Diane Gorny

Flow (Shear Stress)–Induced Endothelium-Dependent Dilation Is Altered in Mice Lacking the Gene Encoding for Dystrophin

Background —Dystrophin has a key role in striated muscle mechanotransduction of physical forces. Although cytoskeletal elements play a major role in the mechanotransduction of pressure and flow in vascular cells, the role of dystrophin in vascular function has not yet been investigated. Thus, we studied endothelial and muscular responses of arteries isolated from mice lacking dystrophin (mdx mice). Methods and Results —Carotid and mesenteric resistance arteries 120 &mgr;m in diameter were isolated and mounted in vitro in an arteriograph to control intraluminal pressure and flow. Blood pressure was not affected by the absence of dystrophin. Pressure-induced (myogenic), phenylephrine-induced, and KCl-induced forms of tone were unchanged. Flow (shear stress)–induced dilation in arteries isolated from mdx mice was decreased by 50% to 60%, whereas dilation to acetylcholine or sodium nitroprusside was unaffected. NG-nitro-L-arginine methyl ester–sensitive flow dilation was also decreased in arteries from mdx mice. Thus, the absence of dystrophin was associated with a defect in signal transduction of shear stress. Dystrophin was present in vascular endothelial and smooth muscle cells, as shown by immunolocalization, and localized at the level of the plasma membrane, as seen by confocal microscopy of perfused isolated arteries. Conclusions —This is the first functional study of arteries lacking the gene for dystrophin. Vascular reactivity was normal, with the exception of flow-induced dilation. Thus, dystrophin could play a specific role in shear-stress mechanotransduction in arterial endothelial cells. Organ damage in such diseases as Duchenne dystrophy might be aggravated by such a defective arterial response to flow.

Combination of Doxazosin and Sildenafil Exerts an Additive Relaxing Effect Compared with Each Compound Alone on Human Cavernosal and Prostatic Tissue

INTRODUCTION Phosphodiesterase 5 inhibitors (PDE5) such as sildenafil are first-line treatment for erectile dysfunction (ED). Alpha1 (alpha1)-adrenoceptor antagonists such as doxazosin are indicated for the treatment of patients with lower urinary tract symptoms (LUTS)/benign prostatic hyperplasia (BPH). ED and LUTS/BPH are conditions that are often associated. Accordingly, alpha1-adrenoceptor antagonists and PDE5 inhibitors will be often prescribed in real life setting together. AIM To evaluate the effects of the combination of sildenafil and doxazosin on human cavernosal and prostatic tissue. METHODS Prostatic and erectile tissues were obtained from nine to 12 patients, respectively. Patients underwent cystoprostatectomy for infiltrating bladder cancer or penile surgery for penile implant, congenital curvature or Peyronies disease. MAIN OUTCOME MEASURES In organ baths, prostatic and cavernosal strips were submitted to either concentration-response curves (CRC) to phenylephrine (Phe) or norepinephrine (NE), respectively, in presence of vehicle, sildenafil (10(-6) M, 10(-5) M), doxazosin (10(-8) M, 3.10(-8) M, or 10(-7) M), or a combination of both. Continuous electrical field stimulation (EFS; 32 Hz, 5 ms, 5 seconds, 300 mA) was performed on prostatic strips which were incubated with sildenafil 10(-6) M or vehicle before the successive addition of doxazosin (10(-7) M, 10(-6) M) or vehicle. Cavernosal strips were pre-incubated with doxazosin (10(-9) M, 10(-8) M) or vehicle, then CRC to sildenafil were constructed on NE (3.10(-6) M) precontracted cavernosal strips. RESULTS Combination of sildenafil and doxazosin exerted a greater relaxing effect on CRC to Phe or NE compared with each compound alone in both tissues. Sildenafil significantly enhanced the relaxing effect of doxazosin on EFS-induced contractions in prostatic strips. Doxazosin significantly increased the ability of sildenafil to inhibit NE-induced contractions in cavernosal strips. CONCLUSIONS Sildenafil and doxazosin reduced adrenergic tone of prostatic and cavernosal smooth muscle and their combination provided a significant benefit when targeting relaxation of both tissues. These experiments provide support for further clinical evaluation of the sildenafil and doxazosin combination in ED patients with LUTS/BPH.

