Erectile Dysfunction in Young Non-Obese Type II Diabetic Goto-Kakizaki Rats is Associated with Decreased eNOS Phosphorylation at Ser1177

IntroductionDiabetes mellitus (DM) is a risk factor for erectile dysfunction (ED). Although type 2 DM is responsible for 90–95% diabetes cases, type 1 DM experimental models are commonly used to study diabetes-associated ED.AimGoto-Kakizaki (GK) rat model is relevant to ED studies since the great majority of patients with type 2 diabetes display mild deficits in glucose-stimulated insulin secretion, insulin resistance, and hyperglycemia. We hypothesized that GK rats display ED which is associated with decreased nitric oxide (NO) bioavailability.MethodsWistar and GK rats were used at 10 and 18 weeks of age. Changes in the ratio of intracavernosal pressure/mean arterial pressure (ICP/MAP) after electrical stimulation of cavernosal nerve were determined in vivo. Cavernosal contractility was induced by electrical field stimulation (EFS) and phenylephrine (PE). In addition, nonadrenergic-noncholinergic (NANC)- and sodium nitroprusside (SNP)-induced relaxation were determined. Cavernosal neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS) mRNA and protein expression were also measured.Main Outcome MeasureGK diabetic rats display ED associated with decreased cavernosal expression of eNOS protein.ResultsGK rats at 10 and 18 weeks demonstrated impaired erectile function represented by decreased ICP/MAP responses. Ten-week-old GK animals displayed increased PE responses and no changes in EFS-induced contraction. Conversely, contractile responses to EFS and PE were decreased in cavernosal tissue from GK rats at 18 weeks of age. Moreover, GK rats at 18 weeks of age displayed increased NANC-mediated relaxation, but not to SNP. In addition, ED was associated with decreased eNOS protein expression at both ages.ConclusionAlthough GK rats display ED, they exhibit changes in cavernosal reactivity that would facilitate erectile responses. These results are in contrast to those described in other experimental diabetes models. This may be due to compensatory mechanisms in cavernosal tissue to overcome restricted pre-penile arterial blood supply or impaired veno-occlusive mechanisms. Carneiro FS, Giachini FRC, Carneiro ZN, Lima VV, Ergul A, Webb RC, and Tostes RC. Erectile dysfunction in young non-obese type II diabetic Goto-Kakizaki rats is associated with decreased eNOS phosphorylation at Ser¹¹⁷⁷.     

AGE-Breaker ALT-711 Plus Insulin Could Restore Erectile Function in Streptozocin-Induced Type 1 Diabetic Rats

