Nebivolol Potentiates the Efficacy of PDE5 Inhibitors to Relax Corpus Cavernosum and Penile Arteries from Diabetic Patients by Enhancing the NO/cGMP Pathway

IntroductionThe efficacy of oral pharmacotherapy for erectile dysfunction (ED) (i.e., type 5 phosphodiesterase [PDE5] inhibitors) is significantly reduced in diabetic patients. Nebivolol is a selective β₁‐blocker used for treating hypertension that has been shown to increase the efficacy of sildenafil to reverse ED in diabetic rats.AimTo evaluate the effects of nebivolol on the efficacy of the PDE5 inhibitors, sildenafil, tadalafil, and vardenafil to relax human corpus cavernosum (HCC) and vasodilate human penile resistance arteries (HPRA) from diabetic patients with ED (DMED). The influence of nebivolol on the capacity of these three PDE5 inhibitors to stimulate cyclic guanosine monophosphate (cGMP) production in HCC was also evaluated.MethodsHCC and HPRA were obtained from organ donors without ED (NEND; n = 18) or patients with diabetes undergoing penile prosthesis implantation (DMED; n = 19). Relaxations of HCC strips and HPRA to sildenafil, tadalafil, and vardenafil were evaluated in organ chambers and wire myographs. cGMP content in HCC was determined by ether extraction and quantification by ELISA.Main Outcome MeasuresEffects of nebivolol on PDE5 inhibitor‐induced relaxation of HCC, vasodilation of HPRA and cGMP accumulation in HCC.ResultsTreatment with nebivolol (1 μM) significantly potentiated sildenafil‐, tadalafil‐ and vardenafil‐induced relaxations of HCC and vasodilations of HPRA from both NEND and DMED. Enhancement of relaxant capacity by nebivolol resulted in reversion of the impairment of PDE5 inhibition‐induced responses in DMED and it was accompanied by enhancing the ability of PDE5 inhibitors to increase cGMP in HCC restoring reduced cGMP levels in HCC from DMED.ConclusionsNebivolol potentiated the capacity of PDE5 inhibitors to relax vascular structures of erectile tissue from diabetic patients by enhancing the nitric oxide (NO)/cGMP pathway in these tissues. These effects suggest a potential therapeutic utility of nebivolol as an adjunct to PDE5 inhibitors for the treatment of ED associated with diabetes. Martínez‐Salamanca JI, La Fuente JM, Cardoso J, Fernández A, Cuevas P, Wright HM, and Angulo J. Nebivolol potentiates the efficacy of PDE5 inhibitors to relax corpus cavernosum and penile arteries from diabetic patients by enhancing the NO/cGMP pathway. J Sex Med 2014;11:1182–1192.     

The Novel Antioxidant,AC3056 (2,6-di-t-butyl-4-((Dimethyl-4-Methoxyphenylsilyl)Methyloxy)Phenol), Reverses Erectile Dysfunction in Diabetic Rats and Improves NO-mediated Responses in Penile Tissue from Diabetic Men

IntroductionDiabetes is associated with a high incidence of erectile dysfunction (ED) and poor response to standard treatments. Oxidative stress could be relevant in the pathophysiology of diabetic ED.AimTo evaluate the effects of the antioxidant, AC3056 (2,6-di-t-butyl-4-((dimethyl-4-methoxyphenylsilyl)methyloxy)phenol), on diabetic ED.MethodsErectile responses to cavernosal nerve electrical stimulation were determined in streptozotocin-induced diabetic rats. Relaxation of human corpus cavernosal (HCC) tissue and penile resistance arteries (HPRA) from human cavernosal specimens was evaluated in organ chambers and myographs, respectively.Main Outcome MeasuresThe influence of AC3056 on erectile responses, lipid peroxidation, and nitrite plus nitrate serum content, and nuclear factor-κB (NF-κB) expression in penile tissue, in diabetic rats, and on endothelium-dependent and neurogenic relaxation of HCC and HPRA from diabetic patients was determined.ResultsEight weeks of diabetes caused ED in rats that was prevented by oral AC3056 (0.3% w/w in rat chow) when given from the induction of diabetes. AC3056 also prevented the diabetes-induced elevation of serum thiobarbituric acid-reactive substances (TBARS), the reduction of serum nitric oxide (NO) derivatives, and the increase of NF-κB expression. Acute oral administration of AC3056 (450 mg/kg) partially reversed ED in 8-week diabetic rats. Complete reversion of ED was achieved after 3 days of treatment with 0.3% AC3056. This effect remained after 5 weeks of treatment, but it disappeared after withdrawing for 1 week. Erectile function in diabetic rats was inversely related to serum TBARS. AC3056- (30 µM) reversed endothelial dysfunction in diabetic HCC and enhanced endothelium-dependent relaxation in diabetic HPRA and significantly potentiated neurogenic relaxation of both tissues. The reduced cGMP content in HCC from diabetic patients after exposure to acetylcholine (10 µM) was corrected by AC3056 (30 µM).ConclusionsThese results suggest that oxidative stress has a relevant role in pathophysiology of diabetic ED and provide a rationale for the use of antioxidant therapy in the treatment of ED in diabetes. Angulo J, Peiró C, Cuevas P, Gabancho S, Fernández A, González-Corrochano R, La Fuente JM, Baron AD, Chen KS, and Sáenz de Tejada I. The novel antioxidant, AC3056 (2,6-di-t-butyl-4-([dimethyl-4-methoxyphenylsilyl] methyloxy) phenol), reverses erectile dysfunction in diabetic rats and improves NO-mediated responses in penile tissue from diabetic men. J Sex Med 2009;6:373–387.     

