Female Type 1 Diabetic Akita Mice Demonstrate Increased Bladder Contractility via FP Receptor Activation due to NLRP3-Mediated Inflammation.

BACKGROUND: Diabetic bladder dysfunction (DBD) is driven in part by inflammation which dysregulates prostaglandin release in the bladder. Precise inflammatory mechanisms responsible for such dysregulation have been elusive. Since prostaglandins impact bladder contractility, elucidating these mechanisms may yield potential therapeutic targets for DBD. In female Type 1 diabetic Akita mice, inflammation mediated by the nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3 (NLRP3) inflammasome is responsible for DBD. Here, we utilized female Akita mice crossbred with NLRP3 knock-out mice to determine how NLRP3-driven inflammation impacts prostaglandin release within the bladder and prostaglandin-mediated bladder contractions. METHODS: Akita mice were crossbred with NLRP3-⁣/- mice to yield four groups of non-diabetics and diabetics with and without the NLRP3 gene. Females were aged to 30 weeks when Akitas typically exhibit DBD. Urothelia and detrusors were stretched ex vivo to release prostaglandins. Prostaglandin E2 (PGE2) and prostaglandin F2α (PGF2α) were quantified using enzyme linked immunosorbent assays (ELISA). In separate samples, ex vivo contractile force to PGE2 and PGF2α +/- the prostaglandin F (FP) receptor antagonist, AL8810, was measured. FP receptor protein expression was determined via western blotting. RESULTS: Stretch-induced PGE2 release increases in urothelia but decreases in detrusors of diabetics. However, PGE2-mediated bladder contractions are not impacted. Conversely, diabetics show no changes in PGF2α release, but PGF2α-mediated contractions increase significantly. This is likely due to signaling through the FP receptors as FP receptor antagonism prevents this increase and diabetics demonstrate a four-fold increase in FP receptor proteins. Without NLRP3-mediated inflammation, changes in prostaglandin release, contractility, and receptor expression do not occur. CONCLUSION: NLRP3-dependent inflammation dysregulates prostaglandin release and prostaglandin-mediated bladder contractions in diabetic female Akita mice via FP receptor upregulation.

Recurrent infections drive persistent bladder dysfunction and pain via sensory nerve sprouting and mast cell activity.

Urinary tract infections (UTIs) account for almost 25% of infections in women. Many are recurrent (rUTI), with patients frequently experiencing chronic pelvic pain and urinary frequency despite clearance of bacteriuria after antibiotics. To elucidate the basis for these bacteria-independent bladder symptoms, we examined the bladders of patients with rUTI. We noticed a notable increase in neuropeptide content in the lamina propria and indications of enhanced nociceptive activity. In mice subjected to rUTI, we observed sensory nerve sprouting that was associated with nerve growth factor (NGF) produced by recruited monocytes and tissue-resident mast cells. Treatment of rUTI mice with an NGF-neutralizing antibody prevented sprouting and alleviated pelvic sensitivity, whereas instillation of native NGF into naïve mice bladders mimicked nerve sprouting and pain behavior. Nerve activation, pain, and urinary frequency were each linked to the presence of proximal mast cells, because mast cell deficiency or treatment with antagonists against receptors of several direct or indirect mast cell products was each effective therapeutically. Thus, our findings suggest that NGF-driven sensory sprouting in the bladder coupled with chronic mast cell activation represents an underlying mechanism driving bacteria-independent pain and voiding defects experienced by patients with rUTI.

Enzyme-induced hypoxia leads to inflammation in urothelial cells in vitro.

