Streamlined identification of clinically and functionally relevant genetic regulators of lower-tract urogenital development.

Congenital anomalies of the lower genitourinary (LGU) tract are frequently comorbid due to genetically linked developmental pathways, and are among the most common yet most socially stigmatized congenital phenotypes. Genes involved in sexual differentiation are prime candidates for developmental anomalies of multiple LGU organs, but insufficient prospective screening tools have prevented the rapid identification of causative genes. Androgen signaling is among the most influential modulators of LGU development. The present study uses SpDamID technology in vivo to generate a comprehensive map of the pathways actively regulated by the androgen receptor (AR) in the genitalia in the presence of the p300 coactivator, identifying wingless/integrated (WNT) signaling as a highly enriched AR-regulated pathway in the genitalia. Transcription factor (TF) hits were then assayed for sexually dimorphic expression at two critical time points and also cross-referenced to a database of clinically relevant copy number variations to identify 252 TFs exhibiting copy variation in patients with LGU phenotypes. A subset of 54 TFs was identified for which LGU phenotypes are statistically overrepresented as a proportion of total observed phenotypes. The 252 TF hitlist was then subjected to a functional screen to identify hits whose silencing affects genital mesenchymal growth rates. Overlap of these datasets results in a refined list of 133 TFs of both functional and clinical relevance to LGU development, 31 of which are top priority candidates, including the well-documented renal progenitor regulator, Sall1. Loss of Sall1 was examined in vivo and confirmed to be a powerful regulator of LGU development.

PIEZO2 in sensory neurons and urothelial cells coordinates urination.

Henry Miller stated that "to relieve a full bladder is one of the great human joys". Urination is critically important in health and ailments of the lower urinary tract cause high pathological burden. Although there have been advances in understanding the central circuitry in the brain that facilitates urination1-3, there is a lack of in-depth mechanistic insight into the process. In addition to central control, micturition reflexes that govern urination are all initiated by peripheral mechanical stimuli such as bladder stretch and urethral flow4. The mechanotransduction molecules and cell types that function as the primary stretch and pressure detectors in the urinary tract mostly remain unknown. Here we identify expression of the mechanosensitive ion channel PIEZO2 in lower urinary tract tissues, where it is required for low-threshold bladder-stretch sensing and urethral micturition reflexes. We show that PIEZO2 acts as a sensor in both the bladder urothelium and innervating sensory neurons. Humans and mice lacking functional PIEZO2 have impaired bladder control, and humans lacking functional PIEZO2 report deficient bladder-filling sensation. This study identifies PIEZO2 as a key mechanosensor in urinary function. These findings set the foundation for future work to identify the interactions between urothelial cells and sensory neurons that control urination.

Expression and localization of HSD17B13 along mouse urinary tract.

17ß-Hydroxysteroid dehydrogenase-13 (HSD17B13), a newly identified lipid droplet-associated protein, plays an important role in the development of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH). Emerging evidence demonstrates that NASH is an independent risk factor for chronic kidney disease, which is frequently accompanied by renal lipid accumulation. In addition, the HSD17B13 rs72613567 variant is associated with lower levels of albuminuria in patients with biopsy-proven NAFLD. At present, the role of HSD17B13 in lipid accumulation in the kidney is unclear. This study utilized bioinformatic and immunostaining approaches to examine the expression and localization of HSD17B13 along the mouse urinary tract. We found that HSD17B13 is constitutively expressed in the kidney, ureter, and urinary bladder. Our findings reveal for the first time, to our knowledge, the precise localization of HSD17B13 in the mouse urinary system, providing a basis for further studying the pathogenesis of HSD17B13 in various renal and urological diseases.NEW & NOTEWORTHY HSD17B13, a lipid droplet-associated protein, is crucial in nonalcoholic fatty liver disease (NAFLD) development. NAFLD also independently raises chronic kidney disease (CKD) risk, often with renal lipid buildup. However, HSD17B13's role in CKD-related lipid accumulation is unclear. This study makes the first effort to examine HSD17B13 expression and localization along the urinary system, providing a basis for exploring its physiological and pathophysiological roles in the kidney and urinary tract.

