ESPU Meeting on Wednesday 3, September 2025, 12:45 - 13:50
12:45 - 12:49
S01-1 (OP)
Kevin CAO 1, Jessica TRUNDLE 1, David LONG 1 and Navroop JOHAL 2
1) UCL Great Ormond Street Institute of Child Health, Developmental Biology, London, UNITED KINGDOM - 2) UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, Developmental Biology, London, UNITED KINGDOM
PURPOSE
Primary epispadia(PE) is rare and despite surgical advances often results in urinary incontinence. The morphometric characteristics of the PE bladder are unknown. This study compares bladder histology and biomechanics in PE against classical bladder exstrophy (CBE) and normal paediatric bladders.
MATERIAL AND METHODS
Between 2018 and 2023, bladder muscle strips were collected from patients undergoing surgery for PE (n=17), CBE (n=37; neonates n=9), and control (no bladder dysfunction) conditions (n=15). Histological analysis measured the smooth muscle-to-connective tissue ratio (SM:CTr), where lower ratios indicate greater fibrosis. Bladder tissue compliance was assessed through stress-strain experiments. Statistical significance was determined using Mann-Whitney U tests.
RESULTS
Median SM:CTr was highest in controls (2.21), lowest in newborn CBE (0.20) and similar between older children with CBE (0.63) and PE (0.67). Compliance was lowest in controls (27.75) and similar among newborn CBE (37.10), CBE (41.71), and PE (36.07)(all mN.mm-2). Only normal vs. CBE was statistically significant (p=0.022).
The SM:CTr was significantly reduced in PE compared to controls and similar to older children with CBE (p=0.79), suggesting that PE has a degree of fibrosis comparable to CBE beyond the neonatal period. Bladder compliance in PE was also similar to both newborns (p=0.71) and older children with CBE (p=0.56), indicating that passive stretch properties do not significantly differ between the two condition.
CONCLUSIONS
PE exhibits histological impairment and altered passive biomechanics comparable to classical exstrophy. These findings suggest PE has a complex pathophysiology and should be clinically interrogated as rigorously as exstrophy, rather than assuming a higher likelihood of achieving continence.
12:49 - 12:53
S01-2 (OP)
Jason YANG, Alexander HIRSCH, Wayland WU, David HEAP, Catherine ROBEY, Heather DI CARLO, Larissa SHIMODA, John GEARHART and Chad CRIGGER
Johns Hopkins University, Urology, Baltimore, USA
PURPOSE
Approximately one-third of exstrophic bladders demonstrate inadequate growth and reduced capacity following surgical closure. To elucidate potential underlying mechanisms, we investigated the mitochondrial architecture and function in smooth muscle cells from exstrophic bladders compared to normal bladders.
MATERIAL AND METHODS
Full-thickness bladder biopsies were collected from 5 classic bladder exstrophy patients and from 5 normal bladder patients undergoing ureteral reimplantation. Mitochondrial morphology was evaluated by staining cells with Mitotracker dye. Fluorescence microscopy images were then captured using Micro-Manager software and analyzed with proprietary software. Moreover, ultrastructural analysis was performed using transmission electron microscopy (TEM) at 80 kV. For protein expression assessment, cell lysates were prepared with Laemmli lysis buffer, and levels of phosphorylated DRP1 (pDRP1) were quantified by western blot.
RESULTS
TEM and fluorescence imaging revealed distinct phenotypic differences in mitochondrial structure between exstrophic and normal bladder samples. Mitotracker fluorescence analysis showed decreasing trends towards shorter median mitochondrial branch lengths in exstrophy cells compared to controls (9.60 vs. 10.45 um, p=0.19) with a more comparable median branch number (2.81 vs. 3.14, p=0.56). Western blot analysis indicated a trend towards an elevated pDRP1/DRP1 ratio in exstrophic bladder cells (1.77 vs. 1.07, p=0.10).
CONCLUSIONS
Smooth muscle cells from exstrophic bladders exhibit distinct mitochondrial abnormalities, including a trend toward shorter branch lengths, coupled with a trend toward increased mitochondrial fission activity. These findings implicate mitochondrial dysfunction in exstrophic bladders and identify mitochondrial dynamics as a potential target for improving outcomes in this congenital condition. Of note, with over 100 biopsy samples, we anticipate achieving statistical significance by the time of conference as we continue processing additional exstrophy samples.
