Bladder Outlet Obstruction Rats Research Articles

MP09-16 PHARMACOLOGICAL INHIBITION OF THE NLRP3 INFLAMMASOME PREVENTS BLADDER DECOMPENSATION IN A RAT MODEL OF CHRONIC BLADDER OUTLET OBSTRUCTION

You have accessJournal of UrologyBladder & Urethra: Anatomy, Physiology & Pharmacology I1 Apr 2018MP09-16 PHARMACOLOGICAL INHIBITION OF THE NLRP3 INFLAMMASOME PREVENTS BLADDER DECOMPENSATION IN A RAT MODEL OF CHRONIC BLADDER OUTLET OBSTRUCTION Francis Hughes, Stephanie Sexton, Patrick Leidig, Huixia Jin, and J. Purves Francis HughesFrancis Hughes More articles by this author , Stephanie SextonStephanie Sexton More articles by this author , Patrick LeidigPatrick Leidig More articles by this author , Huixia JinHuixia Jin More articles by this author , and J. PurvesJ. Purves More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2018.02.342AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Bladder outlet obstruction (BOO), most often the result of benign prostatic hyperplasia, may lead to progressive voiding dysfunction. BOO can create an overactive bladder (OAB) phenotype defined by increased frequency of voiding and decreased voiding volume (VV)). We have previously shown in the rat that acute BOO (12 days) activates the NLRP3 inflammasome which mediates inflammation and the OAB phenotype. Initially, the bladder can compensate for the obstruction by generating higher voiding pressures. However, over time the bladder decompensates which results in an increase in post-void residual volume (PVR) and decreased voiding efficiency. In this study we have examined the role of NLRP3 on rat bladder function after chronic BOO (42 days) and investigated the utility of an NLRP3 inhibitor in preventing decompensation. METHODS 4 groups were examined: 1) control, 2) sham-operated, 3) BOO and 4) BOO + gly (glyburide; an NLRP3 inhibitor). BOO was created by inserting a 1 mm transurethral catheter, tying a suture around the urethra, and removing the catheter. Gly was provided via 50 mg, 21 day release pellet (s.c.) that was replaced after 21 days. Suprapubic tubes were implanted on Day 35 and urodynamics performed on Day 42. PVR was assessed immediately after the final micturition by drawing back on the catheter with a syringe. The rat was immediately sacrificed and any additional post void liquid remaining in the bladder removed by needle puncture and added to the total PVR volume. RESULTS Voiding pressures were increased and flow rates decreased in BOO and BOO + gly groups, demonstrating physical obstruction of the urethra. No difference in frequency or VV were detected among groups. However, PVRs were greatly increased in BOO rats while BOO + gly rats were not different than controls (Figure 1). Moreover, also shown in Figure 1, there was a dramatic decrease in voiding efficiency in the chronically obstructed rats which was completely prevented by glyburide treatment. CONCLUSIONS The results suggest a critical role for NLRP3 in the progression to bladder decompensation during chronic BOO and that treatment with an NLRP3 inhibitor preserves voiding efficiency. © 2018FiguresReferencesRelatedDetails Volume 199Issue 4SApril 2018Page: e112-e113 Advertisement Copyright & Permissions© 2018MetricsAuthor Information Francis Hughes More articles by this author Stephanie Sexton More articles by this author Patrick Leidig More articles by this author Huixia Jin More articles by this author J. Purves More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Effects of combined treatment of tadalafil and tamsulosin on bladder dysfunction via the inhibition of afferent nerve activities in a rat model of bladder outlet obstruction.

To investigate the effects of combined treatment of tadalafil (a phosphodiesterase-5 inhibitor) and tamsulosin (an α1-adrenoceptor antagonist) on bladder dysfunction in a rat model of bladder outlet obstruction (BOO). Cystometry was performed in conscious female BOO rats 6weeks after partially ligation of the urethra. Either tadalafil (0.03, 0.1 and 0.3mg/kg) or tamsulosin (0.001, 0.003 and 0.01mg/kg) was cumulatively applied intravenously at 30-min intervals to examine changes in cystometric parameters and blood pressures. Changes in cystometric parameters and blood pressures were also checked when tadalafil (0.3mg/kg), tamsulosin (0.003mg/kg) or both were intravenously applied. In BOO rats, application of either tadalafil (0.3mg/kg) or tamsulosin (0.003, 0.01mg/kg) alone significantly increased threshold pressures and intercontraction intervals whereas there were no significant changes in other cystometric parameters. In addition, because a significant reduction in blood pressures was detected after the administration of tamsulosin (0.01mg/kg), tamsulosin at a lower dose (0.003mg/kg) was used for the combined treatment. The combination therapy of tadalafil and tamsulosin induced a significantly larger rate of increase in intercontraction intervals (1.7 times) compared with monotherapy of either drug (1.3 times each) although the combined therapy did not affect blood pressures. These results suggest that the combination therapy of tadalafil and tamsulosin can induce the additive inhibitory effects on urinary frequency compared with monotherapy, more likely via inhibition of the afferent limb of micturition reflex rather than the efferent function as evidenced by the increases in threshold pressures and intercontraction intervals without affecting bladder contractile function.

