Effect of BKCa Channel Opener LDD175 on Erectile Function in an In Vivo Diabetic Rat Model

IntroductionThe development of novel therapeutic options is imperative in patients with erectile dysfunction, especially those non-responsive to phosphodiesterase type 5 inhibitors. LDD175, a potent BKCa channel opener, has a relaxation effect on the in vitro cavernosal smooth muscle strip.AimTo investigate the effect of LDD175 on erectile function using in vivo animal disease model.MethodsMale Sprague-Dawley rats were assigned to a normal control group and seven diabetic groups: diabetic control, sildenafil (1 and 5 mg/kg), LDD175 (5 and 10 mg/kg), LDD175 5 mg/kg plus sildenafil 1 mg/kg, and LDD175 10 mg/kg plus tetraethylammonium.Main Outcome MeasuresIntracavernosal pressure (ICP), ratio of ICP to mean arterial pressure (MAP), and the area under curve of ICP/MAP of eight groups were compared using in vivo pelvic nerve stimulation.ResultsThe ICP, ICP/MAP ratio, and area under curve of the ICP/MAP ratio of the normal control rats increased with an increase in electrical field stimulation voltage. All parameters in the diabetic control group were significantly lower than those in the normal control rats, with an electrical field stimulation ranging from 1 to 5 V (P < .05). LDD175 improved the erectile response in diabetic rats in a dose-dependent manner. The combination of sildenafil (1 mg/kg) and LDD175 (5 mg/kg) showed a significant additive effect (P < .05) on the improvement of erectile function compared with sildenafil (1 mg/kg) alone. The enhancement of erectile function by LDD175 was completely blocked by tetraethylammonium.ConclusionThe results showed that the BKCa channel opener LDD175 improved erectile function in an in vivo diabetic rat model. Furthermore, combination therapy of LDD175 and sildenafil had an additive effect on the improvement of erectile function in diabetic rats. LDD175 could be a new candidate for the treatment of erectile dysfunction.     

Udenafil Enhances the Recovery of Erectile Function and Ameliorates the Pathophysiological Consequences of Cavernous Nerve Resection

IntroductionRadical prostatectomy is the treatment of choice for prostate cancer patients. Despite the introduction of nerve-sparing surgical techniques, its success is not entirely guaranteed and the majority of patients report compromised erectile function following surgical procedures.AimThis study was performed to investigate the effect of repeated dosing of udenafil, a novel phosphodiesterase type 5 inhibitor, on penile hypoxia and fibrosis induced by bilateral cavernous nerve resection (BCNR) in rats.MethodsThirty male Sprague-Dawley rats (300–320 g) were used in this study. The animals were divided into three groups; group I consisted of sham-operated animals (N = 10), animals in group II underwent BCNR alone (N = 10), and animals in group III were orally treated with 10 mg/kg udenafil b.i.d. for 8 weeks following BCNR (N = 10).Main Outcome MeasuresThe expression of transforming growth factor-β1, hypoxia-inducible factor-1α, endothelial nitric oxide synthase, neuronal nitric oxide synthase, and endothelin B receptor in penile tissue was examined at gene level. Additionally, erectile function, measured by intracavernous pressure (ICP), and pathological changes in the corpus cavernosum were examined.ResultsWhile fibrosis, apoptosis, and the expression of TGF-β1, HIF-1α, and ET_B were significantly increased, and the expression of eNOS and nNOS were significantly decreased in group II, compared with the sham-operated animals, repeated dosing of udenafil significantly ameliorated these changes. Erectile function was profoundly impaired in animals that underwent BCNR alone, and udenafil treatment significantly attenuated this impairment as measured by ICP.ConclusionsThese results demonstrate that long-term administration of udenafil ameliorates penile hypoxia and fibrosis induced by cavernous nerve resection. This study also suggests the potential beneficial role of repeated dosing of udenafil in the recovery of erectile function in patients with neuronal erectile dysfunction. Lee C-H, Shin J-H, Ahn G-J, Kang K-K, Ahn B-O, and Yoo M. Udenafil enhances the recovery of erectile function and ameliorates the pathophysiological consequences of cavernous nerve resection.     

H2S Relaxes Vas Deferens Smooth Muscle by Modulating the Large Conductance Ca2+‐Activated K+ (BKCa) Channels via a Redox Mechanism

IntroductionHydrogen sulfide (H₂S) is generated in mammalian cells mainly by one of the two pyridoxal‐5′‐phosphate‐dependent enzymes, cystathione‐γ‐lyase (CSE), and cystathione‐β‐synthase (CBS) using L‐cysteine as the main substrate. In previous studies, we found that CBS and CSE were functionally expressed in vas deferens (VD) and H₂S‐mediated VD smooth muscle relaxation. However, the detail mechanisms that H₂S‐relaxed VD smooth muscle were unknown so far.AimThe aim of this study is to explore the molecular target sites of H₂S in VD smooth muscle.MethodsIsolated rat VD smooth muscle strips were used for tension recording in vitro. Double immunofluorescence staining was used to identify the localization of large conductance Ca²⁺‐activated K⁺ (BK_Ca) channels.Main Outcome MeasuresChanges in tonic contraction of isolated rat VD smooth muscle strip were measured after the treatment of drugs. The expression of BKca channels in rat VD smooth muscle cells was also assessed.ResultsThe results showed that L‐NG‐nitroarginine methyl ester (a nitric oxide synthase inhibitor) did not affect the response of VD to sodium hydrosulphide (NaHS), suggesting that nitric oxide pathway was not involved. Further studies revealed that transient receptor potential (TRP) channels did not contribute to the NaHS‐induced relaxant effect. Glibenclamide, an ATP‐sensitive K channel blocker, did the same thing, whereas BK_Ca channel blockers iberiotoxin or tetraethylammonium largely reversed the relaxant effect, suggesting that H₂S may target BK_Ca channels. We also confirmed that BK_Ca channels were localized in VD smooth muscle cells. Then, studies revealed that NaHS‐induced VD smooth muscle relaxation was abolished by N‐ethylmaleimide, which was widely used as a sulfhydryl alkylation compound protecting thiols from oxidation, whereas DL‐Dithiothreitol, a strong reducing agent, did not affect the response of VD to NaHS.ConclusionsWe concluded that H₂S relaxed the VD smooth muscle by targeting BK_Ca channels via redox‐mediated mechanism. Li Y, Zang Y, Fu S, Zhang H, Gao L, and Li J. H₂S relaxes vas deferens smooth muscle by modulating the large conductance Ca²⁺‐activated K⁺ (BK_Ca) channels via a redox mechanism. J Sex Med 2012;9:2806–2813.     

