Johanna L. Hannan

Beneficial Impact of Exercise and Obesity Interventions on Erectile Function and Its Risk Factors

INTRODUCTION Erectile dysfunction (ED) is a multifaceted disease involving cardiovascular, metabolic, and hormonal factors and affects over 100 million men worldwide. ED has been shown to be a harbinger of underlying cardiovascular diseases (CVD), as there are common risk factors (aging, hypertension, obesity) and mechanistic basis. AIM To provide an update on clinical and experimental evidence regarding the impact of lifestyle modifications, such as exercise and diet, with respect to changes in erectile function. MAIN OUTCOME MEASURES Published evidence regarding the impact of aging, hypertension, and obesity on ED and CVD, as well as new experimental data linking obesity and diminished erectile responses. METHODS We reviewed the literature regarding common risk factors of ED and CVD, particularly involving obesity, as well as performed new analysis on the findings of other experimental studies involving diet and exercise interventions. RESULTS Physical inactivity negatively impacts on erectile function, and experimental and clinical exercise interventions have been shown to improve sexual responses and overall cardiovascular health. Mediterranean-style diets and a reduction in caloric intake have been found to improve erectile function in men with the aspects of the metabolic syndrome. In addition, both clinical and experimental studies have confirmed that combining the two interventions provides additional benefit to erectile function, likely via reduced metabolic disturbances (e.g., inflammatory markers, insulin resistance), decreased visceral adipose tissue, and improvement in vascular function (e.g., increased endothelial function). CONCLUSIONS Lifestyle modifications provide significant benefits to vascular health and erectile function in a population that is increasingly aged and more obese.

Inhibition of Rho-Kinase Improves Erectile Function, Increases Nitric Oxide Signaling and Decreases Penile Apoptosis in a Rat Model of Cavernous Nerve Injury

PURPOSE Bilateral cavernous nerve injury results in up-regulation of ROCK signaling in the penis. This is linked to erectile dysfunction in an animal model of post-prostatectomy erectile dysfunction. We evaluated whether daily treatment with the ROCK inhibitor Y-27632 (Tocris Bioscience, Ellisville, Missouri) would prevent erectile dysfunction in a rat model of bilateral cavernous nerve injury. MATERIALS AND METHODS Sprague-Dawley® rats underwent surgery to create sham (14) or bilateral (27) cavernous nerve injury. In the injury group 13 rats received treatment with Y-27632 (5 mg/kg twice daily) and 14 received vehicle. At 14 days after injury, rats underwent cavernous nerve stimulation to determine erectile function. Penes were assessed for neuronal and nitric oxide synthase membrane-endothelial nitric oxide synthase. ROCK2 was assessed by Western blot. Cyclic guanosine monophosphate was determined by enzyme-linked immunosorbent assay. Cavernous homogenates were tested for ROCK and protein kinase G enzymatic activity. Penile apoptosis was evaluated using the Apostain technique (Alexis, San Diego, California). Data were analyzed on ROCK using ANOVA and the t test. RESULTS While erectile function was decreased in rats with bilateral cavernous nerve injury, daily administration of Y-27632 improved erectile responses. Injury decreased neuronal and nitric oxide synthase membrane-endothelial nitric oxide synthase but ROCK2 was significantly increased. Y-27632 treatment restored neuronal nitric oxide synthase, nitric oxide synthase membrane-endothelial nitric oxide synthase and cyclic guanosine monophosphate levels, and protein kinase G activity. Treatment significantly decreased ROCK2 protein and ROCK activity. There were significantly fewer apoptotic cells after treatment than in injured controls. CONCLUSIONS These results provide evidence for up-regulation of the RhoA/ROCK signaling pathway with detrimental effects on erectile function after bilateral cavernous nerve injury. ROCK inhibition improved erectile dysfunction associated with bilateral cavernous nerve injury by preserving penile nitric oxide bioavailability and decreasing penile apoptosis.

