Copy Number Variant Analysis and Genome-wide Association Study Identify Loci with Large Effect for Vesicoureteral Reflux.

BACKGROUND: Vesicoureteral reflux (VUR) is a common, familial genitourinary disorder, and a major cause of pediatric urinary tract infection (UTI) and kidney failure. The genetic basis of VUR is not well understood. METHODS: A diagnostic analysis sought rare, pathogenic copy number variant (CNV) disorders among 1737 patients with VUR. A GWAS was performed in 1395 patients and 5366 controls, of European ancestry. RESULTS: Altogether, 3% of VUR patients harbored an undiagnosed rare CNV disorder, such as the 1q21.1, 16p11.2, 22q11.21, and triple X syndromes ((OR, 3.12; 95% CI, 2.10 to 4.54; P=6.35×10-8) The GWAS identified three study-wide significant and five suggestive loci with large effects (ORs, 1.41-6.9), containing canonical developmental genes expressed in the developing urinary tract (WDPCP, OTX1, BMP5, VANGL1, and WNT5A). In particular, 3.3% of VUR patients were homozygous for an intronic variant in WDPCP (rs13013890; OR, 3.65; 95% CI, 2.39 to 5.56; P=1.86×10-9). This locus was associated with multiple genitourinary phenotypes in the UK Biobank and eMERGE studies. Analysis of Wnt5a mutant mice confirmed the role of Wnt5a signaling in bladder and ureteric morphogenesis. CONCLUSIONS: These data demonstrate the genetic heterogeneity of VUR. Altogether, 6% of patients with VUR harbored a rare CNV or a common variant genotype conferring an OR >3. Identification of these genetic risk factors has multiple implications for clinical care and for analysis of outcomes in VUR.

Teratogenesis in the chick embryo following post-gastrulation exposure to Y-27632 -effect of Y-27632 on embryonic development.

The pyridine derivative Y-27632 inhibits Rho-associated coiled-coil-containing protein kinase (ROCK) signaling, which is involved in numerous developmental processes during embryogenesis, primarily by controlling actin-cytoskeleton assembly and cell contractility. Somite formation requires rearrangement of the cytoskeleton and assists in major morphological mechanisms, including ventral body wall formation. Administration of Y-27632 impairs cytoskeletal arrangements in post-gastrulation chick embryos leading to ventral body wall defects (VBWD) at later stages of development. The aim of this study was to investigate the effect of Y-27632 on somite development in post-gastrulation chick embryos during early embryogenesis. After 60 h incubation, embryos in shell-less culture were treated with Y-27632 or vehicle for controls. Following administration, abnormality rates were assessed. In treatment groups, embryos showed a kinked longitudinal body axis. Western blot confirmed impaired ROCK downstream signaling by decreased expression of phosphorylated cofilin-2. Histology, Lysotracker studies and RT-PCR demonstrated increased cell death in somites, the neural tube and the ectoderm. RT-PCR and Western blot of factors known to be involved during somitogenesis revealed reduced expression in the treatment group compared to controls. We hypothesize that administration of Y-27632 disrupts somite development causing axial kinking and embryo malformation, which may lead to VBWD.

Concurrent Hirschsprung's disease and anorectal malformation: a systematic review.

