Cannabidiol Alleviates Chronic Prostatitis and Chronic Pelvic Pain Syndrome via CB2 Receptor Activation and TRPV1 Desensitization.

PURPOSE: This study elucidates the mechanism of the physiological effect of cannabidiol (CBD) by assessing its impact on lipopolysaccharide (LPS)-induced inflammation in RWPE-1 cells and prostatitis-induced by 17ß-estradiol and dihydrotestosterone in a rat model, focusing on its therapeutic potential for chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS). MATERIALS AND METHODS: RWPE-1 cells were stratified in vitro into three groups: (1) controls, (2) cells with LPS-induced inflammation, and (3) cells with LPS-induced inflammation and treated with CBD. Enzyme-linked immunosorbent assays and western blots were performed on cellular components and supernatants after administration of CBD. Five groups of six Sprague-Dawley male rats were assigned: (1) control, (2) CP/CPPS, (3) CP/CPPS and treated with 50 mg/kg CBD, (4) CP/CPPS and treated with 100 mg/kg CBD, and (5) CP/CPPS and treated with 150 mg/kg CBD. Prostatitis was induced through administration of 17ß-estradiol and dihydrotestosterone. After four weeks of CBD treatment, a pain index was evaluated, and prostate tissue was collected for subsequent histologic examination and western blot analysis. RESULTS: CBD demonstrated efficacy in vivo for CP/CPPS and in vitro for inflammation. It inhibited the toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) pathway by activating the CB2 receptor, reducing expression of interleukin-6, tumor necrosis factor-alpha, and cyclooxygenase-2 (COX2) (p<0.01). CBD exhibited analgesic effects by activating and desensitizing the TRPV1 receptor. CONCLUSIONS: CBD inhibits the TLR4/NF-κB pathway by activating the CB2 receptor, desensitizes the TRPV1 receptor, and decreases the release of COX2. This results in relief of inflammation and pain in patients with CP/CPPS, indicating CBD as a potential treatment for CP/CPPS.

Non-Invasive Radiofrequency Hyperthermia Attenuates HMGB1/TLR4/NF-κB Inflammatory Axis in a Chronic Prostatitis/Chronic Pelvic Pain Syndrome Rat Model.

PURPOSE: The primary goal of this study is to evaluate the effect of the non-invasive radiofrequency hyperthermia (RFHT) device on chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) rat model and investigate the underlying mechanism. MATERIALS AND METHODS: In this study, Sprague-Dawley rats were randomly distributed into three groups: (1) normal control group, (2) CP/CPPS group, and (3) RFHT group. CP/CPPS rat models were induced by 17ß-estradiol and dihydrotestosterone for 4 weeks and RFHT was administered for 5 weeks after model establishment. During RFHT administration, core body temperatures were continuously monitored with a rectal probe. After administering RFHT, we assessed pain index for all groups and collected prostate tissues for Western blot analysis, immunofluorescence, and immunohistochemistry. We also collected adjacent organs to the prostate including urinary bladder, testes, and rectum for safety assessment via H&E staining along with a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling assay. RESULTS: After administering RFHT, pain in rats was significantly alleviated compared to the CP/CPPS group. RFHT reduced high-mobility group box 1 (HMGB1) expression and improved inflammation by downregulating subsequent proinflammatory cytokines through inhibition of the toll-like receptor 4 (TLR4)-nuclear factor kappa B (NF-κB) pathway. In prostate-adjacent organs, no significant histological alteration or inflammatory infiltration was detected. The area of cell death also did not increase significantly after RFHT. CONCLUSIONS: In conclusion, RFHT demonstrated anti-inflammatory effects by inhibiting the HMGB1-TLR4-NF-κB pathway in CP/CPPS rat models. This suggests that RFHT could serve as a safe and promising therapeutic strategy for CP/CPPS.

Urinary Microbiome Dysbiosis and Immune Dysregulations as Potential Diagnostic Indicators of Bladder Cancer.