Morphological and Functional Evidence for the Contribution of the Pudendal Artery in Aging-Induced Erectile Dysfunction

INTRODUCTION Aging increases the risk of both erectile dysfunction (ED) and cardiovascular disease. These conditions have similar etiologies and commonly coexist. One unifying concept is the role of arterial insufficiency which is a primary factor in the onset of age-related ED. AIM Based on the novel finding that the pudendal arteries contribute 70% of the total penile vascular resistance, our objective was to morphometrically and functionally characterize this vessel in young and old normotensive rats. METHODS Erectile function was monitored in 15- and 77-week Sprague-Dawley rats using the apomorphine bioassay (80mg/kg, s.c.). Anesthetized animals were perfusion-fixed, aortic, renal, and internal pudendal arteries were excised, embedded, sectioned, stained, and morphometrically assessed using light microscopy. Hearts were excised, separated, and weighed prior to perfusion. Contractile and relaxation responses to acetylcholine (ACh) and phenylephrine (PE) were assessed by wire myograph. MAIN OUTCOME MEASURES Erectile function, morphological measurements, concentration response curves to ACh and PE. RESULTS With age, there were marked decreases in erectile responses compared to younger rats (2.8±0.87 vs. 0.3±0.58). The pudendal arteries had a relatively small lumen (303±13.8µm) and a thick medial layer (47±2.2µm). In aged pudendal arteries, the lumen diameter did not change, and yet the medial layer, cross sectional area, and extracellular matrix were markedly increased. In contrast, the lumen diameter and wall thickness of the aorta and renal arteries in aged rats increased proportionally. An increase in small, round, smooth muscle cells was seen in aged pudendal arteries. Functionally, there were no differences in contractile responses to PE; however, ACh-induced relaxation decreased with age. CONCLUSIONS In aged rats, erectile function was severely diminished when pudendal arteries had undergone marked phenotypic changes. Specifically, there was endothelial dysfunction and pathological remodeling of this vessel with age, characterized by medial thickening, impaired vasodilation and significantly reduced capacity for penile blood flow.

Understanding and targeting the Rho kinase pathway in erectile dysfunction

Erectile dysfunction (ED) is a common disorder that affects a quarter of US men, and has many causes, including endothelial impairment, low testosterone levels, prior surgical manipulation, and/or psychogenic components. Penile erection is a complex process requiring neurally mediated relaxation of arteriolar smooth muscle and engorgement of cavernosal tissues, mediated by nitric oxide (NO). Current medical therapies for ED largely seek to maximize endogenous NO signalling. Certain aetiologies, including diabetes, are difficult to treat with current modalities, emphasizing the need for new molecular targets. Research has demonstrated the importance of RhoA–Rho-associated protein kinase (ROCK) signalling in maintaining a flaccid penile state, and inhibition of RhoA–ROCK signalling potentiates smooth-muscle relaxation in an NO-independent manner. The mechanisms and effects of RhoA–ROCK signalling and inhibition suggest that the RhoA–ROCK pathway could prove to be a new therapeutic target for the treatment of ED.

Chronic Oral Administration of the Arginase Inhibitor 2(S)-amino-6-boronohexanoic Acid (ABH) Improves Erectile Function in Aged Rats

Arginase expression and activity have been noted to be heightened in conditions associated with erectile dysfunction, including aging. Previously, arginase inhibition by chronic administration of the arginase inhibitor 2-(S)-amino-6-boronohexanoic acid (ABH) has been shown to improve endothelial dysfunction in aged rats. The objective of this study was to assess whether chronic oral ABH administration affects cavernosal erectile function. Rats were divided into 4 groups: young control, young treated with arginase inhibitor, aged control, and aged treated with arginase inhibitor. Arginase activity was measured and presented as a proportion of young untreated rats. In vivo erectile responses to cavernous nerve stimulation were measured in all cohorts. The cavernous nerve was stimulated with a graded electrical stimulus, and the intracavernosal/mean arterial pressure ratios and total intracavernosal pressure were recorded. Arginase activity was elevated in the aged rats compared with young controls; however, arginase activity was significantly decreased in aged rats treated with ABH. With the addition of ABH, erectile responses improved in the aged rats (P < .05). Oral inhibition of arginase with ABH results in improved erectile function in aged rats, resulting in erectile hemodynamics similar to young rats. This represents the first documentation of systemic arginase inhibition positively affecting corporal cavernosal function.