PURPOSE: To determine the contributions of different durations of hypoxia to NLRP3 inflammasome activation in urothelial cells and how ischemic changes in bladder tissues is an important chemical que that leads to pathological changes seen in BOO. METHODS: A rat urothelial cell line (MYP3) was exposed to either a short duration (2 h) or long duration (6 h) of enzyme-induced hypoxia. Following exposure to a short duration of hypoxia, NO and ATP concentrations were measured from supernatant media and caspase-1 levels were measured from cell lysates. In a separate experiment, cells were fixed following hypoxia exposure and immunostained for HIF-1α stabilization. RESULTS: Although short exposure of low oxygen conditions resulted in a hypoxic response in MYP3 cells, as indicated by HIF-1α stabilization and increased NO activity, NLRP3 inflammasome activation was not observed as caspase-1 activity remained unchanged. However, exposure of MYP3 cells to a longer duration of hypoxia resulted in an increase in intracellular caspase-1 activity. Furthermore, treatment with antioxidant (GSH) or TXNIP inhibitor (verapamil) attenuated the hypoxia-induced increase in caspase-1 levels indicating that hypoxia primarily drives inflammation through a ROS-mediated TXNIP/NLRP3 pathway. CONCLUSION: We conclude that hypoxia induced bladder damage requires a duration that is more likely related to elevated storage pressures/hypoxia, seen in later stages of BOO, as compared to shorter duration pressure elevation/hypoxia that is encountered in normal micturition cycles or early in the BOO pathology where storage pressures are still normal.

Male Akita mice develop signs of bladder underactivity independent of NLRP3 as a result of a decrease in neurotransmitter release from efferent neurons.

Diabetic bladder dysfunction (DBD) is a prevalent diabetic complication that is recalcitrant to glucose control. Using the Akita mouse model (type 1) bred to be NLR family pyrin domain containing 3 (NLRP3)+/+ or NLRP3-/-, we have previously found that females (mild hyperglycemia) progress from an overactive to underactive bladder phenotype and that this progression was dependent on NLRP3-induced inflammation. Here, we examined DBD in the male Akita mouse (severe hyperglycemia) and found by urodynamics only a compensated underactive-like phenotype (increased void volume and decreased frequency but unchanged efficiency). Surprisingly, this phenotype was still present in the NLRP3-/- strain and so was not dependent on NLRP3 inflammasome-induced inflammation. To examine the cause of the compensated underactive-like phenotype, we assessed overall nerve bundle density and afferent nerve bundles (Aδ-fibers). Both were decreased in density during diabetes, but denervation was absent in the diabetic NLRP3-/- strain so it was deemed unlikely to cause the underactive-like symptoms. Changes in bladder smooth muscle contractility to cell depolarization and receptor activation were also not responsible as KCl (depolarizing agent), carbachol (muscarinic agonist), and α,ß-methylene-ATP (purinergic agonist) elicited equivalent contractions in denuded bladder strips in all groups. However, electrical field stimulation revealed a diabetes-induced decrease in contractility that was not blocked in the NLRP3-/- strain, suggesting that the bladder compensated underactive-like phenotype in the male Akita mouse is likely through a decrease in efferent neurotransmitter release.NEW & NOTEWORTHY In this study, we show that diabetic bladder dysfunction (the most common diabetic complication) manifests through different mechanisms that may be related to severity of hyperglycemia and/or sex. Male Akita mice, which have severe hyperglycemia, develop bladder underactivity as a result of a decrease in efferent neurotransmitter release that is independent of inflammation. This contrasts with females, who have milder hyperglycemia, where diabetic bladder dysfunction progresses from overactivity to underactivity in an inflammation-dependent manner.

Why Are Some People with Lower Urinary Tract Symptoms (LUTS) Depressed? New Evidence That Peripheral Inflammation in the Bladder Causes Central Inflammation and Mood Disorders.

Anecdotal evidence has long suggested that patients with lower urinary tract symptoms (LUTS) develop mood disorders, such as depression and anxiety, at a higher rate than the general population and recent prospective studies have confirmed this link. Breakthroughs in our understanding of the diseases underlying LUTS have shown that many have a substantial inflammatory component and great strides have been made recently in our understanding of how this inflammation is triggered. Meanwhile, studies on mood disorders have found that many are associated with central neuroinflammation, most notably in the hippocampus. Excitingly, work on other diseases characterized by peripheral inflammation has shown that they can trigger central neuroinflammation and mood disorders. In this review, we discuss the current evidence tying LUTS to mood disorders, its possible bidirectionally, and inflammation as a common mechanism. We also review modern theories of inflammation and depression. Finally, we discuss exciting new animal studies that directly tie two bladder conditions characterized by extensive bladder inflammation (cyclophosphamide-induced hemorrhagic cystitis and bladder outlet obstruction) to neuroinflammation and depression. We conclude with a discussion of possible mechanisms by which peripheral inflammation is translated into central neuroinflammation with the resulting psychiatric concerns.