There are a total of 82,290 new cases and 16,710 deaths estimated for bladder cancer in the United States in 2023. Currently, urine cytology tests are widely used for bladder cancer diagnosis, though they suffer from variable sensitivity, ranging from 45 to 97%. More recently, the microbiome has become increasingly recognized for its role in human diseases, including cancers. This study attempts to characterize urinary microbiome bladder cancer-specific dysbiosis to explore its diagnostic potential. RNA-sequencing data of urine samples from patients with bladder cancer (n = 18) and matched controls (n = 12) were mapped to bacterial sequences to yield species-level abundance approximations. Urine samples were analyzed at both the population and species level to reveal dysbiosis associated with bladder cancer. A panel of 35 differentially abundant species was discovered, which may be useful as urinary biomarkers for this disease. We further assessed whether these species were of similar significance in a validation dataset (n = 81), revealing that the genera Escherichia, Acinetobacter, and Enterobacter were consistently differentially abundant. We discovered distinct patterns of microbial-associated immune modulation in these samples. Several immune pathways were found to be significantly enriched with respect to the abundance of these species, including antigen processing and presentation, cytosolic DNA sensing, and leukocyte transendothelial migration. Differential cytokine activity was similarly observed, suggesting the urinary microbiome's correlation to immune modulation. The adherens junction and WNT signaling pathways, both implicated in the development and progression of bladder cancer, were also enriched with these species. Our findings indicate that the urinary microbiome may reflect both microbial and immune dysregulations of the tumor microenvironment in bladder cancer. Given the potential biomarker species identified, the urinary microbiome may provide a non-invasive, more sensitive, and more specific diagnostic tool, allowing for the earlier diagnosis of patients with bladder cancer.

Correlation Analysis between Hypogonadal Symptoms and Changes in Body Composition and Physical Fitness after Testosterone Treatment in Men with Testosterone Deficiency.

PURPOSE: This study analyzed changes in body composition and physical fitness in men with testosterone deficiency (TD) after testosterone treatment (TT) and examined the correlations of body composition and physical fitness with serum testosterone levels and hypogonadal symptoms. MATERIALS AND METHODS: Seventy patients with TD were divided into control (group I, n=23) and experimental (group II, n=47) groups. Patients in the experimental group were administered intramuscular testosterone enanthate (250 mg) for six months. The aging males symptom scale (AMS) score, international prostate symptom score (IPSS), body mass index, waist circumference, and serum laboratory values were measured at baseline and at the end of the study. Bioelectrical impedance analysis was used to assess the patients' body composition. Seven types of basic exercise tests were used to evaluate the patients' physical fitness. RESULTS: After six months, there were no significant differences in group I, while group II had significantly improved IPSS and AMS scores; increased hemoglobin, hematocrit, prostate-specific antigen, and testosterone levels and skeletal muscle mass; and waist circumference, and body fat mass. All elements of the physical fitness test were significantly improved in group II, with the exceptions of flexibility and endurance. Decreased waist circumference was correlated with changes in testosterone levels in group II, and the IPSS, cardiorespiratory fitness, and agility were correlated with improved hypogonadal symptoms. CONCLUSIONS: TT improved the hypogonadal and lower urinary tract symptoms in patients with TD and improved body composition, physical fitness, and metabolic syndrome parameters. Increased testosterone and improved hypogonadal symptoms were correlated with a decrease in waist circumference and an improvement in cardiorespiratory fitness and agility. As such, when implementing TT, we should consider whether these areas may be improved, as this can help to predict the effect.

Effect of Li-ESWT on Testicular Tissue and Function in Androgen-Deficient Rat Model.

Low-intensity extracorporeal shockwave therapy (Li-ESWT), as a microenergy therapy, has the effects of inhibiting oxidative stress, antiapoptosis, and tissue repair, which is increasingly applied to a variety of diseases. Our research aims to explore the protective effects of Li-ESWT in the aging rat model and its possible molecular mechanism through in vivo and in vitro experiments. In vitro, TM3 Leydig cells incubated with H2O2 were treated with Li-ESWT at 4 energy levels (0.01, 0.05, 0.1, and 0.2 mJ/mm2). In vivo, we employed an androgen-deficient rat model to simulate male aging and treated it with Li-ESWT at three different energy levels (0.01, 0.05, and 0.2 mJ/mm2). Li-ESWT increased the expression of vascular endothelial growth factor (VEGF) in TM3 cells, improved antioxidant capacity, and reduced apoptosis, with the effect being most significant at 0.05 mJ/mm2 energy level. In androgen-deficient rat model, LI-ESWT can improve sperm count, motility, and serum testosterone level, enhancing tissue antioxidant capacity and antiapoptotic ability, and the effect is most significant at 0.05 mJ/mm2 energy level. Therefore, Li-ESWT at an appropriate energy level can improve sperm count, motility, and serum testosterone levels in androgen-deficient rat models, reduce oxidative stress in the testis, and increase antioxidant capacity and antiapoptotic abilities. The mechanism of this condition might be related to the increased VEGF expression in Leydig cells by Li-ESWT.