Atorvastatin Ameliorates Sildenafil-Induced Penile Erections in Experimental Diabetes by Inhibiting Diabetes-Induced RhoA/Rho-Kinase Signaling Hyperactivation

IntroductionOne of the proposed mechanisms responsible for diabetes-related erectile dysfunction (ED) is overactivity of RhoA/ROCK signaling, as seen in experimental models of chemical diabetes.AimBecause statins may interfere with RhoA/Rho-kinase (ROCK) signaling through the reduction of geranyl-geranyl pyrophosphate (GGPP), required for RhoA activation, we investigated whether atorvastatin ameliorated diabetes-related ED.MethodsStreptozotocin-induced (8 weeks) diabetic rats and alloxan-induced (8 weeks) diabetic rabbits received atorvastatin (5 mg/kg daily) for the last 2 weeks. In vitro contractility studies were conducted in the rabbit model. In the rat model, sildenafil effect on electrical stimulation (ES)-induced erection was investigated. Atorvastatin action was also analyzed using human fetal penile smooth muscle cells (hfPSMCs) exposed to low (5 mM), high (22 mM), and very high (40 mM) glucose.Main Outcome MeasuresAtorvastatin effect on hyperglicemia-induced RhoA/ROCK signaling was evaluated using the ROCK inhibitor Y-27632 in both animal models and by analyzing functional effects downstream to RhoA activation in hfPSMCs.ResultsIn both diabetic models, atorvastatin did not affect glycemia, lipid plasma levels, and the hypogonadal state. In diabetic rats, atorvastatin ameliorated the erectile response to the ES of the cavernous nerve and normalized sildenafil effect on erectile function, strongly decreased by diabetes. In penile tissue from diabetic animals, atorvastatin completely restored the diabetes-induced hypersensitivity to Y-27632 and prevented RhoA membrane translocation/activation. In hfPSMCs, high glucose significantly increased not only membrane RhoA expression, but also ROCK activity (increased phosphorylation of the ROCK substrate myosin phosphatase target subunit 1) and several RhoA-dependent functions such as proliferation, migration, and smooth muscle-related gene expression. Atorvastatin restored all the high-glucose-induced effects, an action specifically reverted by GGPP.ConclusionAtorvastatin improves diabetes-related ED and restores sildenafil responsiveness, most probably by inhibiting RhoA/ROCK signaling, which underlies several high-glucose-induced derangements in penile smooth muscle cell commitment. Morelli A, Chavalmane AK, Filippi S, Fibbi B, Silvestrini E, Sarchielli E, Zhang X-H, Vignozzi L, Vannelli GB, Forti G, and Maggi M. Atorvastatin ameliorates sildenafil-induced penile erections in experimental diabetes by inhibiting diabetes-induced RhoA/Rho-kinase signaling hyperactivation. J Sex Med 2009;6:91–106.     

STIM/Orai Inhibition as a Strategy for Alleviating Diabetic Erectile Dysfunction Through Modulation of Rat and Human Penile Tissue Contractility and in vivo Potentiation of Erectile Responses

BackgroundStromal interaction molecule (STIM)/Orai calcium entry system appears to have a role in erectile dysfunction (ED) pathophysiology but its specific contribution to diabetic ED was not elucidated.AimTo evaluate STIM/Orai inhibition on functional alterations associated with diabetic ED in rat and human penile tissues and on in vivo erectile responses in diabetic rats.MethodsRat corpus cavernosum (RCC) strips from nondiabetic (No DM) and streptozotocin-induced diabetic (DM) rats and human penile resistance arteries (HPRA) and corpus cavernosum (HCC) from ED patients undergoing penile prosthesis insertion were functionally evaluated in organ chambers and wire myographs. Erectile function in vivo in rats was assessed by intracavernosal pressure (ICP) responses to cavernous nerve electrical stimulation (CNES). Expression of STIM/Orai elements in HCC was determined by immunofluorescence and immunoblot.Main Outcome MeasuresFunctional responses in RCC, HCC and HPRA and STIM/Orai protein expression in HCC. In vivo erectile responses to CNES.ResultsInhibition of Orai channels with YM-58483 (20 µM) significantly reduced adrenergic contractions in RCC but more effectively in DM. Thromboxane-induced and neurogenic contractions were reduced by STIM/Orai inhibition while defective endothelial, neurogenic and PDE5 inhibitor-induced relaxations were enhanced by YM-58483 (10 µM) in RCC from DM rats. In vivo, YM-58483 caused erections and attenuated diabetes-related impairment of erectile responses. YM-58483 potentiated the effects of PDE5 inhibition. In human tissues, STIM/Orai inhibition depressed adrenergic and thromboxane-induced contractions in ED patients more effectively in those with type 2 diabetes. Diabetes was associated with increased expression of Orai1 and Orai3 in ED patients.Clinical TranslationTargeting STIM/Orai to alleviate diabetes-related functional alterations of penile vascular tissue could improve erectile function and potentiate therapeutic effects of PDE5 inhibitors in diabetic ED.Strengths and LimitationsImproving effects of STIM/Orai inhibition on diabetes-related functional impairment was evidenced in vitro and in vivo in an animal model and validated in human tissues from ED patients. Functional findings were complemented with expression results. Main limitation was low numbers of human experiments due to limited human tissue availability.ConclusionsSTIM/Orai inhibition alleviated alterations of functional responses in vitro and improved erectile responses in vivo in diabetic rats, potentiating the effects of PDE5 inhibition. STIM/Orai inhibition was validated as a target to modulate functional alterations of human penile vascular tissue in diabetic ED where Orai1 and Orai3 channels were upregulated. STIM/Orai inhibition could be a potential therapeutic strategy to overcome poor response to conventional ED therapy in diabetic patients.Sevilleja-Ortiz A, El Assar M, García-Gómez B, et al. STIM/Orai Inhibition as a Strategy for Alleviating Diabetic Erectile Dysfunction Through Modulation of Rat and Human Penile Tissue Contractility and in vivo Potentiation of Erectile Responses. J Sex Med 2022;19:1733–1749.     