IntroductionThe interaction between advanced glycation end-products (AGEs) and its receptors for AGEs (RAGEs) elicits oxidative stress and mediates the development of erectile dysfunction (ED). ALT-711, an AGE cross-link breaker, has the therapeutic potential for ED but has been less intensively investigated.AimThe aim of this study was to investigate the effects of an AGEs breaker 3-phenacyl-4,5-dimethylthiazolium chloride (ALT-711) plus insulin on erectile function in streptozocin (STZ)-induced type 1 diabetic rats.MethodsFifty 8-week-old Sprague-Dawley rats were randomly distributed into five groups: normal control (C), diabetic (D), insulin-treated diabetic (D + I), ALT-711-treated diabetic (D + ALT-711) and insulin plus ALT-711-treated diabetic (D + I + ALT-711) rats. Diabetes was induced by a single intraperitoneal injection of STZ. Eight weeks after induction of diabetes, ALT-711 was administered by intraperitoneal injection. Two to six units of intermediate-acting insulin were utilized to achieve normal levels of glycemic control. After treatment for 6 weeks, erectile function was determined via measurement of intracavernous pressures (ICPs) following electrostimulation of the cavernous nerve. The deposition of AGEs, expression of RAGEs, superoxide dismutase activity, and lipid peroxidation were measured. We also evaluated penile histological changes such as smooth muscle contents, endothelial cells contents, and apoptotic activity.Main Outcome MeasuresThe main outcome measures were the ratio of ICP/mean arterial pressure (MAP), penile endothelial cells, smooth muscle cells, neuronal nitric oxide synthase, AGE and RAGE expression, malondialdehyde concentration, SOD activity, and apoptosis index.ResultsDiabetic rats demonstrated significantly reduced ICP/MAP ratio, penile endothelial cells, smooth muscles cells, increased AGEs and RAGE expression, and increased apoptosis. Insulin and ALT-711 monotherapy partially restored erectile function and histological changes. However, the combination therapy group showed erectile parameters and components similar to those in C. ALT-711-treated group demonstrated less deposition of AGEs and lower expression of RAGE than those in insulin-treated group.ConclusionThese results suggest that although insulin can effectively control glycemic levels, it does not completely alter the pathological changes in erectile tissues. Better efficacy could be expected with tight glycemic control plus ALT-711, an AGEs cross-link breaker. The combination therapy might have the potential to eliminate metabolic memory by down-regulating the AGEs–RAGE oxidative stress axis. Wang L, Tian W, Uwais Z, Li G, Li H, Guan R, Gao Z, and Xin Z. AGE-breaker ALT-711 plus insulin could restore erectile function in streptozocin-induced type 1 diabetic rats. J Sex Med 2014;11:1452–1462.     

Erectile Dysfunction Precedes Coronary Artery Endothelial Dysfunction in Rats Fed a High‐Fat,High‐Sucrose,Western Pattern Diet

IntroductionIt is suggested that erectile dysfunction (ED) may be an early risk factor for cardiovascular disease.AimThe goal of this study was to determine whether development of ED precedes the onset of coronary artery endothelial dysfunction in response to a Western diet (WD), thereby establishing whether the WD differentially impacts the endothelium in a time‐dependent manner. Additionally, a goal was to determine if diet‐induced ED is reversible with intracavernosal sepiapterin treatment.MethodsMale Sprague‐Dawley rats were fed a WD for 4, 8, or 12 weeks, or a control diet for 8 weeks. Erectile function was evaluated by measuring the mean arterial pressure (MAP) and intracavernosal pressure (ICP) in response to electrical field stimulation of the cavernosal nerve near the major pelvic ganglion, in the absence and presence of sepiapterin. Coronary artery endothelial function was evaluated ex vivo with cumulative doses of acetylcholine (ACh) applied to segments of the left anterior descending coronary artery preconstricted with serotonin.Main Outcome MeasuresErectile function was assessed as the ICP response to electrical field stimulation (EFS), normalized to MAP. Coronary artery endothelial function was assessed as the effective concentration producing 50% of a maximal response (EC₅₀) of the ACh response.ResultsThe ICP/MAP response to EFS was significantly attenuated following both 8 and 12 weeks of the WD compared with the control diet (P < 0.05). Sepiapterin treatment augmented the ICP/MAP response in all WD groups (P < 0.05). The coronary artery EC₅₀ of the ACh response was not different from control following 4 or 8 weeks but was significantly elevated following 12 weeks of the WD (P < 0.01).ConclusionsThese data suggest that erectile function is reduced prior to coronary artery endothelial function in response to the WD. Improvement of erectile function with sepiapterin in WD rats indicates that nitric oxide synthase uncoupling is a key mechanism in diet‐induced ED. La Favor JD, Anderson EJ, Hickner RC, and Wingard CJ. Erectile dysfunction precedes coronary artery endothelial dysfunction in rats fed a high‐fat, high‐sucrose, Western pattern diet. J Sex Med 2013;10:694–703.     

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.     