Transpelvic Magnetic Stimulation Enhances Penile Microvascular Perfusion in a Rat Model: A Novel Interventional Strategy to Prevent Penile Fibrosis after Cavernosal Nerve Injury.

PURPOSE: Penile microvascular dysfunction is a known contributor to erectile dysfunction (ED) and penile fibrosis has been shown to impair microvascular perfusion (MVP). Our objectives were to: (i) determine beneficial effects of TPMS to modulate penile MVP, (ii) determine its mechanism, (iii) evaluate impact of cavernosal nerve injury (CNI) on penile MVP, and (iv) determine time-course of cavernosal tissue elastin changes after CNI in rats. MATERIALS AND METHODS: Adult male rats (n=5) were anesthetized and subjected to TPMS (13%, 15%, and 17%) and MVP changes were recorded using laser speckle contrast imaging (LSCI). Another group of male rats were subjected to either bilateral cavernosal nerve injury (CNI; n=7) or sham surgery (n=7). After recovery, animals were monitored for MVP using LSCI before and after TPMS. Rat penile tissues were harvested and analyzed for fibrosis using a marker for elastin. RESULTS: Rat TPMS resulted in a stimulus dependent increase in MVP; maximal perfusion was observed at 17%. L-N(G)-Nitroarginine methyl ester (L-NAME) resulted in a marked decrease in TPMS induced MVP increase (393.33 AU vs. 210.67 AU). CNI resulted in 40% to 50% decrease in MVP. CNI produced a remarkable increase in elastin deposits that are noticeable throughout the cavernosal tissues post injury. CONCLUSIONS: TPMS is a novel and non-invasive intervention to improve penile MVP after CNI. Potential application includes treatment of ED and sexual function preservation following cancer treatment, possibly through improved penile hemodynamics that might help prevent penile hypoxia and fibrosis.

The Bladder Microbiome Is Associated with Epithelial-Mesenchymal Transition in Muscle Invasive Urothelial Bladder Carcinoma.

The intra-tumor microbiome has recently been linked to epithelial-mesenchymal transition (EMT) in a number of cancers. However, the relationship between EMT and microbes in bladder cancer has not been explored. In this study, we profiled the abundance of individual microbe species in the tumor samples of over 400 muscle invasive bladder carcinoma (MIBC) patients. We then correlated microbe abundance to the expression of EMT-associated genes and genes in the extracellular matrix (ECM), which are key players in EMT. We discovered that a variety of microbes, including E. coli, butyrate-producing bacterium SM4/1, and a species of Oscillatoria, were associated with expression of classical EMT-associated genes, including E-cadherin, vimentin, SNAI2, SNAI3, and TWIST1. We also found significant correlations between microbial abundance and the expression of genes in the ECM, specifically collagens and elastin. Lastly, we found that a large number of microbes exhibiting significant correlations to EMT are also associated with clinical prognosis and outcomes. We further determined that the microbes we profiled were likely not environmental contaminants. In conclusion, we discovered that the intra-tumoral microbiome could potentially play a significant role in the regulation of EMT in MIBC.

Identification of Lung and Blood Microbiota Implicated in COVID-19 Prognosis.

