Endogenous vasoactive peptides and the human vagina--a molecular biology and functional study.

INTRODUCTION: Endogenous peptides, such as vasoactive intestinal polypeptide (VIP), C-type natriuretic peptide (CNP), and bradykinin (BK), have been proposed to play a role in the female sexual arousal response by exerting relaxation of clitoral, labial, and vaginal smooth muscle. While the effects of endogenous peptides on the human male erectile tissue have already been described, only very few studies have been conducted to investigate the peptidergic control of female genital tissues, including the vagina. AIMS: To elucidate the expression of mRNA specifically encoding for peptide receptors in the human vagina and the effects of VIP, CNP, and BK on the tension induced by endothelin-1 (ET-1) of isolated human vaginal wall smooth muscle. The production of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) in response to exposure of the tissue to the peptides was also measured. METHODS: The expression of mRNA encoding for receptor proteins specific for VIP, CNP, and BK were investigated by means of molecular biology (reverse transcriptase polymerase chain reaction [RT-PCR] analysis). Using the organ bath technique, the effects of VIP, CNP, and BK (0.1 nM to 1 µM) on the tension induced by 0.1 µM ET-1 of human vaginal strips were investigated. The tissue was also exposed to three different concentrations of VIP, CNP, and BK (0.01 µM, 0.1 µM, 1 µM) and the production of cAMP and cGMP determined by means of radioimmunoassays. MAIN OUTCOME MEASURES: Characterize the expression of peptide receptors in the human vagina and measure the relaxation exerted by BK, CNP, and VIP on the contraction induced by ET-1 of isolated human vaginal tissue. In addition, the effects of the peptides on the production of cAMP and cGMP were also elucidated. RESULTS: RT-PCR analysis revealed the expression of mRNA transcripts encoding for the VIP receptors VIP1R/vasoactive intestinal polypeptide receptor type 1 (VPAC1) and VIP2R/VPAC2, CNP receptors natriuretic peptide receptor type A (NPRA), natriuretic peptide receptor type B (NPRB) and natriuretic peptide receptor type C (NPRC), and BK receptor B2R. The tension induced by ET-1 was reversed by the peptides with the following rank order of efficacy: BK (21.7%) > VIP (20.9%) > CNP (13.3%). The relaxing effects of VIP and BK were paralleled by a 4.8-fold and fivefold increase in cAMP, while the production of cGMP was stimulated 38-fold and 119-fold in the presence of CNP or BK, respectively. CONCLUSION: Our results are in support of the hypothesis that endogenous peptides may contribute to the control of human vaginal smooth muscle tone through the involvement of the cyclic nucleotide-dependent pathways.

Evaluating the role of the serotoninergic system in the control of human seminal vesicle smooth muscle-an in vitro approach.

INTRODUCTION: It has been suggested that serotonin re-uptake inhibitors (SRIs) may retard the ejaculatory response by acting directly on the seminal vesicle (SV) and ductus deferens smooth muscle. However, until now, only a very few experimental studies have investigated such potential local (peripheral) effects. AIM: To elucidate the effects of serotonin (5-HT) and the SRIs clomipramine, fluoxetine and imipramine on the tension induced by norepinephrine (NE) of isolated human SV smooth muscle, as well as on the production of tissue cyclic AMP and cyclic GMP. MAIN OUTCOME MEASURES: To measure the inhibition exerted by serotonin and SRIs clomipramine, fluoxetine, and imipramine on the contractile response of isolated SV tissue. In addition, the effects of the drugs on the turn-over of cyclic nucleotides cAMP and cGMP were also elucidated. METHODS: The effects of the cumulative addition of serotonin and the SRIs clomipramine, fluoxetine and imipramine (1 nM-10 microM) on the tension induced by the alpha(1)-adrenoceptor agonist NE (10 microM) of SV strip preparations were studied using the organ bath technique. Cyclic AMP and cyclic GMP were measured by means of specific radioimmunoassays. RESULTS: The tension induced by NE was dose-dependently reversed by the drugs tested. The rank order of efficacy was: imipramine > or = fluoxetine > or = clomipramine > serotonin. Mean reversion of tension was measured between 66 +/- 6.6% and 52 +/- 6.6%. These effects were paralleled by a 1.3-fold to 2.7-fold increase in tissue cAMP in response to exposure to the drugs. In contrast, no significant enhancement in cGMP was noted. CONCLUSIONS: The findings, for the first time, present evidence that SRIs may antagonize the sympathetic contraction of SV smooth muscle via stimulation of tissue cyclic AMP.