Investigation of the Effects of the Level of Glycemic Control on Erectile Function and Pathophysiological Mechanisms in Diabetic Rats

IntroductionPoor glycemic control is associated with erectile dysfunction (ED); however, differences in ED according to the level of glycemic control have been poorly investigated.AimThe aim of this paper is to investigate the change in erectile function according to the level of glycemic control and to clarify the pathophysiological mechanism of diabetes-associated ED.MethodsStreptozotocin was injected into 55 male Sprague-Dawley rats classified into four groups: control (group 1), diabetes with multiple insulin injections (group 2), diabetes with a single injection (group 3), and untreated diabetes (group 4). Daily insulin injections in groups 2 and 3 were administered for 4 weeks after 10 weeks of diabetic induction.Main Outcome MeasuresThe main outcome measures are the anova or Kruskal–Wallis tests to evaluate glycosylated hemoglobin (HbA1c), testosterone levels, the ratios of intracavernosal pressure to mean arterial pressure (ICP/MAP), area under the ICP curve to MAP (AUC/MAP), and changes in cavernous tissue and protein expression related to Rho kinase and nitric oxide pathways.ResultsHbA1c levels were different between pairs of groups. Group 4 showed the lowest erectile parameters and group 2 showed near normal level. No differences in erectile parameters were found between groups 1 and 2 or between groups 3 and 4, except the ratio of AUC to MAP for group 1 was significantly higher than that of group 2 (20 Hz stimulation). Decrease in erectile function of group 2 was related to decreased expression of nitrergic nitric oxide synthase or decreased testosterone level compared with group 1. Groups 2 and 3 showed significant differences in erectile parameters, which were associated with difference in apoptotic index. Groups 3 and 4 showed no differences in erectile parameters, although these groups had significant differences in apoptotic index, smooth muscle component, and protein expression ratios of phosphorylated to total myosin phosphatase target subunit 1, endothelial nitric oxide synthase, and Akt.ConclusionsImprovement in glycemic control assists recovery from diabetes-associated ED; however, only tight glycemic control can provide recovery from ED to a near normal status. Cho SY, Chai JS, Lee SH, Park K, Paick J-S, and Kim SW. Investigation of the effects of the level of glycemic control on erectile function and pathophysiological mechanisms in diabetic rats. J Sex Med 2012;9:1550–1558.     

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.     

Impact of systemically active neurohumoral factors on the erectile response of the rat

IntroductionMean arterial pressure (MAP) and specific regulation of penile blood flow are the primary determinants of an erection. While this concept is well recognized, the differential relationship between systemically acting vasoactive factors on arterial pressure and erectile responses is not well described.AimThe aim of this study was to determine how the modification of systemic levels of neurohumoral factors impacts on the magnitude and efficiency of the erectile response.Main Outcome MeasuresThe main outcome measures for this study are changes in MAP and intracavernosal pressure (ICP) following electrostimulation of the cavernous nerve.MethodsAnesthetized adult, male Sprague‐Dawley rats were catheterized for measuring MAP (carotid), ICP, and drug administration (vena cava). Erections were induced via cavernous nerve electrostimulation. Vasoactive drug infusions were used to produce changes in MAP levels including: hexamethonium, angiotensin II (ANGII) ± hexamethonium, methoxamine ± hexamethonium, losartan, MAHMA NONOate, and terbutaline.ResultsIn general, ICP and MAP were linearly correlated regardless of treatment. Hexamethonium markedly dropped MAP and proportionately decreased the magnitude of the erectile response. ANGII or methoxamine given to hexamethonium‐pretreated or untreated rats increased MAP similarly, but produced contrasting effects on erectile responses. ANGII‐induced pressor responses were associated with increased erectile responses whereas all methoxamine treatments markedly decreased erectile responses. Depressor changes with losartan or terbutaline, but not MAHMA NONOate, also impacted negatively on the efficiency of the erectile responses at lower arterial pressures.ConclusionsIn general, the magnitude of the erectile responses was found to be dependent upon the level of MAP, although the mechanism by which arterial pressure was changed impacted substantially on the characteristics of the relationship. The major finding was that circulation‐wide α‐adrenoceptor stimulation was extremely deleterious to erectile responses whereas global stimulation of ANG II receptors was actually proerectile. Overall, the results indicate that neurohumoral specificity in systemic hemodynamic control is also critical in establishing the optimal erectile environment in rats. MacKenzie LD, Heaton JPW, and Adams MA. Impact of systemically active neurohumoral factors on the erectile response of the rat. J Sex Med 2011;8:2461–2471.     

