. Recognizing that SCD is a disease of hypoxia, we investigated the effect of hypoxia on gene expression in corporal smooth muscle (CSM) cells. Rat CSM cells in vitro were treated with CoCl2 or low oxygen tension to mimic hypoxia. Hypoxic conditions increased expression of genes previously associated with priapism in animal models. Variable coding sequence a1 (Vcsa1; the rat opiorphin homologue, sialorphin), hypoxia-inducible factor 1a (Hif-1a), and A2B adenosine receptor (a2br) were increased by 10-, 4-, and 6-fold, respectively, by treatment with CoCl2, whereas low oxygen tension caused increases in expression of 3-, 4-, and 1.5-fold, respectively. Sialorphin-treated CSM cells increased expression of Hif-1a and a2br by 4-fold, and vcsa1-siRNA treatment reduced expression by ∼50%. Using a Hif-1a inhibitor, we demonstrated up-regulation of a2br by sialorphin is dependent on Hif-1a, and knockdown of vcsa1 expression with vcsa1-siRNA demonstrated that hypoxic-up-regulation of Hif-1a is dependent on vcsa1. In CSM from a SCD mouse, there was 15-fold up-regulation of opiorphin at a life stage prior to priapism. We conclude that in CSM, opiorphins are master regulators of the hypoxic response. Opiorphin up-regulation in response to SCD-associated hypoxia activates CSM “relaxant” pathways; excessive activation of these pathways results in priapism.—Fu, S., Tar, M. T., Melman, A., Davies, K. P. Opiorphin is a master regulator of the hypoxic response in corporal smooth muscle cells.

What's known on the subject? and What does the study add? Partial urethral obstruction (PUO), a common urologic pathology in men and children, can be associated with detrusor overactivity (DO), but the exact molecular mechanisms responsible for the altered contractile phenotypes have not been fully elucidated. Our study is the first to determine by direct patch-clamp current measurement that PUO, to recapitulate many of the pathological characteristics of benign prostatic hyperplasia (BPH), is associated with an approximate 5-fold decrease in bladder myocyte Maxi-K channel activity. Similar to others we report a 40% decrease in Maxi-K α subunit expression which correlates with the observed decrease in Maxi-K activity, but uniquely as much as a 5-fold increase in expression of Maxi-K β subunit which we hypothesize may be a compensatory response. Based upon our previous finding of increased detrusor overactivity (DO) in similar male PUO rats, we suggest the Maxi-K channel as a therapeutic target to treat BPH associated lower urinary tract symptoms (LUTS). OBJECTIVES • To examine the effect of partial urethral obstruction (PUO) on bladder smooth muscle outward potassium current and the contribution of the large-conductance calcium-activated potassium (Maxi-K, BKCa) channel to this activity in smooth muscle cells isolated from bladders of sham-operated and PUO male rats using whole-cell patch clamp recording techniques. • To determine the effect of PUO on the expression of the Maxi-K channel α and β1 subunits and in vitro detrusor contractility. MATERIALS AND METHODS • Twenty adult male Sprague–Dawley rats were divided equally into two groups and subjected to surgical ligation of the urethra (PUO) or sham surgery (SHAM). • After 2 weeks, the detrusors from PUO and SHAM rats were used for molecular analyses (mRNA and protein quantification of Maxi-K subunits) or organ bath contractility studies, or myocytes were isolated for conventional whole-cell patch clamp analyses. RESULTS • PUO increased bladder mass 2.5-fold and detrusor strips exhibited a more tonic-type contraction and increased contractility compared with controls (SHAM). • Iberiotoxin (300 nM) sensitive Maxi-K channel current comprised about 40% of the outward whole-cell current in SHAM bladders but only about 8% in PUO bladders. • Expression of the α subunit of the Maxi-K channel was significantly decreased ∼40% while the expression of the β1 subunit was increased ∼2-fold at the mRNA level. The increase in β1 expression was confirmed by Western blotting. CONCLUSIONS • Our findings show that obstruction of the rat bladder is associated with decreased Maxi-K channel activity of bladder smooth muscle cells, determined via direct current measurement. • Increased expression of the β1 subunit points to a compensatory reaction to decreased Maxi-K channel activity. • Maxi-K channel openers or gene therapy may therefore provide therapeutic benefit for the overactive bladder.

Diabetes mellitus (DM) is a quite common chronic disease, and the prevalence of erectile dysfunction (ED) is three times higher in this large population. Although diabetes-related ED has been studied extensively, the actin-myosin contractile apparatus was not examined. The mRNAs encoding smooth muscle myosin (SMM) heavy chains (MHC) and essential light chains (LC17) exist as several different alternatively spliced isoforms with distinct contractile properties. Recently, we provided novel data that blebbistatin (BLEB), a specific myosin II inhibitor, potently relaxed corpus cavernosum smooth muscle (CCSM). In this study, we examine whether diabetes alters SMM expression, alternative splicing, and/or functional activities, including sensitivity to BLEB. By using streptozotocin (STZ)-induced 2-mo diabetic rats, functional activities were tested in vivo by intracavernous pressure (ICP) recording during cavernous nerve stimulation and in vitro via organ bath contractility studies. SMM isoform composition was analyzed by competitive RT-PCR and total SMM, myocardin, and embryonic SMM (SMemb) expression by real-time RT-PCR. Results revealed that the blood glucose level of STZ rats was 407.0 vs. 129.5 mg/dl (control). STZ rats exhibited ED confirmed by significantly increased CCSM contractile response to phenylephrine and decreased ICP response. For STZ rats, SM-B, LC17aand SM2 isoforms, total SMM, and myocardin expression increased, whereas SM-A, LC17b, and SM1 isoforms were decreased, with SMemb unchanged. BLEB was significantly more effective in relaxing STZ CCSM both in vitro and in vivo. Thus we demonstrated a novel diabetes-specific effect on alternative splicing of the SMM heavy chain and essential light chain genes to a SMM isoform composition favoring a heightened contractility and ED. A switch to a more contractile phenotype was supported further by total SMM expression increase. Moreover, the change in CCSM phenotype was associated with an increased sensitivity to BLEB, which may serve as a novel pharmacotherapy for ED.

Erectile dysfunction (ED) is a neurovascular phenomenon modulated by hormonal, local biochemical, and biomechanical/structural factors of the penis. The success of the orally active phosphodiesterase inhibitors for the treatment of ED has boosted research activities into the physiology of the erectile mechanism. Peripheral intracellular signal transduction in the penis as well as central brain and spinal cord pathways controlling penile erection have been investigated and are now better understood. The results of this ongoing research have provided the basis for the development and introduction of several novel therapeutic modalities into the management of ED. Many novel pharmacotherapeutic approaches under development including the use of melanocortins and Rho-kinase inhibitors as well as the introduction of gene therapy and tissue engineering have demonstrated efficacy in animal as well as early human trials. This review describes the major new and evolving pharmacological advances in the field of oral pharmacotherapy for the treatment of male ED.