Inhibitory effects of silodosin on the bladder mechanosensitive afferent activities and their relation with bladder myogenic contractions in male rats with bladder outlet obstruction.

We investigated the effects of silodosin, an α1A-adrenoceptor (AR) antagonist, on bladder function, especially on non-voiding contractions (NVCs), in a male rat model of bladder outlet obstruction (BOO) by evaluating cystometry (CMG) findings and bladder mechanosensitive single-unit afferent activities (SAAs), related with microcontractions, which may be similar with NVCs and to be of myogenic origin, in the rat model. BOO was created by partial ligation of the posterior urethra. At 4 days after surgery for BOO, an osmotic pump filled with silodosin (0.12 mg/kg/day) or its vehicle was subcutaneously implanted. At 10 days after surgery, CMG and SAAs measurements were taken under conscious and urethane-anesthetized conditions, respectively. The SAAs of Aδ- and C-fibers, which were identified by electrical stimulation of the pelvic nerve and by bladder distention, and intravesical pressure were recorded during constant bladder-filling with saline. Microcontractions were divided into three phases: "ascending," "descending," and "stationary." The silodosin-treated group showed a smaller number of NVCs in CMG measurements and lower SAAs of both Aδ- and C-fibers than the vehicle-treated group during bladder-filling. Moreover, in the vehicle-treated groups, the SAAs of both fibers for the ascending phase of microcontractions were significantly higher than those for the other two phases. On the contrary, no significant change was found between any of these three phases in the silodosin-treated group. The present results suggest that silodosin inhibits the SAAs of mechanosensitive Aδ- and C-fibers at least partly due to suppressing myogenic bladder contractions in male BOO rats.

The Antimuscarinic Agent Tolterodine Regulates Bladder Extracellular Matrix in Partial Bladder Outlet Obstruction in Rats

Background/Aims: Antimuscarinic agents can delay the progression of bladder dysfunction caused by bladder outlet obstruction (BOO). To date, the relationship between muscarinic receptor activity and the bladder extracellular matrix (ECM) remains unclear. Thus, an animal model of partial BOO (PBOO) in female rats was established to explore the variation in bladder wall ECM proteins under PBOO conditions with antimuscarinic agent administration. Methods: Rats were randomly divided into three groups: sham, PBOO, and PBOO plus tolterodine. Picrosirius red staining was used to examine the smooth muscle and collagen content of bladder samples. Gene microarray and RT-PCR were performed to survey the expression of ECM proteins, receptors, and metabolism regulators in the rat bladder. Positive results were further evaluated by immunohistochemistry. Results: Picrosirius red staining showed that smooth muscle volume significantly increased in the PBOO and PBOO plus tolterodine groups (p < 0.05), while collagen significantly increased in the PBOO group (p < 0.05) but not in the PBOO plus tolterodine group. Gene microarray and RT-PCR revealed that none of the collagen subtypes exhibited significant changes after PBOO establishment and tolterodine administration. However, matrix metalloproteinases (MMPs) increased significantly in the PBOO plus tolterodine group (p < 0.05). Additionally, PBOO inhibited the expression of non-collagen ECM proteins in the rat bladder wall, while tolterodine induced the expression of non-collagen ECM proteins and ECM receptors. Conclusions: Tolterodine decreased the volume of collagen in PBOO rat bladder wall, possibly via MMPs, and regulated the expression of ECM proteins and receptors.

Epigallocatechin Gallate Attenuates Bladder Dysfunction via Suppression of Oxidative Stress in a Rat Model of Partial Bladder Outlet Obstruction.