The implications of the microbiome on Coronavirus disease 2019 (COVID-19) prognosis has not been thoroughly studied. In this study we aimed to characterize the lung and blood microbiome and their implication on COVID-19 prognosis through analysis of peripheral blood mononuclear cell (PBMC) samples, lung biopsy samples, and bronchoalveolar lavage fluid (BALF) samples. In all three tissue types, we found panels of microbes differentially abundant between COVID-19 and normal samples correlated to immune dysregulation and upregulation of inflammatory pathways, including key cytokine pathways such as interleukin (IL)-2, 3, 5-10 and 23 signaling pathways and downregulation of anti-inflammatory pathways including IL-4 signaling. In the PBMC samples, six microbes were correlated with worse COVID-19 severity, and one microbe was correlated with improved COVID-19 severity. Collectively, our findings contribute to the understanding of the human microbiome and suggest interplay between our identified microbes and key inflammatory pathways which may be leveraged in the development of immune therapies for treating COVID-19 patients.

Novel Emerging Therapies for Erectile Dysfunction.

Currently, several treatments exist for the improvement of erectile dysfunction (ED). These include medical therapies such as phosphodiesterase type 5 inhibitors (PDE5-Is), invasive methods such as intracavernosal injection therapy of vaso-active substances, vacuum erection devices, and penile prosthesis implants. However, the percentage of patients that are unresponsive to available treatments and who drop out from treatments remains high. Current evidence reveals that the pathogenesis of ED is related to multiple factors including underlying comorbidities, previous surgery, and psychological factors. Diverse approaches using novel molecular pathways or new technologies have been tested as potential therapeutic options for difficultto-treat ED populations. Melanocortin receptor agonist, a centrally acting agent, showed promising results by initiating erection without sexual stimulation in non-responders to PDE5-Is. Recent clinical and pre-clinical studies using human tissues suggested that new peripherally acting agents including the Max-K channel activator, guanylate cyclase activator, and nitric oxide donor could be potential therapies either as a monotherapy or in combination with PDE5-Is in ED patients. According to several clinical trials, regeneration therapy using stem cells showed favorable data in men with diabetic or post-prostatectomy ED. Low-intensity shock wave therapy also demonstrated promising results in patients with vasculogenic ED. There are growing evidences which suggest the efficacy of these emerging therapies, though most of the therapies still need to be validated by well-designed clinical trials. It is expected that, should their long-term safety and efficacy be proven, the emerging treatments can meet the needs of patients hitherto unresponsive to or unsatisfied by current therapies for ED.

Influence of Intratumor Microbiome on Clinical Outcome and Immune Processes in Prostate Cancer.

Although 1 in 9 American men will receive a diagnosis of prostate cancer (PC), most men with this diagnosis will not die from it, as most PCs are indolent. However, there is a subset of patients in which the once-indolent PC becomes metastatic and eventually, fatal. In this study, we analyzed microbial compositions of intratumor bacteria in PC to determine the influence of the microbiome on metastatic growth. Using large-scale RNA-sequencing data and corresponding clinical data, we correlated the abundance of microbes to immune pathways and PC risk factors, identifying specific microbes that either significantly deter or contribute to cancer aggressiveness. Interestingly, most of the microbes we found appeared to play anti-tumor roles in PC. Since these anti-tumor microbes were overrepresented in tumor samples, we believe that microbes thrive in the tumor microenvironment, outcompete cancer cells, and directly mitigate tumor growth by recruiting immune cells. These include Listeria monocytogenes, Methylobacterium radiotolerans JCM 2831, Xanthomonas albilineans GPE PC73, and Bradyrhizobium japonicum, which are negatively correlated with Gleason score, Tumor-Node-Metastasis (TNM) stage, prostate-specific antigen (PSA) level, and Androgen Receptor (AR) expression, respectively. We also identified microbes that contribute to tumor growth and are positively correlated with genomic alterations, dysregulated immune-associated (IA) genes, and prostate cancer stem cells (PCSC) genes.

Smoking-Mediated Upregulation of the Androgen Pathway Leads to Increased SARS-CoV-2 Susceptibility.