Diabetes results in a myriad of vascular complications, often referred to as diabetic vasculopathy, which encompasses both microvascular [erectile dysfunction (ED), retinopathy, neuropathy, and nephropathy] and macrovascular complications (hypertension, coronary heart disease, and myocardial infarction). In diabetic animals and patients with ED, there is decreased opiorphin or opiorphin-related gene expression in corporal tissue. Both opiorphin and the rat homologous peptide sialorphin are found circulating in the plasma. In the present study, we investigated if diabetes induced changes in plasma sialorphin levels and if changes in these levels could modulate the biochemistry and physiology of vascular smooth muscle. We show that circulating sialorphin levels are reduced in a rat model of type I diabetes. Intracorporal injection of plasmids expressing sialorphin into diabetic rats restores sialorphin levels to those seen in the blood of nondiabetic animals and results in both improved erectile function and blood pressure. Sialorphin modulated the ability of C-type natriuretic peptide to relax both corporal and aortic smooth muscle strips and of bradykinin to regulate intracellular calcium levels in both corporal and aortic smooth muscle cells. We have previously shown that expression of genes encoding opiorphins is increased when erectile function is improved. Our findings thus suggest that by affecting circulating levels of opiorphin-related peptides, proper erectile function is not only an indicator but also a modulator of overall vascular health of a man.

The MaxiK potassium channel is regulated by voltage and intracellular calcium, and plays a critical role in regulating intracellular calcium concentration ([Ca(2+) ](i)), which is the ultimate determinant of smooth muscle tone. Tight control of corpus cavernosum smooth muscle (CCSM) tone is critically important and misregulation can result in erectile dysfunction.Because of the tight functional linkage of MaxiK and calcium channel activity, the aim of this study was to determine the effects of silencing and pharmacological inhibition of MaxiK on calcium homeostasis and intercellular calcium signaling in CCSM cells.We compared changes in the basal intracellular [Ca(2+) ](i) and parameters defining intercellular calcium wave (ICW) spread in 48 hours MaxiK silenced CCSM cells vs. acute blockade of the channel with iberiotoxin. To analyze changes occurring in gene expression we performed micro-array analysis following MaxiK silencing for 48 hours.Changes in Fura-2 fluorescence intensities were measured to evaluate basal [Ca(2+) ](i) levels and ICW parameters. Microarray analysis of global gene expression was performed.Forty-eight hours after MaxiK silencing the basal [Ca(2+) ](i) , the ICW amplitude and spread among CCSM cells were not markedly different in silenced compared to mock transfected controls, whereas short-term blockade significantly increased basal [Ca(2+) ](i) level and amplified Ca(2+) signaling among CCSM cells. Micro-array analysis showed that several genes within Ca(2+) homeostasis and smooth muscle tone regulation pathways had significantly altered expression.Our results indicate that while short-term blockade of the MaxiK channel is associated with an increase in basal [Ca(2+) ](i), Ca(2+) homeostasis is restored during the 48 hours period following silencing. We hypothesize that the different pathways regulating [Ca(2+) ](i) and CCSM tone are linked through molecular crosstalk and that their coordinated regulation is part of a compensatory mechanism aimed to maintain Ca(2+) homeostasis and CCSM tone.

Introduction Normal erectile function (EF) involves a coordinated relaxation of the arteries that supply the penis and the corpus cavernosum smooth muscle (CCSM), resulting in expansion of the sinusoids and increased intracavernous pressure. But the CCSM spends the majority of its time in the contracted state which is mediated by norepinephrine released from nerve endings and other vasoconstrictors like endothelins released from the endothelium. These agents cause smooth muscle myosin (SMM) phosphorylation by elevating intracellular calcium. When calcium returns to basal levels, the calcium sensitivity increases and prevents myosin dephosphorylation, which involves the RhoA/Rho-kinase (ROK) mechanism, thus maintaining force. Although mounting evidences demonstrate that androgens have a major influence on EF that is not just centrally mediated, this notion remains quite controversial. Aim To summarize the current knowledge on CCSM contractile pathways, the role they play in modulating EF, and the influence of androgens. Methods The article reviews the literature and contains some previously unpublished data on CCSM contraction signaling including the role that androgens are known to play in modulating these pathways. Main Outcome Measures Data from peer-reviewed publications and previously unpublished observations. Results In addition to downregulation of many pro-erectile molecular mechanisms, decreased testosterone (T) levels upregulate CCSM contractility, including hyperresponsiveness to α-adrenergic agonists, increased SMM phosphorylation, alteration of SMM isoform composition, activation of RhoA/ROK signaling and modulation of sphingosine-1-phosphate regulation of CCSM tone. Conclusions Decreased T levels upregulate CCSM contractile signaling. Meanwhile, it downregulates CCSM relaxation pathways synergizing to produce erectile dysfunction (ED). Although some urologists and researchers are still skeptical of the influence of androgens on penile erection, understanding these molecular control mechanisms as well as the influence that androgens have on these pathways should provide new evidence supporting the roles of androgens in EF and enhance the discovery of novel targets for drug development to treat ED.

OBJECTIVE To investigate the in vitro and in vivo effects of blebbistatin (a small cell-permeable molecule with high affinity and selectivity toward the myosin II contractile molecule) on bladder smooth muscle (SM) contractility, as antimuscarinic therapy is only 65-75% effective in treating overactive bladder (OAB) and is associated with considerable side-effects, with a < 25% continuation rate at 1 year. MATERIALS AND METHODS Bladder and aortic strips from adult male rats, and human bladder strips obtained from open prostatectomy, were used for organ-bath studies of blebbistatin. Awake cystometry was also used in rats in both the presence and absence of intravesically delivered blebbistatin. RESULTS Blebbistatin dose-dependently and completely relaxed both KCl- and carbachol-induced rat detrusor and endothelin-1-induced human bladder contraction. Pre-incubation with 10 µm blebbistatin attenuated carbachol responsiveness by ≈ 65% while blocking electrical field stimulation-induced bladder contraction reaching 50% inhibition at 32 Hz. The basal tone and amplitude of spontaneous contraction were also significantly diminished. Urodynamic variables were obviously altered by intravesical infusion with blebbistatin. CONCLUSION Our novel data show that blebbistatin strongly relaxes both rat and human bladder contraction induced by various physiological stimuli. Coupled with our in vivo data showing that nanomole doses of blebbistatin significantly alter urodynamic variables to produce a less active bladder, our results suggest the possibility of intravesically administered blebbistatin binding at myosin II being developed as a therapeutic treatment for OAB via a novel targeting of the SM contractile apparatus.

What’s known on the subject? and What does the study add? Diabetes is a common precursor for ladder pathology, including detrusor overactivity and cystopathy. There is preliminary, but increasing evidence, suggesting that oxidative stress plays a significant role in the development of diabetic complications including its affect on the bladder. In the present study we investigated the effect of streptozotocin induced-diabetes in rats on the global expression of genes in the rat bladder using microarray analysis, and combined this data with our previously reported study looking at changes in protein levels using proteomics. This analysis demonstrated that markers of oxidative stress were significantly increased in the diabetic bladder. Overall, our work adds to the growing body of evidence that diabetic cystopathy is associated with oxidative damage of smooth muscle cells, and results in protein damage and activation of apoptotic pathways which may contribute to a deterioration in bladder function. OBJECTIVE • To investigate the role that oxidative stress plays in the development of diabetic cystopathy. MATERIALS AND METHODS • Comparative gene expression in the bladder of non-diabetic and streptozotocin (STZ)-induced 2-month- old diabetic rats was carried out using microarray analysis. • Evidence of oxidative stress was investigated in the bladder by analyzing glutathione S-transferase activity, lipid peroxidation, and carbonylation and nitrosylation of proteins. • The activity of protein degradation pathways was assessed using Western blot analysis. RESULTS • Analysis of global gene expression showed that detrusor smooth muscle tissue of STZ-induced diabetes undergoes significant enrichment in targets involved in the production or regulation of reactive oxygen species (P= 1.27 × 10−10). The microarray analysis was confirmed by showing that markers of oxidative stress were all significantly increased in the diabetic bladder. • It was hypothesized that the sequelae to oxidative stress would be increased protein damage and apoptosis. • This was confirmed by showing that two key proteins involved in protein degradation (Nedd4 and LC3B) were greatly up-regulated in diabetic bladders compared to controls by 12.2 ± 0.76 and 4.4 ± 1.0-fold, respectively, and the apoptosis inducing protein, BAX, was up-regulated by 6.76 ± 0.76-fold. CONCLUSION • Overall, the findings obtained in the present study add to the growing body of evidence showing that diabetic cystopathy is associated with oxidative damage of smooth muscle cells, and results in protein damage and activation of apoptotic pathways that may contribute to a deterioration in bladder function.

