Stereological comparison of smooth muscle, collagen, and elastic fibers of the clitoris and glans penis in young adults.

OBJECTIVE: To compare the collagen, elastic fibers, and smooth muscle content of the clitoris and the glans penis in young adults. MATERIALS AND METHODS: The clitoris and the glans penis of six women and six men (mean age 25±3) who died as a result of accidents were excised. The samples were placed under a formaldehyde solution and histologically processed. Masson's trichrome and Weigert's resorcin-fuchsin stain was used to highlight the elastic fibers, smooth muscle, and collagen. Stereological analysis was conducted in 5 random fields of 5 slides for each sample. For statistical analysis, the unpaired t-test was used to compare values between groups, and a value of P<0.05 was considered as significant for all analyses. RESULTS: Stereology revealed a mean smooth muscle content of 35.84±6.46% and 31.64±4.74% for the clitoris and glans penis, respectively, while it also revealed collagen content of 26.11±7.41% and 28.44±3.55% and elastic fibers content of 24.12±4.34% and 30.97±6.13% for the clitoris and glans penis, respectively. The statistical analysis showed no significant differences between them. CONCLUSION: Regardless of anatomical differences, the volumetric density of collagen, elastic fibers, and smooth muscle were similar for the clitoris and glans penis in young adults, a feature possibly explained by their embryology.

Evaluation of effectiveness of antiandrogen treatments on cavernosal tissues in rats.

In this study, we aimed to compare changes in cavernosal tissues in rats with antiandrogen treatment and orchiectomy. A total of 42 Wistar albino rats were divided into four groups. Group I, control group, Group II, LH-RH was given for 1 month, Group III-LH-RH + Bicalutamide was given for 1 month, and Group IV was defined as orchiectomy and followed up for 1 month. Measurements of intracavernosal pressure with different electrical stimuli and pathological findings of smooth muscle collagen in cavernosal tissues were examined. While the cavernosal pressure response in all the different electrical stimuli given in the control group and in all other groups was significantly lower than that in the other groups, it was statistically significant at 7.5 and 10 V (p = .005, p < 0001). According to the pathologic evaluation, the density of tissue collagen increased significantly in the other groups according to the control group. In groups 3 and 4, the density of 4+ collagen was found to be increased according to Groups 1 and 2. In the LH-RH alone group, it appears that there are no 4+ colloid density and less damage. According to these findings, the negative effect of LH-RH treatment on cavernosal tissues appears to be less.

Effect of p-coumaric acid on the erectogenic enzyme activities and non-protein thiol level in the penile tissue of normal and doxorubicin-induced oxidative stress male rat.

This study investigated effect of p-coumaric acid (PCA) on erectogenic enzyme activity and non-protein thiol level in the penile tissue of normal and doxorubicin (DOX)-induced oxidative stress male rat. Sixty-four (64) adult male rats weighing between 170 and 180 g were used for this work. After 14 days of acclimatisation, the rats were divided into eight groups (n = 8). Rats were orally pre-treated with PCA dose dependently (50 and 100 mg/kg body weight [b.w.t]) and vitamin E (100 mg/kg b.w.t) for 14 days before induction with a single dose of DOX (15 mg/kg b.w.t, via i.p.). The result revealed that arginase, acetylcholinesterase (AChE), angiotensin-I-converting enzyme (ACE), phosphodiesterase-5 (PDE-5), adenosine monophosphohydrolase (AMPdase) activities were significantly (p < 0.05) higher in the DOX-induced rats as against the control, which was significantly p < 0.05) higher when compared to normal rats treated with PCA. PCA also improved non-protein thiol level in the penile tissue of both normal and DOX-induced rats. Hence, this study revealed that PCA is capable of causing inhibitory effects on the activities of enzymes, associated with oxidative stress-induced erectile dysfunction (ED) and could also be used as an aphrodisiac agent in the management/treatment of ED.

An experimental study: quantitatively evaluating the change of the content of collagen fibres in penis with two-dimensional ShearWave™ Elastography.