The COVID-19 pandemic is marked by a wide range of clinical disease courses, ranging from asymptomatic to deadly. There have been many studies seeking to explore the correlations between COVID-19 clinical outcomes and various clinical variables, including age, sex, race, underlying medical problems, and social habits. In particular, the relationship between smoking and COVID-19 outcome is controversial, with multiple conflicting reports in the current literature. In this study, we aim to analyze how smoking may affect the SARS-CoV-2 infection rate. We analyzed sequencing data from lung and oral epithelial samples obtained from The Cancer Genome Atlas (TCGA). We found that the receptor and transmembrane protease necessary for SARS-CoV-2 entry into host cells, ACE2 and TMPRSS2, respectively, were upregulated in smoking samples from both lung and oral epithelial tissue. We then explored the mechanistic hypothesis that smoking may upregulate ACE2 expression through the upregulation of the androgen pathway. ACE2 and TMPRSS2 upregulation were both correlated to androgen pathway enrichment and the specific upregulation of central pathway regulatory genes. These data provide a potential model for the increased susceptibility of smoking patients to COVID-19 and encourage further exploration into the androgen and tobacco upregulation of ACE2 to understand the potential clinical ramifications.

A Novel Endoluminal Ultrasound Imaging Technique to Determine Urethral Luminal Cross-Sectional Area.

INTRODUCTION AND OBJECTIVES: Retrograde urethrogram (RUG) and voiding cystourethrogram (VCUG) are currently the gold standard imaging technique for diagnosis of urethral stricture and determination of stricture location. However, RUG and VCUG have multiple limitations. These techniques require exposure to ionizing radiation, the quality is operator and patient dependent, there is a moderate degree of invasiveness with urethral catheterization, can have artifacts because of patient positioning that underestimates stricture length. The development of novel imaging modalities without ionizing radiation to accurately evaluate the presence, location, length, and lumen cross-sectional area (CSA) of the urethral stricture would be of great value. The objective of this study was to develop a novel endoluminal ultrasound (ELUS) imaging technique that permits the accurate quantitation of urethral stricture. METHODS: Urethral strictures were created in rabbits (n = 5) by electrocautery and an ELUS technique was developed for subsequent luminal imaging. A 3.2F 40 MHz ultrasound (US) probe was introduced transurethrally and infused with US contrast agent. Images were recorded as the catheter was pulled back at a constant speed to acquire tomographic images. Lumen CSA over the entire urethral length was calculated using a custom methodology and validated in our laboratory. RESULTS: Urethral luminal CSA over the entire length of urethra before and after experimental stricture development was quantified including the length of stenosis. Intra- and interobserver variability (r = 0.99 for both) was excellent. CONCLUSIONS: Feasibility of ELUS as a quantitative technique to determine healthy urethral lumen and stricture CSA was demonstrated. The translational potential for a nonionizing imaging modality to better describe CSA, length, location, and uninvolved urethral CSA of the stricture is a significant improvement over current methodology.

Chronic partial bladder outlet obstruction does not impair erectile function in male rats.

OBJECTIVE: To evaluate, in a well-controlled study, the effect of surgically induced partial bladder outlet obstruction (PBOO) on male erectile function in a rat model. MATERIALS AND METHODS: PBOO was created in 17 adult male Sprague-Dawley rats by partial ligation of the proximal urethra. Sham-operated and PBOO rats were evaluated for urodynamic and erectile function at 4-8 weeks after surgery. Erectile responses to electrical field stimulation (EFS) to the major pelvic ganglion, and to erectogenic agents (1,1-diethyl-2-hydroxy-2-nitroso-hydrazine, DEA-NO, and Y-27632) were evaluated and the area under the curve (AUC, a product of the intracavernous pressure and duration) was used to denote the erectile response. RESULTS: Experimental PBOO in rats significantly increased the mean (sem) bladder weight, to 256 (25) mg in PBOO rats vs 123 (24) mg in sham controls, and the voiding frequency to 1.01 (0.1) voids/min vs 0.72 (0.14) voids/min in sham controls (P < 0.05). There was no significant difference between the erectile response to EFS, with a mean AUC in sham control rats at 1.5, 3.0 and 4.5 V of 2603 (372), 3200 (332) and 3357 (166), respectively, vs 2273 (183), 3794 (211) and 4177 (306) in PBOO rats (P > 0.05); or to the erectogenic agents, the AUC for DEA-NO being 9000 (975) in PBOO rats vs 13 201 (2756) in sham controls, and the AUC for Y-27 632 being 44 915 (2462) and 45 907 (7408), respectively (P > 0.05). There was greater immunoreactivity to RhoA in bladder and penile tissues of PBOO than control rats. CONCLUSION: PBOO does not affect erectile function in rats. Additional mechanisms or pathways might be involved in lower urinary tract symptom-related erectile dysfunction in humans.