Expression and Clinical Significance of Non B Cell-Derived Immunoglobulins in the Urinary System and Male Reproductive System.

The urinary system comprises kidneys, ureters, bladder, and urethra with its primary function being excretion, referring to the physiological process of transporting substances that are harmful or surplus out of the body. The male reproductive system consists of gonads (testis), vas deferens, and accessory glands such as the prostate. According to classical immunology theory, the tissues and organs mentioned above are not thought to produce immunoglobulins (Igs), and any Ig present in the relevant tissues under physiological and pathological conditions is believed to be derived from B cells. For instance, most renal diseases are associated with uncontrolled inflammation caused by pathogenic Ig deposited in the kidney. Generally, these pathological Igs are presumed to be produced by B cells. Recent studies have demonstrated that renal parenchymal cells can produce and secrete Igs, including IgA and IgG. Glomerular mesangial cells can express and secrete IgA, which is associated with cell survival and adhesion. Likewise, human podocytes demonstrate the ability to produce and secrete IgG, which is related to cell survival and adhesion. Furthermore, renal tubular epithelial cells also express IgG, potentially involved in the epithelial-mesenchymal transition (EMT). More significantly, renal cell carcinoma, bladder cancer, and prostate cancer have been revealed to express high levels of IgG, which promotes tumour progression. Given the widespread Ig expression in the urinary and male reproductive systems, continued efforts to elucidate the roles of Igs in renal physiological and pathological processes are necessary.

Immunoexpression Pattern of Autophagy-Related Proteins in Human Congenital Anomalies of the Kidney and Urinary Tract.

The purpose of this study was to evaluate the spatiotemporal immunoexpression pattern of microtubule-associated protein 1 light chain 3 beta (LC3B), glucose-regulated protein 78 (GRP78), heat shock protein 70 (HSP70), and lysosomal-associated membrane protein 2A (LAMP2A) in normal human fetal kidney development (CTRL) and kidneys affected with congenital anomalies of the kidney and urinary tract (CAKUT). Human fetal kidneys (control, horseshoe, dysplastic, duplex, and hypoplastic) from the 18th to the 38th developmental week underwent epifluorescence microscopy analysis after being stained with antibodies. Immunoreactivity was quantified in various kidney structures, and expression dynamics were examined using linear and nonlinear regression modeling. The punctate expression of LC3B was observed mainly in tubules and glomerular cells, with dysplastic kidneys displaying distinct staining patterns. In the control group's glomeruli, LAMP2A showed a sporadic, punctate signal; in contrast to other phenotypes, duplex kidneys showed significantly stronger expression in convoluted tubules. GRP78 had a weaker expression in CAKUT kidneys, especially hypoplastic ones, while normal kidneys exhibited punctate staining of convoluted tubules and glomeruli. HSP70 staining varied among phenotypes, with dysplastic and hypoplastic kidneys exhibiting stronger staining compared to controls. Expression dynamics varied among observed autophagy markers and phenotypes, indicating their potential roles in normal and dysfunctional kidney development.

Endurance-dependent urinary extracellular vesicle signature: shape, metabolic miRNAs, and purine content distinguish triathletes from inactive people.

Extracellular vesicles (EVs) enriched with bioactive molecules have gained considerable attention in nanotechnology because they are critical to intercellular communication while maintaining low immunological impact. Among biological matrices, urine has emerged as a noninvasive source of extracellular-contained liquid biopsy, currently of interest as a readout for physiological adaptations. Therefore, we aimed to evaluate chronic adaptations of endurance sport practice in terms of urinary EV parameters and evaluated by food consumption assessment. Two balanced groups of 13 inactive controls vs. triathlon athletes were enrolled; their urinary EVs were obtained by differential ultracentrifugation and analyzed by dynamic light scattering and transmission electron and atomic force microscopy. The cargo was analyzed by means of purine and miRNA content through HPLC-UV and qRT-PCR. Specific urinary EV signatures differentiated inactive versus endurance-trained in terms of peculiar shape. Particularly, a spheroid shape, smaller size, and lower roughness characterize EVs from triathletes. Metabolic and regulatory miRNAs often associated with skeletal muscle (i.e., miR378a-5p, miR27a-3p, miR133a, and miR206) also accounted for a differential signature. These miRNAs and guanosine in urinary EVs can be used as a readout for metabolic status along with the shape and roughness of EVs, novel informative parameters that are rarely considered. The network models allow scholars to entangle nutritional and exercise factors related to EVs' miRNA and purine content to depict metabolic signatures. All in all, multiplex biophysical and molecular analyses of urinary EVs may serve as promising prospects for research in exercise physiology.