Purpose To investigate the protective effect of epigallocatechin gallate (EGCG), a green tea extract, and its underlying mechanism on bladder dysfunction in a rat model of bladder outlet obstruction (BOO). Materials and Methods Sprague-Dawley rats of BOO were surgically induced and followed by treatment with EGCG (5 mg/kg/day) or saline (control) via intraperitoneal injection. Cystometry was performed on four weeks postoperatively in conscious rats. H&E, Masson trichrome, and TUNEL staining were performed to observe tissue alterations. Oxidative stress markers were measured, and protein expression of Nrf2-ARE pathway was examined by immunohistochemistry and Western blotting. Results Our data showed that EGCG could increase the peak voiding pressure and bladder compliance and prolong micturition interval of BOO rats compared with control and finally reduce the frequency of urinary. EGCG could ameliorate the increase of collagen fibers and ROS induced by obstruction and increase the activity of SOD, GSH-Px, and CAT. The level of cell apoptosis was decreased in BOO rats treated with EGCG compared with control, and caspase-3 expression was reduced as well. Moreover, EGCG could activate the Nrf2 expression with elevation of its target antioxidant proteins. Conclusions EGCG alleviates BOO-induced bladder dysfunction via suppression of oxidative stress and activation of the protein expression of Nrf2-ARE pathway.

缝隙连接蛋白-43在膀胱出口部分梗阻的大鼠逼尿肌细胞膜中的表达

缝隙连接蛋白-43在膀胱出口部分梗阻的大鼠逼尿肌细胞膜中的表达

Near-Normalized Gene Expression Profiles in Bladder With Detrusor Overactivity in Rats With Bladder Outlet Obstruction After Deobstruction.

PurposeThe pathophysiological role of detrusor overactivity (DO) in the bladder, which is commonly observed in various bladder diseases, is not well understood. DO appears in bladder outlet obstruction (BOO), and may continue even after subsequent deobstruction. DO therefore provides an excellent opportunity to observe molecular biological changes.MethodsIn this study, to understand the molecular effects of persistent DO after BOO induction and deobstruction, we performed awake cystometry on female Sprague-Dawley rats divided into 4 groups: a sham group, a BOO group, a deobstructed group with DO after BOO (DDO), and a deobstructed group without DO after BOO (non-DDO). Total RNA was extracted from the bladder samples, and gene expression profiles were compared between the sham and model groups.ResultsDO was observed in 5 of the 6 rats (83%) in the BOO group, and in 6 of the 13 rats (46%) in the deobstructed group. The non-DDO group showed a significantly greater residual volume than the DDO group. Through a clustering analysis of gene expression profiles, we identified 7,532 common upregulated and downregulated genes, the expression of which changed by more than 2 fold. In the BOO group, 898 upregulated and 2,911 downregulated genes were identified. The non-DDO group showed 3,472 upregulated and 4,025 downregulated genes, whereas in the DDO group, only 145 and 72 genes were upregulated and downregulated, respectively.ConclusionsAbnormal function and gene expression profiles in bladders after BOO were normalized in the BOO rats with DO after deobstruction, whereas in those without DO, abnormal function persisted and the gene expression profile became more abnormal. DO may play a protective role against the stress to the bladder induced by BOO and deobstruction as a form of adaptive neuroplasticity.

Expression of cannabinoid 1 and, 2 receptors and the effects of cannabinoid 1 and, 2 receptor agonists on detrusor overactivity associated with bladder outlet obstruction in rats

BackgroundThis study investigated changes in the expression of cannabinoid (CB) receptors and the effects of CB1 and CB2 agonists on detrusor overactivity (DO) associated with bladder outlet obstruction (BOO) in rats.MethodsMale Sprague Dawley rats were randomly assigned to four groups (n = 10) in each group. The control group comprised sham-operated rats. A animals in the BOO, CB1 agonist and CB2 agonist groups all underwent BOO surgery. Three weeks postoperatively, cystometrography (CMG) was performed on all rats. After confirming the presence of DO in the CB1 and CB2 agonist groups, a CB1 agonist (WIN 55,212–2) and a CB2 agonist (CB65) were instilled intravesically, and CMG was repeated. CMG parameters, including the contraction interval (CI) and contraction pressure (CP) were then analyzed. The bladders of rats in all four groups were excised following CMG. Immunofluorescence staining and Western blotting were performed to localize CB1 and CB2 and measure their expression levels in the urothelium and detrusor muscle.ResultsThe CI was significantly longer and the CP was significantly lower in the CB1 agonist group than in the BOO group. CI and CP in the CB2 agonist group showed the same results. CB1 receptor immunofluorescence staining signals and immunoreactive bands in Western blotting were increased in the BOO group compared with results in the control group. Similarly, results for the CB2 receptor were also increased in the BOO group, although this difference was not significant. The CMG parameters in the BOO group were significantly improved by the inhibitory effects of CB1 and CB2 agonists on BOO-associated DO. The expression of CB1 was significantly increased in the urothelium and detrusor muscle in BOO-associated DO, but no significant change in CB2 expression was observed.ConclusionsCB1 and CB2 receptors, especially CB1, play a role in the pathophysiology of BOO-associated DO, and could serve as therapeutic targets.