13:00 - 13:04
S01-3 (OP)
Nathalia AMADO 1, Karunya ALBERTS 1, Nixon RAJ 1, Alexandria FUSCO 2, Jeremy MATHEWS 2, Caitlin COCO 2, Martinez HILL 2, Brandi CANTAREL 2 and Linda BAKER 1
1) Nationwide Children's Hospital, Kidney and Urinary Tract Center, Columbus, USA - 2) University of Texas Southwestern, Urology, Dallas, USA
PURPOSE
In most cases, the cause of PBS is unknown. MYOCD is highly expressed in embryonic detrusor and is a master transcriptional co-activator initiating expression of SMC genes. Recently, human MYOCD mutations were reported with PBS-like phenotypes. We tested our PBS cohort for MYOCD variants and their functional impact.
MATERIAL AND METHODS
With IRB-approval, PBS proband DNA underwent paired-end whole exome sequencing. DNA variants were filtered (function/ minor allele frequency) and Sanger validated. To test identified MYOCD variants for functional impact, site directed mutagenesis on wild-type (WT) GFP-MYOCD plasmid introduced each PBS MYOCD variant. HEK-293t cells were transfected with WT or mutated-plasmid for Sm22α-luciferase reporter assay and qPCR measuring SMC markers. Mann-Whitney tested group differences, with p<0.05 considered significant.
RESULTS
6 of 140 probands carried 5 novel heterozygous MYOCD mutations in conserved residues, including two truncations (p.N125Tfs*9, p.R384*) in two multiplex PBS families and three missense variants (p.G445S, p.V491M, and p.Q647H) in four sporadic PBS cases. Sm22α-luciferase assays and qPCR from transfected cells demonstrated that while V491M showed no dysfunction, the N125Tfs*9, R384*, G445S, and Q647H MYOCD mutations caused an average 12-fold inhibition of Myocd activity and significantly reduced expression (mean 2X decrease) ofMyh11andTagln. Pedigree segregation analysis showed MYOCD variant transmission via asymptomatic carriers, suggesting incomplete penetrance or genetic modifiers.
CONCLUSIONS
Of 5 novel MYOCD mutations in 2 familial and 4 sporadic PBS cases, 4 demonstrated MYOCD loss-of-functionin vitro, decreasing expression of genes necessary for SMC development. This confirms MYOCD exonic mutations account for 3.5% of PBS cases.
13:04 - 13:08
S01-4 (OP)
Nathalia AMADO 1, Nixon RAJ 1, Thomas EGELAND 2, Shane BATIE 2, Shaheer ALI 2, Marco Antonio ANDRADE 3, Caitlin COCO 2, Jeremy MATHEWS 2, Brandi CANTAREL 2 and Linda BAKER 1
1) Nationwide Children's Hospital, Kidney and Urinary Tract Center, Columbus, USA - 2) University of Texas Southwestern, Urology, Dallas, USA - 3) Nationwide Children's Hospital, Urology, Columbus, USA
PURPOSE
Genetic smooth muscle myopathies (SMM) affect neonates to adults and include vascular myopathies (Thoracic aortic aneurysm/ dissection [TAA/D]) and visceral myopathies (Megacystis Microcolon Intestinal Hypoperistalsis Syndrome [MMIHS] and Chronic Intestinal Pseudoobstruction [CIPO]). We investigated PBS MYH11 variants.
MATERIAL AND METHODS
Proband/family DNA underwent whole exome sequencing. MYH11 variants were filtered based on 1) meeting 5 of 6 criteria from in silico functional prediction software (Polyphen-2, SIFT, CADD, VEST, Mutationtaster and AlphaMissense) and 2) <0.001% minor allelic frequency. MYH11 variants were then validated and screened for prior SMM reporting in ClinVar. Each missense mutation underwent AlphaFold3 modeling and protein structural analysis.