Laparoscopy in German urology: changing acceptance among urologists.

BACKGROUND: Laparoscopic techniques have dramatically influenced urologic surgery in the past 2 decades. OBJECTIVES: A questionnaire was distributed in 2006 to analyse laparoscopic practice patterns in Germany. The results were compared with a survey performed in 2002. DESIGN, SETTING, AND PARTICIPANTS: In 2006, 324 German urology departments received a detailed, anonymous, and self-administered questionnaire regarding demographic data, current use, and attitudes concerning laparoscopy. Quantitative evaluation of laparoscopic procedures was performed for 20 indications. MEASUREMENTS: The response rate was 73% (238 of 324 institutions). Thirty-two responders were affiliated with universities; 95 responders were affiliated with urban hospitals; 101 responders were affiliated with general hospitals; and 9 responders were affiliated with private hospitals. Laparoscopy had been implemented as a standard operating procedure in 82% of the departments, an increase of 28% compared with the 2002 questionnaire. Forty-eight percent of participants expected a similar operating time to that of open surgery, an increase of 16% compared with the 2002 questionnaire. Concerns about the learning curve dropped from 92% in 2002 to 80% in 2006, and concerns about economic disadvantages dropped from 70% in 2002 to 45% in 2006. Criticism regarding lack of sufficient scientific data decreased from 76% in 2002 to 13% in 2006. Laparoscopic radical prostatectomies (>40 per year) were performed in 30 hospitals in 2006, an increase of 27% over 2002. Laparoscopic radical nephrectomy was performed 16-40 times per year in 33 of the responding institutions, an increase of 29% over 2002, and laparoscopic radical nephrectomy was performed >40 per year in 10 of the institutions, an increase of 9% over 2002. Laparoscopic pyeloplasty had a reported frequency of 16-40 procedures per year in 11 of the responding institutions, an increase of 10% over 2002, and laparoscopic pyeloplasty had a frequency between 5 and 15 procedures per year in 42 institutions, an increase of 37% over 2002. Only four hospitals performed cystectomy with ileum conduit and with orthotopic bladder substitute (5-15 cases per year). RESULTS AND LIMITATIONS: The results demonstrate the rising acceptance of laparoscopy in urologic surgery (an increase of 28% more departments performing laparoscopy) and an increasing interest in these techniques (an increase of 12% in the response rate). Their value is still limited by the response rate of only 73%. CONCLUSIONS: This survey demonstrates the increasing impact of laparoscopy on surgical patterns in urology and the increasing acceptance of laparoscopic techniques concerning operating time, learning curve, and scientific approval.

The nitric oxide pathway in the human prostate: clinical implications in men with lower urinary tract symptoms.

To date, there is an increasing interest in the nitric oxide (NO) pathway as a potential pharmacological target to treat male lower urinary tract symptomatology (LUTS). In the transition zone of the human prostate, a dense nitrinergic innervation has been shown of the fibromuscular stroma, glandular epithelium and blood vessels. The expression of key proteins of the NO pathway, such as the endothelial and neuronal nitric oxide synthase (eNOS, nNOS), cGMP-degrading phosphodiesterase type 5 (PDE5) and cGMP-binding protein kinase (cGK), has also been demonstrated. The hypothesis that an impaired NO/cGMP-signaling may contribute to the pathophysiology of benign prostatic hyperplasia (BPH) is supported by the results from randomized, placebo-controlled clinical studies, indicating that NO donor drugs and PDE5-inhibitors sildenafil, tadalafil and vardenafil may be useful to treat storage and voiding dysfunctions resulting from LUTS in men. Thus, given a potential role of the NO-pathway in the prostate and/or in other parts of lower urinary tract (e.g. bladder), the enhancement of the NO signaling by NO donor drugs, PDE5 inhibitors or activators of the soluble guanylyl cyclase (sGC) may represent a new therapeutic strategy for the treatment of LUTS. This review serves to focus on the role of NO and the NO-dependent signaling in the control of smooth muscle function in the human prostate. Results from clinical trials in men with LUTS/BPH are also discussed.