NLRP3/IL-1β mediates denervation during bladder outlet obstruction in rats.

Denervation of the bladder is a detrimental consequence of bladder outlet obstruction (BOO). We have previously shown that, during BOO, inflammation triggered by the NLRP3 inflammasome in the urothelia mediates physiological bladder dysfunction and downstream fibrosis in rats. The aim of this study was to assess the effect of NLRP3-mediated inflammation on bladder denervation during BOO. There were five groups of rats: (i) Control (no surgery); (ii) Sham-operated; (iii) BOO rats given vehicle; (iv) BOO rats given the NLRP3 inhibitor glyburide; and (v) BOO rats given the IL-1 receptor antagonist anakinra. BOO was constructed by ligating the urethra over a 1 mm catheter and removing the catheter. Medications were given prior to surgery and once daily for 12 days. Bladder sections were stained for PGP9.5, a pan-neuronal marker. Whole transverse sections were used to identify and count nerves while assessing cross-sectional area. For in vitro studies, pelvic ganglion neurons were isolated and treated with IL-1β. After a 48 h incubation apoptosis, neurite length and branching were assessed. In obstructed bladders, the number of nerves decreased while total area increased, indicating a loss of cell number and/or branching. The decrease in nerve density was blocked by glyburide or anakinra, clearly implicating the NLRP3 pathway in denervation. In vitro analysis demonstrated that IL-1β, a product of the inflammasome, induced apoptosis in pelvic ganglion neurons, suggesting one mechanism of BOO-induced denervation is NLRP3/IL-1β triggered apoptosis. The NLRP3/IL-1β-mediated inflammation pathway plays a significant role in denervation during BOO.

TAC-302 promotes neurite outgrowth of isolated peripheral neurons and prevents bladder denervation related bladder dysfunctions following bladder outlet obstruction in rats.

To evaluate the ability of TAC-302, a cyclohexenoic fatty alcohol derivative, to enhance neurite outgrowth in cultured rat dorsal root ganglion (DRG) neurons, and the preventive effects of TAC-302 on bladder denervation-related storage and voiding dysfunctions in rats with bladder outlet obstruction (BOO). Rat DRG neurons were cultured in the presence of TAC-302. Cell numbers and neurite lengths were quantified after a 24 h culture. BOO was achieved by partial ligature of the proximal urethra in female rats. BOO rats were divided into three groups and orally treated with vehicle of 3 or 30 mg/kg TAC-302 twice a day for 4 weeks. Cystometry was performed under conscious conditions. Immunohistochemical staining using anti-PGP9.5 of the bladder muscle layer was performed, and the innervation area was scored. TAC-302 significantly and dose-dependently increased neurite outgrowth in cultured DRG neurons. BOO rats showed a decreased innervation area in the urinary bladder compared to sham-operated rats. BOO-induced denervation of the urinary bladder was partially prevented by oral treatment with TAC-302. TAC-302 significantly reduced the frequency of non-voiding contraction (NVC) and residual urine volume (RUV) compared with the BOO vehicle group (P < 0.05). The innervation area score exhibited significant negative correlations with NVC and RUV, indicating that they increased according to the progression of denervation. Our data indicate that TAC-302 promotes neurite outgrowth in vitro. In addition, TAC-302 prevents BOO-induced bladder dysfunction in rats, and has a protective effect on bladder denervation.

Bladder fibrosis during outlet obstruction is triggered through the NLRP3 inflammasome and the production of IL-1β.