Endothelium‐Independent Relaxant Effect of Rubus Coreanus Extracts in Corpus Cavernosum Smooth Muscle

IntroductionRubus coreanus is a perennial shrub native to the southern part of the Korean peninsula. Although it is known that R. coreanus has a dose‐dependent relaxation effect on rabbit corpus cavernosum (CC), the exact mechanism of action by which R. coreanus work is not fully known.AimsTo elucidate the direct effects of unripe R. coreanus extract (RCE) on CC smooth muscle cells.MethodsDried unripe R. coreanus fruits were pulverized and extracted with 95% ethanol. Isolated rabbit CC strips were mounted in an organ‐bath system, and the effects of RCE were evaluated. To estimate [Ca²⁺]_i, we used a Fura‐2 fluorescent technique.Main Outcome MeasuresThe effects of unripe RCE on ion channels and the intracellular Ca²⁺ concentration ([Ca²⁺]_i) of CC.ResultsRCE effectively relaxed phenylephrine (PE)‐induced tone in rabbit CC, and removal of the endothelium did not completely abolish the relaxation effect of RCE. Tetraethylammonium (1 mM) did not inhibit RCE‐induced relaxation in strips precontracted by PE in the organ bath. However, CaCl₂‐induced constriction of CC strips, bathed in Ca²⁺‐free buffer and primed with PE, was abolished by RCE. In addition, RCE decreased basal [Ca²⁺]_i in corporal smooth muscle cells. The increases of [Ca²⁺]_i evoked by 60 mM K⁺‐containing solution in A7r5 cells were suppressed by RCE, and RCE relaxed KCl‐induced tone in endothelium‐free CC, which indicated that RCE blocked the voltage‐dependent Ca²⁺ channels (VDCCs). RCE decreased basal [Ca²⁺]_i and the [Arg8]‐vasopressin‐induced [Ca²⁺]_i increases in A7r5 cells, and RCE inhibited the contraction of endothelium‐free CC induced by PE in Ca²⁺‐free solution, which suggested that RCE might act as a modulator of corporal smooth muscle cell tone by inhibiting Ca²⁺ release from sarcoplasmic reticulum.ConclusionRCE acts through endothelium‐independent and endothelium‐dependent pathways to relax CC. RCE may inhibit VDCCs and Ca²⁺ release from sarcoplasmic reticulum. Lee JH, Chae MR, Sung HH, Ko M, Kang SJ, and Lee SW. Endothelium‐independent relaxant effect ofRubus coreanusextracts in corpus cavernosum smooth muscle. J Sex Med **;**:**–**.     

Powerful Relaxation of Phosphodiesterase Type 4 Inhibitor Rolipram in the Pig and Human Bladder Neck

IntroductionPhosphodiesterase type 5 (PDE5) inhibitors act as effective drugs for the treatment of lower urinary tract symptom (LUTS). There is a poor information, however, about the role of the PDE4 inhibitors on the bladder outflow region contractility.AimTo investigate PDE4 expression and the relaxation induced by the PDE4 inhibitor rolipram versus that induced by the PDE5 blockers sildenafil and vardenafil, in the pig and human bladder neck.MethodsImmunohistochemistry for PDE4 expression, myographs for isometric force recordings and fura‐2 fluorescence for simultaneous measurements of intracellular Ca²⁺ concentration ([Ca²⁺]_i) and tension for rolipram in bladder neck samples were used.Main Outcome MeasuresPDE4 expression and relaxations to PDE4 and PDE5 inhibitors and simultaneous measurements of [Ca²⁺]_i and tension.ResultsPDE4 expression was observed widely distributed in the smooth muscle layer of the pig and human bladder neck. On urothelium‐denuded phenylephrine (PhE)‐precontracted strips of pig and human, rolipram, sildenafil and vardenafil produced concentration‐dependent relaxations with the following order of potency: rolipram> > sildenafil>vardenafil. In pig, the adenylyl cyclase activator forskolin potentiated rolipram‐elicited relaxation, whereas protein kinase A (PKA) blockade reduced such effect. On potassium‐enriched physiological saline solution (KPSS)‐precontracted strips, rolipram evoked a lower relaxation than that obtained on PhE‐stimulated preparations. Inhibition of large (BK_Ca) and intermediate (IK_Ca) conductance Ca²⁺‐activated K⁺ channels, neuronal voltage‐gated Ca²⁺ channels, nitric oxide (NO) and hydrogen sulfide (H₂S) synthases reduced rolipram responses. Rolipram inhibited the contractions induced by PhE without reducing the PhE‐evoked [Ca²⁺]_i increase.ConclusionsPDE4 is present in the pig and human bladder neck smooth muscle, where rolipram exerts a much more potent relaxation than that elicited by PDE5 inhibitors. In pig, rolipram‐induced response is produced through the PKA pathway involving BK_Ca and IK_Ca channel activation and [Ca²⁺]_idesensitization‐dependent mechanisms, this relaxation also being due to neuronal NO and H2S release. Ribeiro ASF, Fernandes VS, Martínez‐Sáenz A, Martínez P, Barahona MV, Orensanz LM, Blaha I, Serrano‐Margüello D, Bustamante S, Carballido J, García‐Sacristán A, Prieto D, and Hernández M. Powerful relaxation of phosphodiesterase type 4 inhibitor rolipram in the pig and human bladder neck. J Sex Med 2014;11:930–941.     