Age and bladder outlet obstruction are independently associated with detrusor overactivity in patients with benign prostatic hyperplasia.

BACKGROUND: Detrusor overactivity is one known cause of lower urinary tract symptoms and has been linked to bladder storage symptoms (urgency, frequency, or urge incontinence). OBJECTIVE: To determine clinical and urodynamic parameters associated with detrusor overactivity in patients with suspected benign prostatic hyperplasia. DESIGN, SETTING, AND PARTICIPANTS: During 1993-2003, urodynamic investigations were performed in patients aged 40 yr or older and with lower urinary tract symptoms, benign prostatic enlargement, and/or suspicion of bladder outlet obstruction (maximum flow rate < 15 ml/s or postvoid residual urine > 50 ml). MEASUREMENTS: Detrusor overactivity was defined according to the new International Continence Society classification (2002) as involuntary detrusor contractions during cystometry, which may be spontaneous or provoked, regardless of amplitude. The Schäfer algorithm was used to determine bladder outlet obstruction. RESULTS: In total, 1418 men were investigated (median age: 63 yr) of whom 864 men (60.9%) had detrusor overactivity. In univariate analysis, men with detrusor overactivity were significantly older, more obstructed, had larger prostates, higher irritative International Prostate Symptoms Score subscores, a lower voiding volume at free uroflowmetry, and a lower bladder capacity at cystometry. The prevalence of detrusor overactivity rose continuously with increasing bladder outlet obstruction grade. Multivariate analysis showed that only age and bladder outlet obstruction grade were independently associated with detrusor overactivity. After age adjustment, the odds ratios of detrusor overactivity compared to Schäfer class 0 were 1.2 for class I, 1.4 for class II, 1.9 for class III, 2.5 for class IV, 3.4 for class V, and 4.7 for class VI. CONCLUSIONS: In patients with clinical benign prostatic hyperplasia, detrusor overactivity is independently associated with age and bladder outlet obstruction. The probability of detrusor overactivity rises with increasing age and bladder outlet obstruction grade.

Effects of phosphodiesterase inhibitors on tension induced by norepinephrine and accumulation of cyclic nucleotides in isolated human prostatic tissue.

OBJECTIVES: To further evaluate the mechanism of action of phosphodiesterase (PDE) inhibitors on the human prostate, the effects of PDE4 and PDE5 inhibitors on the tension induced by norepinephrine (NE) and on the intracellular levels of cyclic nucleotides in isolated human prostatic tissue were investigated. METHODS: Using the organ bath technique, the effects of increasing concentrations (1 nM to 10 microM) of the PDE5 inhibitors sildenafil, tadalafil, and vardenafil and the PDE4 inhibitors rolipram and RP 73401 on the tension induced by NE (40 microM) of prostate strip preparations were investigated. The accumulation of cyclic guanosine monophosphate and cyclic adenosine monophosphate in response to drug exposure was determined by radioimmunoassays. RESULTS: The tension induced by NE was dose dependently reversed by the drugs with the following rank order of efficacy: tadalafil greater than RP 73401 greater than rolipram greater than or equal to vardenafil greater than sildenafil. The maximal reversion of tension values ranged from 52.3% (tadalafil) to 17% (sildenafil). Of the PDE inhibitors, only tadalafil induced a 50% reversion of the initial tension. The most prominent enhancement in tissue cyclic adenosine monophosphate was registered in response to RP 73401 (11-fold), and cyclic guanosine monophosphate levels were significantly elevated by tadalafil, vardenafil, and sildenafil (28-fold, 12-fold, and 3-fold, respectively). CONCLUSIONS: Our results have demonstrated that drugs interfering with the cyclic nucleotide-mediated pathways can reverse the tension induced by NE in isolated prostatic tissue and elevate cyclic adenosine monophosphate and cyclic guanosine monophosphate. Our findings serve to explain how PDE inhibitors can affect symptoms of lower urinary tract symptoms and benign prostatic hyperplasia.