BACKGROUND/PURPOSE: Hirschsprung's disease (HSCR) and anorectal malformation (ARM) are often associated with other congenital malformations, but the association of each other is rare. Some studies have reported the incidence of HSCR associated with ARM ranging from 2.0 to 3.4%. The purpose of this study was to update the current epidemiological and therapeutic features of this rare congenital association. METHODS: A systematic literature search for relevant articles was performed in four databases using a combination of the following terms "association of Hirschsprung's disease and anorectal malformation", "aganglionosis and anorectal malformation" and "congenital megacolon and anorectal malformation" for studies published between 1952 and 2019. Reference lists were screened for additional cases. RESULTS: Forty-three studies met the defined inclusion criteria, reporting a total of 126 patients who were diagnosed with HSCR with ARM. Thirty articles reported 42 single case reports of this association. Twelve articles reported 66 cases of HSCR in case series of 3309 ARM patients, resulting in an incidence of 2% of this association. Associated syndrome was found in 25 cases (20%): Currarino syndrome in 11, Down syndrome in 8, Cat eye syndrome in 4 and Pallister-Hall syndrome in 2 patients. Extent of aganglionosis was reported in 62 cases: short or rectosigmoid aganglionosis was reported in 44, long segment aganglionosis in 8, total colonic aganglionosis in 9 and total intestinal aganglionosis in 1 case. CONCLUSION: Although the association of ARM and HSCR is rare, the incidence of HSCR among ARM cases seems to be higher than in the general pediatric population. There was a high incidence of coexistence of ARM and HSCR with severe associated syndromes.

Transgenic animal models of congenital diaphragmatic hernia: a comprehensive overview of candidate genes and signaling pathways.

Congenital diaphragmatic hernia (CDH) is a relatively common and life-threatening birth defect, characterized by incomplete formation of the diaphragm. Because CDH herniation occurs at the same time as preacinar airway branching, normal lung development becomes severely disrupted, resulting almost invariably in pulmonary hypoplasia. Despite various research efforts over the past decades, the pathogenesis of CDH and associated lung hypoplasia remains poorly understood. With the advent of molecular techniques, transgenic animal models of CDH have generated a large number of candidate genes, thus providing a novel basis for future research and treatment. This review article offers a comprehensive overview of genes and signaling pathways implicated in CDH etiology, whilst also discussing strengths and limitations of transgenic animal models in relation to the human condition.

Accidental bladder injury during elective inguinal hernia repair: a preventable complication with high morbidity.

INTRODUCTION: Bladder injury (BI) represents a rare complication of inguinal hernia surgery. Protrusions of the urinary bladder through the deep inguinal ring ("bladder ears") have been reported with an incidence of 9% in infants younger than 6 months of age and may be misinterpreted as the hernia sac. This literature review was designed to determine incidence and outcomes of bladder injuries during pediatric inguinal hernia repair. METHODS: A literature review of the literature (1967-2017) was performed using the keywords "bladder ears", "inguinal hernia", "iatrogenic bladder injury" and "bladder hernia". Publications were reviewed for epidemiology, presentation and extent of injury, treatment and outcome. RESULTS: Thirteen articles reporting on 30 cases of BI during inguinal hernia repair from 1967 to 2017 were included (19 boys, 2 girls, 9 unknown). Median age at herniotomy was 10.5 months (1 month-6 years). Out of 30 children, 14 (47%) experienced mild complications. Sixteen patients (53%) had severe complications after initial surgery and needed revisional surgery. Complications were noticed up to 4 years after the initial surgery. In 9 (56%) of the 16 severe cases, major damage to the bladder wall and impairment of bladder capacity occurred. In seven patients (44%), secondary closure was successful. In ten patients (63%), the bladder was partially resected, and in one child (6%), the entire bladder was removed. CONCLUSIONS: The degree of accidental BI during inguinal hernia repair was severe in in the majority of reported cases in the literature. Surgeons should be aware of the high prevalence of "bladder ears" in infants to prevent injury to the urinary tract.

Ureteral Obstruction After Endoscopic Treatment of Vesicoureteral Reflux: Does the Type of Injected Bulking Agent Matter?