Nitrergic Function Is Lost but Endothelial Function Is Preserved in the Corpus Cavernosum and Penile Resistance Arteries of Men after Radical Prostatectomy

IntroductionRadical prostatectomy (RP) frequently results in erectile dysfunction (ED). It has been hypothesized that alterations of cavernosal tissue subsequent to RP contribute to ED but functional evaluation of the impact of RP on human erectile structures is lacking.AimThis study aims to evaluate endothelial function of human corpus cavernosum (HCC) and human penile resistance arteries (HPRA) and neurogenic responses of HCC from patients with ED secondary to RP (ED‐RP).MethodsHCC strips and HPRA were obtained from organ donors without history of ED (No‐ED) and patients with ED who were segregated depending on ED etiology: ED‐RP or vasculogenic (ED‐VASC). Functional evaluation of HCC and HPRA was performed in organ chambers and wire myographs, respectively. Histological evaluation of cavernosal tissue consisted of trichrome staining for fibrosis quantification and TUNEL assay for determination of apoptosis.Main Outcome MeasuresEndothelium‐dependent and endothelium‐independent relaxation, electrical field stimulation (EFS)‐induced neurogenic contraction and relaxation, and cavernosal fibrosis and apoptosis.ResultsEndothelium‐dependent relaxations were significantly impaired in HCC and HPRA from ED‐VASC patients while these responses in ED‐PR patients were not different to No‐ED. Similarly, sildenafil‐induced relaxations were reduced in HCC and HPRA from ED‐VASC but were preserved in ED‐RP. Adrenergic contractions induced by EFS in HCC were potentiated in both ED‐RP and ED‐VASC. EFS‐induced nitrergic relaxation was significantly reduced in HCC from ED‐VASC but was almost abolished in ED‐RP. Fibrous tissue content and cavernosal apoptosis in HCC from ED‐RP were not significantly different from No‐ED.ConclusionsEndothelial function and cavernosal sensitivity to phosphodiesterase type 5 inhibitors are preserved in erectile tissue from ED‐RP while a marked imbalance in neurogenic modulation of cavernosal tone favoring adrenergic contractile responses over nitrergic relaxation is manifested. Fibrotic and apoptotic processes in cavernosal tissue are not specifically associated to ED‐RP. These evidences could help to retarget therapeutic strategies in the management of ED after RP. Martínez‐Salamanca JI, La Fuente JM, Fernández A, Martínez‐Salamanca E, Pepe‐Cardoso AJ, Carballido J, and Angulo J. Nitrergic function is lost but endothelial function is preserved in the corpus cavernosum and penile resistance arteries of men after radical prostatectomy. J Sex Med 2015;12:590–599.     

Erectile Function Is Improved in Aged Rats by PnTx2‐6, a Toxin from Phoneutria nigriventer Spider Venom

IntroductionAge‐associated erectile dysfunction (ED) involves a decrease in nitric oxide (NO) availability and impaired relaxation. PnTx2‐6, a toxin from the Phoneutria nigriventer spider, has been demonstrated to improve erectile function via NO/cyclic guanosine monophosphate (cGMP) pathway. This spider's venom is characterized by several symptoms, including erection. PnTx2‐6 has been implicated in this phenomenon. Animal venoms have been postulated as potential drugs to treat ED.AimPnTx2‐6 toxin improves erectile function in aged rats via NO/cGMP. We investigated the effect of PnTx2‐6 in the erectile function of aged rats.Main Outcome MeasuresED was evaluated through changes in intracavernosal pressure/mean arterial pressure ratio during electrical field stimulation (EFS) of the pelvic ganglion of aged and adult rats (70 vs. 14 weeks). In functional studies, EFS‐induced relaxation of corpus cavernosum (CC) strips were performed with or without PnTx2‐6 (10‐8M).ResultsThe decrease in erectile function associated with age was partially restored 15–20 minutes after injection of PnTx2‐6 and further improved by sildenafil. PnTx2‐6 enhanced EFS‐induced relaxation, as well as cGMP levels in CC, from young and aged rats. Relaxation due to PnTx2‐6 was further increased after 30 minutes incubation with Y‐27632, a Rho‐kinase inhibitor (10‐6 M), in aging CC. Nitric oxide synthase (NOS) activity in aged and young cavernosal tissue was increased by incubation with PnTx2‐6 (10 minutes). However, this toxin did not modify NOS expression.ConclusionPnTx2‐6 improves penile relaxation in aged rats, via increased NOS activity and NO release, resulting in enhanced cGMP levels. Nunes KP, Toque HA, Borges MH, Richardson M, Webb RC, and de Lima ME. Erectile function is improved in aged rats by PnTx2‐6, a toxin from Phoneutria nigriventer spider venom. J Sex Med **;**:**–**.     