Expression and distribution of cyclic GMP-dependent protein kinase-1 isoforms in human penile erectile tissue.

INTRODUCTION: Besides the bioavailability of nitric oxide (NO), downstream guanine monophosphate (cGMP) effector proteins are also considered to play a significant role in penile vascular disease. In animal studies, a downregulation of the cGMP-dependent protein kinase-1 (cGKI) alpha isoform has been linked to erectile dysfunction and diabetes mellitus. So far, the expression of cGKI alpha and beta isoforms has not been evaluated in human penile erectile tissue. AIM: To evaluate the expression of cGKI alpha and beta isoforms in relation to smooth muscle alpha-actin, cGMP, and endothelial NO synthase (eNOS) in human cavernous arteries (HCAs) and human corpus cavernosum (HCC). METHODS: Cryostat sections of HCA and HCC were incubated with primary antibodies directed against alpha-actin, cGMP, eNOS, cGKI, cGKI alpha, and cGKI beta. Visualization of double-labeled immunofluorescent stainings was achieved by laser microscopy. Western blot analysis was performed in order to confirm the expression of cGKI isoforms. MAIN OUTCOME MEASURES: Expression of cGKI alpha and beta isoforms in relation to smooth muscle alpha-actin, cGMP, and eNOS in human penile erectile tissue. RESULTS: Immunoreactivities specific for cGKI, cGKI alpha, and cGKI beta were observed within the smooth musculature and the endothelium of cavernous arteries and sinusoids. Double stainings revealed the colocalization of alpha-actin, cGMP, eNOS, and cGKI isoforms. The expression of cGKI isoforms was confirmed by Western blot analysis. CONCLUSIONS: Our results demonstrate, for the first time, the expression of both cGKI alpha and beta isoforms in the smooth musculature of HCA and HCC. Corresponding to recent findings from animal studies, the presence of cGKI alpha and beta provides further evidence for a significant role of these enzymes in the control of smooth muscle function in human penile erectile tissue.

Non-genomic effects of androgens on isolated human vascular and nonvascular penile erectile tissue.

OBJECTIVES: To evaluate non-genomic effects of testosterone and dihydrotestosterone (DHT) on isolated human cavernosal arteries (HCA) and corpus cavernosum (HCC) using organ-bath studies and radio-immunoassays (RIA), as non-genomic effects of androgens are reported for vascular smooth musculature and there is evidence that the relaxant response involves a modulation of cyclic nucleotide tissue levels. MATERIALS AND METHODS: The relaxation induced by the cumulative addition of testosterone and DHT (0.01-10 microm) was studied using circular segments of HCA and strip preparations of HCC. To evaluate the effects of testosterone and DHT on tissue levels of cAMP and cGMP, specimens were exposed to increasing concentrations of the hormones. Forskolin and sodium nitroprusside (SNP) served as reference compounds. RESULTS: Testosterone and DHT dose-dependently reversed the noradrenaline-induced tension of vascular segments and HCC strips. At the maximum concentration, testosterone and DHT reduced the mean (sd) tension to 79.8 (4.43)% and 83.9 (10.94)%, respectively. SNP and forskolin significantly stimulated the production of cGMP and cAMP. No effects of testosterone and DHT on cGMP and cAMP levels were detected. CONCLUSION: Rapid androgen-induced relaxation of HCA and HCC occurs via non-genomic mechanisms. In penile erectile tissue, non-genomic relaxant effects of testosterone and DHT are not mediated via modulation of cyclic nucleotide tissue levels. Additional studies are required to establish if non-genomic relaxant effects are important in ensuring a basal level of perfusion to maintain overall penile function.

Urodynamic evaluation of sacral neuromodulation for urge urinary incontinence.