Mutations of the Transcriptional Corepressor ZMYM2 Cause Syndromic Urinary Tract Malformations.

Congenital anomalies of the kidney and urinary tract (CAKUT) constitute one of the most frequent birth defects and represent the most common cause of chronic kidney disease in the first three decades of life. Despite the discovery of dozens of monogenic causes of CAKUT, most pathogenic pathways remain elusive. We performed whole-exome sequencing (WES) in 551 individuals with CAKUT and identified a heterozygous de novo stop-gain variant in ZMYM2 in two different families with CAKUT. Through collaboration, we identified in total 14 different heterozygous loss-of-function mutations in ZMYM2 in 15 unrelated families. Most mutations occurred de novo, indicating possible interference with reproductive function. Human disease features are replicated in X. tropicalis larvae with morpholino knockdowns, in which expression of truncated ZMYM2 proteins, based on individual mutations, failed to rescue renal and craniofacial defects. Moreover, heterozygous Zmym2-deficient mice recapitulated features of CAKUT with high penetrance. The ZMYM2 protein is a component of a transcriptional corepressor complex recently linked to the silencing of developmentally regulated endogenous retrovirus elements. Using protein-protein interaction assays, we show that ZMYM2 interacts with additional epigenetic silencing complexes, as well as confirming that it binds to FOXP1, a transcription factor that has also been linked to CAKUT. In summary, our findings establish that loss-of-function mutations of ZMYM2, and potentially that of other proteins in its interactome, as causes of human CAKUT, offering new routes for studying the pathogenesis of the disorder.

Prevalence and characteristics of patients with upper urinary tract urothelial carcinoma having potential Lynch syndrome identified by immunohistochemical universal screening and Amsterdam criteria II.

BACKGROUND: This study aimed to identify patients with upper urinary tract urothelial carcinoma (UTUC) having potential Lynch syndrome (pLS) by immunohistochemistry (IHC) of DNA mismatch repair gene-related proteins (MMRPs) and Amsterdam criteria II and explore their clinical characteristics. METHODS: We retrospectively collected the clinical data of 150 consecutive patients with UTUC who underwent surgical resection at our institution between February 2012 and December 2020, and immunohistochemistry (IHC) of four MMRPs (MLH1, MSH2, MSH6, and PMS2) on all UTUC specimens was performed. Patients who tested positive for Amsterdam criteria (AMS) II and/or IHC screening were classified as having pLS and others as non-pLS, and their characteristics were explored. RESULTS: In this study, 5 (3%) and 6 (4%) patients were positive for AMS II and IHC screening, respectively. Two patient were positive for both AMS II and IHC screening, resulting in 9 (6%) patients with pLS. The pLS group was predominantly female (67% vs. 36%; p = 0.0093) and had more right-sided tumors (100% vs. 43%; p = 0.0009) than the non-pLS group. Of the 6 patients who were positive for IHC screening, 4 showed a combined loss of MSH2/MSH6 (n = 3) and MLH1/PMS2 (n = 1). Other two patients showed single loss of MSH6 and PSM2. CONCLUSIONS: AMS II and IHC screening identified pLS in 6% of patients with UTUC. The IHC screening-positive group tends to have relatively high rate of combined loss, but some patients have single loss. AMS II may overlook patients with LS, and a universal screening may be required for patients with UTUC as well as those with colorectal and endometrial cancer.

