Cavernous nerve mapping methods for radical prostatectomy.

INTRODUCTION: Preserving the cavernous nerves, the main autonomic nerve supply of the penis, is a major challenge of radical prostatectomy. Cavernous nerve injury during radical prostatectomy predominantly accounts for post-radical prostatectomy erectile dysfunction. The cavernous nerve is a bilateral structure that branches in a weblike distribution over the prostate surface and varies anatomically in individuals, such that standard nerve-sparing methods do not sufficiently sustain penile erection ability. As a consequence, researchers have focused on developing personalized cavernous nerve mapping methods applied to the surgical procedure aiming to improve postoperative sexual function outcomes. OBJECTIVES: We provide an updated overview of preclinical and clinical data of cavernous nerve mapping methods, emphasizing their strengths, limitations, and future directions. METHODS: A literature review was performed via Scopus, PubMed, and Google Scholar for studies that describe cavernous nerve mapping/localization. RESULTS: Several cavernous nerve mapping methods have been investigated based on various properties of the nerve structures including stimulation techniques, spectroscopy/imaging techniques, and assorted combinations of these methods. More recent methods have portrayed the course of the main cavernous nerve as well as its branches based on real-time mapping, high-resolution imaging, and functional imaging. However, each of these methods has distinctive limitations, including low spatial accuracy, lack of standardization for stimulation and response measurement, superficial imaging depth, toxicity risk, and lack of suitability for intraoperative use. CONCLUSION: While various cavernous nerve mapping methods have provided improvements in identification and preservation of the cavernous nerve during radical prostatectomy, no method has been implemented in clinical practice due to their distinctive limitations. To overcome the limitations of existing cavernous nerve mapping methods, the development of new imaging techniques and mapping methods is in progress. There is a need for further research in this area to improve sexual function outcomes and quality of life after radical prostatectomy.

Testosterone Deficiency in Sickle Cell Disease: Recognition and Remediation.

Hypogonadism is common in men with sickle cell disease (SCD) with prevalence rates as high as 25%. Testicular failure (primary hypogonadism) is established as the principal cause for this hormonal abnormality, although secondary hypogonadism and compensated hypogonadism have also been observed. The underlying mechanism for primary hypogonadism was elucidated in a mouse model of SCD, and involves increased NADPH oxidase-derived oxidative stress in the testis, which reduces protein expression of a steroidogenic acute regulatory protein and cholesterol transport to the mitochondria in Leydig cells. In all men including those with SCD, hypogonadism affects physical growth and development, cognition and mental health, sexual function, as well as fertility. However, it is not understood whether declines in physical, psychological, and social domains of health in SCD patients are related to low testosterone, or are consequences of other abnormalities of SCD. Priapism is one of only a few complications of SCD that has been studied in the context of hypogonadism. In this pathologic condition of prolonged penile erection in the absence of sexual excitement or stimulation, hypogonadism exacerbates already impaired endothelial nitric oxide synthase/cGMP/phosphodiesterase-5 molecular signaling in the penis. While exogenous testosterone alleviates priapism, it disadvantageously decreases intratesticular testosterone production. In contrast to treatment with exogenous testosterone, a novel approach is to target the mechanisms of testosterone deficiency in the SCD testis to drive endogenous testosterone production, which potentially decreases further oxidative stress and damage in the testis, and preserves sperm quality. Stimulation of translocator protein within the transduceosome of the testis of SCD mice reverses both hypogonadism and priapism, without affecting intratesticular testosterone production and consequently fertility. Ongoing research is needed to define and develop therapies that restore endogenous testosterone production in a physiologic, mechanism-specific fashion without affecting fertility in SCD men.

Mirabegron improves erectile function in men with overactive bladder and erectile dysfunction: a 12-week pilot study.