OBJECTIVE: To evaluate the urodynamic data before and 6 months after implantation of sacral neuromodulation (SNM, an established treatment for voiding dysfunction, including refractory urge urinary incontinence, UI) and to assess the correlation between the urodynamic data and clinical efficacy in patients with UI. PATIENTS AND METHODS: In all, 111 patients with a >50% reduction in UI symptoms during a percutaneous nerve evaluation test qualified for surgical implantation of SNM. Patients were categorized in two subgroups, i.e. those with UI with or without confirmed detrusor overactivity (DO) at baseline. At the 6-month follow-up all patients had a second urodynamic investigation, with the stimulator switched on. RESULTS: At baseline, there was urodynamically confirmed DO in 67 patients, while 44 showed no DO. A review of filling cystometry variables showed a statistically significant improvement in bladder volumes at first sensation of filling (FSF) and at maximum fill volume (MFV) before voiding for both UI subgroups, compared with baseline. In 51% of the patients with UI and DO at baseline, the DO resolved during the follow-up. However, those patients were no more clinically successful than those who still had DO (P = 0.73). At the 6-month follow-up, 55 of 84 implanted patients showed clinical benefit, having a >or=50% improvement in primary voiding diary variables. Patients with UI but no DO had a higher rate of clinical success (73%) than patients with UI and DO (61%), but the difference was not statistically significant. CONCLUSION: These urodynamic results show a statistically significant improvement in FSF and MFV in patients with UI with or with no DO after SNM. Although there was a urodynamic and clinical improvement in both groups, patients with UI but no DO are at least as successful as patients with UI and DO. Therefore in patients with UI, DO should not be a prerequisite selection criterion for using SNM.

Adiposity and age are statistically related to enhanced RASSF1A tumor suppressor gene promoter methylation in normal autopsy kidney tissue.

Age, adiposity, and smoking are risk factors for the development of renal cell carcinoma. Hypermethylation of the RAS association domain family 1A gene (RASSF1A) promoter belongs to the most frequently detected epigenetic alterations in human cancers including renal cell carcinoma. RASSF1A is functionally involved in cell cycle control in normal cells and depletion promotes a number of cellular changes increasing the risk for neoplastic growth. We investigated the hypothesis that age, modulated by the factors adiposity and anthracosis as a surrogate for smoking, is a predictor of RASSF1A promoter methylation in normal kidney tissue. Using a cross-sectional study design, we quantitatively analyzed RASSF1A methylation in 78 normal autopsy kidney tissues by quantitative combined bisulfite and restriction analysis and bisulfite sequencing, and statistically evaluated the degree of relative methylation for a relationship with the predictor age and study factors adiposity and state of anthracosis. Statistical analysis showed that age (regression analysis; P < 0.001), adiposity (univariate analysis; P = 0.016), and state of anthracosis (t test; P = 0.005) are each significantly associated with an increase of RASSF1A promoter methylation in normal kidney tissue. However, only age (P = 0.008) and adiposity (P = 0.008) were identified as independent predictors of RASSF1A promoter methylation using covariance analysis. This study provides statistical evidence that the common cancer risk factors age and adiposity enhance RASSF1A promoter methylation in nonmalignant kidney tissue.

Modulation of beta-catenin-mediated TCF-signalling in prostate cancer cell lines by wild-type and mutant p53.

BACKGROUND: Deregulation of the canonical Wnt/beta-catenin-pathway is known to play an important role in the progression of various tumour cell types including prostate cancer (PCa). Recently, the tumour-suppressor p53 was shown to down-regulate beta-catenin-signalling in colon cancer. As p53 is frequently mutated in late stage PCa we investigated the effect of wild-type p53 (p53wt) as well as p53-mutants on beta-catenin-signalling in PCa-cell lines. METHODS: The effects of p53wt and p53-mutants on Wnt/beta-catenin-signalling were studied using reporter gene assays. Expression of beta-catenin levels was monitored by Western blotting. RESULTS: Overexpression of p53wt as well as p53(249Ser) (a structural mutant) and p53(273His) (a DNA-contact-mutant) almost completely inhibited beta-catenin-mediated transcriptional activity of the T-cell factor (TCF) whereas p53(175His), a structural mutant, and a p53-mutant with a C-terminal deletion in the tetramerization domain (Deltap53) were unable to do so. Co-transfection experiments with p53wt and a dominant negative p53-mutant reversed the down-regulation of TCF-signalling, while Deltap53 was unable to interfere with p53wt-function. Down-regulation of TCF-signalling by p53wt and p53(273His) was accompanied by a reduction in beta-catenin protein level. CONCLUSIONS: p53wt, p53(273His)- and p53(249Ser)-mutants are able to down-regulate beta-catenin-signalling in PCa-cells probably via degradation of beta-catenin. The degradation of beta-catenin in PCa by p53 is not linked to transcriptional activity of p53. So far the mechanism how p53 interferes with beta-catenin-signalling is unknown. For the first time we provide experimental evidence that the C-terminus of p53 plays an important role in the down-regulation of beta-catenin-mediated TCF-signalling in PCa-cell lines possibly via p53 transrepressional function.