A homozygous truncating ETV4 variant in a Nigerian family with congenital anomalies of the kidney and urinary tract.

Congenital anomalies of the kidney and urinary tract (CAKUT) are the most prevalent cause of chronic kidney disease that manifests in children. To date ~23 different monogenic causes have been implicated in isolated forms of human CAKUT, but the vast majority remains elusive. In a previous study, we identified a homozygous missense variant in E26 transformation-specific (ETS) Variant Transcription Factor 4 (ETV4) causing CAKUT via dysregulation of the transcriptional function of ETV4, and a resulting abrogation of GDNF/RET/ETV4 signaling pathway. This CAKUT family remains the only family with an ETV4 variant reported so far. Here, we describe one additional CAKUT family with a homozygous truncating variant in ETV4 (p.(Lys6*)) that was identified by exome sequencing. The variant was found in an individual with isolated CAKUT displaying posterior urethral valves and renal dysplasia. The newly identified stop variant conceptually truncates the ETS_PEA3_N and ETS domains that regulate DNA-binding transcription factor activity. The variant has never been reported homozygously in the gnomAD database. To our knowledge, we here report the first CAKUT family with a truncating variant in ETV4, potentially causing the isolated CAKUT phenotype observed in the affected individual.

Neural transcription factor Pou4f1 promotes renal fibrosis via macrophage-myofibroblast transition.

Unresolved inflammation can lead to tissue fibrosis and impaired organ function. Macrophage-myofibroblast transition (MMT) is one newly identified mechanism by which ongoing chronic inflammation causes progressive fibrosis in different forms of kidney disease. However, the mechanisms underlying MMT are still largely unknown. Here, we discovered a brain-specific homeobox/POU domain protein Pou4f1 (Brn3a) as a specific regulator of MMT. Interestingly, we found that Pou4f1 is highly expressed by macrophages undergoing MMT in sites of fibrosis in human and experimental kidney disease, identified by coexpression of the myofibroblast marker, α-SMA. Unexpectedly, Pou4f1 expression peaked in the early stage in renal fibrogenesis in vivo and during MMT of bone marrow-derived macrophages (BMDMs) in vitro. Mechanistically, chromatin immunoprecipitation (ChIP) assay identified that Pou4f1 is a Smad3 target and the key downstream regulator of MMT, while microarray analysis defined a Pou4f1-dependent fibrogenic gene network for promoting TGF-ß1/Smad3-driven MMT in BMDMs at the transcriptional level. More importantly, using two mouse models of progressive renal interstitial fibrosis featuring the MMT process, we demonstrated that adoptive transfer of TGF-ß1-stimulated BMDMs restored both MMT and renal fibrosis in macrophage-depleted mice, which was prevented by silencing Pou4f1 in transferred BMDMs. These findings establish a role for Pou4f1 in MMT and renal fibrosis and suggest that Pou4f1 may be a therapeutic target for chronic kidney disease with progressive renal fibrosis.

Nanoparticle Tracking Analysis of Urinary Extracellular Vesicle Proteins as a New Challenge in Laboratory Medicine.

Urinary extracellular vesicle (uEV) proteins may be used as specific markers of kidney damage in various pathophysiological conditions. The nanoparticle-tracking analysis (NTA) appears to be the most useful method for the analysis of uEVs due to its ability to analyze particles below 300 nm. The NTA method has been used to measure the size and concentration of uEVs and also allows for a deeper analysis of uEVs based on their protein composition using fluorescence measurements. However, despite much interest in the clinical application of uEVs, their analysis using the NTA method is poorly described and requires meticulous sample preparation, experimental adjustment of instrument settings, and above all, an understanding of the limitations of the method. In the present work, we demonstrate the usefulness of an NTA. We also present problems encountered during analysis with possible solutions: the choice of sample dilution, the method of the presentation and comparison of results, photobleaching, and the adjustment of instrument settings for a specific analysis. We show that the NTA method appears to be a promising method for the determination of uEVs. However, it is important to be aware of potential problems that may affect the results.