Gene transfer for the treatment of erectile dysfunction has completed phase 1 safety testing and has shown the necessary safety to proceed to the next level of clinical trial. This review focuses on the background of the components of that have led to US Food and Drug Administration acceptance of human gene transfer trials for nonlethal disease.

OBJECTIVES To examine the effect of partial urethral obstruction (PUO) on the sphingosine-1-phosphate (S1P, a bioactive lipid shown to modulate smooth muscle, SM) pathway in the bladders of male rats, and to determine the effect of PUO on the RhoA/Rho-kinase (ROK) pathway, and whether there is a molecular cross-talk with the S1P pathways associated with bladder overactivity (S1P1-S1P3, where S1P1 is associated with nitric oxide-mediated SM relaxation, and S1P2 and S1P3 receptors are associated more with SM contraction via the ROK pathway). MATERIALS AND METHODS In all, 20 male rats were divided into two groups and underwent PUO or a sham operation (control). After 2 weeks all rats were killed humanely and bladder specimens used for in vitro organ-bath physiological contractility studies, and for mRNA and protein analyses of major S1P/ROK pathway constituents via real-time polymerase chain reaction and Western blotting, respectively. In addition, early-passage SM cells were transfected with recombinant sphingosine kinase (SPHK, the enzyme that converts sphingosine to S1P). RESULTS Bladders from PUO rats had greater mRNA expression of the S1P2 and S1P3 receptors, as well as SPHK1, than the sham controls (4.78, 2.04 and 2.72 times, respectively). PUO rats also had significantly greater expression of RhoA and ROKα (1.76 and 2.19 times, respectively). Western blotting and organ-bath contractility studies showed similar changes at the protein and in vitro functional level, with an increased contractility of bladder strips from PUO rats to exogenous S1P. Transfection of SPHK into isolated SM cells increased ROK expression. CONCLUSIONS We show for the first time that the S1P signalling pathway is significantly upregulated in response to PUO in male rats at both the molecular and in vitro functional levels, correlating with an activation of the RhoA/ROK pathway. Further, we provide novel data that SPHK overexpression increases ROK expression in vitro, suggesting a novel hypothesis of S1P-induced bladder overactivity in the mechanism for PUO-induced bladder dysfunction and the S1P signalling pathway as a possible therapeutic target for bladder overactivity.

Background Despite the advent of effective oral therapies for erectile dysfunction (ED), many patients are not successfully treated, and side effects have been documented. Objective To further evaluate the potential utility of naked DNA-based gene transfer as an attractive treatment option for ED. Design, setting and participants The effects of gene transfer on erectile function and sexual behavior were evaluated in eight male cynomolgus monkeys with ED secondary to moderately severe, diet-induced atherosclerosis. Intervention Following establishment of baseline characteristics, animals were subjected to intracavernous injection of a smooth-muscle–specific gene transfer vector (pSMAA- hSlo ) encoding the pore-forming subunit of the human large-conductance, calcium-sensitive potassium channel (Maxi-K). Measurements For the sexual behavior studies, 2 wk of baseline data were obtained, and then animals were placed in the presence of estrogen-implanted females ( n =2) three times per week for 30min, and sexual behavior was recorded. The intracavernous pressure response to papaverine injection was also monitored. Results and limitations Dramatic changes in erectile function and sexual behavior were observed after intracorporal gene transfer. The frequency of partial (6±2 to 10±2) and full (2±1.5 to 5±1.4) erections were significantly increased, with a parallel 2–3-fold increase in the duration of the observed erections. The frequency and latency of ejaculation were increased and decreased, respectively. Frequency and duration of grooming by the female were increased, and the latency decreased. Increased latency and decreased frequency of body contact was also observed, and this is characteristic of the typical drop in consort intimacy that occurs after mating in most macaque species. In addition, an increased responsiveness to intracavernous papaverine injection was observed. Conclusions The data indicate that intracorporal Maxi-K–channel gene transfer enhances erectile capacity and sexual behavior; the data imply that increased erectile function per se may lead to increased sexual function.

OBJECTIVES To provide sensitive physiological endpoints for the onset and long-term progression of deficits induced by diabetes mellitus (DM) in bladder and erectile function in male rats, and to evaluate parallel changes in urogenital and nerve function induced by hyperglycaemia over a protracted period as a model for chronic deficits in patients with diabetes. MATERIALS AND METHODS The study comprised in 877 male, 3-month-old, Fischer 344 rats; 666 were injected intraperitoneally with 35 mg/kg streptozotocin (STZ) and divided into insulin-treated and untreated diabetic groups. The rats were studied over 8 months and measurements made of both erectile and bladder function, as well as nerve conduction studies over the duration of the study. RESULTS There was an early (first month) abnormality of both erectile and bladder function that persisted through the 8 months of the study. The erectile dysfunction was manifest as reduced intracavernous pressure/blood pressure ratio, and the bladder dysfunction as a persistent increase in detrusor overactivity with no detrusor decompensation. Insulin treatment prevented or modified the abnormality in each organ. Hyperglycaemia caused a progressive decrease in caudal nerve conduction velocity. The mean digital sensory and tibial motor nerve conduction velocity did not deteriorate over time. Correlation measurements of nerve and organ function were not consistent. CONCLUSIONS The results of this extensive long-term study show early and profound effects of hyperglycaemia on the smooth muscle of the penis and bladder, that were persistent and stable in surviving rats over the 8 months. The physiological changes did not correlate well with neurological measurements of those organs. Significantly, diverse smooth-muscle cellular and subcellular events antedated the measured neurological manifestations of the hyperglycaemia by several months. Although autonomic diabetic neuropathy is a primary life-threatening complication of long-term diabetes in humans, this rat model of STZ-induced diabetes showed that the rapid onset of physiological manifestations was based on many molecular changes in the smooth muscle cells in this model of type 1 DM.

Intracorporal injection of plasmids encoding opiorphins into retired breeder rats can result in animals developing a priapic-like condition. Microarray analysis demonstrated that following intracorporal gene transfer of plasmids expressing opiorphins the most significantly upregulated gene in corporal tissue was the ornithine decarboxylase gene (ODC). Quantitative RT-PCR confirmed the upregulation of ODC, as well as other genes involved in polyamine synthesis, such as arginase-I and -II, polyamine oxidase, spermidine synthase, spermidine acetyltransferase (SAT), and S-adenosylmethionine decarboxylase. Western blot analysis demonstrated upregulation of arginase-I and -II, ODC, and SAT at the protein level. Levels of the polyamine putrescine were upregulated in animals treated with opiorphin-expressing plasmids compared with controls. A direct role for the upregulation of polyamine synthesis in the development of the priapic-like condition was supported by the observation that the ODC inhibitor 1,3-diaminopropane, when added to the drinking water of animals treated with plasmids expressing opiorphins, prevented experimental priapism. We also demonstrate that in sickle cell mice, another model of priapism, there is increased expression of the mouse opiorphin homologue in corporal tissue compared with the background strain at a life stage prior to evidence of priapism. At a life stage when there is onset of priapism, there is increased expression of the enzymes involved in polyamine synthesis (ODC and arginase-I and -II). Our results suggest that the upregulation of enzymes involved in the polyamine synthetic pathway may play a role in the development of experimental priapism and represent a target for the prevention of priapism.