Pyelovenous backflow or veno-caliceal valve fistula?

A 70-year-old male with urinary bladder carcinoma was admitted for follow-up. Retrograde pyelography demonstrated transfer of contrast medium into the left renal vein in two independent sessions. The absence of hematuria and a negative CT scan ruled out a classical veno-caliceal fistula. The presence of a veno-caliceal valve fistula into the left renal vein was hypothesized.

Expression of messenger ribonucleic acid encoding for phosphodiesterase isoenzymes in human female genital tissues.

OBJECTIVES: The use of inhibitors of phosphodiesterase 5 (PDE5) has been suggested to treat symptoms of female sexual dysfunction (FSD). Nonetheless, there has been a relatively low success rate of PDE5 inhibitors in FSD in comparison with male erectile dysfunction. The elevated expression of PDE5 in the human penile erectile tissue is considered the reason for the high clinical efficacy of PDE5 inhibitors in the pharmacotherapy of male erectile dysfunction. AIM: To evaluate by means of molecular biology the expression of messenger ribonucleic acid expression (mRNA) encoding for cyclic AMP and cyclic GMP PDE isoenzymes in female genital tissues. MAIN OUTCOME MEASURES: The amount of mRNA transcripts specifically encoding for cyclic AMP- and/or cyclic GMP-degrading PDE isoenzymes was determined. METHODS: Human clitoral, labial, and vaginal tissue was obtained from four female cadavers (age at death: 18-42 years). The expression of mRNA specifically encoding for PDE1A, 1B, 1C, 2A, 4A, 5A, 10A, and 11A was elucidated by means of real-time polymerase chain reaction (PCR) analysis (TaqMan). Human penile erectile tissue (corpus cavernosum [HCC]) was used as a reference tissue. RESULTS: mRNA encoding for all PDE isoforms mentioned above is expressed in the female genital tissues. Different magnitudes of mRNA expression were observed: a predominant expression of mRNA encoding for PDE1A but only insignificant amounts of PDE1B, 1C, 4A, 10, and 11A mRNA were registered. With PDE1A being the only exception, the mRNA expression was always higher in the HCC than in the female genital tissues. Especially, the expression of mRNA encoding for PDE5 was several-fold higher in the HCC. CONCLUSION: On the mRNA level, various PDE isoforms are expressed in the clitoris, labia, and vagina. It remains to be established as to whether the low expression of PDE5 in female genital tissue might be a negative predictor for the success of PDE5 inhibitors in the treatment of FSD.

The future of the oral pharmacotherapy of male erectile dysfunction: things to come.

The convincing clinical data on the use of the orally active phosphodiesterase inhibitors sildenafil, vardenafil and tadalafil for the treatment of male erectile dysfunction have boosted research activities on the physiology of the male erectile mechanism. This included both peripheral intracellular signal transduction in the corpus cavernosum as well as central brain and spinal cord pathways controlling penile erection. This work provided the basis for the development and introduction of several new therapeutic modalities into the management of erectile dysfunction, some of which are already offered to the patients. As the concept of 'taking a pill' as a cure for an illness or the relief of symptoms of a disease has become widely accepted by the consumers, the pharmacologic treatment of erectile dysfunction has primarily focussed on selective, orally available drugs acting by influencing intracellular or central regulatory mechanisms, combining a high response rate and the advantage of an on-demand intake. These agents are regarded as more efficacious, and have a faster onset of drug action in the target tissue and an improved effect to side-effect ratio. The purpose of this review is to describe the major novel and evolving pharmacologic advances in the field of oral pharmacotherapy for the treatment of male erectile dysfunction.