PURPOSE OF REVIEW: Endoscopic injection of bulking agents for the treatment of vesicoureteral reflux (VUR) has become a therapeutic alternative to antibiotic prophylaxis and ureteral reimplantation. Although considered as a safe and efficient procedure, several studies have reported cases of ureteral obstruction (UO) after endoscopic correction of VUR. This review article evaluates the present VUR literature to estimate the incidence of UO following endoscopic injection of different substances, while also discussing the impact of injection technique and implant volume. RECENT FINDINGS: Twenty-five publications were identified that provided detailed information on 64 females and 32 males (age range, 7 months-48 years) that developed UO after endoscopic treatment of VUR using dextranomer/hyaluronic acid (Dx/HA), polyacrylate polyalcohol (PP), polydimethylsiloxane (PDMS), calcium hydroxyapatite (CaHA), polytetrafluoroethylene (PTFE), or collagen. There was some variation in the reported incidence of UO among these materials: Dx/HA (0.5-6.1%), PP (1.1-1.6%), PDMS (2.5-10.0%), CaHA (1.0%), and PTFE (0.3%). Postoperative UO was described following subureteric transurethral injection (STING), intraureteric hydrodistension implantation technique (HIT), combined HIT/STING and double HIT. The injected volume ranged widely, also depending on the type of bulking agent: Dx/HA (0.3-3.0 mL), PP (0.3-1.2 mL), PDMS (1.0-2.2 mL), CaHA (0.4-0.6 mL), and PTFE (1.5-2.0 mL). The timing of UO varied from immediately after the procedure to 63 months. Over half of patients showed asymptomatic hydroureteronephrosis on follow-up imaging, whereas the remaining presented with symptoms of acute UO or fever. UO remains a rare complication after endoscopic correction of VUR, generally reported in less than 1% of treated cases, which appears to be independent of the injected substance, volume, and technique. However, long-term follow-up is recommended as asymptomatic or delayed UO can occur, potentially leading to deterioration of renal function.

NADPH oxidase-derived H2O2 subverts pathogen signaling by oxidative phosphotyrosine conversion to PB-DOPA.

Strengthening the host immune system to fully exploit its potential as antimicrobial defense is vital in countering antibiotic resistance. Chemical compounds released during bidirectional host-pathogen cross-talk, which follows a sensing-response paradigm, can serve as protective mediators. A potent, diffusible messenger is hydrogen peroxide (H2O2), but its consequences on extracellular pathogens are unknown. Here we show that H2O2, released by the host on pathogen contact, subverts the tyrosine signaling network of a number of bacteria accustomed to low-oxygen environments. This defense mechanism uses heme-containing bacterial enzymes with peroxidase-like activity to facilitate phosphotyrosine (p-Tyr) oxidation. An intrabacterial reaction converts p-Tyr to protein-bound dopa (PB-DOPA) via a tyrosinyl radical intermediate, thereby altering antioxidant defense and inactivating enzymes involved in polysaccharide biosynthesis and metabolism. Disruption of bacterial signaling by DOPA modification reveals an infection containment strategy that weakens bacterial fitness and could be a blueprint for antivirulence approaches.

Decreased expression of TRAAK channels in Hirschsprung's disease: a possible cause of postoperative dysmotility.

AIM OF THE STUDY: Potassium (K+) channels with a two-pore domain (K2P) are a large family of hyperpolarising ion channels which play a key role in cell excitability. This family comprises three members: TREK-1, TREK-2 and TRAAK. TRAAK channels have previously been reported to be expressed in murine enteric ganglia. To date, no data exist regarding TRAAK channel expression in the human colon. Thus, we designed this study to investigate TRAAK gene expression in the normal human colon and in Hirschsprung's disease (HSCR). METHODS: HSCR tissue specimens (n = 6) were collected at the time of pull-through surgery, while control samples were obtained at the time of colostomy closure in patients with imperforate anus (n = 6). qRT-PCR analysis was undertaken to quantify TRAAK gene expression, and immunolabelling of TRAAK proteins was visualized using confocal microscopy. MAIN RESULTS: Confocal microscopy revealed TRAAK protein expression within both neurons and interstitial cells of Cajal in the myenteric plexus, with a reduction in both ganglionic HSCR colon and aganglionic HSCR colon, compared to controls. qRT-PCR analysis revealed a significant downregulation of the TRAAK gene in both aganglionic and ganglionic HSCR specimens compared to controls (p < 0.05). CONCLUSIONS: TRAAK gene expression is significantly downregulated in HSCR colon, suggesting a role for these ion channels in colonic neurotransmission. TRAAK downregulation within ganglionic specimens highlights the dysfunctional nature of ganglia in this region.