OBJECTIVE To investigate whether ageing and diabetes alter the expression of the gap junction protein connexin43 (Cx43) and of particular purinoceptor (P2R) subtypes in the corpus cavernosum and urinary bladder, and determine whether changes in expression of these proteins correlate with development of erectile and bladder dysfunction in diabetic and ageing rats. MATERIALS AND METHODS Erectile and bladder function of streptozotocin (STZ)-induced diabetic, insulin-treated and age-matched control Fischer-344 rats were evaluated 2, 4 and 8 months after diabetes induction by in vivo cystometry and cavernosometry. Corporal and bladder tissue were then isolated at each of these sample times and protein expression levels of Cx43 and of various P2R subtypes were determined by Western blotting. RESULTS In the corpora of control rats ageing was accompanied by a significant decrease in Cx43 and P2X1R, and increase in P2X7R expression. There was decreased Cx43 and increased P2Y4R expression in the ageing control rat bladder. There was a significant negative correlation between erectile capacity and P2X1R expression levels, and a positive correlation between bladder spontaneous activity and P2Y4R expression levels. There was already development of erectile dysfunction and bladder overactivity at 2 months after inducing diabetes, the earliest sample measured in the study. The development of these urogenital complications was accompanied by significant decreases in Cx43, P2Y2R, P2X4R and increase in P2X1R expression in the corpora, and by a doubling in Cx43 and P2Y2R, and significant increase in P2Y4R expression in the bladder. Changes in Cx43 and P2R expression were largely prevented by insulin therapy. CONCLUSION Ageing and diabetes mellitus markedly altered the expression of the gap junction protein Cx43 and of particular P2R subtypes in the rat penile corpora and urinary bladder. These changes in Cx43 and P2R expression provide the molecular substrate for altered gap junction and purinergic signalling in these tissues, and thus probably contribute to the early development of erectile dysfunction and higher detrusor activity in ageing and in diabetic rats.

Several reports suggest that the rat Vcsa1 gene is down-regulated in models of erectile dysfunction. The Vcsa protein product sialorphin is an endogenous neutral endopeptidase inhibitor and its down-regulation could result in prolonged activation of G-protein activated signaling pathways by their peptide agonists. We investigated whether Vcsa1 down-regulation could result in an adaptive change in GPCR (G-protein coupled receptor) expression.Gene expression in cultured rat corporeal smooth muscle cells following treatment with siRNA directed against Vcsa1 or the neutral endopeptidase gene was analyzed using microarray and quantitative reverse transcriptase-polymerase chain reaction. In rats Vcsa1 is one of the most down-regulated genes following bilateral transection of the cavernous nerves. In that animal model we also investigated whether Vcsa1 down-regulation was accompanied by similar changes in gene expression in corporeal smooth muscle cells in which Vcsa1 was knocked down in vitro.Microarray analysis and quantitative reverse transcriptase-polymerase chain reaction demonstrated that corporeal smooth muscle cells treated in vitro with siRNA against Vcsa1 resulted in GPCR up-regulation as a functional group. In contrast, treatment of corporeal smooth muscle cells that lowered neutral endopeptidase activity resulted in decreased GPCR expression. These results suggest that the peptide product of Vcsa1, sialorphin, can effect GPCR expression by acting on neutral endopeptidase. In animals with bilaterally transected cavernous nerves the decreased Vcsa1 expression is accompanied by increased GPCR expression in cavernous tissue.These experiments suggest that the mechanism by which Vcsa1 modulates erectile function is partly mediated through changes in GPCR expression.

Male sexual dysfunction can be classified in the following categories: erectile dysfunction (ED); orgasmic and ejaculatory dysfunction; priapism; and decreased libido. Of these various dysfunctions, more patients with ED are likely to seek medical attention. ED, or impotence, is defined as the inability to achieve or maintain an erection sufficient for satisfactory sexual function1. In recent years, there has been an escalated awareness of ED, partly attributed to the advent of ViagraTM and its associated promotion. The impact of ED can be appreciated by the estimation that its prevalence in men 40 to 70 years old is 52%, based on the Massachusetts Male Aging Study.2 Based on these data and United States population projections for the year 2005 of more than 50 million men 40 to 70 years old, ED will affect more than 25 million men, and millions more over the age of 70. The projected worldwide prevalence of ED for year 2025 will be staggering at 322 million men. Certain patients are found to have a significantly higher prevalence of ED; for example, diabetic men command a more than threefold increase in risk of ED than their nondiabetic counterparts. Indeed, diabetes mellitus is the single most common cause of ED.2 More than 50% of diabetic patients are afflicted with some degree of ED, and approximately 50% of the patients evaluated at our Center for Male Sexual Dysfunction are diabetic.

OBJECTIVE To determine if the mature peptide product of the Vcsa1 gene, sialorphin, could restore erectile function in ageing rats, and whether these effects are mediated through relaxation of corporal smooth muscle tissue, as we recently reported that Vcsa1 is one of the most down-regulated genes in the corpora of rats in three distinct models of erectile dysfunction, and gene transfer of plasmids expressing Vcsa1 into the corpora of ageing rats restored erectile function. MATERIALS AND METHODS Sialorphin was injected intracorporeally into retired breeder rats, and the effect on the physiology of corporal tissue was analysed by intracorporal/blood pressure (ICP/BP) measurement at different times after injection. In organ-bath studies, the ability of sialorphin (1 µg/mL) to enhance C-type natriuretic peptide (CNP) relaxation of corporal smooth muscle tissue strips was investigated after pre-contraction with 1 µm phenylephrine. RESULTS Intracorporal injection of 100 µg sialorphin into retired breeder rats resulted in a time-dependent increase in the ICP/BP response to electrostimulation of the cavernosal nerve. After 55–65 min the ICP/BP ratio increased to ≈ 0.6, a value associated with normal erectile function. In organ-bath studies after pre-contraction with 1 µm phenylephrine, 1 µm CNP significantly (67%) increased the relaxation rate of corporal tissue. This rate of relaxation was increased by 2.5-fold after incubation with sialorphin (1 µg/mL) compared with carrier alone. CONCLUSION These results show that sialorphin has a role in erectile function, probably through a mechanism that involves relaxation of corporal smooth muscle tissue.

OBJECTIVE To identify early diabetes-related alterations in gene expression in bladder and erectile tissue that would provide novel diagnostic and therapeutic treatment targets to prevent, delay or ameliorate the ensuing bladder and erectile dysfunction. MATERIALS AND METHODS The RG-U34A rat GeneChip® (Affymetrix Inc., Sunnyvale, CA, USA) oligonucleotide microarray (containing ≈8799 genes) was used to evaluate gene expression in corporal and male bladder tissue excised from rats 1 week after confirmation of a diabetic state, but before demonstrable changes in organ function in vivo. A conservative analytical approach was used to detect alterations in gene expression, and gene ontology (GO) classifications were used to identify biological themes/pathways involved in the aetiology of the organ dysfunction. RESULTS In all, 320 and 313 genes were differentially expressed in bladder and corporal tissue, respectively. GO analysis in bladder tissue showed prominent increases in biological pathways involved in cell proliferation, metabolism, actin cytoskeleton and myosin, as well as decreases in cell motility, and regulation of muscle contraction. GO analysis in corpora showed increases in pathways related to ion channel transport and ion channel activity, while there were decreases in collagen I and actin genes. CONCLUSIONS The changes in gene expression in these initial experiments are consistent with the pathophysiological characteristics of the bladder and erectile dysfunction seen later in the diabetic disease process. Thus, the observed changes in gene expression might be harbingers or biomarkers of impending organ dysfunction, and could provide useful diagnostic and therapeutic targets for a variety of progressive urological diseases/conditions (i.e. lower urinary tract symptoms related to benign prostatic hyperplasia, erectile dysfunction, etc.).