Bladder outlet obstruction (BOO) triggers inflammation in the bladder through the NLRP3 inflammasome. BOO also activates fibrosis, which is largely responsible for the decompensation of the bladder in the chronic state. Because fibrosis can be driven by inflammation, we have explored a role for NLRP3 (and IL-1β produced by NLRP3) in the activation and progression of BOO-induced fibrosis. Female rats were divided into five groups: 1) control, 2) sham, 3) BOO + vehicle, 4) BOO + the NLRP3 inhibitor glyburide, or 5) BOO + the IL-1β receptor antagonist anakinra. Fibrosis was assessed by Masson's trichrome stain, collagen secretion via Sirius Red, and protein localization by immunofluorescence. BOO increased collagen production in the bladder, which was blocked by glyburide and anakinra, clearly implicating the NLRP3/IL-1β pathway in fibrosis. The collagen was primarily found in the lamina propria and the smooth muscle, while IL-1 receptor 1 and prolyl 4-hydroylase (an enzyme involved in the intracellular modification of collagen) both localized to the urothelium and the smooth muscle. Lysyl oxidase, the enzyme involved in the final extracellular assembly of mature collagen fibrils, was found to some extent in the lamina propria where its expression was greatly enhanced during BOO. In vitro studies demonstrated isolated urothelial cells from BOO rats secreted substantially more collagen than controls, and collagen expression in control cultures could be directly stimulated by IL-1β. In summary, NLRP3-derived-IL-1β triggers fibrosis during BOO, most likely through an autocrine loop in which IL-1β acts on urothelia to drive collagen production.

Minimal Invasive Cystometry and Intra-Abdominal Pressure Assessments in Rodents: A Novel Animal Study.

BackgroundThe abdominal straining pattern can act as a novel parameter for improving the prediction of bladder outlet obstruction (BOO). To preserve detrusor function in the early stage of urinary system impairment, such as BOO, we establish a novel method for cystometry and Intra-abdominal pressure (IAP) assessments in rodents without cystostomy.Material/MethodsTwenty mice and rats were divided into three groups (control, sham-operated and BOO group) respectively. The cystometry and IAP assessments were measured by the pediatric venous indwelling sheath and coronary dilatation catheter connected to Laborie urodynamic system on postoperative day 7. Data was collected simultaneously through urethra and rectum in each group. In addition, bladder histology was assessed to confirm BOO.ResultsThe novel method can collect the urodynamic parameters successfully, including the BLPP, IAP, MBC, etc. IAP was elevated in BOO rats, but no significantly difference was found between the sham-operated rats and the control rats. The hypertrophy of detrusor muscle in bladder section was observed by Masson trichrome staining in BOO group compared with other groups.ConclusionsOur novel method based on innovative research implement for cystometry and IAP assessments in rodents is a reliable and replicable approach for evaluating the lower urinary tract function. Especially it provides detailed information to evaluate lower urinary tract structures and function in the early stage of BOO.

MP94-13 THE EFFECT OF FREE RADICAL SCAVENGER ON THE OXIDATION STRESS IN PARTIAL BLADDER OUTLET OBSTRUCTION AND ITS RELIEF IN RAT MODEL

You have accessJournal of UrologyBladder & Urethra: Anatomy, Physiology & Pharmacology II1 Apr 2017MP94-13 THE EFFECT OF FREE RADICAL SCAVENGER ON THE OXIDATION STRESS IN PARTIAL BLADDER OUTLET OBSTRUCTION AND ITS RELIEF IN RAT MODEL Min Soo Choo, Songzhe Piao, and Seung-June Oh Min Soo ChooMin Soo Choo More articles by this author , Songzhe PiaoSongzhe Piao More articles by this author , and Seung-June OhSeung-June Oh More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.2916AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES In patients with BPH, de novo UI developed in 7.1-44.0% after HoLEP. Urodynamic involuntary detrusor contraction (IDC) was often observed in these patients. Ischemia reperfusion injury that occurs after de-obstruction has been proposed as a main cause of post-operative IDC. We investigated the effect of free radical scavenger (tempol) after relief of partial bladder outlet obstruction (pBOO) on bladder function in a rat model. METHODS Eight-week-old female Sprague Dawley of 40 rats (200-250g) were induced pBOO, and relieved it 3 weeks later. Rats were divided randomly 4 groups: tempol treated for 1 week (T1 group) and 3 weeks (T3 group), and no treated for 1 week (nT1 group) and 3 weeks (nT3 group). Cystometrograms were obtained in unanesthetized, unrestrained rats in metabolic cages at 1 or 3 weeks after relief, according to grouping, respectively. After completion of the investigation, the bladder was isolated and weighted. H&E, Masson trichrome and TUNEL staining were used to analyze the change of histology of the bladder. RESULTS Tempol treated groups decreased significantly in the number of IDC per voiding cycle (nT1 vs. T1, 1.18±0.82 vs. 0.36±0.40, P=0.010; nT3 vs. T3, 1.51±0.69 vs. 0.23±0.25, P=0.002). In the H&E staining, treated groups decreased significantly in thickness of the detrusor muscle layer (nT1 vs. T1, 1164.17±190.58 vs. 776.45±140.78, p<0.001; nT3 vs. T3, 905.82±161.16 vs. 726.26±162.76, P=0.043). In the Masson Chrome staining, the rates of collagen fiber were significantly lower in the treated groups. Apoptosis detected by TUNEL was observed in the urothelial cell layer mainly. The treated groups decreased significantly in the rate of apoptosis in urothelial cell layer (nT1 vs. T1, 48.9±3.36% vs. 32.7±11.10%, P=0.024; nT3 vs. T3, 25.8±4.67% vs. 15.7±9.83%, P=0.314). CONCLUSIONS Ischemia reperfusion injury that occurred after de-obstruction caused histologic and functional change of the bladder. Free radical scavenger could prevent this oxidative stress. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e1250-e1251 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Min Soo Choo More articles by this author Songzhe Piao More articles by this author Seung-June Oh More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

