Faiz H. Mumtaz

The effect of sildenafil on corpus cavernosal smooth muscle relaxation and cyclic GMP formation in the diabetic rabbit

Sildenafil, a type V phosphodiesterase inhibitor, enhances smooth muscle relaxation in normal human and rabbit corpus cavernosum. We investigated the in vitro effects of sildenafil on non-adrenergic, non-cholinergic and nitric oxide (NO)-mediated cavernosal smooth muscle relaxation in diabetic rabbits, since alterations in this pathway are recognised in diabetic erectile dysfunction. Diabetes mellitus was induced in male New Zealand White rabbits with alloxan. Cavernosal strips from age-matched control, 3- and 6-month diabetic animals were mounted in organ baths. Relaxation responses to electrical field stimulation (1-20 Hz) or sodium nitroprusside (10(-8)-10(-4) M) were assessed in the absence and presence of sildenafil (10(-8) and 10(-7) M). The effect of sildenafil on cGMP formation by the corpus cavernosum was also assessed following stimulation with sodium nitroprusside, A23187 and acetylcholine. Sodium nitroprusside-stimulated relaxations were significantly (P<0.03) impaired in the corpus cavernosum from both diabetic groups, (IC(50)=4.6 x 10(-6) M following 3 months of diabetes mellitus and 4.0 x 10(-6) M following 6 months of diabetes mellitus; compared to 7.5 x 10(-7) M for pooled age-matched controls). Sildenafil (10(-7) M) significantly enhanced sodium nitroprusside-stimulated relaxation in control (P<0.05) and diabetic groups (P<0.03). Electrical field stimulation-mediated relaxations of the corpus cavernosum were significantly impaired after 6-month diabetes mellitus and enhanced by sildenafil (10(-8) M). cGMP formation by the diabetic corpus cavernosum was impaired significantly, but restored towards normal by sildenafil. We suggest that the impairment of NO-mediated relaxation of the corpus cavernosum reflect, at least in part, a defect in guanylyl cyclase activity. These findings support the use of sildenafil as an effective, orally administered, treatment for diabetic erectile dysfunction.

Cancers of the bladder.

Bladder cancer is the fifth most common malignancy in Europe and the fourth most common malignancy in the United States.1 It affects one in 4000 people and accounts for 5% of all diagnosed cancers. The peak incidence is in the fifth and seventh decade. There is a strong association between smoking and bladder cancer. Smokers have a fourfold higher incidence of developing bladder cancer than the general population.2 The disease has a spectrum of clinical severity varying from superficial bladder cancer to muscle invasive or metastatic disease which carries a poor prognosis. Currently the superficial form of the disease is managed by endoscopic resection of the tumour, often followed by the instillation into the bladder of cytotoxic agents. Due to the tendency of bladder cancer to recur repeated cystoscopies and resections are often required. Because of this, one of the main thrusts of research is to find a way of preventing the progression from superficial disease to muscle invasive and metastatic bladder cancer.

The effect of serotonin and serotonin antagonists on bladder cancer cell proliferation

To investigate the role of serotonin (5‐hydroxytryptamine, 5HT) and its antagonists in the proliferation of high‐grade bladder cancer cells (HT1376), as high‐grade bladder cancer has a rapid rate of progression, invasion and recurrence, and 5HT antagonists inhibit the growth of the prostate cancer cell line (PC3).

ALTERATIONS IN THE NITRIC OXIDE SYNTHASE BINDING SITES AND NON-ADRENERGIC, NON-CHOLINERGIC MEDIATED SMOOTH MUSCLE RELAXATION IN THE DIABETIC RABBIT BLADDER OUTLET: POSSIBLE RELEVANCE TO THE PATHOGENESIS OF DIABETIC CYSTOPATHY