Introduction Bicycling is associated with neurological impairment and impotence in men. Similar deficits have not been confirmed in women. Aim To evaluate the effects of bicycling on genital sensation and sexual function in women. Methods Healthy, premenopausal, competitive women bicyclists and runners (controls) were compared. Main Outcome Measures (1) Genital vibratory thresholds (VTs) were determined using the Medoc Vibratory Sensation Analyzer 3000. (2) Sexual function and sexually related distress were assessed by the Dennerstein Personal Experience Questionnaire (SPEQ) and the Female Sexual Distress Scale (FSDS). Results Forty‐eight bicyclists and 22 controls were enrolled. The median age was 33 years. The bicyclists were older, had higher body mass indices (BMIs), were more diverse in their sexual orientation, and were more likely to have a current partner. Bicyclists rode an average of 28.3 ± 19.7 miles/day (range 4–100), 3.8 ± 1.5 days/week, for an average of 2.1 ± 1.8 hours/ride. The mean number of years riding was 7.9 ± 7.1 years (range 0.5–30). Controls ran an average of 4.65 ± 2.1 miles/day (range 1.5–8) and 5.0 ± 1.2 days/week. On bivariate analysis, bicyclists had significantly higher VTs than runners, indicating worse neurological function at all sites ( P Conclusion There is an association between bicycling and decreased genital sensation in competitive women bicyclists. Negative effects on sexual function and quality of life were not apparent in our young, healthy premenopausal cohort. Guess MK, Connell K, Schrader S, Reutman S, Wang A, LaCombe J, Toennis C, Lowe B, Melman A, and Mikhail MK. Genital sensation and sexual function in women bicyclists and runners: Are your feet safer than your seat? J Sex Med 2006;3:1018–1027.

The molecular interaction between smooth muscle (SM) myosin and actin in the corpus cavernosum (CC) determines the erectile state of the penis. A key mechanism regulating this interaction and subsequent development and maintenance of force is alternative splicing of SM myosin heavy chain (MHC) and 17 kDa essential SM myosin light chain (MLC) pre-mRNAs. Our aim was to examine the relative SM myosin isoform composition in human CC. Tissue samples were obtained from 18 patients with erectile dysfunction (ED), Peyronie's disease, or both. One specimen was obtained during a transgender operation. Patients then were stratified according to presence of diabetes mellitus, hypertension, ED, or Peyronie's disease, as well as failure of phosphodiesterase-5 (PDE5) inhibitors and history of previous pelvic or penile surgeries, radiation, or both. Our results revealed that all human CC samples expressed only the SM-A isoform. There was a predominance of SM2 isoform mRNA relative to SM1 across all samples, with a mean of 63.8%, which correlated with protein analysis by gel electrophoresis. A statistically significant difference was found between patients who had undergone previous pelvic surgery, radiation, or both and those who did not. The ratio of LC(17b) to LC(17a) was approximately 1:1 for all patients, with a mean of 48.9% LC(17b). Statistical difference was seen in the relative ratio of LC(17b) to LC(17a) among the group who failed conservative therapy with PDE5 inhibitors compared with all others. In conclusion, we determined the SM myosin isoform composition of human CC and present for the first time differences in relative myosin isoform expression among patients with several risk factors contributing to their cause of ED. Our data reflect the fact that alternative splicing events in the MHC and 17 kDa MLC pre-mRNA may be a possible molecular mechanism involved in the altered contractility of the CCSM in patients with ED.

OBJECTIVE To investigate whether variable coding sequence protein A1 (Vcsa1) is down-regulated in rat models of diabetes and ageing, and to investigate the role of Vcsa1 in erectile function, as Vcsa1 is the most down-regulated gene in the corpora of a rat model of neurogenic erectile dysfunction (ED). MATERIALS AND METHODS Quantitative reverse-transcriptase polymerase-chain reaction was used to determine Vcsa1 expression in the corpora of rats in three models of ED, i.e. streptozotocin-induced diabetes, retired breeder (old), and neurogenic (bilaterally ligated cavernosal nerves), and in control rats. To confirm a physiological role of Vcsa1 in erectile function, we carried out gene transfer studies using a plasmid in which Vcsa1 was expressed from a cytomegalovirus promoter (pVAX-Vcsa1). This plasmid was injected intracorporally into old rats, and the effect on physiology of corporal tissue was analysed by intracorporal/blood pressure (ICP/BP) measurement and histological analysis, and compared with the effects of a positive control plasmid (pVAX-hSlo, which we previously reported to restore erectile function in diabetic and ageing rats) and a negative control plasmid (pVAX). RESULTS In each rat model of ED there was a significant down-regulation of the Vcsa1 transcript of at least 10-fold in corporal tissue. Remarkably, intracorporal injection with 80 µg pVAX-Vcsa1 caused priapism, as indicated by visible prolonged erection, histological appearance, and elevated resting ICP/BP. Lower doses of pVAX-Vcsa1 (5 and 25 µg) increased ICP/BP over that in untreated controls. CONCLUSION These results show that Vcsa1 has a role in erectile function and might be a molecular marker for organic ED. The role of Vcsa1 in erectile function suggests that it could represent a novel therapeutic target for treating ED.

OBJECTIVE To establish the methods, feasibility and utility of evaluating the impact of diabetes on bladder and erectile function in the same rat, as more than half of diabetic patients have bladder dysfunction, and half of diabetic men have erectile dysfunction, but the severity of coincident disease has not been rigorously assessed. MATERIALS AND METHODS In all, 16 F-344 rats had diabetes induced by streptozotocin (STZ), and were divided into insulin-treated (five) and untreated (11), and compared with age-matched controls (10), all assessed in parallel. All STZ rats were diabetic for 8–11 weeks. Cystometric studies were conducted on all rats, with cavernosometric studies conducted on a subset of rats. RESULTS There were insulin-reversible increases in the following cystometric variables; bladder weight, bladder capacity, micturition volume, residual volume, micturition pressure and spontaneous activity (P

The promise of gene therapy to treat diseases remains largely unfulfilled. Past setbacks and the complexity of the delivery systems used, in terms of both targeting the appropriate cells and inducing expression of products at therapeutic levels, thus far have prevented significant success for gene therapy. Smooth muscle disorders represent a unique target for gene therapy. In many cases, smooth muscle is readily accessible and, to induce a therapeutic effect, will not require very high levels of gene product expression. This allows a lower efficiency of gene transfer to be successful. With these important features in mind, we believe that naked DNA transfer of potassium ion channels represents a novel and successful way to treat smooth muscle disorders. Herein, we present a rationale for treating erectile dysfunction, a smooth muscle disorder of the cavernosal bodies of the penis, with naked DNA gene transfer therapy. By inserting the hSlo gene, which codes for Maxipotassium channels, into smooth muscle cells, we can improve smooth muscle relaxation in the corporal bodies and thus improve erectile function. This method of gene transfer has proven to be safe and effective for erectile dysfunction, and human trials are ongoing.