Transdifferentiation of rat fetal brain stem cells into penile smooth muscle cells.

OBJECTIVES: To evaluate whether rat fetal brain stem cells can be induced to acquire cell fates outside the nervous system, hypothesising that cell-based replacement therapy with stem cells can aid in the regeneration of penile smooth musculature and might help to attenuate organic erectile dysfunction (ED), as the degeneration of penile smooth muscle cells leading to subsequent impairment of function is important in organic ED. MATERIALS AND METHODS: Fetal brain stem cells (FBSCs) from embryonal 12-day Fisher 344 rats were isolated and characterized. For in vitro studies, undifferentiated FBSCs were cultured for 21 days in either N2 media (control) or N2 media conditioned in rat penile smooth muscle cell culture. These were then subjected to immunocytochemistry for specific markers of neural stem cells (nestin) and penile smooth muscle cells, i.e. alpha-smooth muscle actin (alphaSMA), penis-specific myosin light chain (MLC) desmin, calponin, vimentin, phosphodiesterase-5 (PDE5) and connexin. For in vivo studies, male adult Fisher 344 rats had an intracavernous injection with saline (five rats, control) or FBSCs that were labelled genetically by an expression construct for green fluorescent protein (GFP, nine rats, experimental) and maintained for 6 weeks. The rats were then killed and penile tissue was harvested and subjected to immunocytochemistry for markers of neural stem cells, smooth muscle cells, and sinusoidal endothelium (vascular endothelial growth factor, VEGF). RESULTS: Undifferentiated cells exposed to N2 media continued to maintain the characteristic morphological and protein marker features of FBSCs, while the cells exposed to the conditioned media acquired the morphological features of smooth muscle cells. In addition, the differentiated cells (30-40%) expressed smooth muscle markers. Rats implanted with FBSCs had cells that showed double-labelling for GFP/alphaSMA, GFP/calponin and GFP/VEGF. The control group had no evidence of such double-labelling. CONCLUSIONS: These results suggest the transdifferentiation of FBSCs into penile smooth muscle cells. Such transdifferentiated cells showed long-term survival when injected into the cavernous tissue, thus raising the possibility of a novel therapeutic option for organic ED.

Rho-kinase inhibition suppresses potassium chloride-induced bladder hyperactivity in a rat model.

OBJECTIVES: The molecular mechanisms by which potassium induces urinary bladder hyperactivity are not clear. In the present study, we tested our hypothesis that potassium chloride (KCl)-induced bladder hyperactivity might be mediated through a calcium-sensitizing RhoA-Rho-kinase pathway in an in vivo animal model using urodynamic parameters. METHODS: Two groups of adult male rats (n = 8) were anesthetized, their bladder exteriorized, and a saline-filled Intracath fixed into the bladder dome. This Intracath was connected to a pressure transducer and an infusion pump. Continuous filling cystometrograms were performed by infusing warm saline (0.04 mL/min) to obtain baseline data on each rat. The number of contractions per unit time (intercontractile intervals in seconds), pressure threshold, and peak pressure during micturition were recorded. To create bladder hyperactivity, protamine sulfate (30 mg/mL) followed by KCl (500 mM) was infused intravesically, and a continuous filling cystometrogram was again recorded. Y-27632, a specific RhoA-Rho-kinase inhibitor, was administered either intra-arterially (group 1) or intravesically (group 2) to each rat, and an additional continuous filling cystometrogram was recorded with KCl (500 mM) to observe the effects of Rho-kinase inhibition on bladder contractility. RESULTS: Intravesical KCl infusion after protamine exposure resulted in significantly greater contractions and decreased the intercontractile interval (P <0.05). Y-27632 administration attenuated the effect of KCl on the contractions and intercontractile interval and decreased the mean pressure threshold. CONCLUSIONS: Suppression of KCl-induced bladder contractility by the Rho-kinase inhibitor Y-27632 confirmed the involvement of this novel calcium-sensitizing RhoA-Rho-kinase pathway in mediating these smooth muscle contractions.