Single-port hidden incision endoscopic (HIdES) pediatric nephrectomy via pfannenstiel incision.

Laparoendoscopic single-site surgery (LESS) and hidden incision endoscopic surgery techniques are increasingly used in pediatric urology. For pediatric nephrectomy, access through a single Pfannenstiel incision is novel and may offer cosmetic benefit. In this retrospective study, we describe this approach and assess operative outcomes associated with this technique. Patients who underwent LESS nephrectomy through a single Pfannenstiel incision had minimal blood loss, short length of stay, low risk of surgical complications, and satisfactory wound healing. The Pfannenstiel approach to LESS nephrectomy is feasible, versatile, and achieves excellent operative and cosmetic outcomes, although direct comparison to other approaches is warranted.

Medical management of neurogenic bladder in patients with spina bifida: A scoping review.

INTRODUCTION: Neurogenic bladder is a common source of morbidity in patients with spina bifida and can cause renal damage. Medical management may include imaging, urodynamic studies (UDS), laboratory testing, clean intermittent catheterization (CIC), and medication. There is ongoing debate regarding the optimal management regimen. Approaches are described by two paradigms: proactive and expectant management. In a proactive approach, invasive interventions like CIC and UDS are initiated before the onset of renal abnormalities. In expectant management, UDS, CIC, and medications are started after abnormalities are identified. In this scoping review, we aim to comprehensively review existing literature on outcomes of proactive and expectant management of neurogenic bladder in patients with spina bifida. METHODS: We searched multiple databases and screened articles for inclusion using PRISMA-ScR guidelines. Included studies reported clinical outcomes of any aspect of proactive or expectant neurogenic bladder management in patients with spina bifida. RESULTS: Ultimately, 74 articles were included for review including 67 cohort studies, 4 cross-sectional studies, 2 sequential cohort studies, and 1 randomized control trial. Eleven studies directly compared management strategies. There was substantial heterogeneity in study designs, management protocols, and reported outcomes. Most studies addressed multiple simultaneous aspects of management without specifically analyzing individual aspects. However, some commented on individual aspects of management including UDS (13), CIC (32), imaging (7), and medication (5). Although there was no consensus about optimal management, all direct comparisons of paradigms supported a proactive approach. CONCLUSION: Our review identified a broad body of literature about optimal management of neurogenic bladder. Existing studies vary greatly in terms of treatment protocols, measured outcomes, and management recommendations. Overall, studies that directly compare management are scarce but favor proactive management. Given the implications on clinical outcomes, it is crucial to focus future work on directly comparing management strategies and isolating the effects of different individual management elements.

Inflammation triggered by the NLRP3 inflammasome is a critical driver of diabetic bladder dysfunction.

Diabetes is a rapidly expanding epidemic projected to affect as many as 1 in 3 Americans by 2050. This disease is characterized by devastating complications brought about high glucose and metabolic derangement. The most common of these complications is diabetic bladder dysfunction (DBD) and estimates suggest that 50-80% of patients experience this disorder. Unfortunately, the Epidemiology of Diabetes Interventions and Complications Study suggests that strict glucose control does not decrease ones risk for incontinence, although it does decrease the risk of other complications such as retinopathy, nephropathy and neuropathy. Thus, there is a significant unmet need to better understand DBD in order to develop targeted therapies to alleviate patient suffering. Recently, the research community has come to understand that diabetes produces a systemic state of low-level inflammation known as meta-inflammation and attention has focused on a role for the sterile inflammation-inducing structure known as the NLRP3 inflammasome. In this review, we will examine the evidence that NLRP3 plays a central role in inducing DBD and driving its progression towards an underactive phenotype.

Framing the picture: Impact of radiographic imaging on parental vesicoureteral reflux management preferences.