Intracavernous transplantation of bone marrow-derived mesenchymal stem cells restores erectile function of streptozocin-induced diabetic rats

IntroductionErectile dysfunction (ED) is a frequent complication of diabetes mellitus. The efficacy of common ED therapies is low for diabetes‐associated ED.AimTo explore the effects of transplantation of bone marrow‐derived mesenchymal stem cells (BM‐MSCs) on improving erectile function of streptozocin (STZ)‐induced diabetic rats.MethodsMale Sprague Dawley rats were injected either with STZ to induce diabetes or with citrate buffer as controls. Rat BM‐MSCs were harvested and labeled with CM‐DiI (Chloromethylbenzamido derivatives of 1,1′‐dioctadecyl‐3,3,3′,3′‐tetramethylindocarbocyanine perchlorate), and then transplanted into corporal cavernosum of STZ‐induced diabetic rats. Four weeks after transplantation, all rats were analyzed for erectile function and penile histology.Main Outcome MeasuresErectile function was evaluated by the ratio between intracavernous pressure (ICP) and mean arterial pressure (MAP) during electrostimulation of cavernous nerve. Fate of transplanted BM‐MSCs was identified using immunofluorescence staining. Smooth muscle and endothelium in corpora cavernosum were assessed using immunohistochemistry.ResultsAfter BM‐MSCs transplantation, the ICP/MAP ratio was increased significantly compared with diabetic controls. Content of smooth muscle and endothelium in corporal cavernosa of BM‐MSCs transplanted rats was significantly increased compared to diabetic controls. Immunofluorescence analysis demonstrated that CM‐DiI‐labeled BM‐MSCs could stay in corporal cavernosa for at least 4 weeks and some of them expressed von Willebrand Factor, CD31, calponin, or α‐smooth muscle actin, cells markers for endothelial cells or smooth muscle cells, respectively.ConclusionIntracavernous transplantation of BM‐MSCs had beneficial effects on erectile function of diabetic rats and increased the content of endothelium and smooth muscle in corporal cavernosum. Qiu X, Lin H, Wang Y, Yu W, Chen Y, Wang R, and Dai Y. Intracavernous transplantation of bone marrow‐derived mesenchymal stem cells restores erectile function of streptozocin‐induced diabetic rats.     

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.     

Apocynin Improves Erectile Function in Diabetic Rats through Regulation of NADPH Oxidase Expression