Consanguinity and its relevance for the incidence of megacystis microcolon intestinal hypoperistalsis syndrome (MMIHS): systematic review.

BACKGROUND/PURPOSE: Megacystis microcolon intestinal hypoperistalsis syndrome (MMIHS) is a rare congenital and generally fatal cause of functional intestinal obstruction in the newborn. The cause of this syndrome is unknown. Familial occurrence and reports of consanguinity in MMIHS implies that genetic factors may have an important role in the pathogenesis of this syndrome. The aim of the study was to determine the consequence of consanguinity for the incidence of MMIHS. METHODS: A literature search was performed using the keywords "megacystis microcolon intestinal hypoperistalsis" for studies published between 1976 and 2018. Retrieved articles, including additional studies from reference lists, were reviewed for consanguinity between parents and recurrence of MMIHS between siblings. Data were extracted for cases where familial MMIHS was present. RESULTS: A total of 450 patients with the diagnosis of MMIHS have been reported in the literature. There were 56 (12%) cases in which familial MMIHS was confirmed, 25 families with multiple siblings and 3 families with single affected infant. Of the 25 families with multiple siblings, 22 families had 2 siblings with confirmed MMIHS and 3 families had 3 children each with MMIHS. Consanguinity between parents was confirmed in 30 cases (18 siblings and 12 individual cases). Female-to-male ratio in the 30 patients was 4.4:1. CONCLUSION: The occurrence of MMIHS in the offspring of consanguineous parents and recurrence in siblings of healthy parents suggest that MMIHS is an autosomal recessive disorder. Pre-marital and pre-conception counselling of consanguineous populations is recommended to prevent harmful consequences.

Expression of dispatched RND transporter family member 1 is decreased in the diaphragmatic and pulmonary mesenchyme of nitrofen-induced congenital diaphragmatic hernia.

PURPOSE: Congenital diaphragmatic hernia (CDH) and associated pulmonary hypoplasia (PH) are thought to be caused by a malformation of the diaphragmatic and pulmonary mesenchyme. Dispatched RND transporter family member 1 (Disp-1) encodes a transmembrane protein that regulates the release of cholesterol and palmitoyl, which is critical for normal diaphragmatic and airway development. Disp-1 is strongly expressed in mesenchymal compartments of fetal diaphragms and lungs. Recently, Disp-1 mutations have been identified in patients with CDH. We hypothesized that diaphragmatic and pulmonary Disp-1 expression is decreased in the nitrofen-induced CDH model. METHODS: Time-mated rats received nitrofen or vehicle on gestational day 9 (D9). Fetal diaphragms and lungs were microdissected on selected endpoints D13, D15 and D18; and divided into control and nitrofen-exposed specimens (n = 12 per sample, time-point and experimental group). Diaphragmatic and pulmonary Disp-1 expression was evaluated by qRT-PCR. Immunofluorescence double staining for Disp-1 was combined with diaphragmatic and pulmonary mesenchymal markers Wt-1 and Sox-9 to localize protein expression in fetal diaphragms and lungs. RESULTS: Relative mRNA levels of Disp-1 were significantly decreased in pleuroperitoneal folds/primordial lungs on D13 (0.18 ± 0.08 vs. 0.46 ± 0.41; p < 0.05/1.06 ± 0.27 vs. 1.34 ± 0.79; p < 0.05), developing diaphragms/lungs on D15 (0.18 ± 0.06 vs. 0.44 ± 0.23; p < 0.05/0.73 ± 0.36 vs. 1.16 ± 0.27; p < 0.05) and fully muscularized diaphragms/differentiated lungs on D18 (0.22 ± 0.18 vs. 0.32 ± 0.23; p < 0.05/0.56 ± 0.16 vs. 0.77 ± 0.14; p < 0.05) of nitrofen-exposed fetuses compared to controls. Confocal laser scanning microscopy demonstrated markedly diminished Disp-1 immunofluorescence predominately in the diaphragmatic and pulmonary mesenchyme of nitrofen-exposed fetuses on D13, D15 and D18, associated with a clear reduction of proliferating mesenchymal cells. CONCLUSIONS: Decreased Disp-1 expression during diaphragmatic development and lung branching morphogenesis may interrupt mesenchymal cell proliferation, thus leading to diaphragmatic defects and PH in the nitrofen-induced CDH model.