Prolonged Therapy with the Soluble Guanylyl Cyclase Activator BAY 60-2770 Restores the Erectile Function in Obese Mice

IntroductionCardiovascular and endocrine-metabolic diseases associated with increased oxidative stress such as obesity lead to erectile dysfunction (ED). Activators of soluble guanylyl cyclase (sGC) such as BAY 60-2770 reactivate the heme-oxidized sGC in vascular diseases.AimThis study aimed to evaluate the effects of 2-week oral intake with BAY 60-2270 on a murine model of obesity-associated ED.MethodsC57BL/6 male mice were fed for 12 weeks with standard chow or high-fat diet. Lean and obese mice were treated with BAY 60-2770 (1 mg/kg/day, 2 weeks).Main Outcome MeasuresMeasurements of intracavernosal pressure (ICP), along with acetylcholine (10⁻⁹ to 10⁻⁵ M) and electrical field stimulation (EFS; 4–10 Hz)-induced corpus cavernosum relaxations in vitro, were obtained. Levels of cyclic guanosine monophosphate (cGMP), reactive oxygen species (ROS), and sGC protein expressions in cavernosal tissues were measured.ResultsCavernous nerve stimulation caused frequency-dependent ICP increases, which were significantly lower in obese compared with lean mice (P < 0.05). Two-week therapy with BAY 60-2770 fully reversed the decreased ICP in obese group. Acetylcholine-induced cavernosal relaxations were 45% lower (P < 0.001) in obese mice, which were fully restored by BAY 60-2770 treatment. Likewise, the EFS-induced relaxations in obese mice were restored by BAY 60-2770. Basal cGMP content in erectile tissue was 68% lower (P < 0.05) in obese mice, an effect normalized by BAY 60-2770. Levels of ROS were 52% higher (P < 0.05) whereas protein expression of α1 sGC subunit was reduced in cavernosal tissue of obese mice, both of which were normalized by BAY 60-2770. In lean group, BAY 60-2770 did not significantly affect any functional, biochemical, or molecular parameter analyzed.ConclusionsTwo-week therapy with BAY 60-2770 restores the erectile function in obese mice that is associated with reduced ROS levels, up-regulation of α1 sGC subunit, and increased cGMP levels in the erectile tissue. Silva FH, Leiria LO, Alexandre EC, Davel APC, Mónica FZ, De Nucci G, and Antunes E. Prolonged therapy with the soluble guanylyl cyclase activator BAY 60-2770 restores the erectile function in obese mice. J Sex Med 2014;11:2661–2670.     

Endothelin A (ETA) Receptors Are Involved in Augmented Adrenergic Vasoconstriction and Blunted Nitric Oxide-Mediated Relaxation of Penile Arteries from Insulin-Resistant Obese Zucker Rats

IntroductionEndothelin 1 (ET-1) levels and receptors are up-regulated in the erectile tissue of diabetic patients and animal models of erectile dysfunction (ED).AimThe present study assessed the role of ET-1 receptors in the impaired adrenergic vasoconstriction and nitrergic relaxation of penile arteries from a rat model of insulin resistance.MethodsThe effect of ET receptor antagonists was evaluated on the contractile responses to electrical field stimulation (EFS) of penile arteries from obese Zucker rats (OZRs) compared with lean Zucker rats (LZRs). ET receptor expression was determined by immunohistochemistry.Main Outcome MeasuresChanges in neural nitrergic relaxation and adrenergic vasoconstriction and the expression of ET receptors in perivascular nerves were assessed.ResultsET-1 (10⁻¹⁰ M) enhanced EFS-induced vasoconstriction, and treatment with the adrenergic neurotoxin guanethidine reduced the contractions induced by ET-1 in penile arteries from both LZRs and OZRs, thus supporting the hypothesis that ET-1 releases noradrenaline from adrenergic nerves. ET-1 antagonized neural nitric oxide (NO)-mediated relaxant responses in LZR arteries, antagonizing relaxations induced by the NO donor S-nitroso-N-acetylpenicillamine to a larger extent in arteries from OZRs. ET_A and ET_B receptors were expressed in perivascular fibers colocalized with the neuronal marker protein gene product 9.5 in penile arteries from OZRs. The ET_A receptor antagonist BQ-123 reversed the enhancing effect of ET-1 on the vasoconstriction elicited by EFS and the ET-1-induced inhibition of nitrergic relaxations in LZRs, restoring them to control levels in penile arteries of OZRs.ConclusionsET-1 enhances adrenergic vasoconstriction through presynaptic ET_A receptors and antagonizes neural NO-mediated relaxation through postsynaptic smooth muscle ET_A receptors in penile arteries from OZRs, which likely contributes to the augmented vasoconstriction and blunted nitrergic relaxation of erectile tissue under conditions of insulin resistance. Sánchez A, Contreras C, Martínez P, Muñoz M, Martínez AC, García-Sacristán A, Hernández M, and Prieto D. Endothelin (ET_A) receptors are involved in augmented adrenergic vasoconstriction and blunted nitric oxide-mediated relaxation of penile arteries from insulin-resistant obese Zucker rats. J Sex Med 2014;11:1463–1474.     