IntroductionDiabetes is a risk factor for erectile dysfunction (ED). The proposed mechanisms responsible for diabetic ED are associated with an increase in reactive oxygen species (ROS) production, overactivity of RhoA/ROCK signaling pathway and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, as seen in experimental models of diabetic rats.AimThe aim of this study was to investigate whether NADPH oxidase inhibitor apocynin can ameliorate Streptozotocin (STZ)‐induced diabetes‐related ED by reducing the ROS production and inhibiting the activity of RhoA/ROCK signaling pathway.MethodsThe diabetic rats were treated with and without the NADPH oxidase inhibitor apocynin.Main Outcome MeasuresErectile responses were evaluated by determining mean arterial blood pressure (MAP) and intracavernosal pressure (ICP) with electrical stimulation of the cavernous nerve. Levels of mRNA expression were measured by real‐time polymerase chain reaction (RT‐PCR). Levels of protein expression were examined by Western Blot. ROS production was measured by dihydroethidium (DHE) staining and thiobarbituric acid reactive substances assay.ResultsThe ratio of Maximum ICP‐to‐MAP (MaxICP/MAP) was significantly decreased in diabetic ED rats, compared to that of age‐matched control rats (P < 0.05). Apocynin improved erectile function of diabetic rats (P < 0.05). Expression levels of RhoA (cytosol), nNOS and eNOS were reduced, compared to those of control rats (P < 0.05). Apocynin significantly elevated their expression levels in diabetic rats (P < 0.05). Expression levels of ROCK1, RhoA (membrane fraction), p‐MYPT1 and NADPH oxidase subunits p47^phox and p67^phox were increased in diabetic rats when compared to those of control rats (P < 0.05), and it was observed that apocynin significantly reduced their expression levels in diabetic rats (P < 0.05). ROS production was increased in diabetic rats when compared to that of control rats (P < 0.05), the effect of apocynin was a reduction in the ROS production in diabetic rats (P < 0.05).ConclusionNADPH oxidase inhibitor apocynin can ameliorate diabetes‐related ED by reducing the ROS production and inhibiting the activity of RhoA/ROCK signaling pathway. Li M, Zhuan L, Wang T, Rao K, Yang J, Yang J, Quan W, Liu J, and Ye Z. Apocynin improves erectile function in diabetic rats through regulation of NADPH oxidase expression. J Sex Med 2012;9:3041–3050.     

Oxaliplatin,an Anticancer Agent,Causes Erectile Dysfunction in Rats due to Endothelial Dysfunction

BackgroundChemotherapeutics, one of the standard treatment options for cancer worldwide, have various adverse effects, including erectile dysfunction (ED).AimTo investigate erectile function in an animal model after administration of the anticancer agent oxaliplatin (L-OHP).MethodsMale Wistar/ST rats were divided into 2 groups: L-OHP rats (n = 21), which were intravenously administered L-OHP (4 mg/kg; twice a week for 4 weeks), and Control rats (n = 21), which were injected with the same volume of 5% glucose solution, using the same dosing schedule. At the end of the study period, erectile function was evaluated by measuring intracavernous pressure (ICP) and mean arterial pressure (MAP) after cavernous nerve stimulation (n = 9–10). Endothelial function was evaluated with an isometric tension study using corpus cavernosum strips (n = 11). Western blot analysis was used to assess neuronal nitric oxide (nNOS) and endothelial NO synthase (eNOS) protein levels (n = 7). Real-time quantitative polymerase chain reaction (qRT-PCR) was used to assess the expression of inflammation- and oxidative stress-related markers (nicotinamide adenine dinucleotide phosphate oxidase-1, p22^phox, interleukin [IL]-6, and nuclear factor-kappa B) (n = 6). Statistical significance was determined using the Student's t-test.OutcomesThe L-OHP group had a significantly lower ICP:MAP ratio than the control group (P < .05). Compared to the Control group, the L-OHP group exhibited significantly lower responses to ACh and eNOS protein levels and significantly higher inflammatory biomarker levels.Clinical TranslationThe results based on this animal model indicate that use of the anticancer agent L-OHP should be considered as a risk factor for ED occurring via reduction of NO bioavailability in humans; our results provide possible treatment strategies for maintaining the erectile function of cancer survivors.Strengths and limitationsOur study showed that the anticancer agent L-OHP has the propensity to cause ED in rats. A major limitation of this study is the lack of an established cure for ED associated with L-OHP and the lack of clinical evidence.ConclusionsL-OHP causes ED in rats via reduction of NO bioavailability caused by endothelial dysfunction.Kataoka T, Mori T, Suzuki J, et al. Oxaliplatin, an Anticancer Agent, Causes Erectile Dysfunction in Rats due to Endothelial Dysfunction. J Sex Med 2021;18:1337–1345.     

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.     