PURPOSEnTo investigate the effect of diabetes mellitus (DM) on the density and distribution of nitric oxide synthase (NOS) and the smooth muscle responses to non-adrenergic, non-cholinergic (NANC) nerve stimulation and exogenous nitric oxide (NO) in the rabbit lower urinary tract.nnnMATERIALS AND METHODSnTransverse sections of detrusor, bladder neck and urethra, from control and six months alloxan-induced DM New Zealand White rabbits were incubated with a radioligand for NOS ([3H]-L-N(G)-nitroarginine). Densitometric analysis was performed on the autoradiographs. NADPH diaphorase histochemistry was also used as a marker for NOS activity. Responses to NANC nerve stimulation (5 to 20 Hz) and to NO (10(-6) to 3x10(-4) M.) on smooth muscle strips from detrusor, bladder neck and urethra were measured in organ baths.nnnRESULTSnNOS binding sites were significantly (p<0.03) more dense in the bladder neck than in the detrusor in both DM and control groups. In DM bladder neck, NOS binding sites were significantly (p<0.04) increased compared with the controls. NADPH diaphorase activity appeared markedly increased in the detrusor, bladder neck and urethra of DM animals compared with controls. The mean IC50 for exogenous NO in control versus DM were not statistically different in the bladder neck (1.03x10(-4) M versus 9.8x10(-5) M) and urethra (8.1x10(-5) M versus 8.8x10(-5) M), but the relaxations to 5x10(-6) M of NO were significantly impaired (p<0.04) in the DM urethral smooth muscle. NANC nerve-mediated relaxations were significantly impaired (p<0.001) in the DM urethral smooth muscle.nnnCONCLUSIONSnAlterations of both the NOS binding sites and functional responses to NANC nerve stimulation suggest that NO may have a pathophysiological role in the urinary bladder dysfunction associated with DM.

Down-regulation of nitric oxide synthase in the diabetic rabbit kidney: potential relevance to the early pathogenesis of diabetic nephropathy

SUMMARY Objectives: Nephropathy is a well-recognised complication of diabetes mellitus (DM). The aim of this study was to investigate the effect of DM on the density and distribution of nitric oxide (NO) synthase (NOS) in the rabbit kidney. Quantification of the NOS radioligand on slide-mounted sections was compared with the nitroblue tetrazolium reaction, where the intensity of the reaction varies with the nicotinamide adenine dinucleotide diaphorase (NADPH-d) activity of NOS. Materials and methods: DM was induced with alloxan in six New Zealand White (NZW) rabbits. Plasma creatinine, urea and electrolytes were monitored at monthly intervals. The kidneys were removed following 6 months of DM. Transverse serial sections were cut and low-resolution autoradiography was performed using a radioligand for NOS ([3H]-NOARG). Histochemical localisation of NADPH-d activity was also performed. Densitometric analysis was performed on the autoradiographs and the results compared with those obtained from six age-matched control rabbits. Results: There was a significant (p < 0.01) rise in plasma creatinine levels in the diabetic rabbits, although the mean values remained within the reference range. There was a significant (p < 0.0001) down-regulation of NOS binding sites in both the cortex and medulla of the DM kidney when compared with the controls. A similar decrease in NADPH-d activity was seen in the diabetic renal cortex and medulla. In addition, NADPH-d activity also appeared to be reduced in the diabetic glomeruli when compared with controls. Conclusions: NOS binding sites and NADPH-d activity are significantly decreased in the DM renal cortex and medulla. These changes are associated with a mild deterioration in renal function and may be an early event that could subsequently play a role in the progression of DM nephropathy. Manipulating the NO pathway during the early stages of DM nephropathy may be beneficial.

Understanding prostate cancer

Prostate cancer is the second most common cancer affecting men in Europe and the USA. The incidence of prostate cancer has risen by 60-75% in the Western world in the last 15 years. One in twelve men over the age of 60 develop prostate cancer and this figure is expected to rise to three in twelve in the next 20 years. Early prostate cancer often does not cause symptoms. However, patients may present with lower urinary tract symptoms (LUTS) and therefore, such patients should be investigated. Effective treatment in the form of surgery and radiotherapy is available for individuals with localised disease, and the effectiveness of different combination therapy is being assessed to improve the outcome further. Approximately 20% of the patients have metastatic disease on presentation. The mainstay of treatment for these patients is androgen ablation therapy; however patients on this regime eventually relapse and develop an androgen independent tumour. This aggressive stage of the disease carries a high morbidity and mortality. At present the treatment for such hormone refractory prostate cancer is inadequate and the desperate search for alternative forms of therapy continues.

Doxazosin modifies serotonin-mediated rabbit urinary bladder contraction. Potential clinical relevance.