The purpose of this study was to explore the value of two-dimensional ShearWave™ Elastography (2D-SWE) on quantitatively evaluating the change of the content of collagen fibres in penis. Twenty male Sprague Dawley rats were divided into the pre-sexual maturity group (Group 1) and the sexual decline group (Group 2) according to age. The ultrafast ultrasound device Aixplorer® (SuperSonic Imagine, Aix-en-Provence, France) was used for 2D-SWE imaging of penis, and the measurement index was shear wave stiffness (SWS). The immunohistochemistry was used to analyse the content of collagen fibres in penis, and the measurement index was positive area percentage (PAP). The differences of SWS between the two groups and PAP between the two groups were analysed. SWS of Group 1 and Group 2 was 10.18 ± 1.09 and 8.02 ± 1.34 kPa, and SWS of Group 2 was significantly lower than Group 1 (p < .01). PAP of Group 1 and Group 2 was 4.83 ± 3.61% and 16.41 ± 10.02%, and PAP of Group 2 was significantly higher than Group 1 (p < .01). Our results indicate that when the content of collagen fibres changes, SWS of penis measured with 2D-SWE would change significantly as well. Two-dimensional SWE can be used to quantitatively evaluate the change of the content of collagen fibres in penis.

Modulation of some markers of erectile dysfunction and malonaldehyde levels in isolated rat penile tissue with unripe and ripe plantain peels: identification of the constituents of the plants using HPLC.

CONTEXT: Plantain fruit pulp has been used as a natural remedy to manage erectile dysfunction (ED) in traditional medicine. However, the potency of the peel has not been examined with respect to ED management. OBJECTIVE: This study investigated and compared the inhibitory potential of unripe (UPP) and ripe (RPP) plantain peels on some enzymes associated with ED and Fe2+-induced oxidative stress in albino rat penile homogenate in vitro. MATERIALS AND METHOD: Aqueous extract of the peels was prepared and the effect on phosphodiesterase-5 (PDE-5), arginase, acetylcholinesterase (AChE), angiotensin-I converting enzyme (ACE) and Fe2+-induced malonyladehyde in isolated albino rat penile homogenate were investigated. Phenolic constituents of the peels powder were characterized using high-performance liquid chromatography coupled with diode array detector (HPLC-DAD). RESULT: Extract from UPP had higher PDE-5 (IC50 = 3.10 µg/mL), arginase (IC50 = 0.96 µg/mL), AChE (IC50 = 6.30 µg/mL) and ACE (IC50 = 0.41 µg/mL) inhibitory ability compared with RPP (PDE-5, IC50 = 4.33 µg/mL; arginase, IC50 = 1.34 µg/mL; AChE, IC50 = 8.64 µg/mL; ACE, IC50 = 0.63 µg/mL). The extract from UPP also had higher inhibition of Fe2+-induced lipid peroxidation. HPLC-DAD analysis revealed that gallic and caffeic acids, rutin, quercitrin and quercetin were abundant in UPP, while catechin, kaempferol, chlorogenic and ellagic acids were the dominant phenolic compounds in RPP. DISCUSSION AND CONCLUSION: Inhibition of enzymes associated with ED and lipid peroxidation could be linked with the phenolic compounds. However, UPP appeared to be more potent.

Molecular signalling of a novel curcumin derivative versus Tadalafil in erectile dysfunction.

The efficacy of a novel curcumin derivative (NCD) versus tadalafil in erectile signalling was assessed. Ten control male rats and 50 diabetic male rats were used and divided into the following: diabetic (DM), curcumin (CURC), NCD, tadalafil and NCD combined with tadalafil rat groups. Cavernous tissue gene expression of heme oxygenase-1 (HO-1), Nrf2, NF-B and p38, enzyme activities of heme oxygenase (HO) and nitric oxide synthase (NOS), cGMP and intracavernosal pressure (ICP)/mean arterial pressure (MAP) were assessed. Results showed that 12 weeks after induction of diabetes, erectile dysfunction (ED) was confirmed by the significant decrease in ICP/MAP, a significant decrease in cGMP, NOS, HO enzyme activities, a significant decrease in HO-1 gene and a significant increase in NF-Ò ß, p38 genes. Administration of all therapeutic interventions led to a significant increase in ICP/MAP, cGMP levels, a significant increase in HO-1 and NOS enzymes, a significant increase in HO-1, and Nrf2 gene expression, and a significant decrease in NF-Ò ß, p38 gene expression. NCD or its combination with tadalafil showed significant superiority and more prolonged duration of action. In conclusion, a tendency was observed that CURC and NCD have high efficacy and more prolonged duration of action in enhancing erectile function.

Effect of avanafil on rat and human corpus cavernosum.