AB298. SPR-25 NLRP3/IL-1β mediates denervation during bladder outlet obstruction in rats

ObjectiveBladder outlet obstruction (BOO) is a common condition resulting from benign prostatic hyperplasia, neurologic pathology, organ prolapse, etc. Long-term, obstruction is well-established to evoke denervation in the bladder which causes the detrusor to become hypocontractile, resulting in inefficient bladder emptying and consequent infections, continence issues or even renal failure. Recently, considerable attention has been paid to a role for inflammation in bladder deterioration during BOO and we have shown a central role for the NLRP3 inflammasome in triggering this inflammation. In the present study we explore a possible connection between this NLRP3-induced inflammation and bladder denervation.MethodsRats were divided into five groups: (I) control; (II) sham operated; (III) BOO + vehicle (1 mL, 40% ethanol in PBS, p.o.); (IV) BOO + glyburide (Gly, NLRP3 inhibitor; 10 mg/kg, p.o.); (V) BOO + anakinra (Ana, IL-1 receptor antagonist; 25 mg/kg, i.p.). BOO is constructed in female rats by inserting a 1 mm outer diameter transurethral catheter, tying a silk ligature around the urethra and removing the catheter. Medications were administered prior to surgery and once daily. At 12 days animals were sacrificed and the bladders processed for histological analysis. Transverse sections (5 µm) were stained for PGP9.5 expression (a pan-neuronal marker) using standard immunohistochemistry techniques. Entire sections were scanned, using a 10× objective, into TIFF files using Zen software (Zeiss Inc.). Images were imported into Elements software (Nikon Inc.) and the area of individual neurons designated as well as total bladder area (exclusive of the urothelium and lumen). The number of neurons and respective areas were used to calculate nerve density.ResultsDenervation in the bladder wall during BOO was significant, as measured by nerve density. This effect was attenuated by either preventing NLRP3 activation with Gly or blocking IL-1β’s action at its receptor by treatment with Ana, clearly indicating a role for NLRP3/IL-1β in bladder denervation during BOO. The effect was also apparent with the total number of nerves despite considerable changes in bladder wall area (increased in BOO, maintained by Gly or Ana). Interestingly, the mean area of individual nerves was increased in BOO. This effect was blocked by Gly or Ana suggesting a loss of smaller neurons and/or retraction of neuronal branching during BOO as a result of NLRP3/IL-1β.ConclusionsNLRP3/IL-1b-induced inflammation leads to bladder denervation during BOO and blocking this pathway, either by preventing NLRP3 activation or inhibiting the action of IL-1B, prevents nerve loss.Funding Source(s)NIDDK—R01DK103534 (PI-Purves)

AB259. TRPV1 receptor associated with detrusor overactivity induced by partial bladder outlet obstruction in rats