Oral l‐Citrulline Supplementation Improves Erectile Function in Rats with Acute Arteriogenic Erectile Dysfunction

IntroductionOral l‐citrulline supplementation increases serum l‐arginine levels more efficiently than l‐arginine itself and increases nitric oxide (NO) production.AimTo investigate whether oral l‐citrulline supplementation improves erectile function in rats with acute arteriogenic erectile dysfunction (ED).MethodsWe divided 8‐week‐old male Wistar‐ST rats into 3 groups: sham‐operated rats (control group), arteriogenic ED rats who underwent ligation of both internal iliac arteries (ligation group), and arteriogenic ED rats receiving oral 2% l‐citrulline water supplementation (citrulline group). Citrulline water was given to arteriogenic ED rats for 3 weeks from 1 week after surgery. Erectile function was evaluated by maximum intracavernous pressure/mean arterial pressure (ICP/MAP) ratios via cavernous nerve stimulation at 4 weeks after surgery. Then, the penises were resected, stained with Masson's trichrome, and observed microscopically. Serum nitrogen oxides (NOx) levels were measured by high‐performance liquid chromatography. Bonferroni's multiple t‐test was used for statistical analysis.Main Outcome MeasuresThe main outcome measures were changes in ICP/MAP, smooth muscle (SM)/collagen ratios, and NOx levels following l‐citrulline supplementation.ResultsThe ICP/MAP ratio in the ligation group was significantly lower than that in the control group (P < 0.05), denoting ED. The ICP/MAP ratio of the citrulline group was significantly higher than that of the ligation group (P < 0.05), indicating ED amelioration. Levels of NOx in the ligation group were significantly lower than in the control group (P < 0.05), while those in the citrulline group were significantly higher than in the ligation group (P < 0.05). SM/collagen ratios in the ligation group were significantly lower than in the control group (P < 0.05), while ratios in the citrulline group were significantly higher than those in the ligation group (P < 0.05).ConclusionsOral l‐citrulline supplementation improved ICP/MAP and SM/collagen ratios and increased NOx. Therefore, oral l‐citrulline supplementation might be a useful novel therapy for acute arteriogenic ED. Shiota A, Hotta Y, Kataoka T, Morita M, Maeda Y, and Kimura K. Oral l‐citrulline supplementation improves erectile function in rats with acute arteriogenic erectile dysfunction. J Sex Med 2013;10:2423–2429.     

Change of erectile function and responsiveness to phosphodiesterase type 5 inhibitors at different stages of streptozotocin-induced diabetes in rats

IntroductionIt has been suggested that risk of erectile dysfunction (ED) increases with duration of diabetes and phosphodiesterase type 5 inhibitors (PDE5I) are not as effective in treatment of diabetes-associated ED. However, few studies have investigated time-dependent change in erectile function during the course of diabetes.AimTo investigate time-dependent change in erectile function and responsiveness to PDE5I in streptozotocin-induced diabetic rats and to understand the pathophysiology of diabetic ED.Main Outcome MeasuresAt 6, 8, 10, 12, and 14 weeks after diabetic induction, erectile function was assessed by cavernous nerve stimulation before and after administration of DA-8159, a novel PDE5I. Penile tissue was assessed for apoptosis with immunohistochemistry. Protein expression of Rho-kinase 2 (ROCK2), myosin phosphatase targeting subunit 1 (MYPT1), and endothelial nitric oxide synthase (eNOS) was evaluated by Western blot.MethodsStreptozotocin was injected into 50 8-week-old male Sprague-Dawley rats, which were then classified into five diabetic groups according to the observation period.ResultsDiabetic rats maintained normal erectile responses until 6 weeks of diabetes. Following 8 weeks, the rats showed lower erectile responses at higher frequencies of nerve stimulation, which were normalized to control by administration of DA-8159. In contrast, erectile responses were significantly decreased in 10-week diabetic rats, and administration of DA-8159 resulted in partial recovery of normal responses. At more than 12 weeks, rats demonstrated severe deterioration of erectile function, which did not fully respond to PDE5I. Corporal apoptosis was significantly increased after 10 weeks. Upregulation of ROCK2 was found at 6 weeks, and was followed by an increase of MYPT1 phosphorylation. Phosphorylation of eNOS showed marked suppression at 6 weeks and remained lower during the experimental period.ConclusionsImpairment of erectile function was followed by decreased responsiveness to PDE5I during the course of diabetes. The RhoA/ROCK pathway played an important role in diabetes-associated ED. Cho SY, Park K, Paick J-S, and Kim SW. Change of erectile function and responsiveness to PDE5 (Type 5 phosphodiesterase) inhibitors at different stages of streptozotocin-induced diabetes in rats.     