Valproic acid prevents penile fibrosis and erectile dysfunction in cavernous nerve-injured rats

INTRODUCTION Bilateral cavernous nerve injury (BCNI) causes profound penile changes such as apoptosis and fibrosis leading to erectile dysfunction (ED). Histone deacetylase (HDAC) has been implicated in chronic fibrotic diseases. AIMS This study will characterize the molecular changes in penile HDAC after BCNI and determine if HDAC inhibition can prevent BCNI-induced ED and penile fibrosis. METHODS Five groups of rats (8-10 weeks, n = 10/group) were utilized: (i) sham; (ii and iii) BCNI 14 and 30 days following injury; and (iv and v) BCNI treated with HDAC inhibitor valproic acid (VPA 250 mg/kg; 14 and 30 days). All groups underwent cavernous nerve stimulation (CNS) to determine intracavernosal pressure (ICP). Penile HDAC3, HDAC4, fibronectin, and transforming growth factor-β1 (TGF-β1) protein expression (Western blot) were assessed. Trichrome staining and the fractional area of fibrosis were determined in penes from each group. Cavernous smooth muscle content was assessed by immunofluorescence to alpha smooth muscle actin (α-SMA) antibodies. MAIN OUTCOME MEASURES We measured ICP; HDAC3, HDAC4, fibronectin, and TGF-β1 protein expression; penile fibrosis; penile α-SMA content. RESULTS There was a voltage-dependent decline (P < 0.05) in ICP to CNS 14 and 30 days after BCNI. Penile HDAC3, HDAC4, and fibronectin were significantly increased (P < 0.05) 14 days after BCNI. There was a slight increase in TGF-β1 protein expression after BCNI. Histological analysis showed increased (P < 0.05) corporal fibrosis after BCNI at both time points. VPA treatment decreased (P < 0.05) penile HDAC3, HDAC4, and fibronectin protein expression as well as corporal fibrosis. There was no change in penile α-SMA between all groups. Furthermore, VPA-treated BCNI rats had improved erectile responses to CNS (P < 0.05). CONCLUSION HDAC-induced pathological signaling in response to BCNI contributes to penile vascular dysfunction. Pharmacological inhibition of HDAC prevents penile fibrosis, normalizes fibronectin expression, and preserves erectile function. The HDAC pathway may represent a suitable target in preventing the progression of ED occurring post-radical prostatectomy.

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

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

Impact of antihypertensive treatments on erectile responses in aging spontaneously hypertensive rats

Objective We previously demonstrated that brief, aggressive antihypertensive therapy recovered erectile function in 40-week-old spontaneously hypertensive rats (SHR). The present study examined the impact of antihypertensive and testosterone treatments on erectile function in aging SHR. Design and methods Centrally initiated erections were determined in response to apomorphine throughout. At 30 and 49 weeks, SHR were treated for 2 weeks with enalapril or hydralazine. A third more aggressive treatment (68 weeks) involved enalapril or losartan plus a low salt diet or a triple therapy (hydralazine, nifedipine, hydrochlorothiazide). In a separate study, cross-over kidney transplantations were performed between untreated and losartan-treated SHR. Arterial pressure was assessed post-transplantation using radio-telemetric transducers. Results There was an age-related decrease in erections between 30 and 68 weeks (3.1 ± 0.79 versus 0.2 ± 0.38) that was not improved by testosterone administration. Early treatment with enalapril or hydralazine did not prevent this decline, although the second treatment resulted in significant improvements (enalapril, 0.8 ± 0.70; hydralazine, 0.8 ± 0.41 versus control, 0.3 ± 0.60). A 2-week aggressive antihypertensive treatment at 68 weeks increased erections approximately two-fold, with the previously treated rats receiving triple therapy having markedly improved erectile responses (0.2 ± 0.53 versus 1.1 ± 1.67). In the transplantation study, previously losartan-treated SHR given an untreated kidney had higher arterial pressure but twice the number of erections in comparison with the SHR with lower arterial pressure resulting from transplanting a treated kidney. Conclusions Aggressive antihypertensive treatments may be more beneficial in improving erectile function in aged SHR, via an effect that appears to be tissue specific, and not based on changes in blood pressure.