INTRODUCTION/OBJECTIVE: Given the variety of treatment options for vesicoureteral reflux (VUR), shared decision making between clinicians and parents is essential. Despite its importance, shared decision making is limited by the framing effect - people process the same information differently depending on how it is presented. Studies have also demonstrated that showing patients their radiology images can impact behaviors. In this pilot study, we sought to determine if showing parents radiographic images could serve as a framing tool that impacts the decision of whether to pursue surgery, endoscopic intervention, or conservative management for VUR. METHODS: We designed a survey instrument which provided background on VUR and a hypothetical scenario of a 2-year-old child with VUR who had a breakthrough febrile urinary tract infection (UTI). Guideline-concordant management options were presented: (1) change antibiotics, (2) endoscopic management, or (3) open or laparoscopic surgery. All options were similarly presented regarding risks, benefits, and length of stay. Respondents were randomized into a group with no image accompanying the clinical scenario or a group which had a labeled image of a voiding cystourethrogram (VCUG) demonstrating unilateral VUR. Respondents also answered demographic and health experience questions. The instrument was published on Amazon's Mechanical Turk online work interface which provides reliable and validated results in VUR experiments. Parents aged 18-60 years old were eligible. Responses with failed attention questions, duplicate internet addresses, or submission times <1 or >30 min were disqualified. Data were analyzed using t-test, chi-square, and multinomial logistic regression. Sensitivity analyses were performed after excluding all responses submitted under 2, 3, and 5 min. RESULTS: There were a total of 914 responses, 426 met inclusion criteria. The presence or absence of a VCUG image did not result in a statically significant difference in the management decision (p = 0.081). Multinomial logistic regression demonstrated that prior UTI experience influenced the management decision (p = 0.027). Sensitivity analyses revealed a significant difference in the management decision when excluding responses <5 min (p = 0.039). CONCLUSION: In this analysis, there was no statistically significant framing effect by radiographic images on parental management decision for VUR. Multinomial analyses suggested that prior experience with UTI has an impact on VUR management decisions. These results need to be considered within the limitations of this pilot study - the respondents were given a hypothetical clinical scenario and the survey instrument cannot replace an in-office discussion. Further analyses on framing and its role in pre-operative counseling is warranted.

Diabetes causes NLRP3-dependent barrier dysfunction in mice with detrusor overactivity but not underactivity.

Approximately half of the patients with diabetes develop diabetic bladder dysfunction (DBD). The initiation and progression of DBD is largely attributed to inflammation due to dysregulated glucose and the production of toxic metabolites that activate the NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome. NLRP3 activation leads to the production and release of proinflammatory cytokines and causes urothelial pyroptosis, a form of programmed cell necrosis, which we hypothesize compromises urothelial barrier integrity. Here, we investigated how NLRP3-dependent inflammation impacts barrier function during the progression of diabetes using a type 1 diabetic female Akita mouse model that progresses from an early overactive to a late underactive detrusor phenotype at 15 and 30 wk, respectively. To determine the specific role of NLRP3, Akita mice were crossbred with mice lacking the NLRP3 gene. To determine barrier function, permeability to small molecules was assessed, ex vivo using Evans blue dye and in vivo using sulfo-NHS-biotin. Both ex vivo and in vivo permeabilities were increased in diabetic mice at 15 wk. Expression of uroplakin and tight junction components was also significantly downregulated at 15 wk. Interestingly, diabetic mice lacking the NLRP3 gene showed no evidence of barrier damage or downregulation of barrier genes and proteins. At the 30-wk time point, ex vivo and in vivo barrier damage as well as barrier component downregulation was no longer evident in diabetic mice, suggesting urothelial repair or remodeling occurs between the overactive and underactive stages of DBD. Collectively, these findings demonstrate the role of NLRP3-mediated inflammation in urothelial barrier damage associated with detrusor overactivity but not underactivity.NEW & NOTEWORTHY This is the first study to demonstrate that NLRP3-mediated inflammation is responsible for urothelial barrier damage in type 1 diabetic female Akita mice with an overactive bladder. Eliminating the NLRP3 gene in these diabetic mice prevented barrier damage as a result of diabetes. By the time female Akita mice develop an underactive phenotype, the urothelial barrier has been restored, suggesting that inflammation is a critical causative factor early in the development of diabetic bladder dysfunction.

Diabetic bladder dysfunction progresses from an overactive to an underactive phenotype in a type-1 diabetic mouse model (Akita female mouse) and is dependent on NLRP3.