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.     

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.     

Incomplete Recovery of Erectile Function in Rat after Discontinuation of Dual 5‐Alpha Reductase Inhibitor Therapy

AimThe association of 5‐alpha reductase inhibitor (5ARI) therapy and sexual dysfunction has been reported. Some patients claim persistent erectile dysfunction despite long‐term discontinuation of 5ARI treatment. The aim of this study was to assess erectile function after cessation of 5ARI therapy using a rat model.MethodsTwenty‐six adult male Sprague‐Dawley rats were randomized into three groups: (i) control (N = 10); (ii) 8‐week dutasteride treatment (0.5 mg/rat/day, in drinking water, N = 8); and (iii) 6‐week dutasteride treatment followed by a 2‐week washout period (N = 8). The experiments were performed after 8 weeks from the initiation of treatment in all groups. In vivo erectile activity and in vitro contractile and relaxant responses of cavernosal smooth muscle were investigated.ResultsIn vivo erectile activity (intracavernosal pressure [ICP]/mean arterial pressure [MAP] and total ICP) in treatment groups were significantly decreased compared with controls (ICP/MAP: P < 0.001 for 2.5 v, 5 v, and 7.5 v; total ICP: P < 0.001 for 5 v and P < 0.01 for 7.5 v). Acetylcholine‐induced relaxations were diminished in treatment groups (P < 0.05). Relaxant responses to electrical field stimulation (EFS) were decreased in the 8‐week treatment group (P < 0.05) but were similar to controls in the washout group. Sodium nitroprusside (SNP)‐induced endothelium‐independent relaxations were reduced in the 8‐week dutasteride treatment group (P < 0.01), while these responses were restored in the washout group. The contractile responses to the alpha1‐adrenergic agonist phenylephrine were decreased in treatment groups compared with controls (P < 0.01). Direct neurogenic contractile responses in the dutasteride groups were significantly lower than controls between 1 and 15 Hz frequencies (but not at 20 Hz) and washout partially restored the responses at 10 and 15 Hz.ConclusionDiscontinuation of dutasteride improved the relaxant responses to EFS and SNP, while cholinergic and adrenergic responses remained depressed. Our findings suggest a time‐dependent detriment of dutasteride on erectile function. The withdrawal/washout effect of 5ARIs on parameters of human sexual function warrants further investigation. Öztekin ÇV, Gur S, Abdulkadir NA, Lokman U, Akdemir AÖ, Cetinkaya M, and Hellstrom WJG. Incomplete recovery of erectile function in rat after discontinuation of dual 5‐alpha reductase inhibitor therapy. J Sex Med 2012;9:1790–1798.     

Chronic vardenafil treatment improves erectile function via structural maintenance of penile corpora cavernosa in rats with acute arteriogenic erectile dysfunction