BackgroundTo investigate the role of TRPV1 in the pathological process of DO induced by partial bladder outlet obstruction (pBOO).MethodsIn 40 female Wistar rats, pBOO was achieved by partial urethral ligation and urodynamic analysis was taken to filter the DO rats six weeks later. Urinary bladder and dorsal root ganglia (DRG) were removed and RT-PCR, Western Blot and IHC were performed to investigate the expression and location of TRPV1 in control and DO rats. Effect of different concentrations of TRPV1 agonist, as well as TRPV1 antagonist, was also evaluated with isolated detrusor strips.ResultsForty female Wistar rats received pBOO surgery and 24 of them developed DO. Expression of TRPV1 mRNA and protein in urinary bladder and dorsal root ganglia (DRG) significantly increased in pBOO induced OAB rats. While that did not significantly change after de-urothelial treatment both in OAB and control rats. Immunohistochemistry observed linear TRPV1-reactive staining mainly in suburothlial and muscular layer. In isolated detrusor strips studies, the amplitude and frequency of contractions of DO tissues were significantly higher than that of control ones (P<0.05). Capsaicin significantly increased the amplitude but not frequency of detrusor intrinsic contractility and this effect was enhanced in OAB conditions. All changes induced by capsaicin were blocked by capsazepine pre-incubated.ConclusionsIn pBOO induced DO rats, over-expressed TRPV1 was involved in DO pathological process by directly sensitizing bladder afferent fibers or indirectly enhancing detrusor intrinsic properties.

Prostaglandin-targeting agents and spectral heart rate variability in experimental partial bladder outlet obstruction in rats.

The purpose of this study was to determine the activity of the autonomic nervous system (ANS), using spectral analysis of the heart rate variability (HRV) in the model of partial bladder outlet obstruction (PBOO) in rats treated with selected non-steroidal anti-inflammatory drugs (NSAID): piroxicam (PRX) or meloxicam (MLX), and following administration of PGF2a prostaglandin analogue (Enzaprost F5). Neither the use of PGF2a analogue nor of MLX, caused significant changes in the HRV spectrum (except for HRV spectrum total power reduction with MLX). The use of PRX caused reduction of the total power and powers of all components of the HRV spectrum (except for VLF). Moreover, increased nLF and reduced nHF were observed. The obtained results suggest that the total prostaglandin synthesis block with a non-selective cyclooxygenase inhibitor (PRX) results in reduced ANS total activity, with decreased parasympathetic activity and a relative sympathetic predominance. The preferential cyclooxygenase-2 block (MLX) caused reduction of the total ANS activity as well, however with no clear disproportion of any part of the ANS. Therefore, prostaglandin synthesis inhibition and associated decrease of parasympathetic activity may constitute an additional and favourable feature of NSAID pharmacodynamics in the treatment of BPH.

Sulforaphane Ameliorates Bladder Dysfunction through Activation of the Nrf2-ARE Pathway in a Rat Model of Partial Bladder Outlet Obstruction.

Purpose. We evaluated the effect of sulforaphane (SFN) treatment on the function and changes of expression of Nrf2-ARE pathway in the bladder of rats with bladder outlet obstruction (BOO). Materials and Methods. A total of 18 male Sprague-Dawley rats at age of 8 weeks were divided into 3 groups (6 of each): the sham operated group, the BOO group, and the BOO+SFN group. We examined histological alterations and the changes of oxidative stress markers and the protein expression of the Nrf2-ARE pathway. Results. We found that SFN treatment could prolong micturition interval and increase bladder capacity and bladder compliance. However, the peak voiding pressure was lower than BOO group. SFN treatment can ameliorate the increase of collagen fibers induced by obstruction. SFN treatment also increased the activity of SOD, GSH-Px, and CAT compared to the other groups. The level of bladder cell apoptosis was decreased in BOO rats with SFN treatment. Moreover, SFN could reduce the ratio of Bax/Bcl-2 expression. Furthermore, SFN could activate the Nrf2 expression with elevation of its target antioxidant proteins. Conclusions. The sulforaphane-mediated decrease of oxidative stress and activation of the Nrf2-ARE pathway may ameliorate bladder dysfunction caused by bladder outlet obstruction.

Synergic Suppressive Effect of Silodosin and Imidafenacin on Non‐Voiding Bladder Contractions in Male Rats with Subacute Bladder Outlet Obstruction