AIMS: Diabetic bladder dysfunction (DBD) is a prevalent diabetic complication thought to progress from overactive (OAB) to underactive (UAB) bladder. Previously we found OAB at 15 weeks in the Akita mouse, a genetic model of Type 1 diabetes. The first aim of this study assesses bladder function at 30 weeks to assess progression. In addition, inflammation triggered by the NLRP3 inflammasome is implicated in DBD. In a second aim we assessed a role for NLRP3 by crossing Akita mice with NLRP3-/- mice. MAIN METHODS: Akita mice were bred with NLRP3-/- mice. The effect of diabetes was assessed by comparing nondiabetic to diabetic mice (all NLRP3+/+). The effect of diabetes in the absence of the NLRP3 inflammasome was assessed by comparing nondiabetic/NLRP3-/- to diabetic/NLRP3-/- mice. Mice were assessed at 30 weeks for blood glucose (glucometer), inflammation (Evans blue), bladder morphology (histology) and bladder function (urodynamics). KEY FINDINGS: At 30 weeks blood glucose of nondiabetics and diabetics was not affected by the presence of absence of NLRP3. Diabetic/NLRP3+/+ mice showed bladder inflammation and detrusor hypertrophy which was blocked in the diabetic/NLRP3-/- mice, clearly showing a role for NLRP3. When bladder function was examined, diabetic/NLRP3+/+ showed an increase in voiding volume and a decrease in frequency, two signs of underactive bladder. However, in the NLRP3-/- mice, diabetes was unable to effectuate these changes, demonstrating that NLRP3-induced inflammation is responsible for UAB symptoms in these mice. SIGNIFICANCE: Akita diabetic mice progress from OAB to UAB. NLRP3 is a possible target to treat DBD.

A Multidimensional Bioinformatic Platform for the Study of Human Response to Surgery.

OBJECTIVE: To design and establish a prospective biospecimen repository that integrates multi-omics assays with clinical data to study mechanisms of controlled injury and healing. BACKGROUND: Elective surgery is an opportunity to understand both the systemic and focal responses accompanying controlled and well-characterized injury to the human body. The overarching goal of this ongoing project is to define stereotypical responses to surgical injury, with the translational purpose of identifying targetable pathways involved in healing and resilience, and variations indicative of aberrant peri-operative outcomes. METHODS: Clinical data from the electronic medical record combined with large-scale biological data sets derived from blood, urine, fecal matter, and tissue samples are collected prospectively through the peri-operative period on patients undergoing 14 surgeries chosen to represent a range of injury locations and intensities. Specimens are subjected to genomic, transcriptomic, proteomic, and metabolomic assays to describe their genetic, metabolic, immunologic, and microbiome profiles, providing a multidimensional landscape of the human response to injury. RESULTS: The highly multiplexed data generated includes changes in over 28,000 mRNA transcripts, 100 plasma metabolites, 200 urine metabolites, and 400 proteins over the longitudinal course of surgery and recovery. In our initial pilot dataset, we demonstrate the feasibility of collecting high quality multi-omic data at pre- and postoperative time points and are already seeing evidence of physiologic perturbation between timepoints. CONCLUSIONS: This repository allows for longitudinal, state-of-the-art geno-mic, transcriptomic, proteomic, metabolomic, immunologic, and clinical data collection and provides a rich and stable infrastructure on which to fuel further biomedical discovery.

Specialized pro-resolution mediators in the bladder: Receptor expression and recovery of bladder function from cystitis.

Inflammation is a central process in most benign bladder disorders, and its control is a delicate balance between initiating factors and resolving factors. While recent discoveries have shown a central role for the NLRP3 inflammasome in initiation, the resolving pathways remain unexplored. Resolution is controlled by specialized pro-resolution mediators (SPMs) functioning through seven receptors (six in rodents). Here we demonstrate expression of all seven in humans (six in mice) through immunocytochemistry. Expression was universal in urothelia with most also expressed in smooth muscle. We next explored the therapeutic potential of three SPMs; Resolvin E1 (RvE1), Maresin 1 (MaR1), and Protectin D1 (PD1). SPMs promote epithelial wound/barrier repair and RvE1 triggered dose-dependent wound closure in urothelia in vitro (scratch assay) (EC90 = 12.5 nM). MaR1 and PD1 were equally effective at this concentration. In vivo analyses employed a cyclophosphamide (CP) model of bladder inflammation (Day 0-CP [150 mg/kg], Day 1 to 3 SPM [25 µg/kg/day], Day 4 - analysis). All three SPMs reduced bladder inflammation (Evans blue) and bladder weights to control levels. Effects of RvE1 were also examined by urodynamics. CP decreased void volume, increased frequency and decreased bladder capacity while RvE1 restored values to control levels. Finally, SPMs reduce fibrosis and RvE1 reduced urothelial expression of TGF-ß and collagen I to control values. Together these results expand the known SPMs active in the bladder tissue and provide promising therapeutic targets for controlling inflammation in a wide variety of inflammation-associated benign bladder diseases.