Phosphodiesterase type 5 inhibitors (PDE5i) is the only approved oral treatment for erectile dysfunction (ED) in the US, and alternative management remains necessary when this treatment fails or is contraindicated. Targeting other pathways than the NO-cGMP pathway and/or combining this approach with PDE5i may introduce new treatments for men who are unresponsive to PDE5i. This study aims to evaluate whether Mirabegron improves erectile function in men with concurrent overactive bladder and mild to moderate ED. Twenty subjects, 40-70 years old, registering International Index of Erectile Function (IIEF) score 11-25 and International Prostate Symptom Score 8-20, were treated with Mirabegron therapy for 12 weeks. Study participants were re-administered IIEF and OAB-q questionnaires on weeks 2, 4, 8, and 12 and assessed for adverse events. The primary and secondary endpoints were an increase in the IIEF-5 score of 4 units and a decrease in the Overactive Bladder questionnaire (OAB-q) symptom severity score of 10 units between study time points. Thirteen men completed the 12-week study. Mirabegron treatment improved the IIEF-5 scores in five patients (38.4%) by 4 points or more, whereas IIEF-5 scores were not affected by Mirabegron treatment in eight patients (61.5%). There were no clinically relevant decreases in the IIEF-5 score. Significant improvements were observed in intercourse satisfaction at week eight compared to baseline (p = 0.01). Orgasmic function and sexual desire were not affected by Mirabegron treatment. As expected, Mirabegron treatment reduced OAB symptoms based on OAB-q short form (p = 0.006) and OAB-q total health-related quality of life (HRQL) scores compared to baseline (p = 0.03). Residual bladder volumes were not affected by treatment. No serious side effects were reported during the study period. This study suggests that Mirabegron may improve both EF and OAB-related symptoms in some individuals without causing serious adverse events.

TSPO ligand FGIN-1-27 controls priapism in sickle cell mice via endogenous testosterone production.

Priapism, a prolonged penile erection in the absence of sexual arousal, is common among patients with sickle cell disease (SCD). Hypogonadism is also common in patients with SCD. While the administration of exogenous testosterone reverses hypogonadism, it is contraceptive. We hypothesized that the stimulation of endogenous testosterone production decreases priapism by normalizing molecular signaling involved in penile erection without decreasing intratesticular testosterone production, which would affect fertility. Treatment of SCD mice with FGIN-1-27, a ligand for translocator protein (TSPO) that mobilizes cholesterol to the inner mitochondrial membrane, resulted in eugonadal levels of serum testosterone without decreasing intratesticular testosterone production. Normalized testosterone levels, in turn, decreased priapism. At the molecular level, TSPO restored phosphodiesterase 5 activity and decreased NADPH oxidase-mediated oxidative stress in the penis, which are major molecular signaling molecules involved in penile erection and are dysregulated in SCD. These results indicate that pharmacologic activation of TSPO could be a novel, targetable pathway for treating hypogonadal men, particularly patients with SCD, without adverse effects on fertility.

NO-Releasing Nanoparticles Ameliorate Detrusor Overactivity in Transgenic Sickle Cell Mice via Restored NO/ROCK Signaling.

Sickle cell disease (SCD) is associated with overactive bladder (OAB). Detrusor overactivity, a component of OAB, is present in an SCD mouse, but the molecular mechanisms for this condition are not well-defined. We hypothesize that nitric oxide (NO)/ ras homolog gene family (Rho) A/Rho-associated kinase (ROCK) dysregulation is a mechanism for detrusor overactivity and that NO-releasing nanoparticles (NO-nps), a novel NO delivery system, may serve to treat this condition. Male adult SCD transgenic, combined endothelial NO synthases (eNOSs) and neuronal NOS (nNOS) gene-deficient (dNOS-/-), and wild-type (WT) mice were used. Empty nanoparticle or NO-np was injected into the bladder, followed by cystometric studies. The expression levels of phosphorylated eNOS (Ser-1177), protein kinase B (Akt) (Ser-473), nNOS (Ser-1412), and myosin phosphatase target subunit 1 (MYPT1) (Thr-696) were assessed in the bladder. SCD and dNOS-/- mice had a greater (P < 0.05) number of voiding and nonvoiding contractions compared with WT mice, and they were normalized by NO-np treatment. eNOS (Ser-1177) and AKT (Ser-473) phosphorylation were decreased (P < 0.05) in the bladder of SCD compared with WT mice and reversed by NO-np. Phosphorylated MYPT1, a marker of the RhoA/ROCK pathway, was increased (P < 0.05) in the bladder of SCD mice compared with WT and reversed by NO-np. nNOS phosphorylation on positive (Ser-1412) regulatory site was decreased (P < 0.05) in the bladder of SCD mice compared with WT and was not affected by NO-np. NO-nps did not affect any of the measured parameters in WT mice. In conclusion, dysregulation of NO and RhoA/ROCK pathways is associated with detrusor overactivity in SCD mice; NO-np reverses these molecular derangements in the bladder and decreases detrusor overactivity. SIGNIFICANCE STATEMENT: Voiding abnormalities commonly affect patients with sickle cell disease (SCD) but are problematic to treat. Clarification of the science for this condition in an animal model of SCD may lead to improved interventions for it. Our findings suggest that novel topical delivery of a vasorelaxant agent nitric oxide into the bladder of these mice corrects overactive bladder by improving deranged bladder physiology regulatory signaling.