Inflammation-like changes in the urothelium of Lifr-deficient mice and LIFR-haploinsufficient humans with urinary tract anomalies.

Congenital anomalies of the kidney and urinary tract (CAKUT) are the most common cause of end-stage kidney disease in children. While the genetic aberrations underlying CAKUT pathogenesis are increasingly being elucidated, their consequences on a cellular and molecular level commonly remain unclear. Recently, we reported rare heterozygous deleterious LIFR variants in 3.3% of CAKUT patients, including a novel de novo frameshift variant, identified by whole-exome sequencing, in a patient with severe bilateral CAKUT. We also demonstrated CAKUT phenotypes in Lifr-/- and Lifr+/- mice, including a narrowed ureteric lumen due to muscular hypertrophy and a thickened urothelium. Here, we show that both in the ureter and bladder of Lifr-/- and Lifr+/- embryos, differentiation of the three urothelial cell types (basal, intermediate and superficial cells) occurs normally but that the turnover of superficial cells is elevated due to increased proliferation, enhanced differentiation from their progenitor cells (intermediate cells) and, importantly, shedding into the ureteric lumen. Microarray-based analysis of genome-wide transcriptional changes in Lifr-/- versus Lifr+/+ ureters identified gene networks associated with an antimicrobial inflammatory response. Finally, in a reverse phenotyping effort, significantly more superficial cells were detected in the urine of CAKUT patients with versus without LIFR variants indicating conserved LIFR-dependent urinary tract changes in the murine and human context. Our data suggest that LIFR signaling is required in the epithelium of the urinary tract to suppress an antimicrobial response under homeostatic conditions and that genetically induced inflammation-like changes underlie CAKUT pathogenesis in Lifr deficiency and LIFR haploinsufficiency.

Gut Microbiota of Pigs Metabolizes Extracts of Filipendula ulmaria and Orthosiphon aristatus-Herbal Remedies Used in Urinary Tract Disorders.

Urinary tract infections influence the mortality rate in pigs and are linked to extensive antibiotic usage in the farm industry. Filipendula ulmaria (L.) Maxim. and Orthosiphon aristatus (Blume) Miq. are widespread medicinal plants traditionally used to treat urinary tract disorders. As their preparations are orally administered, the metabolism of their constituents by gut microbiota before absorption should be considered. Until now, no experiments had been performed to describe the biotransformation of tthose plants' extracts by animal gut microbiota. The study evaluates the influence of pig intestinal microbiota on the structure of active compounds in flowers of F. ulmaria and leaves of O. aristatus. The incubations of the extracts with piglet gut microbiota were performed in anaerobic conditions, and the samples of the batch culture were collected for 24 h. In F. ulmaria, the main metabolites were quercetin and kaempferol, which were products of the deglycosylation of flavonoids. After 24 h incubation of O. aristatus extract with the piglet gut microbiota, 2 main metabolites were observed. One, tentatively identified as 3-(3-dihydroxyphenyl)propionic acid, is likely the primary metabolite of the most abundant depsides and phenolic acids. The results confirm the formation of the compounds with anti-inflammatory and diuretic activity in the microbiota cultures, which might suggest F. ulmaria and O. aristatus for treating urinary tract disorders in piglets. Based on the similarities of human and pig gut microbiota, the pig model can help estimate the metabolic pathways of natural products in humans.

Non-carbapenemase producing carbapenem-resistant Klebsiella pneumoniae isolated from the urinary tract of a dog.