IntroductionChronic phosphodiesterase type 5 inhibitor treatment may be useful in reversing erectile dysfunction (ED). However, the mechanisms of this improvement remain unknown.AimThe aim of this article was to determine the mechanisms of the improvement by chronic vardenafil treatment for acute arteriogenic ED in rats.MethodsEight‐week‐old male Wistar‐ST rats were divided into four groups: sham‐operated rats (Control group) and rats with acute arteriogenic ED induced by ligating bilateral internal iliac arteries (Ligation group), subsequently treated with low‐dose (0.4 mg/kg/day; VL group) or high‐dose (4.0 mg/kg/day; VH group) vardenafil for 20 days from 1 week after ligature.Main Outcome MeasuresErectile function was assessed based on changes of intracavernous pressure (ICP) followed by electrostimulation of the cavernous nerves and was evaluated by the area under the curve of ICP/area under the curve of mean arterial pressure (area of ICP/MAP). Transforming growth factor (TGF)‐β₁, vascular endothelial growth factor‐A, endothelial nitric oxide synthase (eNOS), inducible NOS, and neuronal NOS mRNA expression levels in penile corpus cavernosum were determined by real‐time PCR. Western blotting for TGF‐β₁ protein levels and Masson trichrome staining of penile tissues were performed in each at group 4 weeks after surgery.ResultsIn the VH group, area of ICP/MAP was significantly improved when compared with the Ligation group (P < 0.01). The smooth muscle (SM)/collagen ratio in the VH group was significantly higher than in the Ligation group (P < 0.05), and was comparable with that in the Control group. TGF‐β₁ mRNA and protein levels in the VH group were significantly lower when compared with the Ligation group (P < 0.05).ConclusionsChronic vardenafil administration ameliorates impairment of penile hemodynamics and maintains normal SM to collagen ratio in cavernous tissues after acute arterial injury in rats. Hotta Y, Hattori M, Kataoka T, Ohno R, Mikumo M, Maeda Y, and Kimura K. Chronic vardenafil treatment improves erectile function via structural maintenance of penile corpora cavernosa in rats with acute arteriogenic erectile dysfunction. J Sex Med 2011;8:705–711.     

Role of Oxidative Stress in a Rat Model of Radiation-Induced Erectile Dysfunction

IntroductionChronic oxidative stress is one of the major factors playing an important role in radiation-induced normal tissue injury. However, the role of oxidative stress in radiation-induced erectile dysfunction (ED) has not been fully investigated.AimsTo investigate role of oxidative stress after prostate-confined irradiation in a rat model of radiation-induced ED.MethodsFifty-four young adult male rats (10–12 weeks of age) were divided into age-matched sham radiotherapy (RT) and RT groups. Irradiated animals received prostate-confined radiation in a single 20 Gy fraction.Main Outcome MeasuresIntracavernous pressure (ICP) measurements with cavernous nerve electrical stimulation were conducted at 2, 4, and 9 weeks following RT. The protein expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits (Nox4 and gp91^phox), markers of oxidative DNA damage (8-hydroxy-2′-deoxyguanosine [8-OHdG]), lipid peroxidation (4-hydroxynonenal [4HNE]), and inflammatory response including inducible nitric oxide synthase, macrophage activation (ED-1), and nitrotyrosine, and endogenous antioxidant defense by nuclear factor erythroid 2-related factor (Nrf2) were evaluated in irradiated prostate tissue and corpora cavernosa (CC). In addition, we investigated the relationships between results of ICP/mean arterial pressure (MAP) ratios and expression level of oxidative stress markers.ResultsIn the RT group, hemodynamic functional studies demonstrated a significant time-dependent decrease in ICP. Increased expression of Nox4, gp91^phox, 8-OHdG, and 4HNE were observed in the prostate and CC after RT. Similarly, expressions of inflammatory markers were significantly increased. There was a trend for increased Nrf2 after 4 weeks. ICP/MAP ratio negatively correlated with higher expression level of oxidative markers.ConclusionsNADPH oxidase activation and chronic oxidative stress were observed in irradiated prostate tissue and CC, which correlated with lower ICP/MAP ratio. Persistent inflammatory responses were also found in both tissues after RT. These findings suggest that oxidative stress plays a crucial role in the development of radiation-induced ED. Kimura M, Rabbani ZN, Zodda AR, Yan H, Jackson IL, Polascik TJ, Donatucci CF, Moul JW, Vujaskovic Z, and Koontz BF. Role of oxidative stress in a rat model of radiation-induced erectile dysfunction. J Sex Med 2012;9:1535–1549.