To investigate single or combined effect of silodosin, an α1A-adrenoceptor antagonist, and imidafenacin, an antimuscarinic agent, on bladder function in a subacute bladder outlet obstruction (BOO) model of male rats. Partial BOO was created in male Wistar rats by ligating the urethra with a steel rod. On day 10 after surgery, when frequent voiding was most remarkable on voiding behavior measurement in a metabolic cage, cystometric investigations in a conscious restrained condition were conducted with cumulative intravenous administration of silodosin alone (0.1, 1, 10, and 100 µg/kg), imidafenacin alone (1, 3, and 10 µg/kg), or a combination of the two drugs. When compared with the Sham rats, the BOO rats showed an increase in bladder capacity, residual volume, mean amplitude and the number of non-voiding contractions (NVCs), accompanied with an increase in bladder weight. In the BOO rats, silodosin alone at 100 µg/kg significantly decreased the number of NVCs, whereas imidafenacin alone at 3 and 10 µg/kg significantly decreased both the number and mean amplitude of NVCs. The combination administration with lower doses (silodosin at 10 µg/kg and imidafenacin at 1 µg/kg) significantly suppressed both the number and mean amplitude of NVCs. The present results indicate a suppressive effect of silodosin or imidafenacin alone and a synergic effect of the combination of these two drugs on NVCs in a subacute BOO model of male rats.

Effect of silodosin, a selective α1A-adrenoceptor antagonist, on voiding behavior and bladder blood flow in a rat model of bladder outlet obstruction

This study was performed to investigate the effects of silodosin (selective α1A-adrenoceptor antagonist) on bladder blood flow (BBF) and bladder function in a rat model of bladder outlet obstruction (BOO) and to determine the expression of α1-adrenoceptor subtype mRNA in human and rat bladder microvessels.BOO was produced by partial ligature of the proximal urethra, which was maintained for 2 weeks. The BOO rats received either silodosin at a rate of 0.3mg/kg/day or vehicle subcutaneously via an osmotic pump for 2 weeks after BOO surgery. A metabolic cage study was performed in conscious animals. BBF was measured using a Laser Speckle Blood Flow Imager. Urinary levels of 8-hydroxy-2′-deoxyguanosine (8-OHdG) and nerve growth factor (NGF) were measured. Immunohistological examinations of nerve distribution and NGF expression in the rat bladder were conducted. The expression of each α1-adrenoceptor subtype mRNA in human and rat bladder microvessels was determined by in situ hybridization. Silodosin ameliorated the increase in voiding frequency and decrease in mean voided volume in BOO rats in the metabolic cage study. Silodosin also abrogated the decrease in BBF in BOO rats. The levels of 8-OHdG and NGF in BOO rats were significantly decreased by administration of silodosin. Silodosin prevented the decrease in nerve distribution and increase in NGF expression. Human and rat bladder microvessels showed expression of all α1-adrenoceptor subtype mRNAs. The results presented here suggest that silodosin improves voiding behavior in rat models with BOO by inducing recovery of BBF.

Expression of brain derived-neurotrophic factor and granulocyte-colony stimulating factor in the urothelium: relation with voiding function.

BackgroundWe designed this experiment to elucidate the relationship between the expression of brain derived-neurotrophic factor (BDNF), the expression of granulocyte-colony stimulating factor (G-CSF), and the development of overactive bladder (OAB). In our previous study, the urothelium was observed to be more than a simple mechanosensory receptor and was found to be a potential therapeutic target for OAB. Moreover, neuregulin-1 and BDNF were found to be potential new biomarkers of OAB. Here, we investigated the relationship between changes in the voiding pattern and the expression of BDNF and G-CSF in the urothelium and evaluated the effects of 5-hydroxymethyl tolterodine (5-HMT) on rats with bladder outlet obstruction (BOO).MethodsA total of 100 Sprague–Dawley rats were divided into the following groups: 20 control rats; 40 BOO rats; and 40 BOO rats administered 5-HMT (0.1 mg/kg). After BOO was induced for 4 weeks, the rats were assessed by cystometrography. The changes in BDNF and G-CSF expression were examined in both separated urothelial tissues and in cultured urothelial cells by reverse transcription polymerase chain reaction (RT-PCR).ResultsBOO rats showed increased non-voiding activity [NVA; (number/10 voidings)] and bladder weight and decreased micturition volume (MV), micturition interval (MI), and micturition time (MT) relative to the controls. Moreover, the 5-HMT administration rats showed decreased NVA and bladder weight and increased MV and MI in comparison to the BOO rats. BDNF and G-CSF expression was increased in BOO rats and decreased following 5-HMT administration. In this model, voiding dysfunction developed as a result of BOO. As a therapeutic agent for OAB, the administration of 5-HMT improved the voiding dysfunction.ConclusionsBDNF and G-CSF might modulate voiding patterns through micturition pathways and might be involved only in the urothelium. Moreover, the expression of both genes in the urothelium might be related to voiding dysfunction in OAB patients. Thus, the urothelium has an important role in the manifestation of voiding symptoms.