Effects of potassium channel modulators on myogenic spontaneous phasic contractile activity in human detrusor from neurogenic patients.

To characterize the spontaneous contractile activity (SCA) developed by detrusor from patients with neurogenic detrusor overactivity (NDO) because the alteration of detrusor properties plays a critical role in the pathogenesis of detrusor overactivity, as well as to evaluate the role of KATP and KCa channels on this SCA because these channels regulate detrusor SCA in many species, including humans without overactive bladder (OAB).

Combination of Alfuzosin and Tadalafil Exerts an Additive Relaxant Effect on Human Detrusor and Prostatic Tissues In Vitro

BACKGROUND Lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) and erectile dysfunction (ED) are highly prevalent in aging men and are strongly linked. Alpha₁-blockers such as alfuzosin are effective monotherapies for LUTS. Phosphodiesterase type 5 (PDE5) inhibitors such as tadalafil are the first-line treatment for ED. Both drugs act by two different mechanisms of action on common urogenital target organs and, thus, may have additive effects. OBJECTIVES We evaluated in vitro the effects of alfuzosin, tadalafil, and the combination of both on human detrusor and prostatic smooth muscle. DESIGN, SETTING, AND PARTICIPANTS Prostatic and bladder tissue were obtained from patients (n=20 and n=17, respectively) undergoing cystoprostatectomy for bladder cancer. MEASUREMENTS In organ baths, isolated prostatic strips and isolated bladder strips were incubated with vehicle, tadalafil (10⁻⁶ M and 10⁻⁵ M), alfuzosin (3×10⁻⁸ M or 10⁻⁶ M and 10⁻⁵ M) or a combination. Concentration-response curves (CRCs) to norepinephrine were generated on prostatic strips and detrusor strips precontracted with carbachol. Strips were also submitted to electrical field stimulation (EFS). RESULTS AND LIMITATIONS When alfuzosin and tadalafil were combined, the maximal relaxation to norepinephrine on carbachol-precontracted detrusor strips was significantly increased compared with tadalafil alone, and EFS-induced detrusor contractions were better inhibited compared with each compound alone. Tadalafil significantly inhibited norepinephrine-induced prostatic strip contractions by reducing the maximal effect, whereas alfuzosin shifted the CRC of norepinephrine to the right. Combining both tadalafil and alfuzosin resulted in a greater relaxant effect. Likewise, the combination was more effective at reducing EFS-induced contractions compared with each compound alone. CONCLUSIONS The combination of alfuzosin and tadalafil exerts an additive effect of inhibiting adrenergic smooth muscle tone of prostatic tissue and EFS-induced detrusor contractions and conversely, of enhancing adrenergic relaxation of detrusor precontracted with carbachol. These experiments provide experimental support for the clinical investigation of the combination of α1-blockers and PDE5 inhibitors in the treatment of LUTS.

Combination of alfuzosin and tadalafil exerts in vitro an additive relaxant effect on human corpus cavernosum.