Mechanisms underlying priapism in sickle cell disease: targeting and key innovations on the preclinical landscape.

Introduction: Priapism is prolonged penile erection in the absence of sexual arousal or desire and is a devastating condition affecting millions of patients with sickle cell disease (SCD) globally. Available drug treatments for SCD-related priapism remain limited and have been primarily reactive rather than preventive. Hence, there is an unmet need for new drug targets and pharmacologic therapies.Areas covered: We examine the molecular mechanisms underlying SCD-associated priapism evaluated mostly in animal models. In mouse models of SCD, molecular defects of priapism operating at the cavernous tissue level include reduced tonic NO/cGMP signaling, elevated oxidative/nitrosative stress, vascular adhesion molecule derangements, excessive adenosine and opiorphin signaling, dysregulated vasoconstrictive RhoA/ROCK signaling, and testosterone deficiency. We discuss the consequences of downregulated cGMP-dependent phosphodiesterase type 5 (PDE5) activity in response to these molecular signaling derangements, as the main effector mechanism causing unrestrained cavernous tissue relaxation that results in priapism.Expert opinion: Basic science studies are crucial for understanding the underlying pathophysiology of SCD-associated priapism. Understanding the molecular mechanisms could unearth new therapeutic targets for this condition based on these mechanisms. Treatment options should aim to improve deranged erection physiology regulatory signaling to prevent priapism and potentially restore or preserve erectile function.

Dysregulated NO/PDE5 signaling in the sickle cell mouse lower urinary tract: Reversal by oral nitrate therapy.

AIMS: Nitric oxide (NO) has a critical, but not well understood, influence in the physiology of the lower urinary tract. We evaluated the effect of NO/phosphodiesterase (PDE)5 signaling in voiding dysfunction in the sickle cell disease (SCD) mouse, characterized by low NO bioavailability. MAIN METHODS: Adult SCD (Sickle) and wild-type (WT) male mice were treated daily with sodium nitrate (10 mM) or vehicle. After 18 days, blood was obtained for nitrite measurement, urethra was collected for organ bath study, and bladder and urethra were collected for Western blot analysis of PDE5 phosphorylation (Ser-92) (activated form). Non-anesthetized mice underwent evaluation of urine volume by void spot assay. eNOS phosphorylation (Ser-1177) and nNOS phosphorylation (Ser-1412) (positive regulatory sites) were evaluated in the bladder and urethra of untreated mice. KEY FINDINGS: Sickle mice exhibited decreased eNOS, nNOS, and PDE5 phosphorylation in the bladder and urethra, decreased plasma nitrite levels, increased relaxation of phenylephrine-contracted urethral tissue to an NO donor sodium nitroprusside, and increased total urine volume, compared with WT mice. Nitrate treatment normalized plasma nitrite levels, relaxation of urethra to sodium nitroprusside, PDE5 phosphorylation in the urethra and bladder, and urine volume in Sickle mice. SIGNIFICANCE: Derangement in PDE5 activity associated with basally low NO bioavailability in the bladder and urethra contributes to the molecular basis for voiding abnormalities in Sickle mice. Inorganic nitrate supplementation normalized voiding in Sickle mice through mechanisms likely involving upregulation of PDE5 activity. These findings suggest that interventions targeting dysregulatory NO/PDE5 signaling may ameliorate overactive bladder in SCD.

Urinary dysfunction in transgenic sickle cell mice: model of idiopathic overactive bladder syndrome.

Voiding abnormalities are common among the sickle cell disease (SCD) population, among which overactive bladder (OAB) syndrome is observed at rates as high as 39%. Although detrusor overactivity is the most common cause of OAB, its molecular pathophysiology is not well elucidated. The nitric oxide (NO) signaling pathway has been implicated in the regulation of lower genitourinary tract function. In the present study, we evaluated the role of the NO signaling pathway in voiding function of transgenic SCD mice compared with combined endothelial and neuronal NO synthase gene-deficient mice, both serving as models of NO deficiency. Mice underwent void spot assay and cystometry, and bladder and urethral specimens were studied using in vitro tissue myography. Both mouse models exhibited increased void volumes; increased nonvoiding and voiding contraction frequencies; decreased bladder compliance; increased detrusor smooth muscle contraction responses to electrical field stimulation, KCl, and carbachol; and increased urethral smooth muscle relaxation responses to sodium nitroprusside compared with WT mice. In conclusion, our comprehensive behavioral and functional study of the SCD mouse lower genitourinary tract, in correlation with that of the NO-deficient mouse, reveals NO effector actions in voiding function and suggests that NO signaling derangements are associated with an OAB phenotype. These findings may allow further study of molecular targets for the characterization and evaluation of OAB.