Functional responses of isolated human seminal vesicle tissue to selective phosphodiesterase inhibitors.

OBJECTIVES: To further elucidate the significance of the cyclic nucleotide-mediated signal transduction, we examined the in vitro functional responses of isolated seminal vesicle (SV) smooth muscle tissue to selective phosphodiesterase (PDE) inhibitors. METHODS: Using the organ bath technique, the effects of increasing concentrations (1 nM to 10 microM) of the PDE inhibitors vinpocetine (PDE1 inhibitor), rolipram (PDE4 inhibitor), and sildenafil and vardenafil (PDE5 inhibitors) on the tension induced by 10 microM of norepinephrine on SV tissue strips were investigated. To examine the drug effects on the tissue levels of cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP), the SV strips were exposed to different concentrations of the compounds (0.1, 1, and 10 microM). After freezing, homogenization, and extraction of cyclic nucleotides, cAMP and cGMP were measured using radioimmunoassays. In the experiments, sodium nitroprusside and forskolin were used as reference compounds. RESULTS: The norepinephrine-induced tension was reversed by the drugs in a dose-dependent manner. The rank order of efficacy was rolipram greater than sildenafil greater than vardenafil greater than or equal to vinpocetine greater than sodium nitroprusside greater than forskolin. The reversion of the norepinephrine-induced tension at maximum drug concentration ranged from 79% (rolipram) to 32% (forskolin). Only rolipram and sildenafil reached a median effective concentration. The effects of the PDE inhibitors were paralleled by a 1.7-fold to 173-fold increase in tissue cGMP or cAMP. CONCLUSIONS: Our results have demonstrated that PDE inhibitors can reverse the adrenergic tension of human SV tissue and increase levels of cyclic nucleotides. This outlines the potential significance of cAMP and cGMP in the control of SV smooth muscle function. These findings might be of importance with regard to the pharmacologic treatment of premature ejaculation.

RASSF1A promoter methylation and expression analysis in normal and neoplastic kidney indicates a role in early tumorigenesis.

BACKGROUND: Epigenetic silencing of the RAS association domain family 1A (RASSF1A) tumor suppressor gene promoter has been demonstrated in renal cell carcinoma (RCC) as a result of promoter hypermethylation. Contradictory results have been reported for RASSF1A methylation in normal kidney, thus it is not clear whether a significant difference between RASSF1A methylation in normal and tumor cells of the kidney exists. Moreover, RASSF1A expression has not been characterized in tumors or normal tissue as yet. RESULTS: Using combined bisulfite restriction analysis (COBRA) we compared RASSF1A methylation in 90 paired tissue samples obtained from primary kidney tumors and corresponding normal tissue. Bisulfite sequence analysis was carried out using both pooled amplicons from the tumor and normal tissue groups and subclones obtained from a single tissue pair. Expression of RASSF1A was analyzed by the use of tissue arrays and immunohistochemistry. We found significantly increased methylation in tumor samples (mean methylation, 20%) compared to corresponding normal tissues (mean methylation, 11%; P < 0.001). Densely methylated sequences were found both in pooled and individual sequences of normal tissue. Immunohistochemical analysis revealed a significant reduced expression of RASSF1A in most of the tumor samples. Heterogeneous expression patterns of RASSF1A were detected in a subgroup of histologically normal tubular epithelia. CONCLUSION: Our methylation and expression data support the hypothesis that RASSF1A is involved in early tumorigenesis of renal cell carcinoma.

Immunohistochemical description of cyclic nucleotide phosphodiesterase (PDE) isoenzymes in the human labia minora.