Association between race and in-hospital outcomes in children with spina bifida following inpatient urologic surgery.

PURPOSE: Post-operative complication rates may vary among racial and/or ethnic groups and have not been previously described in individuals with spina bifida (SB) undergoing urologic surgery. The aim of this study was to compare in-hospital complication frequencies of individuals with SB following urologic surgery by race/ethnicity. METHODS: The Nationwide Inpatient Sample was used to identify pediatric patients with SB who underwent inpatient urologic procedures. A pediatric cohort (<18 years old) with SB that underwent urologic surgery were assessed. All analyses report weighted descriptive statistics, outcomes, and race/ethnicity was the primary predictor variable. The primary outcome of interest was post-operative complications which were defined using NSQIP ICD-9 code definitions. Secondary analysis included length of stay (LOS), and encounter cost was estimated using the cost-to-charge ratio files provided by the Healthcare Cost and Utilization Project. RESULTS: The unadjusted model showed no differences in complications, LOS, and cost. In the adjusted model there were no differences in complications, LOS, and cost between Black and White encounters. However, Hispanic ethnicity was associated with a 20%(95%CI: 4-40%) increase in LOS and 18%(95%CI: 2-35%, p = 0.02) increase in cost compared to White encounters. CONCLUSION: There was no evidence of variation for in-hospital complication rates among racial/ethnic groups undergoing urologic surgery. Hispanic ethnicity was associated with higher costs and longer LOS in pediatric SB encounters.

The Importance of Early Diagnosis and Management of Pediatric Neurogenic Bladder Dysfunction.

Neurogenic bladder dysfunction is a major source of urologic morbidity in children, especially in those with spina bifida (SB). Complications from progression of bladder dysfunction can include urinary tract infections (UTIs), urinary incontinence, upper tract deterioration, and renal dysfunction or failure. In these children, there has been a recent trend toward proactive rather than expectant management of neurogenic bladder. However, there is a lack of consensus on how to best achieve the three main goals of neurogenic bladder management: 1) preserving kidney function, 2) achieving continence (if desired by the family/individual), and 3) achieving social and functional urologic independence (if appropriate). Hence, our objective was to perform a narrative literature review to evaluate the approaches to diagnosis and management of pediatric neurogenic bladder dysfunction, with special focus on children with SB. The approach strategies vary across a spectrum, with a proactive strategy on one end of the spectrum and an expectant strategy at the other end. The proactive management strategy is characterized by early and frequent labs, imaging, and urodynamic (UDS) evaluation, with early initiation of clean intermittent catheterization (CIC) and proceeding with pharmacotherapy, or surgery if indicated. The expectant management strategy prioritizes surveillance labs and imaging prior to proceeding with invasive assessments and interventions such as UDS or pharmacotherapy. Both treatment strategies are currently utilized and data have historically been inconclusive in demonstrating efficacy of one regimen over the other. We performed a narrative literature evaluating proactive and expectant treatment strategies as they relate to diagnostics and management of Spina Bifida. From the available literature and our practice, a proactive strategy favors greater benefit in preventative management and may decrease risk of renal dysfunction compared with expectant management.

Specialized proresolution mediators in the bladder: annexin-A1 normalizes inflammation and bladder dysfunction during bladder outlet obstruction.