Objectives To perform a comparison to determine which of two methods of partial urethral ligation produces the most consistent outcome and fewest side effects. Such a study has not been previously reported. Partial urethral ligation is a means of causing reproducible bladder outlet obstruction. In the male rat model, partial urethral obstruction can be performed either by perineal incision and bulbous urethral ligation or retropubic incision and midprostatic obstruction. Methods Fifteen male Sprague-Dawley rats were studied. Five were selected for bulbous urethral obstruction through a perineal incision, five for midprostatic obstruction using a retropubic approach, and five for a sham operation through a perineal incision. Results The operative time was shorter and morbidity lower with the perineal approach compared with the retropubic approach. Inflammation or infection, or both, were seen in the prostate, bladder, proximal urethra, ureters, and kidneys in the rats in which a midprostatic obstruction was performed. The proximal urethra and prostate were mildly inflamed in those rats that underwent bulbous obstruction. Sham-operated rats exhibited mild prostatitis only. Conclusions The perineal approach to the bulbous urethra is the method of choice for creating a partial urethral obstruction model of bladder outlet obstruction in the male rat.

Objective: To test the safety of a single intracavernous injection of a plasmid vector (hMaxi-K) that expresses the hSlo gene, that encodes the a-subunit of the Maxi-K channel, for the treatment of erectile dysfunction (ED). Methods: Six men, thus far have fulfilled the entry criteria of the protocol and had gene transfer with hMaxi-K. Three received a dose of 500 mg and three received a dose 1000 mg of the gene product, injected intracavernously as naked DNA. Dosing at 5000 mg and higher is planned. Results: The primary end point of the phase I trial is safety. No drug-related adverse events or significant laboratory changes have occurred after the gene transfer. Moreover, there is no evidence of the gene in semen at the one copy per mg total DNA in any of the participants. Conclusion: Preliminary results indicate that, in a single dose escalation study, ion channel gene transfer with hMaxi-K can be administered safely to men with ED without adverse events. # 2005 Elsevier B.V. All rights reserved.

To evaluate penile sensory thresholds in neonatally circumcised and uncircumcised men.We evaluated 125 patients, 62 uncircumcised men and 63 neonatally circumcised men. All patients completed the Erectile Function domain of the International Index of Erectile Function questionnaire. Of the 125 patients, 29 (International Index of Erectile Function score of between 25 and 30) were placed in the functional group, and 96 in the dysfunctional group. The patients were tested on the dorsal midline glans of the penis (foreskin retracted). Quantitative somatosensory testing was performed and included vibration, pressure, spatial perception, and warm and cold thermal thresholds.In the functional group, t-test analysis demonstrated a significant (P0.001) difference, with worse vibration and better pressure sensation for uncircumcised men. When controlling for age, hypertension, and diabetes, all t-test significance was lost. In the dysfunctional group, circumcised men (49 +/- 16 years) were significantly younger (P0.01) than uncircumcised men (56 +/- 13 years). For the dysfunctional group, t-test analysis also demonstrated worse vibration sensation for uncircumcised men (P0.01). Again, when controlling for age, hypertension, and diabetes, all t-test significance was lost.We present a comparative analysis of uncircumcised and circumcised men using a battery of quantitative somatosensory tests that evaluate the spectrum of small to large axon nerve fiber function. Our study controlled for factors, including age, erectile function status, diabetes, and hypertension, that have been shown to alter neurologic testing. In our study of neonatally circumcised men, we demonstrated that circumcision status does not significantly alter the quantitative somatosensory testing results at the glans penis.

OBJECTIVE To investigate the feasibility of perineal radical prostatectomy (RP) in renal transplant recipients with localized prostate cancer. PATIENTS AND METHODS The study comprised seven consecutive renal transplant patients who had a perineal RP between May 1991 and February 2004. All available clinicopathological data were reviewed. Results All seven patients successfully tolerated RP with no major complications. The mean (sd, range) age at surgery was 62.3 (2.5, 55–74) years and the mean interval from renal transplant to RP 86.5 (25.25, 24–192) months. There was no evidence of increased blood loss, operative duration, transfusion requirement, hospital stay or deterioration of graft function. The presence of an allograft did not alter the surgical approach or management of the patients after RP. The mean follow-up was 22 (2–130) months and all seven patients were followed. One patient had evidence of biochemical recurrence with no radiographic evidence of metastatic disease. Serum prostate-specific antigen was undetectable in the remaining patients. CONCLUSION A perineal RP in renal transplant recipients for treating localized prostate cancer offers many advantages over other treatments.

To evaluate the influence of intensive laparoscopic skills training and self-critical video review on the learning curve for laparoscopic radical prostatectomy (LRP).The initial 40 patients who underwent a transperitoneal LRP (groups 1-4) and the subsequent 20 who underwent LRP by the extraperitoneal approach (group 5) were studied. Eight weeks prior to initiating the LRP program, intensive laparoscopic skills training at a minimally invasive surgery center was undertaken for an average of 4 hours per week. This self-training was continued for 12 weeks into the program, with self-critical review of videotapes of each procedure. The groups were compared with respect to total operative time (ORT), anastomosis time, and blood loss.There were significant differences in the ORT and anastomosis times between each of the first two groups and the last two groups (P0.001). The learning curve for ORT was overcome after approximately 35 cases, as there were no significant differences in ORT between group 3 and the subsequent groups. The anastomosis took longer to master, as significant time decreases were observed up to group 4, after which, the mean reached a plateau (group 4 v 5 P = NS). The differences in blood loss were not significant. Overall, there were 7 intraoperative (12.7%) and 8 postoperative (14.5%) complications.The use of similar facilities and training tools can help overcome the steep learning curve of LRP. Longer follow-up is needed to assess these means of attaining better functional results after LRP.

Introduction Clinical trials on sexual dysfunctions in women are limited in spite of the fact that sexual dysfunctions are likely more common in women than in men. Currently there are no medications approved for treatment in women, and limited data on drug efficacy or psychological efficacy in well-controlled studies. Aim To provide recommendations/guidelines concerning state-of-the-art knowledge for the research design and outcome assessment standards for clinical trials in women's sexual dysfunctions. Methods An International Consultation in collaboration with the major urology and sexual medicine associations assembled over 200 multidisciplinary experts from 60 countries into 17 committees. Committee members established specific objectives and scopes for various male and female sexual medicine topics. The recommendations concerning state-of-the-art knowledge in the respective sexual medicine topic represent the opinion of experts from five continents developed in a process over a 2-year period. Concerning the Standards for Clinical Trials in Women's Sexual Dysfunctions Committee, there were seven experts from two countries. Main Outcome Measure Expert opinion was based on grading of evidence-based medical literature, widespread internal committee discussion, public presentation and debate. Results A comprehensive update was created which included references and recommended guidelines for rationale and design of clinical trials, study populations, outcome assessments, protocol design and implementation, data analysis and reporting, as well as ethical and clinical issues related to sexual dysfunction research. Conclusions There is a need for more research in developing standards to be used when performing clinical trials and outcomes assessment research in sexual dysfunctions of women.