Excessive Penile Norepinephrine Level Underlies Impaired Erectile Function in Adenosine A1 Receptor Deficient Mice

IntroductionPenile erection is a complex neurovascular physiological event controlled by multiple factors and signaling pathways. A considerable amount of evidence indicates that adenosine plays a significant role in cavernosal smooth muscle relaxation. However, the specific role of adenosine and its receptors in erectile physiology and pathology is not fully understood.AimTo determine the role of the adenosine A₁ receptor (ADORA1) in penile erection.MethodAdenosine A₁ receptor deficient (Adora1^‐/‐) mice and aged‐matched wild‐type (WT) mice were utilized. We evaluated the in vivo erectile function by measuring the intracavernosal pressure (ICP) in response to cavernous nerve stimulation (CNS). Enzyme‐linked immunosorbent assay was used to measure the norepinephrine (NE) plasma concentration in the corpus cavernosum and systemic circulation. We also evaluated the myosin light chain phosphorylation (p‐MLC) in penile tissue pre‐ and post‐CNS.Main Outcome MeasurementThe main outcome measurement of this research was the evaluation of in vivo erectile response to CNS by measuring the ICP in Adora1^‐/‐ mice and WT mice and to identify the localization and specific neuron types of ADORA1 expression by dual immunostaining and immunofluorescence co‐localization.ResultIn vivo, both the ratio of CNS‐induced Maximum ICP to mean arterial pressure and CNS‐induced slope in Adora1^‐/‐ mice were significantly lower than WT mice. At the cellular level in penile tissue, we determined that ADORA1 was highly abundant in neuronal cells. During penile erection, Adora1^‐/‐ mice exhibited a higher level of NE plasma concentration in the penis than WT mice. And WT mice had a significantly greater reduction in p‐MLC compared to Adora1^‐/‐ mice.ConclusionOur results show that ADORA1 is enriched on neuron cells where it functions to control NE release. Activation of this receptor during penile erection results in reduced NE release and reduced cavernosal smooth muscle contraction, therefore facilitating penile erection. Ning C, Qi L, Wen J, Zhang Y, Zhang W, Wang W, Blackburn M, Kellems R, and Xia Y. Excessive penile norepinephrine level underlies impaired erectile function in adenosine A1 receptor deficient mice.     

Chronic administration of udenafil, a selective phosphodiesterase type 5 inhibitor, promotes erectile function recovery in an animal model of bilateral cavernous nerve crush injury

IntroductionPreservation of the cavernous nerves (CNs) during radical prostatectomy is crucial for the patient's erectile function. Despite advances in operative technique, the majority of men report compromised erectile function postprostatectomy or complete loss of potency due to CN trauma even with nerve-sparing modifications.AimThis study was designed to investigate whether repeated dosing of udenafil, a phosphodiesterase type 5 inhibitor, helps to improve erectile function after CN injury.MethodsUsing the CN crush injury model, 8-week-old male Sprague Dawley rats were divided into the following groups; sham-operated group, bilateral CN crush injury exposed to either no udenafil (vehicle) or udenafil (5, 20 mg/kg) daily for two different durations (4 and 8 weeks, p.o.).Main Outcome MeasuresAt both time points, CN electrical stimulation was used to assess erectile function by measuring the intracavernous pressure. The expressions of hypoxia-inducible factor 1-alpha (HIF-1α), transforming growth factor-beta (TGF-β1), nerve growth factor (NGF), endothelin B receptor (ET_B), endothelial nitric oxide synthase (eNOS), neuronal nitric oxide synthase (nNOS), and sonic hedgehog homolog (SHH) in penile tissue were examined. Immunohistochemical antibody staining was performed for NGF, eNOS, nNOS, CD31, and alpha-smooth muscle actin (α-SMA). Additionally, terminal deoxynucleotidyl transferase-mediated nick-end labeling assay was performed to quantify apoptosis and the tissue slides were stained for Masson's trichrome to assess the smooth muscle/collagen ratio.ResultsUdenafil improved erectile function in a dose- and time-dependent manner with the maximum erectile function recovery achieved by 20 mg/kg udenafil at an 8-week time point. CN injury increased the expression of HIF-1α, TGF-β1, NGF, and ET_B, however, decreased the expression of eNOS, nNOS, and SHH. Udenafil significantly suppressed these alterations. The results from the histological analyses show that udenafil markedly reduces apoptosis induced by CN injury and augments the smooth muscle/collagen ratio.ConclusionsCN injury induces significantly impaired erectile function and altered gene/protein expression. Chronic administration of udenafil preserves erectile function and has a beneficial role against the pathophysiological consequences of CN injury. Lee C-H, Kim H-S, Goo M-J, Kang K-K, Ahn B-O, Kim SH, and Yang D-Y. Chronic administration of udenafil, a selective phosphodiesterase type 5 inhibitor, promotes erectile function recovery in an animal model of bilateral cavernous nerve crush injury.     