We compared the activity of a new phosphodiesterase-5 inhibitor (PDE5i) avanafil with sildenafil and tadalafil in human and rat corpus cavernosum (CC) tissues. The effect of avanafil with several inhibitors and electrical field stimulation (EFS) was evaluated on CC after pre-contraction with phenylephrine. With the PDE5i, sildenafil and tadalafil, concentration-response curves were obtained and cyclic guanosine monophosphate (cGMP) levels were measured in tissues. Avanafil induced relaxation with maximum response of 74 ± 5% in human CC. This response was attenuated by NOS inhibitor and soluble guanylate cyclase (sGC) inhibitor. Avanafil potentiated relaxation responses to acetylcholine and EFS in human CC and enhanced SNP-induced relaxation and showed 3-fold increase in cGMP levels. When compared with sildenafil, avanafil and tadalafil were effective at lower concentrations in human CC. In addition, Sprague-Dawley rats underwent in vivo intracavernosal pressure (ICP) and mean arterial pressure (MAP) measurements. Avanafil increased ICP/MAP that was enhanced by SNP and cavernous nerve (CN) stimulation in rat CC tissues. Also avanafil showed maximum relaxation response of 83 ± 7% in rat CC with 3-fold increase in cGMP concentration. Taken together, these results of our in vivo and in vitro studies in human and rat suggest that avanafil promotes the CC relaxation and penile erection via NO-cGMP pathway.

Intracellular calcium concentration of corpus cavernosum smooth muscle cells is decreased by the overexpression of PnNOS gene in adipose tissue-derived stem cells.

The study investigated the effects of adipose tissue-derived stem cells (ADSCs) modified with penile neuronal nitric oxide synthase (PnNOS) gene on intracellular calcium concentration in rat corpus cavernosum smooth muscle cells (CCSMCs). ADSCs and CCSMCs of Sprague-Dawley (SD) rats were isolated and cultured in vitro respectively. The rat PnNOS gene was transferred into the ADSCs mediated by a recombinant adenovirus vector. The expression of the PnNOS gene was detected. At the same time, the concentration of nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) was assayed. After coculturing with the CCSMCs of SD rats, which were isolated and expanded ex vivo, the cGMP and NO levels of ADSCs and CCSMCs were measured. Intracellular calcium concentration ([Ca(2+) ]i ) in rat CCSMCs was measured with Fluo-3/AM by flow cytometer after cocultured with ADSCs overexpressing PnNOS gene. The mRNA and protein expression of PnNOS gene mediated by recombinant adenovirus vector significantly overexpressed and lasted at least 2 weeks. Meanwhile, the concentration of NO and cGMP in ADSCs was greatly increased. The concentration of cGMP was significantly increased, and [Ca(2+) ]i was obviously decreased in CCSMCs compared with the control groups (P < 0.05) after cocultured with ADSCs for 3 days. These findings demonstrated that ADSCs overexpressing PnNOS gene might decrease [Ca(2+) ]i in CCSMCs by up-regulating NO-cGMP signalling pathway.

A pilot study to determine penile oxygen saturation before and after vacuum therapy in patients with erectile dysfunction after radical prostatectomy.

INTRODUCTION: Provoked and spontaneous nocturnal erections are thought to play a role in maintenance of male sexual health through oxygenation of the corpus cavernosa. Conversely, hypoxia is thought to be an etiological factor in the pathogenesis of cavernosal fibrosis and long-term erectile dysfunction. It has been hypothesized that the early penile hypoxia after radical prostatectomy (RP) may lead to fibrosis and consequently a decrease in stretched penile length and long-term erectile dysfunction. AIM: The aim of this study was to assess the changes in penile tissue oxygenation with vacuum erection device (VED) use. METHODS: Twenty men between 2 and 24 months following RP were enrolled prospectively. Each man cycled a VED to achieve full erection 10 consecutive times over a period of approximately 2 minutes without constriction ring. MAIN OUTCOME MEASURES: Tissue oximetry was measured at baseline and immediately after VED using a tissue oximeter at five sites: right thigh, right corpora, glans, left corpora, and left thigh. Additional measurements were captured over the course of an hour. RESULTS: Mean age and time from surgery was 58.2 years and 12.6 months, respectively, and the average Sexual Health Inventory for Men score was 7. Use of the VED significantly increased both glanular and corporal oximetry relative to the baseline values for the entire 60 minutes. An initial increase of 55% was seen in corporal oxygenation with VED use. CONCLUSIONS: This is the first study demonstrating that a single, brief application of the VED without a constriction ring results in significant improvement in penile oxygen saturation. The use of a VED has significant benefits for patients both with regard to cost and invasiveness when compared with other penile rehabilitation protocols.