Objective: Carbapenems are broad-spectrum ß-lactams with excellent activity against multidrug-resistant (MDR) Enterobacterales. Unfortunately, resistance to carbapenems within this bacterial family, known as carbapenem-resistant Enterobacterales (CRE), occurs and challenges the ability to treat difficult MDR infections. Although the impact of carbapenem-resistance has been greatest in human medicine, reports in the veterinary literature are increasing especially as national veterinary antimicrobial resistance surveillance programs are now in place. In this brief communication, we report the isolation of a non-carbapenemase-producing, carbapenem-resistant Klebsiella pneumoniae from the urine of a dog, discuss the likely mechanism of resistance, and wider implications. Animal: Canine. Procedure: Whole genome sequencing and phenotypic antimicrobial susceptibility testing was performed on a K. pneumoniae isolated from the urine of a dog. Results: Antimicrobial susceptibility testing identified phenotypic resistance to imipenem and meropenem. Phenotypic detection of carbapenemase production was negative. Whole genome sequencing identified efflux pump genes associated with carbapenem resistance and point mutations in membrane porin genes. No carbapenemase gene was identified. Conclusion: Phenotypic antimicrobial susceptibility testing identified the K. pneumoniae as a non-carbapenemase producing carbapenem-resistant organism with the proposed genotypic mechanism including alteration of efflux pumps and membrane porin activity and/or expression. Clinical significance: Currently, there is limited use of carbapenem antimicrobial drugs in veterinary medicine, and practitioners may be unfamiliar or unaware of this type of resistance, its significance on routine antimicrobial susceptibility test reports, and implications for antimicrobial therapy and public health. Carbapenem-resistant Enterobacterales are infrequently isolated from companion animals; however, due to increasing adoption of advanced medical and surgical interventions, they may become more prevalent.

Objectif: Les carbapénèmes sont des ß-lactamines à large spectre avec une excellente activité contre les Enterobacterales multirésistantes (MDR). Malheureusement, la résistance aux carbapénèmes au sein de cette famille bactérienne, connue sous le nom d'Enterobacterales résistantes aux carbapénèmes (CRE), se produit et remet en question la capacité de traiter les infections MDR difficiles. Bien que l'impact de la résistance aux carbapénèmes ait été plus important en médecine humaine, les rapports dans la littérature vétérinaire se multiplient, d'autant plus que des programmes nationaux de surveillance de la résistance aux antimicrobiens vétérinaires sont désormais en place. Dans cette brève communication, nous rapportons l'isolement d'une Klebsiella pneumoniae non-productrice de carbapénémase et résistante aux carbapénèmes à partir de l'urine d'un chien, discutons du mécanisme probable de résistance et des implications plus larges. Animal: Canin. Procédure: Le séquençage du génome entier et les tests de sensibilité phénotypique aux antimicrobiens ont été effectués sur un isolat de K. pneumoniae provenant de l'urine d'un chien. Résultats: Les tests de sensibilité aux antimicrobiens ont identifié une résistance phénotypique à l'imipénème et au méropénème. La détection phénotypique de production de carbapénèmase était négative. Le séquençage du génome entier a identifié des gènes de pompe à efflux associés à la résistance aux carbapénèmes et à des mutations ponctuelles dans les gènes des porines membranaires. Aucun gène de carbapénémase n'a été identifié. Conclusion: Les tests de sensibilité phénotypique aux antimicrobiens ont identifié cet isolat de K. pneumoniae comme un organisme résistant aux carbapénèmes ne produisant pas de carbapénémase avec le mécanisme génotypique proposé, y compris l'altération des pompes à efflux et l'activité et/ou l'expression de porines membranaires. Signification clinique: Actuellement, l'utilisation des médicaments antimicrobiens à base de carbapénème en médecine vétérinaire est limitée, et les praticiens peuvent ne pas être familiers ou ne pas être au fait de ce type de résistance, de son importance dans les rapports de routine sur les tests de sensibilité aux antimicrobiens et de ses implications pour la thérapie antimicrobienne et la santé publique. Les Enterobacterales résistantes aux carbapénèmes sont rarement isolées des animaux de compagnie; cependant, en raison de l'adoption croissante d'interventions médicales et chirurgicales avancées, elles peuvent devenir plus répandues.(Traduit par Dr Serge Messier).

The clonal relation of primary upper urinary tract urothelial carcinoma and paired urothelial carcinoma of the bladder.