INTRODUCTION: Up until now, only minimal research has been carried out on those female genital organs known to contribute to the normal cycle of sexual arousal and orgasm. Some findings indicated that there might be a significance of cyclic nucleotide-mediated pathways in the control of the normal function of female genital tissues. AIM: To elucidate, by means of immunohistochemistry, the distribution of the phosphodiesterase (PDE) isoenzymes 1, 3, 4, 5, 10, and 11 in the human labia minora. MAIN OUTCOME MEASURES: The amount of immunohistochemical staining specific for cyclic adenosine monophosphate (cAMP)- and/or cyclic guanosine monophosphate (cGMP)-degrading PDE isoenzymes was detected. METHODS: Human labial tissue was obtained from four female cadavers (age at death: 18-42 years). Vibratome sections prepared from formaldehyde-fixated tissue specimens were incubated with primary antibodies directed against the respective PDE isoenzymes. Sections were then incubated with fluorochrome (fluorescein isothiocyanate, Texas Red)-labeled secondary antibodies. Visualization was commenced by means of a laser fluorescence microscope. RESULTS: Immunostaining indicating the expression of PDE4 and PDE5 was abundantly observed in the smooth musculature of vessels interspersing the tissue. Immunoreactions specific for PDE3 were recognized in epithelial and subepithelial layers, sebaceous glands, and interstitial or neuroendocrine-like single cells located in the epithelium. Signals related to PDE10 and PDE11 were limited to the epithelium or glandular-like structures, respectively. CONCLUSIONS: Our results, for the first time, demonstrate the presence of cAMP- and cGMP-PDE isoenzymes in the human labia minora and give a hint to a significance of PDE4 and PDE5 in the control of labial vascular tissue function.

Urinary tract injuries in patients with multiple trauma.

This article mainly reviews urinary tract injuries in patients with multiple trauma. Approximately 10% of all traumatic injuries resulting from an external force will involve the genitourinary system. The article discusses mechanisms of injury, diagnosis, and therapeutical approaches for renal, ureteral, bladder, and urethral trauma. Due to the complexity of such injuries--despite several attempts to provide a standard strategy in trauma patients with urinary tract involvement--an individual and patient-specific-therapeutic approach is mandatory in most cases. However, the availability of classified guidelines may help the surgeon to reach the most accurate decision. Because of the similarity of American and European guidelines on urological trauma, this article adapts injury severity scales and classification from the American Association for the Surgery of Trauma.

Immunohistochemical distribution of cyclic GMP-dependent protein kinase-1 in human prostate tissue.

OBJECTIVES: Phosphodiesterase 5 (PDE5) inhibitors improve smooth muscle relaxation and therefore are considered for pharmacotherapy of benign prostatic hyperplasia (BPH) and lower urinary tract symptoms (LUTS). Cyclic guanosine monophosphate (cGMP)-dependent protein kinase-1 (cGKI) has been identified as one of the downstream targets for cGMP. The aim of the present study was to evaluate, by means of immunohistochemistry and Western blot analysis, the expression and localization of cGKI isoforms in relation to smooth muscle alpha-actin and cGMP in the human prostate. METHODS: Cryostat sections of tissue segments excised from the transition zone of human prostates from 11 patients (aged 54-68 yr) were incubated with primary antibodies directed against smooth muscle alpha-actin, cGMP, cGKI, cGKIalpha, and cGKIbeta. Visualization of double-labelled immunofluorescent staining was achieved by laser microscopy. Western blot analysis was performed to confirm the expression of cGKI isoforms. RESULTS: Immunoreactivities specific for cGKI, cGKIalpha, and cGKIbeta were observed in the smooth musculature of the transition zone. Double-staining revealed the colocalization of smooth muscle alpha-actin, cGMP, and cGKI isoforms in smooth muscle cells of the fibromuscular stroma. The expression of cGKI isoforms was confirmed by Western blot analysis. CONCLUSIONS: Our results confirm the presence of cGKI isoforms alpha and beta in the transition zone of human prostate tissue. In addition, the colocalization of alpha-actin, cGMP, and cGKI isoforms provides further evidence for a significant role of the nitric oxide/cGMP pathway in the regulation of smooth muscle contractility in human prostate tissue and therefore could provide additional targets for pharmacotherapy of BPH and LUTS.