Poly(ADP‐Ribose) Polymerase Inhibition Improves Erectile Function by Activation of Nitric Oxide/Cyclic Guanosine Monophosphate Pathway in Diabetic Rats

IntroductionEndothelial dysfunction‐induced abnormalities of the nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) signaling pathway in the corpus cavernosum are thought to be the main factors involved in the pathogenesis of diabetes‐induced erectile dysfunction (ED). Recent studies have shown that the poly(adenosine diphosphate ribose) polymerase (PARP) pathway plays a critical role in diabetic endothelial dysfunction.AimThe aim of this study is to determine whether activation of the PARP pathway is involved in diabetic cavernosal endothelial dysfunction and abnormalities of the NO/cGMP pathway.MethodsMale Sprague‐Dawley rats were randomly divided into three groups: age‐matched controls, diabetic controls (DM), and the 3‐aminobenzamide (3‐AB, a PARP inhibitor)‐treated diabetic group (DM+3‐AB). Diabetes was induced by intraperitoneal injection of streptozotocin. Eight weeks after inducing diabetes, the DM+3‐AB group was treated with 3‐AB for 4 weeks.Main Outcome MeasuresErectile function was assessed at 12 weeks after inducing diabetes by stimulating the cavernous nerve. Expression of poly(ADP‐ribose), protein kinase B (Akt), phospho‐Akt, endothelial nitric oxide synthase (eNOS), phospho‐eNOS, and neuronal nitric oxide synthase (nNOS) were evaluated by Western blot. Cavernous NO generation and cGMP levels were also determined.ResultsThe DM group showed impaired erectile function and significantly increased PARP activity. Expression of total eNOS and nNOS, phospho‐Akt, and eNOS decreased significantly in the DM group compared with those in the control group. In addition, cavernous NO generation and cGMP levels decreased significantly in the DM group compared with those in the control group. Treatment with 3‐AB restored erectile function and significantly reversed all molecular alterations except decreased nNOS expression.ConclusionOveractivation of the PARP pathway in the corpus cavernosum of diabetic rats was involved in cavernosal endothelial dysfunction and abnormalities of the NO/cGMP pathway resulting in ED. These findings may be applied to develop novel therapies for patients with diabetic ED. Li WJ, Peng Y, Zhou J, Li B, Wang H, Zhang J, and Wang Z. Poly(ADP‐ribose) polymerase inhibition improves erectile function by activation of nitric oxide/cyclic guanosine monophosphate pathway in diabetic rats. J Sex Med 2012;9:1319–1327.     

P144, A TGF‐β1 Antagonist Peptide,Synergizes with Sildenafil and Enhances Erectile Response via Amelioration of Cavernosal Fibrosis in Diabetic Rats

IntroductionPatients with diabetes exhibit more severe erectile dysfunction (ED) and are less responsive to first‐line oral phosphodiesterase type 5 inhibitor (PDE5i). It has been suggested that increased collagen deposition and reduced smooth muscle content in the corpus cavernosum are important mechanisms for diabetes‐associated ED and that transforming growth factor‐β1 (TGF‐β1) is a potent fibrotic factor responsible for the structural alterations in the corpus cavernosum.AimsThe aims of this study are to determine whether activation of TGF‐β1 and its downstream pathways is responsible for the reduced efficacy of the PDE5is in diabetic ED via abnormalities in cavernosal structures and to investigate the synergistic effects of the TGF‐β1 antagonist P144 and sildenafil on erectile response.MethodsSix weeks after inducting diabetes with streptozotocin in male Sprague‐Dawley rats, age‐matched control and diabetic rats were treated with vehicle, sildenafil, or P144 alone or in combination for 4 weeks, respectively.Main Outcome MeasuresIntracavernous pressure, dynamic infusion cavernosometry, and histological and molecular alterations of the corpus cavernosum were analyzed.ResultsDiabetic rats exhibited a decreased erectile response, severe corporal veno‐occlusive dysfunction (CVOD), and structural alterations including cavernosal fibrosis and decreased smooth muscle content. Expression and activation of TGF‐β1 and its downstream Smad and non‐Smad pathways increased in diabetic rats. Treatment with sildenafil showed modest effect on erectile response and a less suppressive effect on CVOD, cavernosal fibrosis, and molecular alterations. Treatment with P144 had lower effect on erectile response, even greatly improved the histological and molecular alterations and CVOD than sildenafil. The combined treatment with P144 and sildenafil effectively restored erectile response, CVOD, and histological and molecular alterations.ConclusionAn insufficient suppressive effect of sildenafil on cavernosal fibrosis, severe CVOD, and TGF‐β1 pathways was implicated in reduced efficacy of the PDE5i in diabetic ED. Treatment with P144 synergized sildenafil and significantly increased erectile response by the potential antifibrotic activity. Li WJ, Wang H, Zhou J, Li B, Zhang J, Lu M, and Wang Z. P144, a TGF‐β1 antagonist peptide, synergizes with sildenafil and enhances erectile response via amelioration of cavernosal fibrosis in diabetic rats. J Sex Med 2013;10:2942–2951.     