The risk of developing urothelial carcinoma of the bladder (UCB) in patients treated by radical nephroureterectomy (RNU) for an upper urinary tract urothelial carcinoma (UTUC) is 22% to 47% in the 2 years after surgery. Subject of debate remains whether UTUC and the subsequent UCB are clonally related or represent separate origins. To investigate the clonal relationship between both entities, we performed targeted DNA sequencing of a panel of 41 genes on matched normal and tumor tissue of 15 primary UTUC patients treated by RNU who later developed 19 UCBs. Based on the detected tumor-specific DNA aberrations, the paired UTUC and UCB(s) of 11 patients (73.3%) showed a clonal relation, whereas in four patients the molecular results did not indicate a clear clonal relationship. Our results support the hypothesis that UCBs following a primary surgically resected UTUC are predominantly clonally derived recurrences and not separate entities.

Homozygous WNT9B variants in two families with bilateral renal agenesis/hypoplasia/dysplasia.

WNT9B plays a key role in the development of the mammalian urogenital system. It is essential for the induction of mesonephric and metanephric tubules, the regulation of renal tubule morphogenesis, and the regulation of renal progenitor cell expansion and differentiation. To our knowledge, WNT9B has not been associated with renal defects in humans; however, WNT9B-/- mice have renal agenesis/hypoplasia and reproductive tract abnormalities. We report four individuals from two unrelated consanguineous families with bilateral renal agenesis/hypoplasia/dysplasia and homozygous variants in WNT9B. The proband from Family 1 has bilateral renal cystic dysplasia and chronic kidney disease. He has two deceased siblings who presented with bilateral renal hypoplasia/agenesis. The three affected family members were homozygous for a missense variant in WNT9B (NM_003396.2: c.949G>A/p.(Gly317Arg)). The proband from Family 2 has renal hypoplasia/dysplasia, chronic kidney disease, and is homozygous for a nonsense variant in WNT9B (NM_003396.2: c.11dupC/p.(Pro5Alafs*52)). Two of her siblings died in the neonatal period, one confirmed to be in the context of oligohydramnios. The proband's unaffected brother is also homozygous for the nonsense variant in WNT9B, suggesting nonpenetrance. We propose a novel association of WNT9B and renal anomalies in humans. Further study is needed to delineate the contribution of WNT9B to genitourinary anomalies in humans.

Role of α- and ß-adrenergic signaling in phenotypic targeting: significance in benign and malignant urologic disease.

The urinary tract is highly innervated by autonomic nerves which are essential in urinary tract development, the production of growth factors, and the control of homeostasis. These neural signals may become dysregulated in several genitourinary (GU) disease states, both benign and malignant. Accordingly, the autonomic nervous system is a therapeutic target for several genitourinary pathologies including cancer, voiding dysfunction, and obstructing nephrolithiasis. Adrenergic receptors (adrenoceptors) are G-Protein coupled-receptors that are distributed throughout the body. The major function of α1-adrenoceptors is signaling smooth muscle contractions through GPCR and intracellular calcium influx. Pharmacologic intervention of α-and ß-adrenoceptors is routinely and successfully implemented in the treatment of benign urologic illnesses, through the use of α-adrenoceptor antagonists. Furthermore, cell-based evidence recently established the antitumor effect of α1-adrenoceptor antagonists in prostate, bladder and renal tumors by reducing neovascularity and impairing growth within the tumor microenvironment via regulation of the phenotypic epithelial-mesenchymal transition (EMT). There has been a significant focus on repurposing the routinely used, Food and Drug Administration-approved α1-adrenoceptor antagonists to inhibit GU tumor growth and angiogenesis in patients with advanced prostate, bladder, and renal cancer. In this review we discuss the current evidence on (a) the signaling events of the autonomic nervous system mediated by its cognate α- and ß-adrenoceptors in regulating the phenotypic landscape (EMT) of genitourinary organs; and (b) the therapeutic significance of targeting this signaling pathway in benign and malignant urologic disease. Video abstract.

In Vivo and In Vitro Evaluation of Urinary Biomarkers in Ischemia/Reperfusion-Induced Kidney Injury.