Abstract 5-Hydroxytryptamine (5-HT) induces rabbit detrusor contractions via 5-HT3 receptors. Similarly, 5-HT4 receptors are known to be present in the human bladder. Doxazosin, a non-selective α1 antagonist, is used for the symptomatic relief of bladder outflow obstruction. Previous work has shown that doxazosin inhibits 5-HT2-mediated platelet shape change. Hence, the aim of this study was to assess, using organ baths and autoradiography, whether doxazosin has any 5-HT-inhibiting activity in the rabbit detrusor. Detrusor strips from adult New Zealand White rabbits were placed in organ baths; phenoxybenzamine (10−5u2009M) was added to block alpha-receptors. After KCl responses were assessed, the tissues were exposed to 10−3u2009M 5-HT. Subsequently, the strips were incubated with doxazosin or ondansetron (10−5u2009M; 5-HT3 antagonist) followed by a further exposure to 5-HT. In some experiments, after the initial 5-HT-induced contractions, the tissues were washed and then re-exposed to 5-HT. These latter experiments acted as controls. Low-resolution autoradiography was performed on detrusor sections to assess the effect of doxazosin on 5-HT binding. These sections were analyzed densitometrically. Doxazosin and ondansetron produced a significant reduction in 5-HT-mediated contractions. Inhibition by doxazosin was in a concentration-dependent manner. Autoradiography demonstrated a significant reduction in [3H]-5-HT binding by doxazosin. Doxazosin significantly inhibits 5-HT-mediated contractions in the rabbit detrusor. This effect appears to be mainly mediated via 5-HT3 receptor inhibition. Autoradiographic evidence suggests that doxazosin reduces 5-HT binding in the rabbit detrusor. The beneficial effects of doxazosin in bladder outflow obstruction may be due, at least in part, to 5-HT antagonism.

Relaxation of rabbit lower urinary tract smooth muscle by nitric oxide and carbon monoxide: modulation by hydrogen peroxide.

Recent studies suggest that the body produces two gaseous messengers, nitric oxide (NO) and carbon monoxide (CO), both of which activate soluble guanylyl cyclase and thus modulate the activity of smooth muscle cells. In the present study, the effects of NO and CO on the smooth muscle of the lower urinary tract were compared. In addition, the modulation of tissue NO- and CO-induced relaxation by hydrogen peroxide was examined. NO, produced endogenously by electrical field stimulation (EFS) or applied exogenously as a solution, induced a concentration-dependent relaxation of rabbit cavernosal and urethral smooth muscle strips, but not of bladder tissues. The cavernosal tissue was found to be three times more sensitive to the actions of NO than the urethra. CO also induced relaxation of both tissue types, but with no apparent difference in sensitivity between the tissues. However, CO was much less potent than NO with respect to smooth muscle relaxation. The mechanism of action of the two mediators was cyclic guanosine monophosphate (cGMP)-dependent, as evidenced by enhanced formation of cGMP and inhibition of relaxation by the guanylyl cyclase inhibitor, oxadiazoloquinoxaline-1-one (ODQ.) The data suggests that NO is the dominant messenger in these tissues, but does not exclude a role for CO. In the presence of hydrogen peroxide, the relaxation responses induced by both NO and CO were significantly increased, regardless of tissue type. The mechanism for this effect is unclear, but evidence points to a requirement for the activation of guanylyl cyclase and enhanced formation of cGMP, since potentiation by the peroxide was blocked by a specific guanylyl cyclase inhibitor. We suggest that H(2)O(2) may play a positive role in the amplification or NO and CO-mediated responses.

Serotonin Induces a Biphasic Response in Rabbit Cavernosal Smooth Muscle: Relevance to the Erectile Process

Introduction: Serotonin (5-hydroxytryptamine; 5-HT) can cause contraction in cavernosal smooth muscle. We further evaluated this effect of 5-HT. Methods: Organ bath studies were used. Results: 5-HT induced a sustained contraction occasionally accompanied by a transient relaxation (in 30% of rabbit cavernosal tissues) that preceded the contraction. Ondansetron and Y-25130 (both 5-HT3 receptor antagonists) but not SB-269970 (a 5-HT7 receptor antagonist) significantly inhibited or abolished this transient relaxation. Doxazosin (dox, an α1-receptor antagonist) and ketanserin (ketan, a 5-HT2A receptor antagonist) significantly inhibited or abolished the sustained contraction. The effects of dox on 5-HT-mediated contraction were concentration-dependent. Conclusions: Our findings further confirm that the peripheral serotonergic pathway may play a part in the erectile process via 5-HT2A receptor-mediated contractile and 5-HT3 receptor-mediated relaxant activities. Our results also support the findings of human studies, which suggest that both ketan and dox may exert beneficial effects on the erectile process.

Potential Role of Endothelin and Nitric Oxide in Physiology and Pathophysiology of the Lower Urinary Tract

Endothelium-derived vasoactive mediators (endothelin-1 with its vasoconstrictive and mitogenic properties and nitric oxide with its vasodilatory and antiproliferative properties) play an important role in the regulation of vascular smooth muscle tone and cellular proliferation. Several recent studies have now demonstrated the presence of these vasoactive agents in the urinary tract where they are thought to play a prominent role in urinary tract physiology and disease. This article reviews the synthesis, localisation and actions of endothelin and nitric oxide in the lower urinary tract and examines the possible role of these mediators in disease.