Metrics for Evaluation of Age-Related Changes in Erectile Capacity in a Rodent Model

IntroductionStimulation of the cavernous nerve elicits increases in intracavernosal pressure (ICP) and visible penile erection (VPE) in rats. The observed penile erection can be characterized by the change in ICP and the ratio of ICP over blood pressure (BP; systemic blood pressure).AimWe evaluated correlations between ICP, BP, ICP/BP, and VPE, and the magnitude of the nerve stimulation required to elicit those responses in order to evaluate age-related changes in erectile function in a rodent model.MethodsNine young (3 months) and 10 old (18 months) rats were used. Under anesthesia cavernous nerve stimulation was performed at stimulation strengths ranging from 0.1 to 10 mA.Main Outcome MeasuresThe ICP, BP, ICP/BP, and VPE responses to cavernous nerve stimulation were quantified and compared.ResultsSignificant correlations were observed between ICP and ICP/BP in both young (r² = 0.886) and old (r² = 0.962) rats. If the ratio ICP/BP was taken as a metric of VPE and non-erection, then the threshold value for observing a VPE in the majority of animals was ≥0.5 in both young and old rats. However, higher current stimulation was required in the old than in the young animals to achieve erections. Moreover, both the maximal ICP and ICP/BP ratios achieved during erection were also lower in the old (85 ± 2 cmH₂O and 0.6 ± 0.02, respectively) than young (106 ± 3 cmH₂O and 0.75 ± 0.02, respectively) rats (P < 0.0001).ConclusionOld rats without obvious comorbidities were capable of penile erections, but those erections were accompanied by reduced ICP responses to cavernous nerve stimulation, implying apparently reduced penile rigidity. Taken together our current observations indicate that ICP, ICP/BP, and VPE provide useful metrics/indices of erectile capacity/function, and, moreover, that improved understanding of the inter-relationships among these parameters provides a frame work for further exploration of the mechanistic basis for age-related erectile dysfunction. Zhao W, Sato Y, Melman A, Andersson K-E, and Christ G. Metrics for evaluation of age-related changes in erectile capacity in a rodent model. J Sex Med 2009;6:1885–1892.     

The Correlation Between Intracavernosal Pressure and Cavernosal Blood Oxygenation

IntroductionGiven that regular nocturnal erections are physiological, it has been suggested that erections are pivotal to the maintenance of erectile tissue health. It has been postulated that a critical element to erectile tissue protection is cavernosal oxygenation. It is accepted that the corpora cavernosa are oxygenated fully during a rigid erection. However, it remains unknown what degree of penile rigidity is required to achieve cavernosal oxygenation at the arterial level.AimThis analysis was undertaken to define the correlation between intracavernosal pressure (ICP) and cavernosal oxygen partial pressure (pO₂).Main Outcome MeasuresCavernosal pO₂ at various ICPs.MethodsThe study population was comprised of patients undergoing dynamic infusion cavernosometry (DIC) in the evaluation of erectile dysfunction or prior to penile reconstructive surgery. DIC was conducted with a standard vasoactive agent redosing schedule. One milliliter of corporal blood was aspirated at various ICPs into a heparinized syringe for later pO₂ analysis. Blood was placed on ice immediately and transported to the laboratory upon completion of the DIC.ResultsTwenty-one blood samples were analyzed from 13 patients. Mean patient age was 43 ± 18 years. Blood specimens were collected at an ICP range of 6–90 mm Hg. Mean ± SD pO₂ was 39 ± 11 mm Hg at ICP < 10 mm Hg, 87 ± 3 at ICP 11–20 mm Hg, 89 ± 6 at ICP 21–45 mm Hg and 96 ± 13 at ICP > 45 mm Hg.ConclusionsSignificant increases in cavernosal oxygenation occur in the earliest stages of erection at relatively low ICP. These findings suggest that partial erections may be sufficient to oxygenate erectile tissue and protect it from prolonged hypoxia-induced damage. Tal R, Mueller A, and Mulhall JP. The correlation between intracavernosal pressure and cavernosal blood oxygenation. J Sex Med 2009;6:2722–2727.     

A therapeutic confirmatory study to assess the safety and efficacy of Zydena (udenafil) for the treatment of erectile dysfunction in male patients with diabetes mellitus

IntroductionPatients with diabetes mellitus (DM) are reported to experience more severe erectile dysfunction (ED) symptoms and respond less to ED treatments compared with patients with ED of other etiologies.AimThis study was undertaken to evaluate the safety and efficacy of udenafil for the treatment of ED in a larger number of patients with DM.MethodsA placebo‐controlled, randomized, double‐blind, double‐dummy, parallel‐group design multicenter study, fixed‐dose trial was conducted. The trial involved seven study sites in Korea, with 174 ED patients with DM. The subjects, treated with placebo, 100 mg, or 200 mg of udenafil for 12 weeks, were asked to complete the International Index of Erectile Function (IIEF), the Sexual Encounter Profile (SEP) diary, and the Global Assessment Question (GAQ) during the study period.Main Outcome MeasuresThe primary efficacy parameter was the change in the erectile function domain (EFD) score of IIEF from baseline. Secondary parameters were IIEF questions 3 (Q3) and Q4, SEP Q2 and Q3, rate of achieving normal erectile function (EFD ≥ 26), and the response to GAQ.ResultsCompared with the placebo, patients receiving both doses of udenafil showed statistically significant improvements in the IIEF‐EFD score, respectively. However, statistically significant difference was not observed between the udenafil 100 mg and the udenafil 200 mg groups. Similar results were observed in the comparison of Q3 and Q4 of IIEF, SEP diary, and GAQ. The percentages of subjects experiencing at least one adverse event related to the study drugs were 3.6%, 15.8%, and 22.4% for the placebo, udenafil 100 mg, and udenafil 200 mg groups, respectively. However, these events were all mild in severity. Major adverse events were flushing, headache, nausea, and conjunctival hyperemia.ConclusionUdenafil was significantly effective for the treatment of ED, demonstrating statistically significant improvement in erectile function in patients with DM. The incidence of adverse events was relatively low and well tolerated in patients with DM. Moon DG, Yang DY, Lee CH, Ahn TY, Min KS, Park K, Park CK, and Kim JJ. A therapeutic confirmatory study to assess the safety and efficacy of Zydena^® (udenafil) for the treatment of erectile dysfunction in male patients with diabetes mellitus. J Sex Med 2011;8:2048–2061.     