Phosphodiesterase type 5 in men with vasculogenic and post-radical prostatectomy erectile dysfunction: is there a molecular difference?

OBJECTIVES: To clarify the molecular basis of penile erection at the human level and distinguish the mechanisms underlying vasculogenic and post-radical prostatectomy (RP) erectile dysfunction (ED) subtypes. PATIENTS AND METHODS: Erectile tissue was obtained from men without history of ED who underwent penile surgery for Peyronie's disease (control group, n = 5) and from men with ED who underwent penile prosthesis implantation (n = 17). ED was categorized into vasculogenic (n = 8) and post-RP (n = 9) subtypes. Penile erectile tissue samples were collected for molecular analyses of protein expressions of neuronal and endothelial isoforms of nitric oxide synthase (nNOS and eNOS, respectively), phospho-nNOS (Ser-1412), phospho-eNOS (Ser-1177), phospho-protein kinase B (Ser-473), phosphodiesterase type 5 (PDE5), α-smooth muscle actin, phospho-myosin phosphatase target subunit 1, RhoA/Rho-associated protein kinase (ROCK)-α, ROCK-ß, 4-hydroxy-2-nonenal, and nNOS and eNOS uncoupling by Western blot. RESULTS: Vasculogenic ED was characterized by decreased eNOS protein expression and eNOS and nNOS phosphorylation on their activatory sites (Ser-1177 and Ser-1412, respectively), uncoupled eNOS, upregulated PDE5 protein expression, increased ROCK activity, and increased oxidative stress in erectile tissue. Post-RP ED was characterized by decreased nNOS protein expression, increased nNOS phosphorylation on its activatory site (Ser-1412), uncoupled nNOS, downregulated PDE5 protein expression, and increased oxidative stress in erectile tissue. CONCLUSION: The mechanisms of vasculogenic and post-RP ED in the human penis involve derangements in constitutive nitric oxide synthase function, PDE5 protein expression and ROCK activity, and increased oxidative stress, which conceivably provide a molecular basis for chronically reduced nitric oxide bioavailability and increased smooth muscle contraction contributing to erectile impairment. Selective differences in PDE5 protein expression suggest distinct molecular mechanisms are in play for these ED subtypes.

S-nitrosylation of NOS pathway mediators in the penis contributes to cavernous nerve injury-induced erectile dysfunction.

cGMP-independent nitric oxide (NO) signaling occurs via S-nitrosylation. We evaluated whether aberrant S-nitrosylation operates in the penis under conditions of cavernous nerve injury and targets proteins involved in regulating erectile function. Adult male Sprague-Dawley rats underwent bilateral cavernous nerve crush injury (BCNI) or sham surgery. Rats were given a denitrosylation agent N-acetylcysteine (NAC, 300 mg/kg/day) or vehicle in drinking water starting 2 days before BCNI and continuing for 2 weeks following surgery. After assessment of erectile function (intracavernous pressure), penes were collected for measurements of S-nitrosylation by Saville-Griess and TMT-switch assays and PKG-I function by immunoblotting of phospho (P)-VASP-Ser-239. Erectile function was decreased (P < 0.05) after BCNI, and it was preserved (P < 0.05) by NAC treatment. Total S-nitrosothiols and total S-nitrosylated proteins were increased (P < 0.05) after BCNI, and these were partially prevented by NAC treatment. S-nitrosylation of sGC was increased (P < 0.05) after BCNI, and it was prevented (P < 0.05) by NAC treatment. S-nitrosylation of eNOS was increased (P < 0.05) after BCNI, and showed a trend towards decrease by NAC treatment. Protein expression of P-VASP-Ser-239 was decreased (P < 0.05) after BCNI, and showed a trend towards increase by NAC treatment. In conclusion, erectile dysfunction following BCNI is mediated in part by S-nitrosylation of eNOS and its downstream signaling mediator GC, while denitrosylation protects erectile function by preserving the NO/cGMP signaling pathway.

cAMP-dependent post-translational modification of neuronal nitric oxide synthase neuroprotects penile erection in rats.