Oxidative stress plays an important role in the pathophysiology of acute kidney injury (AKI). Previously, we reported that vanin-1, which is involved in oxidative stress, is associated with renal tubular injury. This study was aimed to determine whether urinary vanin-1 is a biomarker for the early diagnosis of AKI in two experimental models: in vivo and in vitro. In a rat model of AKI, ischemic AKI was induced in uninephrectomized rats by clamping the left renal artery for 45 min and then reperfusing the kidney. On Day 1 after renal ischemia/reperfusion (I/R), serum creatinine (SCr) in I/R rats was higher than in sham-operated rats, but this did not reach significance. Urinary N-acetyl-ß-D-glucosaminidase (NAG) exhibited a significant increase but decreased on Day 2 in I/R rats. In contrast, urinary vanin-1 significantly increased on Day 1 and remained at a significant high level on Day 2 in I/R rats. Renal vanin-1 protein decreased on Days 1 and 3. In line with these findings, immunofluorescence staining demonstrated that vanin-1 was attenuated in the renal proximal tubules of I/R rats. Our in vitro results confirmed that the supernatant from HK-2 cells under hypoxia/reoxygenation included significantly higher levels of vanin-1 as well as KIM-1 and NGAL. In conclusion, our results suggest that urinary vanin-1 might be a potential novel biomarker of AKI induced by I/R.

5-HT3 Signaling Alters Development of Sacral Neural Crest Derivatives That Innervate the Lower Urinary Tract.

The autonomic nervous system derives from the neural crest (NC) and supplies motor innervation to the smooth muscle of visceral organs, including the lower urinary tract (LUT). During fetal development, sacral NC cells colonize the urogenital sinus to form pelvic ganglia (PG) flanking the bladder neck. The coordinated activity of PG neurons is required for normal urination; however, little is known about the development of PG neuronal diversity. To discover candidate genes involved in PG neurogenesis, the transcriptome profiling of sacral NC and developing PG was performed, and we identified the enrichment of the type 3 serotonin receptor (5-HT3, encoded by Htr3a and Htr3b). We determined that Htr3a is one of the first serotonin receptor genes that is up-regulated in sacral NC progenitors and is maintained in differentiating PG neurons. In vitro cultures showed that the disruption of 5-HT3 signaling alters the differentiation outcomes of sacral NC cells, while the stimulation of 5-HT3 in explanted fetal pelvic ganglia severely diminished neurite arbor outgrowth. Overall, this study provides a valuable resource for the analysis of signaling pathways in PG development, identifies 5-HT3 as a novel regulator of NC lineage diversification and neuronal maturation in the peripheral nervous system, and indicates that the perturbation of 5-HT3 signaling in gestation has the potential to alter bladder function later in life.

Intrauterine low-protein diet aggravates developmental abnormalities of the urinary system via the Akt/Creb3 pathway in Robo2 mutant mice.

The offspring of Robo2 mutant mice usually present with variable phenotypes of congenital anomalies of the kidney and urinary tract (CAKUT). An intrauterine low-protein diet can also cause CAKUT in offspring, dominated by the duplicated collecting system phenotype. A single genetic or environment factor can only partially explain the pathogenesis of CAKUT. The present study aimed to establish an intrauterine low-protein diet roundabout 2 (Robo2) mutant mouse model and found that the intrauterine low-protein diet led to significantly increased CAKUT phenotypes in Robo2PB/+ mice offspring, dominant by a duplicated collecting system. At the same time, more ectopic and lower located ureteric buds (UBs) were observed in the intrauterine low-protein diet-fed Robo2 mutant mouse model, and the number of UB branches was reduced in the serum-free culture. During UB protrusion, intrauterine low-protein diet reduced the expression of Slit2/Robo2 in Robo2 mutant mice and affected the expression of glial cell-derived neurotrophic factor/Ret, which is a key molecule for metanephric development, with increasing phospho-Akt and phospho-cAMP responsive element-binding protein 3 activity and a reduction of apoptotic cells in embryonic day 11.5 UB tissues. The mechanism by which an intrauterine low-protein diet aggravates CAKUT in Robo2 mutant mice may be related to the disruption of Akt/cAMP responsive element-binding protein 3 signaling and a reduction in apoptosis in UB tissue.