Radical perineal prostatectomy was historically the surgical treatment of choice for localized adenocarcinoma of the prostate until the 1980s when radical retropubic prostatectomy began to gain popularity. Nevertheless, the perineal approach possesses advantages that prompt resurgence in the interest of this classic operation. We review the relevant anatomy and our modified technique of performing a successful radical perineal prostatectomy.The English literature pertaining to the different surgical approaches to radical perineal prostatectomy was reviewed through PubMed. Attention was paid to its indications, anatomical significance and various surgical techniques.Studies demonstrate no difference in the incidence of positive surgical margins and biochemical recurrence between radical retropubic and perineal prostatectomies. Furthermore, the perineal approach avoids the dorsal venous complex and better facilitates the vesicourethral anastomosis in the face of minimal pain and requirement for transfusion. We use a modified Belt approach, aiming to yield the most optimal outcome with minimal morbidity. A meticulous anatomical approach is warranted if complications such as rectal injury, incontinence and erectile dysfunction are to be minimized.With careful preoperative evaluation, selected patients should benefit from radical perineal prostatectomy for the management of localized prostate cancer. Familiarity with this specialized technique should be an immeasurable addition to any armamentarium in the therapy of prostatic diseases.

Quantitative somatosensory testing, including vibration, pressure, spatial perception and thermal thresholds of the penis, has demonstrated neuropathy in patients with a history of erectile dysfunction of all etiologies. We evaluated which measurement of neurological function of the penis was best at predicting erectile dysfunction and examined the impact of location on the penis for quantitative somatosensory testing measurements.A total of 107 patients were evaluated. All patients were required to complete the erectile function domain of the International Index of Erectile Function (IIEF) questionnaire, of whom 24 had no complaints of erectile dysfunction and scored within the "normal" range on the IIEF. Patients were subsequently tested on ventral middle penile shaft, proximal dorsal midline penile shaft and glans penis (with foreskin retracted) for vibration, pressure, spatial perception, and warm and cold thermal thresholds.Mixed models repeated measures analysis of variance controlling for age, diabetes and hypertension revealed that method of measurement (quantitative somatosensory testing) was predictive of IIEF score (F = 209, df = 4,1315, p0.001), while site of measurement on the penis was not. To determine the best method of measurement, we used hierarchical regression, which revealed that warm temperature was the best predictor of erectile dysfunction with pseudo R(2) = 0.19, p0.0007. There was no significant improvement in predicting erectile dysfunction when another test was added. Using 37C and greater as the warm thermal threshold yielded a sensitivity of 88.5%, specificity 70.0% and positive predictive value 85.5%.Quantitative somatosensory testing using warm thermal threshold measurements taken at the glans penis can be used alone to assess the neurological status of the penis. Warm thermal thresholds alone offer a quick, noninvasive accurate method of evaluating penile neuropathy in an office setting.

PURPOSE The use of estrogen replacement therapy for treating postmenopausal urinary incontinence is a controversial topic. We examined the behavioral, cystometric and histological changes that occur with long-term estrogen depletion and supplementation in rat bladders to determine the role of menopause in lower urinary tract dysfunction. MATERIALS AND METHODS A total of 40 female Sprague-Dawley rats were placed into 1 of 3 groups, including bilateral ovariectomy, bilateral ovariectomy plus estrogen replacement and control. The estrogen replaced group received a 0.25 mg. 16-week sustained release pellet (Innovative Research of America, Sanasota, Florida) placed subcutaneously. After surgery voiding frequency and volume were measured in 24-hour periods by placing animals in metabolic cages. After 16 weeks the rats underwent catheterization and continuous cystometry. The bladder was then removed and stained with Gomori trichrome. The collagen-to-smooth muscle density ratio was calculated for each specimen using current imaging software. RESULTS There was no significant difference in voiding patterns in the 3 groups, as measured by volume and voiding frequency. Cystometric data showed a trend toward higher voiding pressure, threshold pressure, baseline pressure and mean inter-voiding pressure in the ovariectomy group compared with the estrogen and control groups, although there was no statistical significance. Histological studies showed a higher mean collagen-to-smooth muscle ratio plus or minus standard deviation in the ovariectomy group (0.807 +/- 0.204) than in the ovariectomy plus estrogen replacement (0.709 +/- 0.118) and control (0.700 +/- 0.129) groups (p0.05). Furthermore, when histological and cystometric data were compared for individual samples, we found a direct correlation of mean inter-voiding pressure (a measure of bladder instability) with the collagen-to-smooth muscle ratio (p0.05). CONCLUSIONS Long-term estrogen replacement is beneficial for treating postmenopausal urinary incontinence.

Initiation, maintenance, and modulation of corporal smooth muscle tone are critically dependent upon agonist-induced changes in intracellular calcium levels and mobilization as well as transmembrane calcium flux. The transient control of myocyte excitability and contractility at the cellular level is inextricably linked to membrane potential, which, in turn, is modulated by potassium ion efflux through one of the four known corporeal smooth muscle potassium ion channels. Corporal tissue responses are subsequently coordinated by means of the movement of intracellular second messenger molecules (i.e., IP3, cAMP, cGMP) and ions (i.e., K+ and Ca2+) among the corporal myocytes by means of intercellular communication through gap junction channels. Knowledge of the critical contribution of these interlinking cellular (nonjunctional ion channels [e.g., maxi-K]) and tissue (gap junction channels [e.g., connexin 43]) systems to the modulation of erectile capacity has provided the scientific rationale for the promulgation of the successful preclinical testing of hSlo ion channel gene therapy for the normalization of erectile status in both aged and diabetic rats.

Purpose: We evaluated the effects of vitamin A and E supplementation alone or in combination with non-steroidal anti-inflammatory drugs (NSAIDs) on the development of inflammation in an animal model of ascending pyelonephritis.Materials and Methods: Ascending pyelonephritis was induced in adult rats by surgical bladder inoculation with P-pili-forming Escherichia coli. Treatment of pyelonephritic rats was initiated at 72 hours post-infection. Treatment groups included no treatment, or a five day regimen of antibiotic only, antibiotic plus vitamins A and E, or antibiotic, vitamins and either of two NSAIDs. Kidneys were harvested at six weeks post-infection and assessed for histopathologic inflammation.Results: Antibiotic treatment of pyelonephritic rats with vitamins A and E alone or in combination with NSAIDs resulted in significantly less kidney inflammation, as compared with untreated rats or rats treated with antibiotic alone. There was no significant difference in inflammation between animals t...

Modulation of the sexual behavior of male rats by the anxiolytic buspirone (S-20499) and its analog gepirone were compared to the effects of 8-OH-DPAT (or DPAT, a selective 5-HT1A reference agonist), and BMY-7378 (a selective 5-HT1A partial agonist). Long-Evans rats were used; modulation of copulatory behavior and alteration of penile reflexes were examined. Modulation of copulatory behavior was assessed by three indices: frequency and length of intromission, and latency of ejaculation. DPAT, at doses of 1-8 mg/kg, reduced these three indices in a time dependent manner such that the effects peaked at 45 min and normalized at 90 min. The dose-effect relationship (assessed 45 min after DPAT injection) is bell-shaped with an ED50 approximately 1 mg/kg on the ascending limb of the curve. The effects of buspirone (2 mg/kg) and gepirone (2 mg/kg) on copulatory behavior were indistinguishable from control. BMY-7378 alone and in combination with these other 5-HT1A agonists reduced copulatory behavior, though not statistically significant. Penile reflexes, including number of erections, cups and flips, were inhibited by these agents: DPAT>buspirone>gepirone (inactive at 2 mg/kg). Furthermore, the latency period to erection was at least doubled by DPAT (2 mg/kg). Buspirone and gepirone, however, reduced the latency period to erection. BMY-7378 inhibited penile reflexes when administered alone and even more in combination with DPAT or buspirone. Two butyrophenone analogs, spiperone (a 5-HT1A and dopamine D2 antagonist) and haloperidol (a D2 antagonist), were also tested for their interaction with DPAT. Both of these drugs (at 0.25 mg/kg, 60 min after administration) reduced all indices of penile reflexes and copulation. Furthermore, in combination with DPAT (2 mg/kg, 45 min), the effects were synergistic such that sexual activity came nearly to a standstill. These opposing effects on putatively brain originated copulatory behavior and spinal mediated penile reflexes indicate that the effects of buspirone and DPAT on sexual behavior in the male rat may be possible at different parts of the central nervous system. If a tentative shared target site by DPAT and buspirone is the 5-HT1A receptor, than the same 5-HT receptor sub-type at different locations (brain, raphe nuclei, spinal cord and autonomic ganglia) may modulate rat sexual behavior in opposing ways.