L-arginine and tetrahydrobiopterin, but not sodium nitrite partially restored erectile dysfunction in aged rats.

Aging is associated with erectile dysfunction (ED), in which nitric oxide synthase (NOS) activity and NO bioavailability are reduced due to deficiencies of NOS cofactor (tetrahydrobiopterin, BH(4)) and substrate (L-arginine). We determined whether the prolonged treatment with sodium nitrite (NaNO(2)) as a storage form of NO ameliorates ED in aged rats. Male Sprague-Dawley rats were divided: younger, aged and NaNO(2)-treated (20 mg/kg per day) aged groups. The erectile (intracavernosal pressure [ICP]/mean arterial pressure [MAP]) and corpus cavernous (CC) responses were evaluated after 12 weeks. The ICP/MAP in aged rats was lower than in young controls, which was not improved by the NaNO(2) treatment. Immunohistochemical (IHC) staining for endothelial NOS and collagen deposition was performed. We assayed NO indirectly by measuring the level of its stable end products, nitrite/nitrate, using the Griess reagent. The relaxations to ACh and EFS in the aged group were considerably less than in the younger group, which were normalized by acute incubations of l-arginine or BH(4) of aged CC. In conclusion, NaNO(2) treatment did not restore erectile response while nitrate levels were enhanced in aged penis. The cofactor or substrate administrations, but not chronic exogenous modulation of NO system may be beneficial in aged men with ED.

Antiviral antibodies and T cells are present in the foreskin of simian immunodeficiency virus-infected rhesus macaques.

No information exists regarding immune responses to human immunodeficiency virus (HIV) infection in the foreskin or glans of the human penis, although this is a key tissue for HIV transmission. To address this gap, we characterized antiviral immune responses in foreskin of male rhesus macaques (RMs) inoculated with simian immunodeficiency virus (SIV) strain SIVmac251 by penile foreskin exposure. We found a complete population of immune cells in the foreskin and glans of normal RMs, although B cells were less common than CD4(+) and CD8(+) T cells. IgG-secreting cells were detected by enzyme-linked immunospot (ELISPOT) assay in cell suspensions made from the foreskin. In the foreskin and glans of SIV-infected RMs, although B cells were less common than CD4(+) and CD8(+) T cells, SIV-specific IgG antibody was present in foreskin secretions. In addition, cytokine-secreting SIV-specific CD8(+) T cells were readily found in cell suspensions made from the foreskin. Although potential HIV target cells were found in and under the epithelium covering all penile surfaces, the presence of antiviral effector B and T cells in the foreskin suggests that vaccines may be able to elicit immunity in this critical site to protect men from acquiring HIV.

Proteomic and Clinicopathologic Assessment of Penile Amyloidosis: A Single Institutional Review of 12 Cases.

OBJECTIVES: There is a paucity of data on penile amyloidosis. We aimed to assess the frequency of different amyloid types in surgical specimens from the penis involved by amyloidosis and correlate relevant clinicopathologic parameters with proteomic findings. METHODS: Since 2008, our reference laboratory has performed liquid chromatography/tandem mass spectrometry (LC-MS/MS) for amyloid typing. The institutional pathology archive and reference laboratory database were queried to retrospectively identify all penile surgical pathology specimens with LC-MS/MS results between January 1, 2008, and November 23, 2022. Archived H&E-stained and Congo red-stained sections were re-reviewed. RESULTS: Twelve cases of penile amyloidosis were identified, which represented 0.35% (n = 3,456) of penile surgical specimens. AL-type amyloid was most frequent (n = 7), followed by keratin-type amyloid (n = 3) and ATTR (transthyretin)-type amyloid (n = 2). AL-type amyloid cases often showed diffuse dermal/lamina propria deposition, whereas all keratin-type amyloid cases were localized to the superficial dermis. Two cases with keratin-type amyloid had concomitant cutaneous findings (penile intraepithelial neoplasia and condyloma). CONCLUSIONS: This series, the largest to date, demonstrates that penile amyloidosis has a heterogeneous proteomic landscape. To the best of our knowledge, this is the first study describing ATTR (transthyretin)-type penile amyloid.