OBJECTIVES: To evaluate neuronal nitric oxide (NO) synthase (nNOS) phosphorylation, nNOS uncoupling, and oxidative stress in the penis and major pelvic ganglia (MPG), before and after the administration of the cAMP-dependent protein kinase A (PKA) agonist colforsin in a rat model of bilateral cavernous nerve injury (BCNI),which mimics nerve injury after prostatectomy. MATERIALS AND METHODS: Adult male Sprague-Dawley rats were divided into BCNI and sham-operated groups. Each group included two subgroups: vehicle and colforsin (0.1 mg/kg/day i.p.). After 3 days, erectile function (intracavernosal pressure) was measured and penis and MPG were collected for molecular analyses of phospho (P)-nNOS (Ser-1412 and Ser-847), total nNOS, nNOS uncoupling, binding of protein inhibitor of nNOS (PIN) to nNOS, gp91phox subunit of NADPH oxidase, active caspase 3, PKA catalytic subunit α (PKA-Cα; by Western blot) and oxidative stress (hydrogen peroxide [H2 O2 ] and superoxide by Western blot and microdialysis method). RESULTS: Erectile function was decreased 3 days after BCNI and normalized by colforsin. nNOS phosphorylation on both positive (Ser-1412) and negative (Ser-847) regulatory sites, and nNOS uncoupling, were increased after BCNI in the penis and MPG, and normalized by colforsin. H2 O2 and total reactive oxygen species production were increased in the penis after BCNI and normalized by colforsin. Protein expression of gp91phox was increased in the MPG after BCNI and was normalized by colforsin treatment. Binding of PIN to nNOS was increased in the penis after BCNI and was normalized by colforsin treatment. Protein expression of active Caspase 3 was increased in the MPG after BCNI and was normalized by colforsin treatment. Protein expression of PKA-Cα was decreased in the penis after BCNI and normalized by colforsin. CONCLUSION: Collectively, BCNI impairs nNOS function in the penis and MPG by mechanisms involving its phosphorylation and uncoupling in association with increased oxidative stress, resulting in erectile dysfunction. PKA activation by colforsin reverses these molecular changes and preserves penile erection in the face of BCNI.

cAMP-dependent regulation of RhoA/Rho-kinase attenuates detrusor overactivity in a novel mouse experimental model.

OBJECTIVE: To investigate detrusor function and cAMP activation as a possible target for detrusor overactivity in an experimental model lacking a key denitrosylation enzyme, S-nitrosoglutathione reductase (GSNOR). MATERIALS AND METHODS: GSNOR-deficient (GSNOR-/- ) (n = 30) and wild-type (WT) mice (n = 26) were treated for 7 days with the cAMP activator, colforsin (1 mg/kg), or vehicle intraperitoneally. Cystometric studies or molecular analyses of bladder specimens were performed. Bladder function indices and expression levels of proteins that regulate detrusor relaxation (nitric oxide synthase pathway) or contraction (RhoA/Rho-kinase pathway) and oxidative stress were assessed. For statistical analysis the Student's t-test and one-way analysis of variance were used. RESULTS: GSNOR-/- mice had significantly higher (P < 0.05) voiding and non-voiding contraction frequencies compared to WT mice (Cohen's effect size values d = 1.82 and 2.52, respectively). Colforsin normalised these abnormalities (Cohen's effect size values d = 1.85 and 1.28, respectively). Western blot analyses showed an up-regulation of the RhoA/Rho-kinase pathway reflected by significantly higher (P < 0.05) phosphorylated myosin phosphatase target subunit 1 (P-MYPT-1) expression in GSNOR-/- mouse bladders, which was reversed by colforsin treatment. There was a higher level (P < 0.05) of gp91phox expression in the bladders of GSNOR-/- mice without significant change after colforsin treatment. Neuronal and endothelial nitric oxide synthase phosphorylation on Ser-1412 and Ser-1177, respectively, did not differ between GSNOR-/- and WT mouse bladders irrespective of colforsin treatment. CONCLUSION: Impaired denitrosylation is associated with detrusor overactivity, which is linked with upregulated RhoA/Rho-kinase signalling. Colforsin reverses physiological and molecular abnormalities. This study describes a novel model of detrusor overactivity and suggests a possible basis for its treatment.