Role of sialic acid in urinary cytoprotective activity of Tamm-Horsfall protein.

OBJECTIVES: Tamm-Horsfall protein (THP) from normal urine has been shown to protect against the cytotoxic effects of toxic urinary cations (TFs) in vivo and in vitro. This study investigated the effect of desialylation on the cytoprotective activity of THP. METHODS: From pooled 24-hour urine specimens from healthy individuals, both TFs and THP were obtained. Sprague-Dawley rats received intravesical NaCl or KCl, and the baseline urodynamic percentage of nonvoiding contractions (NVCs) was recorded. Then, rehydrated TF, TF plus THP, or TF plus THP-desialylated (THP-d) were instilled, followed by KCl, and the urodynamic measurements were repeated. In vitro, human HTB4 bladder cells were incubated overnight with the rehydrated TF, TF plus THP, TF plus THP-d, or assay media alone, and the cytotoxicity levels were determined. RESULTS: Acid hydrolysis resulted in an 88% loss of sialic acid. TFs consistently demonstrated a greater than 50% toxicity for human HTB4 cells compared with cells incubated in media (P <0.01). TF cytotoxic activity was blocked by preincubation with THP but not by preincubation with THP-d. Similarly, rat bladder NVCs increased significantly over baseline when KCl was infused after TF infusion (1.68 NVCs/min; P <0.0001). NVCs were significantly reduced by preincubating TFs with THP (0.42 NVCs/min) but not by preincubating with THP-d (1.55 NVCs/min, P <0.0001). CONCLUSIONS: The cytoprotective function of urinary THP in the bladder can be compromised when a decrease is present in the sialic acid residues on THP. Such a decrease in THP cytoprotective activity may play a critical role in the pathophysiology of interstitial cystitis.

MN-001, a novel oral anti-inflammatory agent, suppresses bladder hyperactivity in a rat model.

OBJECTIVE: To evaluate the effects of MN-001, a novel orally active anti-inflammatory agent, in suppressing the bladder hyperactivity resulting from ovalbumin (OA)-induced mast-cell stimulation in a rat model. MATERIALS AND METHODS: Sprague-Dawley rats of both sexes were divided into five groups of 10 each, with group 1 as the control and groups 2-5 undergoing OA sensitization to produce mast-cell degranulation using an established method. At 14 days after sensitization, rats were given an acute intravesical challenge: in group 1, by saline (control), and in groups 2-5, with approximately 2 mL of OA (10 mg/mL in sterile saline). Groups 3-5 received the investigational agent MN-001 orally at 10, 30 or 50 mg/kg, respectively, 1 h before intravesical OA challenge. Urodynamics were then evaluated to quantify the frequency of contractions (voids), intercontractile interval (ICI) and non-voiding contractions (NVCs). RESULTS: Acute intravesical OA challenge in rats in group 2 caused contractions of bladder smooth muscle, leading to a significant (P < 0.05) dose-dependent increase in NVCs and a decrease in ICI. Rats pre-treated with MN-001 at 30 and 50 mg/kg (groups 4 and 5) had significantly fewer NVCs and a greater ICI than rats in group 2 (P < 0.05). CONCLUSION: OA challenge in OA-sensitized rats produces bladder hyperactivity, as reflected in significantly more NVCs and a lower ICI. At doses of 30 and 50 mg/kg orally, MN-001 produces significant protection against the OA-induced bladder hyperactivity. MN-001 might have a role in managing mast cell activity and the associated bladder symptoms in patients with interstitial cystitis.

Toxic factors in human urine that injure urothelium.