Altered ryanodine receptor gene expression in Hirschsprung's disease.

AIM OF THE STUDY: Ryanodine receptors are the largest of all ion channels, named after their exogenous ligand, ryanodine. The ryanodine receptor calcium release channel is central to cytoplasmic Ca2+ signalling in skeletal muscle, the heart, and many other tissues, playing a vital role in muscular contraction. Three ryanodine receptors exist, Ryr1, Ryr2 and Ryr3. The ryanodine receptor, Ryr3, is encoded by the Ryr3 gene, which has been reported to be highly specific to colonic smooth muscle cells in mice. We designed this study to investigate Ryr1, Ryr2 and Ryr3 gene expression in the normal human colon and in Hirschsprung's disease (HSCR). METHODS: HSCR tissue specimens (n = 6) were collected at the time of pull-through surgery, while control samples were obtained at the time of colostomy closure in patients with imperforate anus (n = 6). qRT-PCR analysis was undertaken to quantify Ryr1, Ryr2 and Ryr3 gene expression, and immunolabelling of Ryr1, Ryr2 and Ryr3 proteins was visualised using confocal microscopy. MAIN RESULTS: qRT-PCR analysis revealed a significant downregulation of the Ryr1 and Ryr3 genes in both aganglionic and ganglionic HSCR specimens compared to controls (p < 0.05). Confocal microscopy revealed Ryr1, Ryr2 and Ryr3 protein expressions within the smooth muscle, with a reduction in aganglionic and ganglionic HSCR colon compared to controls. CONCLUSIONS: Ryr1 and Ryr3 gene expression is significantly downregulated in HSCR colon, suggesting a role for these genes in colonic smooth-muscle motility. Ryr1 and Ryr3 downregulations within ganglionic specimens highlight the physiologically abnormal nature of this segment which may explain the occurrence of persistent bowel symptoms in some HSCR patients following a properly performed pull-through operation.

Abnormal Scn1b and Fxyd1 gene expression in the pulled-through ganglionic colon may influence functional outcome in patients with Hirschsprung's disease.

PURPOSE: Smooth muscle cells are electrically coupled to ICC and PDGFRα+ cells, to regulate smooth muscle contraction. Recent studies have reported that the voltage-gated sodium channel type 1ß (Scn1b), and the chloride channel subunit, Fxyd1, are highly expressed by both ICC and PDGFRα+ cells in the mouse colon. We designed this study to investigate the expression of the Scn1b and Fxyd1 genes in the normal human colon and in HSCR. METHODS: HSCR tissue specimens (n = 6) were collected at the time of pull-through surgery, while control samples were obtained at the time of colostomy closure in patients with imperforate anus (n = 6). qRT-PCR analysis was undertaken to quantify Scn1b and Fxyd1 gene expression, and immunolabelling of Scn1b and Fxyd1 proteins were visualized using confocal microscopy. RESULTS: qRT-PCR analysis revealed significant downregulation of Scn1b and Fxyd1 genes in both aganglionic and ganglionic HSCR specimens compared to controls (p < 0.05). Confocal microscopy revealed a reduction in Scn1b and Fxyd1 protein expression in both aganglionic and ganglionic HSCR colon compared to controls. CONCLUSION: Scn1b and Fxyd1 expression was significantly downregulated in HSCR colon. These results add to mounting evidence suggesting that the pulled-through ganglionic segment of bowel in these patients is abnormal, despite the presence of ganglion cells.