Estrogen-induced maldevelopment of the penis involves down-regulation of myosin heavy chain 11 (MYH11) expression, a biomarker for smooth muscle cell differentiation.

Cavernous smooth muscle cells are essential components in penile erection. In this study, we investigated effects of estrogen exposure on biomarkers for smooth muscle cell differentiation in the penis. Neonatal rats received diethylstilbestrol (DES), with or without the estrogen receptor (ESR) antagonist ICI 182,780 (ICI) or the androgen receptor (AR) agonist dihydrotestosterone (DHT), from Postnatal Days 1 to 6. Tissues were collected at 7, 10, or 21 days of age. The smooth muscle cell biomarker MYH11 was studied in depth because microarray data showed it was significantly down-regulated, along with other biomarkers, in DES treatment. Quantitative real time-PCR and Western blot analyses showed 50%-80% reduction (P ≤ 0.05) in Myh11 expression in DES-treated rats compared to that in controls; and ICI and DHT coadministration mitigated the decrease. Temporally, from 7 to 21 days of age, Myh11 expression was onefold increased (P ≥ 0.05) in DES-treated rats versus threefold increased (P ≤ 0.001) in controls, implying the long-lasting inhibitory effect of DES on smooth muscle cell differentiation. Immunohistochemical localization of smooth muscle alpha actin, another biomarker for smooth muscle cell differentiation, showed fewer cavernous smooth muscle cells in DES-treated animals than in controls. Additionally, DES treatment significantly up-regulated Esr1 mRNA expression and suppressed the neonatal testosterone surge by 90%, which was mitigated by ICI coadministration but not by DHT coadministration. Collectively, results provided evidence that DES treatment in neonatal rats inhibited cavernous smooth muscle cell differentiation, as shown by down-regulation of MYH11 expression at the mRNA and protein levels and by reduced immunohistochemical staining of smooth muscle alpha actin. Both the ESR and the AR pathways probably mediate this effect.

Poly(ADP-ribose) polymerase inhibition improves erectile function by activation of nitric oxide/cyclic guanosine monophosphate pathway in diabetic rats.

INTRODUCTION: Endothelial 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. AIM: The 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. METHODS: Male 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 MEASURES: Erectile 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. RESULTS: The 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. CONCLUSION: Overactivation 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.

Effects of chronic vardenafil treatment persist after end of treatment in rats with acute arteriogenic erectile dysfunction.

INTRODUCTION: In our previous study, chronic vardenafil treatment improved erectile function soon after the end of the treatment in rats with acute arteriogenic erectile dysfunction (ED). AIM: The aim of this study is to evaluate whether the effects of chronic vardenafil treatment persist after the end of treatment using rats with acute arteriogenic ED. METHODS: Rats were randomly divided into three groups: (i) control; (ii) ligation; and (iii) vardenafil + no treatment. Rats in the ligation and vardenafil + no treatment groups underwent ligature of the bilateral internal iliac arteries to induce acute arteriogenic ED and were subsequently treated with vehicle or vardenafil (4.0 mg/kg/day), respectively, for 3 weeks. Subsequently, all rats were kept for a further 2 weeks with no treatment. Rats in the control group underwent sham surgery. MAIN OUTCOME MEASURES: Erectile function was assessed by changes in intracavernous pressure (ICP). Smooth muscle (SM)/collagen ratios in corpus cavernosum were analyzed by Masson trichrome staining. Transforming growth factor-ß1 (TGF-ß1 ) mRNA and protein levels in corpus cavernosum (CC) were, respectively, evaluated by real-time polymerase chain reaction (PCR) analysis and Western blotting analysis. RESULTS: ICP/mean arterial pressure (MAP) in the ligation group remained significantly lower than that in control group (P < 0.01). Despite no treatment for 2 weeks, ICP/MAP in the var + no treatment group remained significantly higher than that in ligation group (P < 0.05). SM/collagen ratio in the ligation group remained significantly lower when compared with the control group (P < 0.01). The ratio in the var + no treatment group remained significantly higher when compared with the ligation group at 2 weeks after the end of treatment (P < 0.05). TGF-ß(1) mRNA and protein levels did not differ among the groups. CONCLUSIONS: The effects of chronic vardenafil treatment on erectile function and penile structure persist, even after the end of treatment, in acute arteriogenic ED rats.