M1 Macrophages Are Predominantly Recruited to the Major Pelvic Ganglion of the Rat Following Cavernous Nerve Injury

IntroductionNeurogenic erectile dysfunction is a common sequela of radical prostatectomy. The etiology involves injury to the autonomic cavernous nerves, which arise from the major pelvic ganglion (MPG), and subsequent neuroinflammation, which leads to recruitment of macrophages to the injury site. Currently, two macrophage phenotypes are known: neurotoxic M1 macrophages and neuroprotective M2 macrophages.AimTo examine whether bilateral cavernous nerve injury (BCNI) in a rat model of erectile dysfunction would increase recruitment of neurotoxic M1 macrophages to the MPG.MethodsMale Sprague-Dawley rats underwent BCNI and the MPG was harvested at various time points after injury. The corpora cavernosa was used to evaluate tissue myographic responses to electrical field stimulation ex vivo. Quantitative real-time polymerase chain reaction was used to examine the gene expression of global macrophage markers, M1 macrophage markers, M2 macrophage markers, and cytokines and chemokines in the MPG. Mathematical calculation of the M1/M2 index was used to quantify macrophage changes temporally. Western blot of MPG tissues was used to evaluate the protein amount of M1 and M2 macrophage markers quantitatively. Immunohistochemistry staining of MPGs for CD68, CD86, and CD206 was used to characterize M1 and M2 macrophage infiltration.Main Outcome MeasuresCorpora cavernosa responsiveness ex vivo; gene (quantitative real-time polymerase chain reaction) and protein (western blot) expressions of M1 and M2 markers, cytokines, and chemokines; and immunohistochemical localization of M1 and M2 macrophages.ResultsBCNI impaired the corporal parasympathetic-mediated relaxation response to electrical field stimulation and enhanced the contraction response to electrical field stimulation. Gene expression of proinflammatory (Il1b, Il16, Tnfa, Tgfb, Ccl2, Ccr2) and anti-inflammatory (Il10) cytokines was upregulated in the MPG 48 hours after injury. M1 markers (CD86, inducible nitric oxide synthase, interleukin-1β) and M2 markers (CD206, arginase-1, interleukin-10) were increased after BCNI in the MPG, with the M1/M2 index above 1.0 indicating that more M1 than M2 macrophages were recruited to the MPG. Protein expression of the M1 macrophage marker (inducible nitric oxide synthase) was increased in MPGs after BCNI. However, the protein amount of M2 macrophage markers (arginase-1) remained unchanged. Immunohistochemical characterization demonstrated predominant increases in M1 (CD68⁺CD86⁺) macrophages in the MPG after BCNI.ConclusionThese results suggest that an increase in M1 macrophage infiltration of the MPG after BCNI is associated with impaired neurogenically mediated erectile tissue physiology ex vivo and thus has significant implications for cavernous nerve axonal repair. Future studies are needed to demonstrate that inhibition of M1 macrophage recruitment prevents erectile dysfunction after CNI.     

Nitric Oxide-Induced Vasorelaxation in Response to PnTx2-6 Toxin from Phoneutria nigriventer Spider in Rat Cavernosal Tissue

IntroductionPriapism is one of several symptoms observed in accidental bites by the spider Phoneutria nigriventer. The venom of this spider is comprised of many toxins, and the majority has been shown to affect excitable ion channels, mainly sodium (Na⁺) channels. It has been demonstrated that PnTx2-6, a peptide extracted from the venom of P. nigriventer, causes erection in anesthetized rats and mice.AimWe investigated the mechanism by which PnTx2-6 evokes relaxation in rat corpus cavernosum.Main Outcome MeasuresPnTx2-6 toxin potentiates nitric oxide (NO)-dependent cavernosal relaxation.MethodsRat cavernosal strips were incubated with bretylium (3 × 10^?5 M) and contracted with phenylephrine (PE; 10^?5 M). Relaxation responses were evoked by electrical field stimulation (EFS) or sodium nitroprusside (SNP) before and after 4 minutes of incubation with PnTx2-6 (10^?8 M). The effect of PnTx2-6 on relaxation induced by EFS was also tested in the presence of atropine (10^?6 M), a muscarinic receptor antagonist, N-type Ca²⁺ channel blockers (ω-conotoxin GVIA, 10^?6 M) and sildenafil (3 × 10^?8 M). Technetium99m radiolabeled PnTx2-6 subcutaneous injection was administrated in the penis.ResultsWhereas relaxation induced by SNP was not affected by PnTx2-6, EFS-induced relaxation was significantly potentiated by this toxin as well as PnTx2-6 plus SNP. This potentiating effect was further increased by sildenafil, not altered by atropine, however was completely blocked by the N-type Ca²⁺ channels. High concentrated levels of radiolabeled PnTx2-6 was specifically found in the cavernosum tissue, suggesting PnTx2-6 is an important toxin responsible for P. nigriventer spider accident-induced priapism.ConclusionWe show that PnTx2-6 slows Na⁺ channels inactivation in nitrergic neurons, allowing Ca²⁺ influx to facilitate NO/cGMP signalling, which promotes increased NO production. In addition, this relaxation effect is independent of phosphodiesterase enzyme type 5 inhibition. Our data displays PnTx2-6 as possible pharmacological tool to study alternative treatments for erectile dysfunction. Nunes KP, Cordeiro MN, Richardson M, Borges MN, Diniz SOF, Cardoso VN, Tostes R, De Lima ME, Webb RC, and Leite R. Nitric oxide-induced vasorelaxation in response to PnTx2-6 toxin from Phoneutria nigriventer spider in rat cavernosal tissue.     