AIM: Loss of the lower urinary permeability barrier and passive potassium cycling into tissue are an initiating event in interstitial cystitis. We tested whether a low molecular weight cytotoxic fraction from normal urine causes sensitivity to intravesical potassium in rats and whether the sulfated anionic polysaccharide pentosan polysulfate can neutralize this fraction's cytotoxic activity. METHODS: A low molecular weight (> 100 < 3500) toxic urinary fraction was prepared from normal human urine by dialysis and the lyophilized, salt free product (toxic factor) further investigated. Anaesthetized adult male Sprague-Dawley rats received intravesical sodium or potassium, and urodynamic parameters, including number of voids and non-voiding contractions, were recorded. Then protamine sulfate, rehydrated toxic factor, or toxic factor plus pentosan polysulfate was infused, followed by potassium, and urodynamic measurements repeated. The toxic factor was evaluated in a commercial cytotoxicity protocol using cultured rat urothelial cells. RESULTS: Rat bladder non-voiding contractions increased markedly over baseline when potassium was infused after toxic factor (1.681 +/- 0.1131 non-voiding contractions/min; P = 0.0004) but not after toxic factor premixed with pentosan polysulfate. Toxic factor had a significant (P < 0.001) cytotoxic effect in cultured rat bladder epithelial cells; toxic factor plus pentosan polysulfate was significantly less cytotoxic than toxic factor alone (P < 0.007). CONCLUSIONS: Normal urine contains a cationic cytotoxic factor that increases urothelial permeability by injuring the mucosa, allowing potassium to penetrate the urothelium and depolarize the underlying nerves and muscles. Pentosan polysulfate neutralizes the toxic factor, attenuates urothelial damage, and suppresses potassium-mediated bladder hyperactivity.

Rho-kinase inhibition suppresses bladder hyperactivity in spontaneously hypertensive rats.

AIMS: Spontaneously hypertensive rats (SHR) exhibit overactive bladder (OAB) symptoms and have an up-regulated calcium sensitizing RhoA/Rho-kinase pathway in their vascular smooth muscle tissues. This study examined the role of RhoA/Rho-kinase pathway in bladder hyperactivity by evaluating the effect of a specific Rho-kinase inhibitor (Y-27632) on SHR bladder function. METHODS: Adult male SHR (n = 9) and their normotensive controls (Wistar-Kyoto; WKY) (n = 8) were anesthetized and the carotid artery cannulated for blood pressure monitoring. A catheter was fixed into the bladder dome and connected to a pressure transducer and an infusion pump. After equilibration, systemic and bladder pressure were recorded. Continuous filling cystometrograms (CMGs) were performed and threshold pressure (TP), peak pressure (PP), and number of voids and non-voiding contractions (NVCs) per unit time recorded. Each SHR then received Y-27632, 10 mumol intra-arterially. After 10 min, CMG was repeated and the same measurements recorded. Bladder tissues were evaluated immunohistochemically (IHC) for RhoA protein expression. RESULTS: SHR exhibited significantly higher number of voids and NVCs than normotensive WKY rats (P < 0.05). In SHR, Y-27362 administration significantly decreased the number of voids (29%, from 0.83 +/- 0.3 to 0.63 +/- 0.17 voids/min) and NVCs (61%, from 1.8 +/- 0.54 to 0.64 +/- 0.167 NVC/min). IHC showed significantly higher RhoA protein expression in SHR bladder tissues. CONCLUSIONS: Overexpression of RhoA may play a role in hypertension-related OAB. Inhibition of Rho-kinase activity with Y-27632 produced a significant suppression of bladder overactivity. Identification of Rho-kinase isoforms that are bladder-tissue specific and their selective inhibitors may help to disassociate the unwanted hypotensive effects of this approach.

Curcumin: a potential vaginal contraceptive.

The purpose of this investigation was to evaluate the sperm-immobilizing effects of curcumin, a plant-derived diferuloylmethane compound. Washed human healthy sperm were suspended in Ham's F10 and exposed to varying concentrations of curcumin. Sperm motility was evaluated and changes in sperm mitochondrial transmembrane potential (MTP) was quantified by flow cytometry. Incubation of normal human sperm with curcumin resulted in a dose- and time-dependent loss of sperm motility. At lower concentrations (30 g/mL), curcumin produced a significant (20%) decrease in sperm motility within 30 min without significant effects on sperm viability. An instantaneous (>50%) loss of sperm motility was observed with higher concentrations (300 g/mL) of curcumin and a total loss of sperm motility was achieved within 60 min. A significant reduction in sperm MTP was found with all doses of curcumin tested. Our results indicate that curcumin has a selective sperm-immobilizing effect, in addition to a previously studied anti-HIV property. This compound may have potential clinical applications as a novel intravaginal spermicidal agent for contraception and HIV prevention.