Enhanced Contribution of Orai Channels to Contractility of Human Penile Smooth Muscle in Erectile Dysfunction

BackgroundStore-operated calcium entry and its key players, stromal interaction molecule (STIM) and Orai calcium channels, have been proposed as emergent therapeutic targets in cardiovascular pathophysiology. We hypothesize alteration of STIM/Orai signaling in erectile dysfunction (ED).AimTo evaluate the contribution of STIM/Orai to human penile tissue contraction and to analyze the influence of ED on STIM/Orai signaling at functional and expression levels in human penile vascular tissues.MethodsHuman penile resistance arteries (HPRA) and human corpus cavernosum (HCC) were dissected from cavernosal specimens from 30 organ donors without history of ED (No ED) and from 48 patients with ED undergoing penile prosthesis insertion and functionally evaluated in wire myographs and organ chambers, respectively. Expression of STIM-1, Orai1, and Orai3 in HCC was localized and quantified by immunofluorescence.Main Outcome MeasuresThe main outcome measures are functional responses in HCC and HPRA and STIM/Orai channel protein expression in human cavernosal tissue.ResultsInhibition of Orai channels with YM-58483 (20 μM) significantly reduced norepinephrine–induced contractions in both HCC and HPRA from either No ED or ED subjects, but the effects were more marked in ED (−20.1 ± 5.9% vs −45.5 ± 13.2% and −15.9 ± 4.0% vs −31.4 ± 6.9% reduction in E_max to norepinephrine in HCC and HPRA, respectively). Thromboxane-induced contractions were reduced and neurogenic contractile and relaxant responses modulated by Orai inhibition in penile tissues from patients with ED. In fact, addition of YM-58483 concentration dependently relaxed precontracted HPRA and HCC. These relaxations were significantly more pronounced in tissues from patients with ED (EC₅₀ 7.5 vs 1.3 μM and 10.5 vs 1.3 μM, for HCC and HPRA, respectively). All HCC specimens displayed expression of STIM-1, Orai1, and Orai3. Significantly increased expression of Orai1 and Orai3 but not STIM-1 was observed in patients with ED.Clinical TranslationInhibition of enhanced Orai activity in human penile vascular tissue could facilitate erectile responses, alleviating ED.Strengths and LimitationsEnhanced STIM/Orai activity contribution to penile smooth muscle tone in ED is demonstrated at functional and structural levels in human tissues from a representative sample of patients with ED and in comparison with healthy tissue. We cannot differentiate the specific contribution of risk factors associated with ED to hyperactivity of the Orai system.ConclusionsOrai channels significantly contribute to human penile smooth muscle contraction. Orai contribution to penile smooth muscle tone is functionally enhanced in ED accompanied by increased expression of Orai channels in cavernosal tissue. Orai inhibition could be a potential therapeutic strategy to reduce penile smooth muscle contraction in ED.Sevilleja-Ortiz A, El Assar M, García-Rojo E, et al. Enhanced Contribution of Orai Channels to Contractility of Human Penile Smooth Muscle in Erectile Dysfunction. J Sex Med 2020;17:881–891.     

Nitric Oxide‐Releasing Polymeric Microspheres Improve Diabetes‐Related Erectile Dysfunction

IntroductionWe have used a long‐acting nitric oxide (NO)‐releasing polymer to develop injectable biodegradable microspheres capable of localized NO release over prolonged periods of time.AimThe aim of this study was to evaluate the therapeutic potential of these microspheres for diabetes‐related erectile dysfunction (ED) in the rat model.MethodsNO‐releasing microspheres were incubated in physiologic buffer, and in vitro NO release was measured using a Griess assay. To ensure no migration, microspheres were fluorescently tagged and injected into the corpus cavernosum of adult rats, and fluorescent imaging was performed weekly for 4 weeks, at which point rats were sacrificed. To assess physiologic efficacy, diabetes was induced in 40 rats using streptozotocin (STZ), whereas 10 rats were kept as age‐matched controls. Diabetic rats were divided into four groups: no treatment, sildenafil, NO‐releasing microspheres, and combination therapy. For each rat, the cavernosal nerve (CN) was stimulated at various voltages, and intracavernosal pressure (ICP) and mean arterial pressure (MAP) were measured via corpus cavernosum and carotid artery catheterization, respectively. Long‐term efficacy was determined by injecting diabetic rats with microspheres and measuring erectile response at predetermined intervals for up to 5 weeks.Main Outcome MeasuresErectile response was determined via calculation of mean peak ICP/MAP and area under curve (AUC) for each experimental group.ResultsUnder physiologic conditions in vitro, microspheres continued NO release for up to 4 weeks. Fluorescent imaging revealed no detectable signal in tissues besides cavernosal tissue at 4 weeks postinjection. Upon CN stimulation, peak ICP/MAP ratio and AUC of diabetic rats improved significantly (P < 0.05) in microsphere and combination therapy groups compared with no treatment and sildenafil groups. In long‐term efficacy studies, microspheres augmented the effect of sildenafil for 3 weeks following injection (P < 0.05).ConclusionsNO‐releasing microspheres significantly improved erectile response in diabetic rats for 3 weeks and hence offer a promising approach to ED therapy, either as monotherapy or combination therapy. Soni SD, Song W, West JL, and Khera M. Nitric oxide‐releasing polymeric microspheres improve diabetes‐related erectile dysfunction. J Sex Med 2013;10:1915‐1925.     