INTRODUCTION Phosphodiesterase type 5 (PDE5) inhibitors, such as tadalafil, are a first-line treatment for erectile dysfunction (ED). Nevertheless, some patients do not respond to this treatment. Clinical data suggest that the addition of alpha1-adrenoceptor blocker, such as alfuzosin, commonly prescribed for lower urinary tract symptoms suggestive of benign prostatic hyperplasia, may be of benefit. Aim. Evaluation of the effect of alfuzosin, tadalafil or the combination of both on human corpus cavernosum. METHODS Human cavernosal tissues were obtained from 10 patients undergoing penile surgery. Strips contractility was studied in organ baths. Concentration-response curves to tadalafil were generated on norepinephrine (NE, 1-10 microM)-precontracted strips in the presence of alfuzosin or vehicle. Frequency-response curves (FRC) to electrical field stimulation (EFS, 0-64 Hz, 3 ms, 10 seconds, 300 mA) were generated in the presence of vehicle, alfuzosin, tadalafil, or both drugs combined. EFS (20 Hz, 1 ms, 10 seconds, 300 mM)-induced nitrergic relaxation on NE-precontracted strips was studied in the presence of vehicle, alfuzosin, tadalafil, or both drugs combined. MAIN OUTCOME MEASURES Functional measurement of cavernosal smooth muscle relaxation in the presence of tadalafil and alfuzosin. RESULTS The relaxation induced by tadalafil (10(-10) to 10(-5) M) on precontracted strips was enhanced by alfuzosin at both 10(-8) and 10(-7) M. The combination of alfuzosin (3.10(-8) M) and tadalafil (10(-7) M) was more efficient to inhibit FRC-induced contractions than each compound alone. The combination of tadalafil (10(-6) M) and alfuzosin (10(-8) M) increased the relaxation induced by EFS and its effect was greater than tadalafil alone. In addition, the combination of tadalafil (10(-6) M) and alfuzosin (10(-7) M) prolonged EFS-induced relaxation to a greater extent than each compound alone. CONCLUSIONS In vitro, the combination of alfuzosin and tadalafil is more efficient than each compound alone to relax adrenergic tone or to enhance nitrergic relaxation of the human corpus cavernosum. Such a combination deserves further investigation in placebo-controlled studies to evaluate its benefit in ED patients who are not sufficiently improved by PDE5 inhibitors.

Low Intensity Extracorporeal Shock Wave Therapy Improves Erectile Function in a Model of Type II Diabetes Independently of NO/cGMP Pathway

PURPOSE Erectile dysfunction is highly prevalent in type II diabetes mellitus. Low intensity extracorporeal shock wave therapy improves erectile function in patients with erectile dysfunction of vasculogenic origin, including diabetes. However, its mode of action remains unknown. We investigated the effects of low intensity extracorporeal shock wave therapy compared to or combined with sildenafil on erectile dysfunction in a type II diabetes mellitus model. Our purpose was to test our hypothesis of a mode of action targeting the cavernous nitric oxide/cyclic guanosine monophosphate pathway. MATERIALS AND METHODS GK rats, a validated model of type II diabetes mellitus, and age matched Wistar rats were treated with low intensity extracorporeal shock wave therapy twice weekly for 3 weeks. Treatment was repeated after a 3-week no-treatment interval. The penis was stretched and dipped in a specifically designed water-filled cage. Shock waves were delivered by a calibrated probe yielding a controlled energy flux density (0.09 mJ/mm(2)). The probe was attached to an electrohydraulic unit with a focused shock wave source, allowing for accurate extrapolation to humans. Following a 4-week washout period erectile function was assessed as well as endothelium dependent and independent, and nitrergic relaxations of the corpus cavernosum of GK rats. RESULTS Low intensity extracorporeal shock wave therapy significantly improved erectile function in GK rats to the same extent as sildenafil. Treatment effects were potentiated when combined with sildenafil. Shock wave effects were not associated with improved cavernous endothelium dependent or independent, or nitrergic reactivity. CONCLUSIONS Low intensity extracorporeal shock wave therapy improved erectile function in GK rats. Unexpectedly, this was not mediated by a nitric oxide/cyclic guanosine monophosphate dependent mechanism. Sildenafil increased shock wave efficacy. This preclinical paradigm to deliver low intensity extracorporeal shock wave therapy to the rat penis should help further exploration of the mode of action of this therapy on erectile tissue.

Impact of a Long-Term Sildenafil Treatment on Pressor Response in Conscious Rats With Insulin Resistance and Hypertriglyceridemia