Bladder outlet obstruction (BOO) is ultimately experienced by ≈90% of men, most commonly secondary to benign prostatic hyperplasia. Inflammation is a critical driver of BOO pathology in the bladder and can be divided into two critical steps: initiation and resolution. Although great strides have been made toward understanding the initiation of inflammation in the bladder [through the NLR family pyrin domain containing 3 (NLRP3) inflammasome], no studies have examined resolution. Resolution is controlled by five classes of compounds known as specialized proresolving mediators (SPMs), all of which bind to one or more of the seven different receptors. Using immunocytochemistry, we showed the presence of six of the known SPM receptors in the bladder of control and BOO rats; the seventh SPM receptor has no rodent homolog. Expression was predominantly localized to urothelia, often with some expression in smooth muscle, but little to none in interstitial cells. We next examined the therapeutic potential of the annexin-A1 resolution system, also present in control and BOO bladders. Using the peptide mimetic Ac2-26, we blocked inflammation-initiating pathways (NLRP3 activation), diminished BOO-induced inflammation (Evans blue dye extravasation), and normalized bladder dysfunction (urodynamics). Excitingly, Ac2-26 also promoted faster and more complete functional recovery after surgical deobstruction. Together, the results demonstrate that the bladder expresses a wide variety of potential proresolving pathways and that modulation of just one of these pathways can alleviate many detrimental aspects of BOO and speed recovery after deobstruction. This work establishes a precedent for future studies evaluating SPM effectiveness in resolving the many conditions associated with bladder inflammation.NEW & NOTEWORTHY To our knowledge, this is the first study of proinflammation-resolving pathways in the bladder, which is the basis of a new pharmacological genus-dubbed "resolution pharmacology" aimed at reducing inflammation without creating an immunocompromised state. Inflammation plays a causative or exacerbating role in numerous bladder maladies. We documented proresolution receptors in the rat bladder and the effectiveness of a specialized proresolving mediator, annexin-A1, in alleviating detrimental aspects of bladder outlet obstruction and speeding recovery after deobstruction.

BOO induces fibrosis and EMT in urothelial cells which can be recapitulated in vitro through elevated storage and voiding pressure cycles.

PURPOSE: To determine the unique contributions from elevated voiding and storage pressures in the development of fibrosis and the epithelial-to-mesenchymal transition (EMT) in urothelial cells, and how progressive BOO pressure cycling is an important mechanical cue leading to these pathological changes. MATERIALS AND METHODS: Urothelial cells isolated from control, SHAM, 2 (acute)- or 6 (chronic)-week BOO rats treated with an inflammasome inhibitor or no drug. Total RNA was isolated and RT-PCR was conducted with custom primers for pro-fibrotic and EMT genes. In separate experiments, a rat urothelial cell line was exposed to cyclic pressure regimes characteristic of acute and chronic BOO in the presence or absence of an inflammasome inhibitor. Following exposure, RT-PCR was conducted, collagen content was determined and intracellular caspase-1 activity was measured. RESULTS: Urothelial cells isolated from acute and chronic BOO rat models demonstrated expression of pro-fibrotic and EMT genes. Similarly, MYP3 rat urothelial cells subjected to pressure cycling regimes that reflect intravesical pressures in the acute or chronic BOO bladder also demonstrated increased expression of pro-fibrotic and EMT genes, along with elevated soluble collagen. Treatment with inflammasome inhibitors reduced expression of pro-fibrotic genes in the rat model and pressure cycling model but had a limited effect on EMT. CONCLUSION: These results indicate that acute and chronic BOO pressure cycling are essential in the initiation and progression of fibrosis in the bladder via the NLRP3 inflammasome, but also provide new evidence that there is also an alternative NLRP3-independent pathway leading to EMT and fibrosis.

Immune Expression in Children With Vesicoureteral Reflux: A Pilot Study.

OBJECTIVE: To perform an exploratory, descriptive pilot study of the systemic and local immune environment in patients with vesicoureteral reflux (VUR) and bladder-bowel dysfunction (BBD). METHODS: Consecutive children with VUR undergoing intravesical ureteral reimplantation were enrolled. Patients were assessed for presence of BBD by reported patient history and validated questionnaire. Fresh blood and bladder tissue, collected at the time of surgery, were immediately processed for analysis. Immune cell compositions were determined via flow cytometry. Immune cell activation was also defined at the time of analysis. LegendPlex assay analysis was utilized to define levels of circulating chemokines and cytokines. RESULTS: A total of 7 patients were enrolled. Although percentages of circulating immune cells in the blood of those with VUR/BBD and VUR alone were similar, within bladder tissue, VUR/BBD demonstrated increased immune infiltrates compared to VUR alone. Bladder sample analysis showed that B cells, and Effector Memory and Naïve T cell percentages were significantly increased in VUR/BBD patients compared to VUR patients. T cell expression of PD1 was increased in bladder tissues of BBD/VUR. Additionally, analysis of circulating neutrophils displayed significantly increased upregulation of PDL-1 in patients with VUR/BBD vs those with VUR only. CONCLUSION: These pilot data suggest an immune-rich microenvironment is present within VUR. Severity of inflammation appeared to correlate with presence of BBD. This implies that targeting pelvic inflammation may be a novel therapy for children with VUR- or non-VUR-related BBD. Follow-up studies are currently underway.