TNF-α, erectile dysfunction, and NADPH oxidase-mediated ROS generation in corpus cavernosum in high-fat diet/streptozotocin-induced diabetic rats.

INTRODUCTION: Patients 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. AIM: We 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. METHODS: Four 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 MEASURES: The effect of INF on HFD/STZ-induced diabetic ED and NADPH oxidase-mediated ROS generation was studied in diabetic corpus cavernosum. RESULTS: Untreated 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. CONCLUSIONS: Increased 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.

Molecular targets for diabetes mellitus-associated erectile dysfunction.

Protein expression profiles in rat corporal smooth muscle tissue were compared between animal models of streptozotocin-induced diabetes mellitus (STZ-DM) and age-matched controls (AMCs) at 1 week and 2 months after induction of hyperglycemia with STZ treatment. At each time point, protein samples from four STZ-DM and four AMC rat corpora tissues were prepared independently and analyzed together across multiple quantitative two-dimensional gels using a pooled internal standard sample to quantify expression changes with statistical confidence. A total of 170 spots were differential expressed among the four experimental groups. A subsequent mass spectrometry analysis of the 170 spots identified a total of 57 unique proteins. Network analysis of these proteins using MetaCore suggested altered activity of transcriptional factors that are of too low abundance to be detected by the two-dimensional gel method. The proteins that were down-regulated with diabetes include isoforms of collagen that are precursors to fibril-forming collagen type 1; Hsp47, which assists and mediates the proper folding of procollagen; and several proteins whose abundance is controlled by sex hormones (e.g. CRP1 and A2U). On the other hand, proteins seen or predicted to be up-regulated include proteins involved in cell apoptosis (e.g. p53, 14-3-3-gamma, Serpinf1, Cct4, Cct5, and Sepina3n), proteins that neutralize the biological activity of nerve growth factor (e.g. anti-NGF 30), and proteins involved in lipid metabolism (e.g. apoA-I and apoA-IV). Subsequent Western blot validation analysis of p53, 14-3-3-gamma, and Hsp47 confirmed increased p53 and 14-3-3-gamma and decreased Hsp47 levels in separate samples. According to the results from the Western blot analysis, Hsp47 protein showed a approximately 3-fold decrease at 1 week and was virtually undetectable at 2 months in diabetic versus control. Taken together, our results identify novel candidate proteins playing a role in erectile dysfunction in diabetes resulting from STZ treatment.

Effect of periodontitis on erectile function and its possible mechanism.

INTRODUCTION: Periodontitis is one of the important risk factors resulting in cardiovascular diseases. Erectile dysfunction (ED) is strongly correlated with cardiovascular diseases. The expression of endothelial nitric oxide synthase (eNOS) in penile tissue has an important role in the mechanism of erection. AIM: To investigate the effect of periodontitis on erectile function and the possible mechanism. METHODS: After induction of periodontitis in rat, the ratio of maximum intracavernosal pressure/mean arterial pressure (ICPmax /MAP)×100, the expression of eNOS in penile tissue, the level of serum C-reactive protein (CRP) and tumor necrosis factor-α (TNF-α), and the ultrastructural changes of the cavernous tissue were examined and compared between periodontitis rats (group A) and control rats (group B). MAIN OUTCOME MEASURE: Periodontitis significantly decrease not only the ICPmax/MAP×100 and the expression of eNOS but also the activity of NOS and the level of cyclic guanosine monophosphate (cGMP) in cavernous tissue of rat. RESULTS: After electrostimulation by 3 and 5 voltage, the ratio of ICPmax /MAP×100 in group A was significantly less than that in group B (19.54±6.16 vs. 30.45±3.12; 30.91±5.61 vs. 50.52±9.52, respectively; P<0.05).The level of serum CRP and TNF-α in group A is significantly higher in group B (P<0.05).The quantitative real-time reverse transcription polymerase chain reaction study demonstrated no statistically significant difference in the expression of mRNA of eNOS in cavernous tissue between the two groups (P>0.05). But there was significant decrease in eNOS protein of the cavernous tissue in group A than in group B (P<0.05). Total NOS activity and cGMP level in cavernosal tissue were significantly lower in group A than in group B (P<0.05). There was no significant alternation occurred in the ultrastructures of penile cavernous tissue. CONCLUSIONS: The function of penile erection is impaired by periodontitis. The decreased in the expression of eNOS and NOS activity in penile cavernous tissue caused by mild systemic inflammatory status in periodontitis may be one of the important risk factors of ED.