With the patch clamp method we demonstrate a stretch-sensitive Cl- currents as well as stretch-sensitive Cl- channels in a small group (5%,n 117) of cultured human corpus cavernosal muscle cells. The current and the channel activities had the following characteristics: (1) Their equilibrium potentials changed with extracellular Cl- concentration close to that predicted by Nernst equation provided that the relevant channels had high permeability to Cl- but low permeability to acetate ions; (2) They were blocked by mM concentrations of Zn2+; (3) The i-v relation of single channel current was almost linear for holding potentials varied from -70 to +60 mV; and (4) The channels had unitary conductances of approximately 140-170 pS.

Purpose: Published studies on the epidemiology of erectile dysfunction and the physiology/pathophysiology of erectile function are reviewed.Materials and Methods: A literature search of more than 400 studies of the epidemiology and pathophysiology of impotence and erectile dysfunction published during the last 3 decades was conducted and the most pertinent articles are discussed.Results: It has been estimated that the prevalence of erectile dysfunction of all degrees is 52% in men 40 to 70 years old, with higher rates in those older than 70 years. Erectile dysfunction has a significant negative impact on quality of life. Risk factors for erectile dysfunction include aging, chronic illnesses, various medications and cigarette smoking. A nitric oxide/cyclic guanosine monophosphate mechanism has an important role in mediating the corporal smooth muscle relaxation necessary for erectile function. Other mechanisms involving neuropeptides, gap junctions and ion channels also may modulate corporal smooth...

From 1980 to July 1997 sixty-one male-to-female gender transformation surgeries were performed at our university center by one author (A.M.). Data were collected from patients who had surgery up to 1994 (n = 47) to obtain a minimum follow-up of 3 years; 28 patients were contacted. A mail questionnaire was supplemented by personal interviews with 11 patients and telephone interviews with remaining patients to obtain and clarify additional information. Physical and functional results of surgery were judged to be good, with few patients requiring additional corrective surgery. General satisfaction was expressed over the quality of cosmetic (normal appearing genitalia) and functional (ability to perceive orgasm) results. Follow-up showed satisfied who believed they had normal appearing genitalia and the ability to experience orgasm. Most patients were able to return to their jobs and live a more satisfactory social and personal life. One significant outcome was the importance of proper preparation of patients for surgery and especially the need for additional postoperative psychotherapy. None of the patients regretted having had surgery. However, some were, to a degree, disappointed because of difficulties experienced postoperatively in adjusting satisfactorily as women both in their relationships with men and in living their lives generally as women. Findings of this study make a strong case for making a change in the Harry Benjamin Standards of Care to include a period of postoperative psychotherapy.

During male-to-female gender reassignment surgery we used an abdominal pedicled inverted penile skin technique to create a vagina and extra folds of skin to create a clitoral hood. Although patients had orgasms with that technique, there was a general request for the placement of a sensate, erectile clitoris. Therefore, in 10 such patients undergoing transsexual surgery a neoclitoris was fashioned from the glans penis. Surgical technique and results are described.From 1980 to 1995, 57 male-to-female gender surgeries were performed at our university center. In the last 10 such patients undergoing transsexual surgery a neoclitoris was created. Glans volume is reduced to match that of a normal size clitoris and the entire length of the dorsal neurovascular bundle is preserved. The neoclitoris is placed through a buttonhole in the newly constructed introitus. There was minimal bleeding from the bundle intraoperatively.In 8 of 10 patients the neoclitoris remained intact postoperatively with good sensation to touch, vibration and light pressure (sexual sensation). The cosmetic and functional appearance was that of a normal clitoris, and patients were satisfied. In 2 patients the results were not satisfactory because of necrosis of the neoclitoris.This technique has excellent functional and cosmetic results in the majority of patients. Preservation of the somatic, tactile and vibratory sensation of the neoclitoris is surgically practical, and can afford the patient the possibility of a more functional outcome of gender reassignment surgery.

Increases in cytosolic calcium levels trigger smooth muscle contraction while nuclear calcium increases are thought to regulate gene expression. Endothelin-1 (ET-1) affects both. The goal of these studies was to further investigate the importance of ET-1 to corporal physiology by examining its actions on proliferation and immediate early gene (IEG) expression in cultured human corporal smooth muscle cells.Early passage (1-3) smooth muscle cells were grown in culture and exposed to either phenylephrine (PE) or ET-1 in the absence and presence of serum, the ET(A) or ET(B) selective antagonist BQ123 or IRL1038, or the L-type Ca2+ channel blocker, verapamil. Cell proliferation was assessed with a hemocytometer. The effects of ET-1 on c-myc and c-fos were evaluated using Northern blot analysis. Parametric or nonparametric statistics were used as appropriate.Addition of ET-1 (100 nM) to serum-starved cultured corporal smooth muscle cells was associated with a nearly 2-fold increase in cell number, as well as 2 to 6-fold increases in c-myc and c-fos levels. Cellular proliferation was inhibited by ET(A)- or ET(B)-receptor subtype blockade with BQ123 (1 microM) or IRL1038 (1 microM), respectively, or blockade of Ca2+ channels with verapamil (10 microM). PE (3 microM) had no detectable effect on smooth muscle proliferation.Cell proliferation was mediated by activation of the ET(A) and ET(B) receptor subtypes, dependent on transmembrane Ca2+ flux, and correlated with significant increases in c-myc and c-fos mRNA levels. These studies extend previous observations to indicate the potential pleotropic actions of this peptide in the regulation of human corporal smooth muscle physiology in vivo.

The Ca2+-sensitive K+channel (maxi-K+) is an important modulator of corporal smooth muscle tone. The goal of these studies was twofold: 1) to determine the feasibility of transfecting corporal smooth muscle cells in vivo with the hSlo cDNA, which encodes for the human smooth muscle maxi-K+channel, and 2) to determine whether transfection of the maxi-K+channel would affect the physiological response to cavernous nerve stimulation in a rat model in vivo. Intracorporal microinjection of pCMVβ/Lac Z DNA in 10-wk-old rats resulted in significant incorporation and expression of β-galactosidase activity in 10 of 12 injected animals for up to 75 days postinjection. Moreover, electrical stimulation of the cavernous nerve revealed that, relative to the responses obtained in age-matched control animals ( N = 12), intracavernous injection of naked pcDNA/ hSlo DNA was associated with a statistically significant elevation in the mean amplitude of the intracavernous pressure response at all levels of current stimulation (range 0.5–10 mA) at both 1 mo ( N= 5) and 2 mo ( N = 8) postinjection. Furthermore, qualitatively similar observations were made at 3 mo ( N = 2) and 4 mo ( N = 2) postinjection. These data indicate that naked hSlo DNA is quite easily incorporated into corporal smooth muscle and, furthermore, that expression is sustained for at least 2 mo in corporal smooth muscle cells in vivo. Finally, after expression, hSlo is capable of measurably altering nerve-stimulated penile erection. Taken together, these data provide compelling evidence for the potential utility of gene therapy in the treatment of erectile dysfunction.