Pregnancy reduces RhoA/Rho kinase and protein kinase C signaling pathways downstream of thromboxane receptor activation in the rat uterine artery

During pregnancy, reduced vascular responses to constrictors contribute to decreased uterine and total vascular resistance. Thromboxane A(2) (TxA(2)) is a potent vasoconstrictor that exerts its actions via diverse signaling pathways, and its biosynthesis increases in preeclampsia. In this study, we hypothesized that maternal vascular responses to TxA(2) will be attenuated via Rho kinase, PKC, p38 MAPK, and ERK1/2 signaling pathways. Isolated ring segments of uterine and small mesenteric arteries from late pregnant (19-21 days) and virgin rats were suspended in a myograph, and isometric force was measured. Pregnancy did not affect uterine and mesenteric artery responses to the TxA(2) analog U-46619 (10(-9)-10(-5) M), but transduction signals associated with these contractions were different between pregnant and nonpregnant rats. Inhibition of Rho kinase (10(-6) M Y-27632) reduced sensitivity to U-46619 in virgin uterine vessels but did not inhibit these contractions in pregnant uterine arteries and had no effect on mesenteric vessels. Treatment of arterial segments with a PKC inhibitor (10(-6) M bisindolylmaleimide I) reduced U-46619-induced contractions in virgin uterine and mesenteric arteries and in pregnant mesenteric arteries. Pregnant uterine arteries, however, were unresponsive to PKC inhibition. Inhibition of ERK1/2 (10(-5) M PD-98059) and p38 MAPK (10(-5) M SB-203580) reduced U46619-induced contractions in nonpregnant vessels and in pregnant uterine and mesenteric vessels. These data suggest that normal pregnancy does not affect uterine and mesenteric contractile responses to TxA(2) but reduces the contribution of Rho kinase and PKC signaling pathways to these contractions in the uterine vasculature. In contrast, the role of ERK1/2 and p38 MAPK in U-46619-induced uterine contractions remains unchanged with pregnancy. TxA(2)-associated transduction signals and its regulators might present potential targets for the development of new treatments for preeclampsia and other pregnancy-associated vascular diseases.

Caspase-3 dependent nitrergic neuronal apoptosis following cavernous nerve injury is mediated via RhoA and ROCK activation in major pelvic ganglion

Axonal injury due to prostatectomy leads to Wallerian degeneration of the cavernous nerve (CN) and erectile dysfunction (ED). Return of potency is dependent on axonal regeneration and reinnervation of the penis. Following CN injury (CNI), RhoA and Rho-associated protein kinase (ROCK) increase in penile endothelial and smooth muscle cells. Previous studies indicate that nerve regeneration is hampered by activation of RhoA/ROCK pathway. We evaluated the role of RhoA/ROCK pathway in CN regulation following CNI using a validated rat model. CNI upregulated gene and protein expression of RhoA/ROCK and caspase-3 mediated apoptosis in the major pelvic ganglion (MPG). ROCK inhibitor (ROCK-I) prevented upregulation of RhoA/ROCK pathway as well as activation of caspase-3 in the MPG. Following CNI, there was decrease in the dimer to monomer ratio of neuronal nitric oxide synthase (nNOS) protein and lowered NOS activity in the MPG, which were prevented by ROCK-I. CNI lowered intracavernous pressure and impaired non-adrenergic non-cholinergic-mediated relaxation in the penis, consistent with ED. ROCK-I maintained the intracavernous pressure and non-adrenergic non-cholinergic-mediated relaxation in the penis following CNI. These results suggest that activation of RhoA/ROCK pathway mediates caspase-3 dependent apoptosis of nitrergic neurons in the MPG following CNI and that ROCK-I can prevent post-prostatectomy ED.