Altered expression of KCNG3 and KCNG4 in Hirschsprung's disease.

PURPOSE: Voltage-gated potassium ion channels have long been implicated in gastrointestinal motility. Recent studies have highlighted the role of voltage-gated channel subfamily G member 3 (KCNG3) and 4 (KCNG4) genes in the electrical functioning of interstitial cells of Cajal and PDGFRα+ cells of the mouse colon. We designed this study to investigate KCNG3 and KCNG4 expression in the normal human colon and in Hirschsprung's disease (HSCR). METHODS: HSCR tissue specimens (n = 6) were collected at the time of pull-through surgery, while control samples were obtained at the time of colostomy closure in patients with imperforate anus (n = 6). qRT-PCR analysis was undertaken to quantify KCNG3 and KCNG4 gene expression, and immunolabelling of KCNG3 and KCNG4 proteins was visualized using confocal microscopy. RESULTS: qRT-PCR analysis revealed significant downregulation of the KCNG3 and KCNG4 genes in both aganglionic and ganglionic HSCR specimens compared to controls (p < 0.05). Confocal microscopy revealed KCNG3 and KCNG4 expression within neurons, ICC and PDGFRα+ cells of the myenteric plexus and smooth muscle layers, with a reduction in both proteins in aganglionic and ganglionic HSCR colon compared to controls. CONCLUSION: KCNG3 and KCNG4 gene expression is significantly downregulated in HSCR colon, suggesting a role for these genes in colonic motility. KCNG3 and KCNG4 downregulation within ganglionic specimens highlights the physiologically abnormal nature of this segment in HSCR patients.

Altered expression of inflammasomes in Hirschsprung's disease.

AIM OF THE STUDY: The pathogenesis of Hirschsprung's disease-associated enterocolitis (HAEC) is poorly understood. Inflammasomes are a large family of multiprotein complexes that act to mediate host immune responses to microbial infection and have a regulatory or conditioning influence on the composition of the microbiota. Inflammasomes and the apoptosis-associated speck-like protein (ASC) lead to caspase-1 activation. The activated caspase-1 promotes secretion of pro-inflammatory cytokines (IL-1ß and IL-18) from their precursors (pro-IL-1ß and pro-IL-18). Inflammasomes have been implicated in a host of inflammatory disorders. Among the inflammasomes, NLRP3, NLRP12 and NLRC4 are the most widely investigated. Knock-out mice models of inflammasomes NLRP3, NLRP12, NLRC4, caspase-1 and ASC are reported to have higher susceptibility to experimental colitis. The purpose of this study was to investigate the expression of NLRP3, NLRP12, NLRC4, caspase-1, ASC, pro-IL-1ß and pro-IL-18 in the bowel specimens from patients with HSCR and controls. METHODS: Pulled-through colonic specimens were collected from HSCR patients (n = 6) and healthy controls from the proximal colostomy of children with anorectal malformations (n = 6). The gene expression of NLRP3, NLRP12, NLRC4, caspase-1, ASC, pro-IL-1ß and pro-IL-18 was assessed using qPCR. The protein distribution was assessed using immunofluorescence and confocal microscopy. MAIN RESULTS: qRT-PCR analysis revealed that NLRP3, NLRP12, NLRC4, ASC and pro-IL-1ß gene expressions was significantly downregulated in the aganglionic and ganglionic colon of patients with HSCR compared to controls. Confocal microscopy revealed a markedly decreased expression of NLRP3, NLRP12, NLRC4 and ASC protein in the colonic epithelium of aganglionic and ganglionic bowel of patients with HSCR compared to controls. CONCLUSIONS: To our knowledge, this is the first study analyzing NLRP3, NLRP12, NLRC4, ASC and pro-IL-1ß gene expressions in patients with HSCR. Decreased expression of NLRP3, NLRP12, NLRC4, ASC and pro-IL-1ß in the aganglionic and ganglionic bowel may increase susceptibility of HSCR patients to develop HAEC.