Effect of Mucuna pruriens (Linn.) on oxidative stress-induced structural alteration of corpus cavernosum in streptozotocin-induced diabetic rat

IntroductionErectile dysfunction is one of the major secondary complications of diabetes. Mucuna pruriens (M. pruriens), a leguminous plant identified for its antidiabetic, aphrodisiac, and fertility enhancing properties, has been the choice of Indian traditional medicine.AimThe objective of the present study was to analyze the efficacy of M. pruriens on free radicals‐mediated penile tissue alterations in hyperglycemic male rats.MethodsMale albino rats were divided as group I (sham) control, group II (STZ) diabetes‐induced (streptozotocin 60 mg/kg of body weight [bw] in 0.1 M citrate buffer), group III (STZ + MP) diabetic rats administered with 200 mg/kg bw of ethanolic extract of M. pruriens seed, group IV (STZ + SIL) diabetic rats administered with 5 mg/kg bw of sildenafil citrate, group V (sham + MP) administered with 200 mg/kg bw of extract alone, and group VI (sham + SIL) administered with 5 mg/kg bw of sildenafil citrate. The M. pruriens and sildenafil citrate were given (gavage) once daily for a period of 60 days. At the end of 60 days, the animals were sacrificed and subjected to analysis of reactive oxygen species levels, enzymic and nonenzymic antioxidant levels, levels of NOx, histological, and histomorphometrical study of penile tissue.Main Outcome MeasuresRemedial use of M. pruriens seed extract on diabetes‐induced erectile tissue damage.ResultsSignificantly high levels of oxidative stress and low levels of antioxidants in the penile tissue seem to contribute to the increased collagen deposition and fibrosis of erectile tissue in STZ rats. Relatively, there was increased damage in STZ + SIL group. Supplementation of M. pruriens in STZ + MP group has revealed the potency to overcome oxidative stress, and good preservation of penile histoarchitecture.ConclusionThe ethanolic extract of M. pruriens seed significantly recovered or protected erectile tissue from the oxidative stress‐induced degeneration by its antioxidant potentials. These findings propound to serve mankind by the treatment of diabetes‐induced erectile dysfunction. Suresh S and Prakash S. Effect of Mucuna pruriens (Linn.) on oxidative stress‐induced structural alteration of corpus cavernosum in streptozotocin‐induced diabetic rat. J Sex Med 2011;8:1943–1956.     

Chronic Administration of Sildenafil Modified the Impaired VEGF System and Improved the Erectile Function in Rats with Diabetic Erectile Dysfunction

IntroductionMen frequently develop diabetic erectile dysfunction (DMED), as a result of endothelial dysfunction. DMED patients often have reduced efficacy with phosphodiesterase type 5 inhibitors therapy.AimTo determine whether chronic sildenafil administration can modify the impaired vascular endothelial growth factor (VEGF) system and improve the erectile function in rats with diabetic erectile dysfunction.MethodsA group of Sprague Dawley rats (n = 30) with DMED were induced by intraperitoneal injection of streptozotocin (40 mg/kg) and screened by subcutaneous injection of Apomorphine (100 mg/kg). They were then exposed to either vehicle or sildenafil (prescribed in our hospital, 5 mg/kg and 10 mg/kg, respectively) for 10 weeks. An additional nondiabetic and age-matched control group (n = 10) was also allocated and given the routine diet for the same period. Assessments were performed to both groups at 36 hours after the last dose of sildenafil. Penile intracavernous pressure (ICP), mean arterial pressure (MAP), penile tissue morphology, immunohistologic analysis, and Western blot analysis of VEGF, VEGFR1, and eNOS were determined.Main Outcome MeasureFunctional, morphological, and proteomical changes on penile structures by the chronic Sildenafil (5 mg/kg and 10 mg/kg, respectively) administration were determined.ResultsA significant increase of ICP, ICP/MAP ratio, and area under the curve were observed in the both groups treated by sildenafil (5 mg/kg and 10 mg/kg, respectively), compared with the DMED rats without receiving Sildenafil. Immunohistochemical staining of their penile tissue showed a decrease in VEGF, VEGFR1, and eNOS staining in the controlled group compared with an improvement in the chronic sildenafil administration group. Western blot analysis demonstrated exactly the same results.ConclusionWe demonstrated that daily sildenafil administration can restore the impaired VEGF system in the penis of DMED rats and progressively improve both erectile function and endothelial function, suggesting a potential general mechanism of improved signaling through the VEGF/eNOS signaling cascade. Liu G, Sun X, Dai Y, Zheng F, Wang D, Huang Y, Bian J, and Deng C. Chronic administration of sildenafil modified the impaired VEGF system and improved the erectile function in rats with diabetic erectile dysfunction.     

TNF‐α, Erectile Dysfunction,and NADPH Oxidase‐Mediated ROS Generation in Corpus Cavernosum in High‐Fat Diet/Streptozotocin‐Induced Diabetic Rats

IntroductionPatients with diabetes‐associated erectile dysfunction (ED) are characterized by an increase in circulating tumor necrosis factor‐alpha (TNF‐α). However, no study has indicated whether and how TNF‐α plays a role in the pathogenesis of ED associated with diabetes.AimWe examined the effects and potential mechanism of infliximab (INF), a chimeric monoclonal antibody to TNF‐α, on reactive oxygen species (ROS) generation in corpus cavernosum and ED in diabetic rats.MethodsFour groups of male rats were used: age‐matched normal controls; diabetic rats induced by a high‐fat diet (HFD) combined with a single streptozotocin (STZ) injection (35 mg/kg body weight, intraperitoneal [i.p.]); nondiabetic rats receiving INF (5 mg/kg body weight/week, i.p.), and diabetic rats receiving INF. Erectile function was assessed with electrical stimulation of the cavernous nerve after 8 weeks. The blood and penile tissues were harvested for plasma biochemical determinations, serum TNF‐α measurement, penile ROS detection, and molecular assays of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits, endothelial nitric oxide synthase (eNOS), phospho‐eNOS, and neural nitric oxide synthase (nNOS) in the penis.Main Outcome MeasuresThe effect of INF on HFD/STZ‐induced diabetic ED and NADPH oxidase‐mediated ROS generation was studied in diabetic corpus cavernosum.ResultsUntreated diabetic rats displayed significantly decreased erectile parameters, and increased plasma TNF‐α levels, penile ROS production, p47^phox and gp91^phox expression compared with nondiabetic controls. INF neutralized TNF‐α and significantly reduced ED in diabetic rats, in which marked decreases in p47^phox and gp91^phox expression and ROS generation in corpus cavernosum were noted. The ratio of phospho‐eNOS to eNOS and expression of nNOS in the penis were significantly increased in INF‐treated vs. untreated diabetic rats.ConclusionsIncreased TNF‐α expression associated with diabetes contributes to ED by promoting NAPDH oxidase‐mediated ROS generation in corpus cavernosum. INF protects against diabetic ED by neutralizing TNF‐α. Long T, Liu G, Wang Y, Chen Y, Zhang Y, and Qin D. TNF‐α, erectile dysfunction, and NADPH oxidase‐mediated ROS generation in corpus cavernosum in high‐fat diet/streptozotocin‐induced diabetic rats. J Sex Med 2012;9:1818–1831.     