Early-stage Type 2 Diabetes Mellitus Impairs Erectile Function and Neurite Outgrowth From the Major Pelvic Ganglion and Downregulates the Gene Expression of Neurotrophic Factors.

OBJECTIVE: To assess neurite sprouting and gene expression of neurotrophic factors, nerve markers, and apoptosis in the major pelvic ganglia (MPGs) of rats with type 2 diabetes mellitus (T2DM) as it relates to erectile function. MATERIALS AND METHODS: Male rats were fed high-fat diet for 2 weeks followed by 2 low-dose injections of streptozotocin (20 mg/kg). In 3 groups (controls, 3-week, or 5-week T2DM), erectile function was measured by ratios of intracavernosal pressure to mean arterial pressure after cavernous nerve stimulation. MPGs were harvested, and gene expressions of neurotrophic factor 3, nerve growth factor, glial cell line-derived neurotrophic factor, brain-derived neurotrophic factor, caspase-1, -3, -9, beta tubulin type III, and neuronal nitric oxide synthase were quantified by quantitative polymerase chain reaction. Additional MPGs were harvested and cultured in Matrigel. Neurite outgrowth from the MPG was evaluated at 48 hours after culture. RESULTS: Erectile function was significantly decreased in all rats with T2DM. Gene expressions of neurotrophic factor 3, nerve growth factor, glial cell line-derived neurotrophic factor, and brain-derived neurotrophic factor were slightly lower at 3 weeks and significantly lower at 5 weeks after T2DM induction. Gene expression of apoptotic markers caspase-1, -3, -9, and neuronal markers beta tubulin type III and neuronal nitric oxide synthase remained unchanged. Rats with T2DM had shorter neurite length and less neurite sprouting than did the control MPG. CONCLUSION: Early-stage T2DM downregulates neurotrophic factors, induces erectile dysfunction, and impairs MPG neurite outgrowth, suggesting that erectile dysfunction may be prevented by supplementing neurotrophic factors at early-stage T2DM.

Beneficial Effect of the Nitric Oxide Donor Compound 3-(1,3-Dioxoisoindolin-2-yl)Benzyl Nitrate on Dysregulated Phosphodiesterase 5, NADPH Oxidase, and Nitrosative Stress in the Sickle Cell Mouse Penis: Implication for Priapism Treatment.

Patients with sickle cell disease (SCD) display priapism, and dysregulated nitric oxide (NO) pathway may contribute to this condition. However, current therapies offered for the prevention of priapism in SCD are few. The 3-(1,3-dioxoisoindolin-2-yl)benzyl nitrate (compound 4C) was synthesized through molecular hybridization of hydroxyurea and thalidomide, which displays an NO-donor property. This study aimed to evaluate the effects of compound 4C on functional and molecular alterations of erectile function in murine models that display low NO bioavailability, SCD transgenic mice, and endothelial NO synthase and neuronal NO synthase double gene-deficient (dNOS-/) mice, focusing on the dysregulated NO-cGMP- phosphodiesterase type 5 (PDE5) pathway and oxidative stress in erectile tissue. Wild-type, SCD, and dNOS-/- mice were treated with compound 4C (100 µmol/kg/d, 3 weeks). Intracavernosal pressure in anesthetized mice was evaluated. Corpus cavernosum tissue was dissected free and mounted in organ baths. SCD and dNOS-/- mice displayed a priapism phenotype, which was reversed by compound 4C treatment. Increased corpus cavernosum relaxant responses to acetylcholine and electrical-field stimulation were reduced by 4C in SCD mice. Likewise, increased sodium nitroprusside-induced relaxant responses were reduced by 4C in cavernosal tissue from SCD and dNOS-/- mice. Compound 4C reversed PDE5 protein expression and reduced protein expressions of reactive oxygen species markers, NADPH oxidase subunit gp91phox, and 3-nitrotyrosine in penises from SCD and dNOS-/- mice. In conclusion, 3-week therapy with the NO donor 4C reversed the priapism in murine models that display lower NO bioavailability. NO donor compounds may constitute an additional strategy to prevent priapism in SCD.

Translational Perspective on the Role of Testosterone in Sexual Function and Dysfunction.