Impaired erectile function in CD73-deficient mice with reduced endogenous penile adenosine production.

INTRODUCTION: Adenosine has been implicated in normal and abnormal penile erection. However, a direct role of endogenous adenosine in erectile physiology and pathology has not been established. AIM: To determine the functional role of endogenous adenosine production in erectile function. METHODS: CD73-deficient mice (CD73(-/-)) and age-matched wild-type (WT) mice were used. Some WT mice were treated with alpha, beta-methylene adenosine diphosphate (ADP) (APCP), a CD73-specific inhibitor. High-performance liquid chromatography was used to measure adenosine levels in mouse penile tissues. In vivo assessment of intracorporal pressure (ICP) normalized to mean arterial pressure (MAP) in response to electrical stimulation (ES) of the cavernous nerve was used. MAIN OUTCOME MEASUREMENT: The main outcome measures of this study were the in vivo assessment of initiation and maintenance of penile erection in WT mice and mice with deficiency in CD73 (ecto-5'-nucleotidase), a key cell-surface enzyme to produce extracellular adenosine. RESULTS: Endogenous adenosine levels were elevated in the erected state induced by ES of cavernous nerve compared to the flaccid state in WT mice but not in CD73(-/-) mice. At cellular levels, we identified that CD73 was highly expressed in the neuronal, endothelial cells, and vascular smooth muscle cells in mouse penis. Functionally, we found that the ratio of ES-induced ICP to MAP in CD73(-/-) mice was reduced from 0.48 ± 0.03 to 0.33 ± 0.05 and ES-induced slope was reduced from 0.30 ± 0.13 mm Hg/s to 0.15 ± 0.05 mm Hg/s (both P < 0.05). The ratio of ES-induced ICP to MAP in APCP-treated WT mice was reduced from 0.49 ± 0.03 to 0.38 ± 0.06 and ES-induced slope was reduced from 0.29 ± 0.11 mm Hg/s to 0.19 ± 0.04 mm Hg/s (both P < 0.05). CONCLUSION: Overall, our findings demonstrate that CD73-dependent production of endogenous adenosine plays a direct role in initiation and maintenance of penile erection.

Increased cavernous expression of transforming growth factor-ß1 and activation of the Smad signaling pathway affects erectile dysfunction in men with spinal cord injury.

INTRODUCTION: Transforming growth factor-ß1 (TGF-ß1) is implicated in bladder fibrosis after spinal cord injury (SCI) and in the fibrosis in the corpus cavernosum tissue after cavernous nerve injury. AIM: We investigated the differential expression of TGF-ß1 and the Smad transcription factor, the key molecule for the initiation of TGF-ß-mediated fibrosis, in cavernous tissue from SCI patients. METHODS: After obtaining informed consent and approval from the patients and our institutional review board, we enrolled 5 patients with psychogenic erectile dysfunction (ED) (mean age 36.8 years; range 20-50 years) and 10 patients with neurogenic ED from SCI (mean age 38.8 years; range 18-50 years). Cavernous tissues were obtained by percutaneous biopsy and stained with Masson trichrome, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL), or antibodies to TGF-ß1 and phospho-Smad2. MAIN OUTCOME MEASURES: Semi-quantitative analysis of TGF-ß1 and phospho-Smad2 was performed, and the numbers of apoptotic cells were counted. We also quantified the cavernous collagen area with the use of an image analyzer system. RESULTS: The expression of TGF-ß1 and phospho-Smad2 protein was significantly higher in the SCI group than in the psychogenic group. The TUNEL assay revealed a higher apoptotic index in the SCI group than in the psychogenic group. Higher TGF-ß1 and phospho-Smad2 expression and more apoptotic cells were noted mainly in endothelial cells, smooth muscle cells, and fibroblasts of the SCI group. Double labeling of cavernous tissue with TUNEL and antibody to phospho-Smad2 revealed that most TUNEL-positive cells showed immunoreactivity to phospho-Smad2 staining. Cavernous collagen content was significantly greater in the SCI group than in the psychogenic group. CONCLUSION: Upregulation of TGF-ß1 and activation of the Smad signaling pathway may play important roles in SCI-induced cavernous fibrosis and deterioration of erectile function, which warrants early pharmacological intervention to protect erectile tissue from irreversible damage.