BACKGROUND Insulin resistance constitutes a risk factor for endothelial dysfunction and subsequent cardiovascular diseases including hypertension. Daily treatment with phosphodiesterase type 5 (PDE5) inhibitors has beneficial effects on endothelial function in men with increased cardiovascular risk. Endothelium-dependent vasomotor function is ultimately linked to blood pressure (BP) regulation. We postulated that sildenafil would ameliorate BP and biological markers of endothelial function in fructose-fed rats (FFRs). METHODS Wistar rats were fed a standard chow or a 60% fructose-enriched diet containing 12% fat for 8 weeks (FFR). From week 6 through 8, sildenafil (twice a day subcutaneously, 20 mg/kg) was administered followed by a 1-week washout period. At the end of the washout period, BP was recorded using radiotelemetry following cumulative infusion of norepinephrine (from 50 to 400 ng/kg/min). RESULTS FFR displayed both an impaired glucose tolerance and elevated triglyceridemia. The latter was corrected by sildenafil treatment. Resting BP was similar in all rats, whereas pressor responses were significantly enhanced in FFR (maximal increase in mean BP to norepinephrine: 25.6 +/- 3.8 vs. 40.8 +/- 4.0 mm Hg, P < 0.05) and normalized by sildenafil treatment (24.9 +/- 5.3 mm Hg, not significant vs. control). Urinary levels of 8-isoprostanes and thromboxane B(2) were increased in FFR and corrected by sildenafil treatment. CONCLUSION Thus, chronic treatment with sildenafil normalized BP regulation in an experimental model of insulin resistance and hypertriglyceridemia while restoring normal excretion of urinary biological markers of oxidative stress and cyclooxygenase-derived vasoconstrictors. The modulation of ROS and cyclooxygenase-derived vasoconstrictors generation by a chronic treatment with sildenafil may represent an added benefit beyond PDE5 inhibition.

An aryloxypropanolamine hβ3-adrenoceptor agonist as bladder smooth muscle relaxant

The relaxant effect of an aryloxypropanolamine β3-adrenoceptor agonist on carbachol pre-contracted human detrusor muscle strips was evaluated and compared with literature results from reference compounds of similar mode of action, including mirabegron. A significant relaxation was observed for rac-4-{2-hydroxy-3-[1-(5-phenylthieno[2,3-d]pyrimidin-4-yl)piperidin-4-ylamino]propoxy}-2-(hydroxymethyl)phenol which was similar to that exerted by mirabegron. In order to allow for a thorough discussion of results in comparison to reference compounds, their affinity, selectivity and efficacy as hβ3-AR agonists have been evaluated and discussed thoroughly. A ranking of hβ3-AR agonists by relative efficacy resulted in the closest analogy to the order of relaxation potential, with only the relaxant effect of mirabegron not reflecting its excellent relative efficacy as such.

SIGNALING PATHWAYS INVOLVED IN SILDENAFIL-INDUCED RELAXATION OF HUMAN DETRUSOR

reduce lower urinary tract symptoms (LUTS) in men. The mechanism of action is not fully understood. Since LUTS comprise urgency, frequency and nocturia which are arising mainly from the bladder, we investigated the effects of PDE5 inhibitors on the dog bladder. We determined the effects of PDE5 inhibitors in vitro, on dog bladder strips and in vivo on micturition frequency and volume per voiding in conscious dogs. METHODS: Bladder strips from male Beagle dogs were pre-

Combination of alfuzosin and tadalafil exerts an additive enhancing effect on norepinephrine-induced relaxation of pre-contracted human detrusor smooth muscle

INTRODUCTION ● Lower urinary tract symptoms (LUTS) and erectile dysfunction (ED) are highly prevalent in aging men and are strongly linked, independently of age and cardiovascular comorbidities1. ● Alpha1-adrenergic blockers such as alfuzosin are considered the most effective monotherapy for LUTS associated with benign prostatic hyperplasia (BPH)2. ● Phophodiesterase 5 (PDE5) inhibitors are a first line treatment for erectile dysfunction (ED)3. ● Recent clinical trials have shown that PDE5 inhibitors (sildenafil, tadalafil, vardenafil) could improve not only ED but also obstructive and irritative LUTS associated with BPH4-6. Interestingly, this beneficial effect on LUTS is not associated with an increased peak flow rate suggesting an extraprostatic mode of action. ● A pilot clinical study also indicates that daily administration of alfuzosin (10mg) in combination with sildenafil (25 mg) for 12 weeks may be more effective than monotherapy on LUTS associated with BPH and ED7. ● Tadalafil is currently the only one long-acting PDE5 inhibitor and the maximum prescribed dose (20 mg) shows no clinically relevant hemodynamic interaction with alfuzosin 10mg once daily8.