INTRODUCTION: The biological importance of testosterone is generally accepted by the medical community; however, controversy focuses on its relevance to sexual function and the sexual response, and our understanding of the extent of its role in this area is evolving. AIM: To provide scientific evidence examining the role of testosterone at the cellular and molecular levels as it pertains to normal erectile physiology and the development of erectile dysfunction and to assist in guiding successful therapeutic interventions for androgen-dependent sexual dysfunction. METHODS: In this White Paper, the Basic Science Committee of the Sexual Medicine Society of North America assessed the current basic science literature examining the role of testosterone in sexual function and dysfunction. RESULTS: Testosterone plays an important role in sexual function through multiple processes: physiologic (stimulates activity of nitric oxide synthase), developmental (establishes and maintains the structural and functional integrity of the penis), neural (development, maintenance, function, and plasticity of the cavernous nerve and pelvic ganglia), therapeutically for dysfunctional regulation (beneficial effect on aging, diabetes, and prostatectomy), and phosphodiesterase type 5 inhibition (testosterone supplement to counteract phosphodiesterase type 5 inhibitor resistance). CONCLUSION: Despite controversies concerning testosterone with regard to sexual function, basic science studies provide incontrovertible evidence for a significant role of testosterone in sexual function and suggest that properly administered testosterone therapy is potentially advantageous for treating male sexual dysfunction.

Transnitrosylation: A Factor in Nitric Oxide-Mediated Penile Erection.

INTRODUCTION: Nitric oxide (NO) signaling can be mediated not only through classic 3',5'-cyclic guanosine monophosphate but also through S-nitrosylation. However, the impact of S-nitrosylation on erectile function and in NO regulation and oxidative stress in the penis remains poorly understood. AIMS: To characterize the role of S-nitrosoglutathione reductase (GSNOR), a major regulator of S-nitrosylation homeostasis, on erection physiology and on endothelial NO synthase (eNOS) function and oxidative-nitrosative stress in the penis. METHODS: Adult GSNOR-deficient and wild-type (WT) mice were used. Erectile function was assessed in response to electrical stimulation of the cavernous nerve. Total NO in penile homogenates was measured by Griess reaction. Protein S-nitrosylation, eNOS phosphorylation on Ser-1177 (positive regulatory site), eNOS uncoupling, and markers of oxidative stress (4-hydroxy-2-nonenal, malondialdehyde, and nitrotyrosine) in the penis were measured by western blot. MAIN OUTCOME MEASURES: Erectile function, eNOS function, and oxidative stress in the penis of GSNOR-deficient mice. RESULTS: Erectile function was intact in GSNOR-deficient mice. Total S-nitrosylated proteins were increased (P < .05) in the GSNOR(-/-) compared with WT mouse penis. Although eNOS phosphorylation on Ser-1177 did not differ between the GSNOR(-/-) and WT mouse penises at baseline, electrical stimulation of the cavernous nerve increased (P < .05) phosphorylated eNOS in the WT mouse penis but failed to increase phosphorylated eNOS in the GSNOR(-/-) mouse penis. Total NO production was decreased (P < .05), whereas eNOS uncoupling, 4-hydroxy-2-nonenal, malondialdehyde, and nitrotyrosine were increased (P < .05) in the GSNOR-deficient mouse penis compared with the WT mouse penis. CONCLUSION: Transnitrosylation mechanisms play an important role in regulating NO bioactivity in the penis. Deficiency of GSNOR leads to eNOS dysfunction and increased oxidative damage, suggesting that homeostatic eNOS function in the penis is governed by transnitrosylation.

Basic Science Evidence for the Link Between Erectile Dysfunction and Cardiometabolic Dysfunction.

INTRODUCTION: Although clinical evidence supports an association between cardiovascular/metabolic diseases (CVMD) and erectile dysfunction (ED), scientific evidence for this link is incompletely elucidated. AIM: This study aims to provide scientific evidence for the link between CVMD and ED. METHODS: In this White Paper, the Basic Science Committee of the Sexual Medicine Society of North America assessed the current literature on basic scientific support for a mechanistic link between ED and CVMD, and deficiencies in this regard with a critical assessment of current preclinical models of disease. RESULTS: A link exists between ED and CVMD on several grounds: the endothelium (endothelium-derived nitric oxide and oxidative stress imbalance); smooth muscle (SM) (SM abundance and altered molecular regulation of SM contractility); autonomic innervation (autonomic neuropathy and decreased neuronal-derived nitric oxide); hormones (impaired testosterone release and actions); and metabolics (hyperlipidemia, advanced glycation end product formation). CONCLUSION: Basic science evidence supports the link between ED and CVMD. The Committee also highlighted gaps in knowledge and provided recommendations for guiding further scientific study defining this risk relationship. This endeavor serves to develop novel strategic directions for therapeutic interventions.