Divergent Molecular Mechanisms Underlay CO‐ and CORM‐2‐Induced Relaxation of Corpora Cavernosa

IntroductionSimilar to nitric oxide (NO), the principal mediator of penile erection, carbon monoxide (CO) possesses vasodilator capacities. However, whether CO could be a therapeutic target for treating erectile dysfunction (ED) is unexplored. The danger associated with systemic administration of CO has led to the development of CO‐releasing molecules (CORMs), releasing CO in a local, safe and controlled way. These CORMs have shown positive outcomes in cardiovascular studies. More knowledge on the (patho)physiological functions of CO in erectile function and the potential therapeutic role of CORMs is required.AimThe present study aims the assessment of the effect of CO and CO donor CORM‐2 on the corporal tension and the underlying molecular mechanisms.MethodsOrgan bath studies were performed measuring isometric tension on isolated mice corpora cavernosa (CC) strips. Responses to CO (10–300 µmol/L) and CORM‐2 (10–100 µmol/L) were measured in the presence/absence of activators/inhibitors of different molecular pathways.Main Outcome MeasuresCO and CORM‐2 relax corporal strips concentration dependently, although the molecular mechanisms behind the corporal relaxation seem to differ completely.ResultsCO induces corporal relaxation by activating soluble guanylyl cyclase (sGC), increasing cyclic guanosine monophosphate (cGMP) concentrations. The molecular mechanism involved in CORM‐2‐induced corporal relaxation is not related to sGC activation and remains obscure.ConclusionsBoth CO and CORM‐2 induce corporal relaxation, although the underlying molecular mechanisms show no resemblance. That CO induces corporal relaxation through a mechanism similar to that of NO could be of importance as it indirectly offers the possibility that endogenous CO might serve as a backup system for insufficient NO availability in cases of ED. Whether CORM‐2 possesses the same capacity remains questionable and requires further research. Decaluwé K, Pauwels B, Boydens C, and Van de Voorde J. Divergent molecular mechanisms underlay CO‐ and CORM‐2‐induced relaxation of corpora cavernosa. J Sex Med 2012;9:2284–2292.     

Efficacy of Once-Daily Administration of Udenafil for 24 Weeks on Erectile Dysfunction: Results from a Randomized Multicenter Placebo-Controlled Clinical Trial

IntroductionThe method of administration of oral phosphodiesterase-5 inhibitors has been expanded to once-daily repeated administration with lower initial dosage than on-demand administration.AimThe aim of this study was to evaluate the efficacy and safety of once-daily udenafil as a treatment for erectile dysfunction (ED) for intermediate-term period.MethodsThis multicenter, randomized, double-blind clinical trial included 346 ED patients (placebo, udenafil 50 mg, udenafil 75 mg). Subjects were treated with each medication once daily for 24 weeks.Main Outcome MeasuresSubjects were asked to complete the International Index of Erectile Function (IIEF)-erectile function (EF) domain at baseline, 12 weeks, and 24 weeks and the development of adverse drug reactions (ADRs) was inspected.ResultsBoth dosages of udenafil induced a significant increase in IIEF-EF compared with placebo at both 12 and 24 weeks. When patients were divided according to the severity of baseline EF score, significant improvement was observed only with udenafil 75 mg regardless of the degree of ED. At 24 weeks, the proportions of patients who reported a return to normal EF (IIEF-EF over 26) were 39.1% for udenafil 50 mg and 47.0% for udenafil 75 mg. In terms of safety, ADRs were observed in 6.1%, 12.9%, and 17.9% for placebo, udenafil 50 mg, and 75 mg, respectively. Although a statistically higher rate of ADRs was observed in the udenafil 75 mg group (P = 0.024), the majority were mild and recovered without treatment.ConclusionsOnce-daily administration of udenafil 50 mg and 75 mg for 24 weeks resulted in improvement of EF. In particular, udenafil 75 mg improves EF regardless of the baseline degree of ED. Moon KH, Ko YH, Kim SW, Moon DG, Kim JJ, Park NC, Lee SW, Paick J-S, Ahn TY, Chung WS, Min KS, Park JK, Yang DY, and Park K. Efficacy of once-daily administration of udenafil for 24 weeks on erectile dysfunction: Results from a randomized multicenter placebo-controlled clinical trial. J Sex Med 2015;12:1194–1201.     

Efficacy of Udenafil for the Treatment of Erectile Dysfunction up to 12 Hours after Dosing: A Randomized Placebo-Controlled Trial

IntroductionUdenafil is a newly developed selective phosphodiesterase type 5 inhibitor for the treatment of men with erectile dysfunction (ED).AimTo evaluate the efficacy of udenafil in treating ED for up to 12 hours after dosing.MethodsThis was a randomized, double-blind, placebo-controlled, parallel-group, fixed dose design, multicenter study. Following a 4-week nondrug baseline period, 104 men with ED of broad etiology and severity were randomized to one of two treatment groups: udenafil 100 mg or placebo. Participants were requested to attempt sexual intercourse at 12 hours after udenafil or placebo dosing during a 4-week treatment period.Main Outcome MeasuresThe primary efficacy variable was the response of patients to question 3 of the Sexual Encounter Profile (SEP Q3). The secondary efficacy measures were the response of patients to question 2 of the Sexual Encounter Profile (SEP Q2). Additional secondary efficacy measures included changes from baseline in the erectile function (EF) domain scores of the International Index of Erectile Function (IIEF) questionnaire.ResultsOf the 104 patients, 103 (50 in the udenafil group, 53 in the placebo group) completed the study. Udenafil significantly enhanced the rate of maintenance of erection (SEP Q3; placebo, 28.3% vs. udenafil, 54.7%; P < 0.0001). Significant change from baseline in the IIEF-EF domain was observed in the udenafil group (placebo, –0.58 ± 0.67; udenafil, 4.40 ± 0.84; P < 0.0001). For SEP Q2, there was no difference from baseline and no difference between the two groups. The overall adverse events rate was 11.3%. Most adverse events were mild or moderate in severity, and no serious adverse events were reported during the study and the follow-up period.ConclusionsUdenafil at 100 mg was effective for relieving ED for up to 12 hours after dosing. This duration of effectiveness could allow for flexibility and spontaneity in the sexual lives of patients. Park HJ, Park JK, Park K, Min K, and Park NC. Efficacy of udenafil for the treatment of erectile dysfunction up to 12 hours after dosing: A randomized placebo-controlled trial.     