Altered expression of caveolin-1 in the colon of patients with Hirschsprung's disease.

BACKGROUND/PURPOSE: The pathogenesis of Hirschsprung's disease-associated enterocolitis (HAEC) is unclear. Caveolin-1 (Cav-1) regulates the functions of different nitric oxide synthase (NOS) isoforms, which play critical roles in inflammation and intestinal epithelial barrier function. We designed this study to investigate the hypothesis that Cav-1 expression is altered in the bowel of patients with Hirschsprung's disease (HSCR). METHODS: HSCR tissue specimens (n = 10) were collected at the time of pull-through surgery and control samples were obtained at the time of colostomy closure in patients with imperforate anus (n = 10). qRT-PCR analysis was undertaken to quantify Cav-1 gene expression, and Western blot analysis was undertaken to determine Cav-1 protein quantification. Immunolabelling of Cav-1 proteins was visualized using confocal microscopy. RESULTS: qRT-PCR and Western blot analysis revealed that Cav-1 was significantly downregulated in the aganglionic and ganglionic colon of patients with HSCR compared to controls (p < 0.01). Confocal microscopy revealed a markedly decreased expression of Cav-1 in colonic epithelium of aganglionic and ganglionic bowel of patients with HSCR compared to controls. CONCLUSION: To our knowledge, this is the first report of significantly decreased Cav-1 expression in patients with HSCR. Decreased expression of Cav-1 in the bowel of HSCR may increase susceptibility to HAEC in HSCR.

Endoscopic Injection of Dextranomer/Hyaluronic Acid as First Line Treatment in 851 Consecutive Children with High Grade Vesicoureteral Reflux: Efficacy and Long-Term Results.

PURPOSE: Endoscopic injection of dextranomer/hyaluronic acid is widely acknowledged as first line treatment of lower grade vesicoureteral reflux. We demonstrate its long-term efficacy and safety in eradicating high grade reflux. MATERIALS AND METHODS: A total of 518 girls and 333 boys with a median age of 2.3 years (range 2 months to 13.7 years) underwent endoscopic correction of high grade vesicoureteral reflux using dextranomer/hyaluronic acid. Reflux was unilateral in 415 cases and bilateral in 436, comprising 1,287 refluxing units. Reflux was grade IV in 1,153 ureters (89.6%) and grade V in 134 (10.4%). 99mTechnetium-dimercaptosuccinic acid scintigraphy identified renal scarring in 317 patients (37.3%). Followup ultrasound and voiding cystourethrogram were performed 3 months after intervention and renal ultrasound yearly thereafter. Median followup was 8.5 years (range 6 months to 16 years). RESULTS: Overall resolution rate after the first endoscopic injection was 69.5% (895 of 1,287 cases), with resolution in 70.4% of grade IV and 61.9% of grade V cases. Reflux resolved after a second injection in 259 cases (20.1%) and after a third injection in 133 (10.4%). Persistent reflux after initial treatment was significantly more common in patients younger than age 1 year and in individuals with renal scarring. No significant postoperative complications were observed and no patient required ureteral reimplantation. Following reflux resolution febrile urinary tract infection developed in 43 children (5.1%), including 24 (55.8%) during the first year, 15 (34.9%) during the second year and 4 (9.3%) during year 3 or later. Of these patients 6 had reflux recurrence and 8 had neocontralateral grade III reflux, which was successfully treated with a single endoscopic injection of dextranomer/hyaluronic acid. CONCLUSIONS: Endoscopic injection of dextranomer/hyaluronic acid is an efficient and safe long-term treatment for grade IV and V vesicoureteral reflux, and can easily be repeated in patients with treatment failure with a high subsequent resolution rate.