Subacute Hemolysis in Sickle Cell Mice Causes Priapism Secondary to NO Imbalance and PDE5 Dysregulation.

INTRODUCTION: Recent research suggests that priapism in sickle cell disease (SCD) is due to dysregulation of penile erection homeostasis including alteration of nitric oxide synthase (NOS) and phosphodiesterase type 5 (PDE5) activities by excessive levels of reactive oxygen species (ROS) released during hemolysis. It is unknown if subacute exposure to hemolysis is sufficient or if chronic reconditioning of erectile tissues is required for perturbation of homeostatic pathways and whether PDE5 inhibitor (PDE5I) treatment can restore erectile homeostasis in the subacute setting. AIMS: The aim of this study was to investigate the effects of subacute hemolysis (3-month exposure) on priapism and NO pathway regulation. METHODS: Mice underwent bone marrow transplantation with either SCD (BM-SS) or wild-type (WT) bone marrow. BM-SS mice were treated with sildenafil 100 mg/kg/day. We measured intracavernous pressure (ICP) measurements with or without cavernous nerve stimulation following bone marrow transplantation to assess for priapism. MAIN OUTCOME MEASURES: ICP and frequency of erections were assessed. Penile tissues were analyzed for NOS, protein kinase G (PKG), PDE5, and ROS activities. RESULTS: BM-SS mice demonstrated a priapism phenotype. PDE5I treatment reduced the frequency of erections in BM-SS mice (1.7 ± 1.1 vs. 5.5 ± 2.8 erections per hour, P < 0.05). Penile tissues from BM-SS mice demonstrated decreased NOS, PKG, PDE5 and elevated ROS activities compared with that of control mice. PDE5I treatment increased NOS (11.6 ± 1.3% vs. 7.8 ± 2.3%, P < 0.05) and PDE5 (76.3 ± 9.8% vs. 52.3 ± 11.1%, P < 0.05) activities and decreased ROS activity (137.8 ± 12.1% vs. 199.1 ± 11.3%, P < 0.05) compared with non-PDE5I treated BM-SS mice. PKG activity was increased beyond control levels with PDE5I treatment (158.4 ± 10.3%, P < 0.05). CONCLUSION: Short-term hemolysis is sufficient to establish a priapism phenotype and results in loss of erectile function. PDE5I treatment ameliorates priapism, in part, because of restored NO balance with decreased ROS generation and increased PDE5 activity.

Mechanism of testosterone deficiency in the transgenic sickle cell mouse.

Testosterone deficiency is associated with sickle cell disease (SCD), but its underlying mechanism is not known. We investigated the possible occurrence and mechanism of testosterone deficiency in a mouse model of human SCD. Transgenic sickle male mice (Sickle) exhibited decreased serum and intratesticular testosterone and increased luteinizing hormone (LH) levels compared with wild type (WT) mice, indicating primary hypogonadism in Sickle mice. LH-, dbcAMP-, and pregnenolone- (but not 22-hydroxycholesterol)- stimulated testosterone production by Leydig cells isolated from the Sickle mouse testis was decreased compared to that of WT mice, implying defective Leydig cell steroidogenesis. There also was reduced protein expression of steroidogenic acute regulatory protein (STAR), but not cholesterol side-chain cleavage enzyme (P450scc), in the Sickle mouse testis. These data suggest that the capacity of P450scc to support testosterone production may be limited by the supply of cholesterol to the mitochondria in Sickle mice. The sickle mouse testis exhibited upregulated NADPH oxidase subunit gp91phox and increased oxidative stress, measured as 4-hydroxy-2-nonenal, and unchanged protein expression of an antioxidant glutathione peroxidase-1. Mice heterozygous for the human sickle globin (Hemi) exhibited intermediate hypogonadal changes between those of WT and Sickle mice. These results demonstrate that testosterone deficiency occurs in Sickle mice, mimicking the human condition. The defects in the Leydig cell steroidogenic pathway in Sickle mice, mainly due to reduced availability of cholesterol for testosterone production, may be related to NADPH oxidase-derived oxidative stress. Our findings suggest that targeting testicular oxidative stress or steroidogenesis mechanisms in SCD offers a potential treatment for improving phenotypic changes associated with testosterone deficiency in this disease.