In Vitro and In Vivo Relaxation of Corpus Cavernosum Smooth Muscle by the Selective Myosin II Inhibitor,Blebbistatin

IntroductionBlebbistatin (BLEB) is a small cell permeable molecule originally reported as a selective inhibitor of myosin II isoforms expressed by striated muscle and non-muscle cells (IC₅₀ = 0.5–5 µM) with poor inhibition of turkey gizzard smooth muscle (SM) myosin II (IC₅₀～80 µM). However, recently it was found that BLEB can potently inhibit mammalian arterial SM (IC₅₀～5 µM).AimTo investigate the effect of BLEB on corpus cavernosum SM (CCSM) tone and erectile function (EF).MethodsCC tissue obtained from penile implant patients along with CC, aorta and bladder from adult male rats were used for BLEB organ bath studies. Intracavernosal BLEB was administered to rats and EF was assessed via intracavernous pressure (ICP).Main Outcome MeasuresEffects of BLEB on agonist-induced CCSM, aorta and bladder contraction in vitro and ICP in vivo.ResultsBLEB completely relaxed human CCSM pre-contracted with phenylephrine (PE) in a dose-dependent manner decreasing tension by 76.5% at 10 µM. BLEB pre-incubation attenuated PE-induced contraction of human CC by ～85%. Human CC strips pre-contracted with endothelin-1 or KCl were almost completely relaxed by BLEB. Rat CCSM pre-contracted with PE showed BLEB relaxation comparable to human CCSM. BLEB inhibition was similar for rat aorta but slower for bladder. Both maximal ICP and ICP/mean arterial pressure were dose-dependently increased by BLEB intracavernous injections with full erection at 1 micromole.ConclusionOur novel data reveals that BLEB nearly completely relaxes rat and human CCSM pre-contracted with a variety of potent agonists and exhibits tissue selectivity. Coupled with our in vivo data in which nanomole doses of BLEB significantly increase ICP, our data substantiates an important role for the SM contractile apparatus in the molecular mechanism for EF and suggests the possibility of BLEB binding at myosin II as a therapeutic treatment for ED by targeting SM contractile pathways. Zhang X, Aydin M, Kuppam D, Melman A, and DiSanto ME. In vitro and in vivo relaxation of corpus cavernosum smooth muscle by the selective myosin II inhibitor, blebbistatin. J Sex Med 2009;6:2661–2671.     

Valproic Acid Prevents Penile Fibrosis and Erectile Dysfunction in Cavernous Nerve‐Injured Rats

IntroductionBilateral cavernous nerve injury (BCNI) causes profound penile changes such as apoptosis and fibrosis leading to erectile dysfunction (ED). Histone deacetylase (HDAC) has been implicated in chronic fibrotic diseases.AimsThis study will characterize the molecular changes in penile HDAC after BCNI and determine if HDAC inhibition can prevent BCNI‐induced ED and penile fibrosis.MethodsFive groups of rats (8–10 weeks, n = 10/group) were utilized: (i) sham; (ii and iii) BCNI 14 and 30 days following injury; and (iv and v) BCNI treated with HDAC inhibitor valproic acid (VPA 250 mg/kg; 14 and 30 days). All groups underwent cavernous nerve stimulation (CNS) to determine intracavernosal pressure (ICP). Penile HDAC3, HDAC4, fibronectin, and transforming growth factor‐β1 (TGF‐β1) protein expression (Western blot) were assessed. Trichrome staining and the fractional area of fibrosis were determined in penes from each group. Cavernous smooth muscle content was assessed by immunofluorescence to alpha smooth muscle actin (α‐SMA) antibodies.Main Outcome MeasuresWe measured ICP; HDAC3, HDAC4, fibronectin, and TGF‐β1 protein expression; penile fibrosis; penile α‐SMA content.ResultsThere was a voltage‐dependent decline (P < 0.05) in ICP to CNS 14 and 30 days after BCNI. Penile HDAC3, HDAC4, and fibronectin were significantly increased (P < 0.05) 14 days after BCNI. There was a slight increase in TGF‐β1 protein expression after BCNI. Histological analysis showed increased (P < 0.05) corporal fibrosis after BCNI at both time points. VPA treatment decreased (P < 0.05) penile HDAC3, HDAC4, and fibronectin protein expression as well as corporal fibrosis. There was no change in penile α‐SMA between all groups. Furthermore, VPA‐treated BCNI rats had improved erectile responses to CNS (P < 0.05).ConclusionHDAC‐induced pathological signaling in response to BCNI contributes to penile vascular dysfunction. Pharmacological inhibition of HDAC prevents penile fibrosis, normalizes fibronectin expression, and preserves erectile function. The HDAC pathway may represent a suitable target in preventing the progression of ED occurring post‐radical prostatectomy. Hannan JL, Kutlu O, Stopak BL, Liu X, Castiglione F, Hedlund P, Burnett AL, and Bivalacqua TJ. Valproic acid prevents penile fibrosis and erectile dysfunction in cavernous nerve‐injured rats. J Sex Med 2014;11:1442–1451.