Vardenafil and resveratrol synergistically enhance the nitric oxide/cyclic guanosine monophosphate pathway in corpus cavernosal smooth muscle cells and its therapeutic potential for erectile dysfunction in the streptozotocin-induced diabetic rat: preliminary findings

IntroductionPhosphodiesterase type 5 (PDE5) inhibitors are very effective agents for erectile dysfunction; however, specific patient populations are hard to treat. The efficacy of PDE5 inhibitors is limited because a minimum amount of nitric oxide (NO) is necessary. Resveratrol, a plant polyphenol, is reported to activate endothelial NO synthase (eNOS) through activation of sirtuin 1. We previously reported that human corpus cavernosal smooth muscle cells (CCSMCs) express eNOS and synthesize cyclic guanosine monophosphate (cGMP) via the NO/cGMP pathway.AimTo investigate the ability of resveratrol and/or vardenafil to increase cGMP in an in vitro model using CCSMCs and to improve erectile function in an in vivo rat model of streptozotocin (STZ)‐induced diabetes.MethodsCCSMCs were treated with resveratrol and/or vardenafil. Twenty male Sprague‐Dawley rats were randomly divided into five groups (N = 4 in each group): age‐matched controls, diabetic controls, and diabetic rats treated with resveratrol, vardenafil, or both in combination for the last 4 weeks of an 8‐week period of diabetes induction.Main Outcome MeasuresIntracellular cGMP measurement, intracovernous pressure (ICP)/mean arterial pressure (MAP) ratio, and smooth muscle/collagen ratio.ResultsIntracellular cGMP level was elevated by resveratrol treatment in CCSMCs. The combination treatment of resveratrol and vardenafil had a synergistic effect. Diabetic rats showed impairment of erectile function. Treatment with either resveratrol or vardenafil improved ICP/MAP ratio, and combination therapy with resveratrol and vardenafil had a synergistic effect in improvement of ICP/MAP.ConclusionsTreatment with either resveratrol or vardenafil elevated cGMP level in CCSMCs and improved erectile function in STZ‐induced diabetic rats. Furthermore, a synergistic effect was observed in vitro and in vivo. Resveratrol or combination therapy of resveratrol and vardenafil can improve erectile function in which NO release is impaired, although further study is needed to confirm the results. Fukuhara S, Tsujimura A, Okuda H, Yamamoto K, Takao T, Miyagawa Y, Nonomura N, and Okuyama A. Vardenafil and resveratrol synergistically enhance the nitric oxide/cyclic guanosine monophosphate pathway in corpus cavernosal smooth muscle cells and its therapeutic potential for erectile dysfunction in the streptozotocin‐induced diabetic rat: Preliminary findings.     

Edaravone ameliorates diabetes-induced dysfunction of NO-induced relaxation in corpus cavernosum smooth muscle in the rat

IntroductionDiabetes mellitus (DM) represents a major risk factor for erectile dysfunction (ED). Although the etiology of diabetes-induced ED is multifactorial and still unknown, reactive oxygen species are thought to be one of the key factors.AimThe aim of this article is to investigate whether administration of edaravone, a free radical scavenger, could prevent type 1 diabetes-induced dysfunction of nitric oxide (NO)-induced relaxation in corpus cavernosum smooth muscle in the rat.MethodsSix-week-old male Wistar rats were randomly divided into three groups. One group was treated with citrate-phosphate buffer plus normal saline (group Cont), whereas in the other two groups, diabetes was induced by streptozotocin (50 mg/kg intraperitoneally [i.p.]). Subsequently, the diabetic rats were treated for 4 weeks either with edaravone (10 mg/kg/day, i.p.; group DM + E) or with normal saline (group DM).Main Outcome MeasuresSerum glucose and malondialdehyde levels as well as penile cyclic guanosine monophosphate (cGMP) concentrations were determined, and penile function was estimated by organ bath studies with norepinephrine-mediated contractions and acetylcholine-mediated relaxations. The participation mRNA levels of muscarinic M₃ receptors, neuronal nitrous oxide synthase (nNOS), endothelial NOS (eNOS) and inducible NOS (iNOS), and participation protein levels of nNOS, eNOS, phosphorylated nNOS, and phosphorylated eNOS were investigated by quantitative real-time polymerase chain reaction (PCR) and immunoblot analysis, respectively.ResultsTreatment with edaravone prevented partially but significantly the decreased body and penile weight induced by diabetes. Treatment with edaravone significantly improved the increased diabetes-induced malondialdehyde levels, the decreased penile cGMP concentrations, the increased diabetes-induced norepinephrine-mediated contractions, and the decreased acetylcholine-mediated relaxation. Although there were no significant differences in expression levels of mRNAs in nNOS, diabetes-induced upregulation of muscarinic M₃ receptor and iNOS mRNAs as well as diabetes-induced downregulations of eNOS, phosphorylated nNOS, and phosphorylated eNOS were significantly prevented by edaravone.ConclusionsEdaravone decreases the oxidative insult in the penile corpus cavernosum by ameliorating the NO–NOS system and thus preventing partially the developing ED in DM in the rat. Ohmasa F, Saito M, Tsounapi P, Dimitriadis F, Inoue S, Shomori K, Shimizu S, Kinoshita Y, and Satoh K. Edaravone ameliorates diabetes-induced dysfunction of NO-induced relaxation in corpus cavernosum smooth muscle in the rat.     