Parental preferences for vesicoureteral reflux treatment: Profile case best-worst scaling.

INTRODUCTION: Vesicoureteral reflux is a common pediatric urologic condition that often has several reasonable treatment options depending on condition severity. In order to choose the best treatment for their child, parents are expected to make judgements that weigh attributes such as treatment cost, effectiveness, and complication rate. Prior research has shown that factors such as treating hospital and surgeon also influence patient treatment choice. OBJECTIVES: This study evaluates parental preferences for reflux treatment using profile case best-worst scaling, an emerging technique in both urologic and health care preference estimation. The study also uses latent class analysis (LCA) to identify parental sub-classes with different preferences. STUDY DESIGN: Data were collected from a community sample of parents via a multimedia best-worst scaling survey instrument published to Amazon's Mechanical Turk online community. After extensive review of the literature, reflux attributes and attribute levels were selected to correspond with available treatments. The profile case best-worst scaling exercise elicited preferences for granular attributes of reflux treatments. Data were analyzed using multinomial logistic regression and class analysis to distinguish preference heterogeneity. Probability scaled values (PSVs) reflected the order of desirability of the attributes. Attribute preference importance was rescaled into dollar units for comparison as well. RESULTS: We analyzed data for 248 respondents. The highest treatment effectiveness was more desirable than all other leveled treatment attributes (PSV 17.8, all p < 0.01) (Table). Low complication rate and doctor recommendation were amongst the other most desirable treatment attributes (PSV 11.3 and 9.0, respectively). Latent class analysis identified a class with more extreme preferences, for whom doctor recommendation and avoiding hospitalization were particularly desirable. DISCUSSION: In this community-based sample, high treatment effectiveness and low complication rate were the most desirable treatment attributes to parents, though parents likely have heterogenous treatment preference structures. Shared parent-physician decision-making that incorporates parental preferences will likely allow more effective, targeted decision-making in the future.

A possible mechanism underlying mood disorders associated with LUTS: Chronic bladder outlet obstruction causes NLRP3-dependent inflammation in the hippocampus and depressive behavior in rats.

AIMS: Reports link urinary dysfunction and mood disorders, such as depression, but a causative mechanism has never been postulated. Contemporary discoveries demonstrate a local inflammatory response in peripheral organs can trigger inflammation in the brain, particularly the hippocampus, mediated through the NLRP3 inflammasome. Critically, central inflammation causes depressive behavior. Since bladder outlet obstruction (BOO) evokes a local inflammatory response in the bladder, we hypothesize it will induce NLRP3-dependent inflammation in the hippocampus and depressive behavior. METHODS: There were four groups of rats: control, sham, BOO, or BOO + glyburide (an NLRP3 inhibitor). BOO was created by urethral ligation over a 1 mm catheter. Sham was tied loosely. Glyburide was provided by slow-release pellet (subcutaneous 50 mg, 21 day, replaced as needed). Rats were analyzed 12 weeks post-op for: hippocampal inflammation, microglial density, neurogenesis, and depression symptoms (open field and sucrose preference). RESULTS: BOO elicited hippocampal inflammation, accompanied by an increase in activated microglia (22%) and a decrease in neurogenesis (35%), which was blocked by glyburide. In addition, BOO rats displayed anxiety (57% decrease in exploratory behavior in the open field assay) and anhedonia (21% decrease in sucrose preference), two symptoms of depression. Like inflammation, these symptoms were diminished by glyburide to levels not statistically significantly different from controls. CONCLUSIONS: BOO, a bladder-localized event, stimulates NLRP3-dependent inflammation in the rat hippocampus after 12 weeks and this inflammation causes depressive behavior. This is the first mechanistic explanation of the link between BOO and depression and provides evidence for a distinct bladder-brain axis.