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.     

Impaired Cavernous Reinnervation after Penile Nerve Injury in Rats with Features of the Metabolic Syndrome

IntroductionThe metabolic syndrome is a cluster of cardiovascular risk factors that predispose toward the development of diseases such as diabetes. Erectile dysfunction (ED) is common in men with metabolic syndrome, but its etiology is poorly understood. Pro‐erectile nitrergic nerves innervating penile erectile tissue are also susceptible to mechanical injury during pelvic surgical procedures, which can lead to sexual dysfunction.AimsThe aims of this article are: (i) to examine erectile function in an experimental model of metabolic syndrome, the phosphoenolpyruvate carboxykinase (PEPCK)‐overexpressing rat; and (ii) to study function and cavernous reinnervation after penile nerve crush injury, which permits regeneration, in transgenic rats.MethodsWe analyzed the density of noradrenergic and nitrergic nerves and performed organ bath pharmacology to assess neurogenic activity.Main Outcome MeasuresBy analyzing changes in neural structure, function, and pharmacologic responses of cavernous tissue after nerve crush injury, we were able to reveal neurologic deficits in rats with metabolic syndrome.ResultsAnimals with features of metabolic syndrome did not develop notable changes in cavernous autonomic nerve density or nerve‐evoked smooth muscle activity. However, regeneration of nitrergic nerves after crush injury in transgenic rats was impaired compared with injured controls. This was manifested as a deficit in axon regrowth and responses to axon activation. However, unlike injured controls, injured PEPCK‐overexpressing rats did not develop a reduced maximal response to the nitric oxide (NO) donor, sodium nitroprusside. This suggests preserved NO responsiveness in tissues from rats with metabolic syndrome, despite impaired regeneration and return of function.ConclusionsThis study revealed that rats with features of metabolic syndrome display impaired cavernous nerve regeneration after penile nerve injury, but the degree of functional impairment may be attenuated due to reduced plasticity of NO signaling. This reinnervation deficit may be of clinical relevance for understanding why ED persists in some (particularly aged) men after pelvic surgery. Nangle MR, Proietto J, and Keast JR. Impaired cavernous reinnervation after penile nerve injury in rats with features of the metabolic syndrome.     

Exercise training improves the defective centrally mediated erectile responses in rats with type I diabetes

IntroductionErectile dysfunction is a serious and common complication of diabetes mellitus. Apart from the peripheral actions, central mechanisms are also responsible for the penile erection.AimThe goal of the present study was to determine the impact of exercise training (ExT) on the centrally mediated erectile dysfunction in streptozotocin (STZ)‐induced type I diabetic (T1D) rats.MethodsMale Sprague–Dawley rats were injected with STZ to induce diabetes mellitus. Three weeks after STZ or vehicle injections, rats were assigned to either ExT (treadmill running for 3–4 weeks) or sedentary groups to produce four experimental groups: control + sedentary, T1D + sedentary, control + ExT, and T1D + ExT.Main Outcome MeasureAfter 3–4 weeks ExT, central N‐methyl‐D‐aspartic acid (NMDA) or sodium nitroprusside (SNP)‐induced penile erectile responses were measured. Neuronal nitric oxide synthase (nNOS) expression in the paraventricular nucleus (PVN) of the hypothalamus was measured by using histochemistry, real time polymerase chain reaction (PCR) and Western blot approaches.ResultsIn rats with T1D, ExT significantly improved the blunted erectile response, and the intracavernous pressure changes to NMDA (50 ng) microinjection within the PVN (T1D + ExT: 3.0 ± 0.6 penile erection/rat; T1D + sedentary: 0.5 ± 0.3 penile erection/rat within 20 minutes, P < 0.05). ExT improved erectile dysfunction induced by central administration of exogenous nitric oxide (NO) donor, SNP in T1D rats. Other behavior responses including yawning and stretching, induced by central NMDA and SNP microinjection were also significantly increased in T1D rats after ExT. Furthermore, we found that ExT restored the nNOS mRNA and protein expression in the PVN in T1D rats.ConclusionThese results suggest that ExT may have beneficial effects on the erectile dysfunction in diabetes through improvement of NO bioavailability within the PVN. Thus, ExT may be used as therapeutic modality to up‐regulate nNOS within the PVN and improve the central component of the erectile dysfunction in diabetes mellitus. Zheng H, Mayhan WG, and Patel KP. Exercise training improves the defective centrally mediated erectile responses in rats with type I diabetes. J Sex Med 2011;8:3086–3097.