RESULTS
6 of 140 PBS probands carry 5 rare, protein-damaging heterozygous MYH11 variants (see Table). All five variants were previously associated to TAA/D and interestingly, 4 out 5 families report cardiac condition in the family, suggesting a potential causal relationship.
| PBS-ID | MYH11 Variant | MYH11 Functional Domain | Inheritance | Minor Allelic Frequency (GNOMAD) | Family history of Miscarriages | Structural Analysis: | |
| Electrostatic interactions | Hydrophobic interactions | ||||||
| PBS-1 | p.R108W | Myosin Head | Maternal | ~0 | Yes | Destabilized | Introduced |
| PBS-2 | p.R676C | Unknown | 0.0008369 | No | Altered | Altered | |
| PBS-3 | p.A699V | De novo | 0.0003897 | ||||
| PBS-4 | p.T1565M | Coil-Colled | Maternal | 0.00068 | Yes | Altered(Inter-monomer) | Altered(Inter-monomer) |
| PBS-5 | |||||||
| PBS-6 | p.K1628Q | 0.0000994 | No changes | No changes | |||
CONCLUSIONS
Heterozygous protein-damaging DNA variants in MYH11 are a genetic cause of PBS in 4.3% of cases. Longitudinal studies are essential to assess life-threatening TAA/D risk and investigate myosin-modulating drugs in PBS patients with MYH11 variants.
13:08 - 13:12
S01-5 (OP)
Nathalia AMADO 1, Justin WOBSER 1, Nixon RAJ 1, Thomas EGELAND 2, Sujay ITHYCHANDA 3, Luis Sifuentes DOMINGOS 2, Nida IQBAL 2, Alexandria FUSCO 2, Monica RIDLON 4, Ashley JACKSON 1, Kimberly KEIL STIETZ 4, Jun QIN 3, Roy ZENT 5 and Linda BAKER 1
1) Nationwide Children's Hospital, Kidney and Urinary Tract Center, New Albany, USA - 2) University of Texas Southwestern, Urology, Dallas, USA - 3) Cleveland Clinic, Cardiovascular and Metabolic Science, Cleveland, USA - 4) University of Wisconsin—Madison, Comparative Biosciences, Madison, USA - 5) Vanderbilt University School of Medicine, Medicine, Nashville, USA
PURPOSE
Previously, we identified 4 PBS males with X-linked, pathogenic FLNA missense mutations, including the severe FLNA p.C2160R gain-of-function mutation disrupting regulatory protein binding sites. The FLNA protein is crucial for smooth muscle cell (SMC) mechanotransduction and hypoxic responses. After creating the Crispr-Cas9-engineered analogous Flna-C2152R mice who have a mild PBS-like phenotype, we exposed them to acute gestational hypoxia (AGH) to assess effects on testis descent and bladder form/function.
MATERIAL AND METHODS
Wild-type (WT) or Flna-C2152R homozygous pregnant dams were exposed to AGH (8hrs of 6% O2 on E12.5). At 2-months, 24-hour metabolic cage studies measured voided urine volume and frequency (UroVoid).Ex-vivo organ bath assays assessed bladder contractility using electrical field stimulation (EFS) and cholinergic and purinergic agonists.
RESULTS
Comparing AGH WT to AGH Flna-C2152R males, mutants had significantly increased bladder weight, detrusor thickness, and collagen deposition at 40%, 50%, and 44%, respectively. Functionally, Flna-C2152R males had 172% higher mean voided volumes per void, and 57% fewer voids per 24hrs. Flna-C2152R bladder contractility was normal on cholinergic and purinergic stimulation, but with a 49% decrease in EFS response, indicating diminished bladder innervation. No abdominal laxity or hydroureteronephrosis was observed in WT or Flna-C2152 AGH mice. Unilateral intra-abdominal cryptorchidism was present in 5% and 75% of WT vs Flna-C2152R mice, respectively.
CONCLUSIONS
When compared to AGH WT mice, the Flna-C2152R AGH mice manifest a PBS-like phenotype, with enlarged, fibrotic, underactive bladders with diminished contractility and unilateral intra-abdominal cryptorchidism. This suggests that PBS may be the result of an interaction between Flna mutation and embryonic hypoxic environment.
13:21 - 13:25
S01-6 (OP)
Enrico DANZER 1, Tomohiro ARAI 2, Eric JOHNSON 3, Marianna SCUGLIA 2, Wasinee TIANTHONG 2, Roland DEVLIEGER 4, Jan DEPREST 5, Francesca RUSSO 2, Yair BLUMENFELD 6 and Kunj SHETH 7
1) Memorial Sloan Kettering Cancer Center, Pediatric Surgery, New York, USA - 2) KU Leuven, Development and Regeneration, Cluster Woman and Child, Group Biomedical Sciences, Leuven, BELGIUM - 3) Vortex Medical, Woodside, USA - 4) KU Leuven, Obstetrics and Gynecoogy, Leuven, BELGIUM - 5) KU Leuven, Development and Regeneration, Cluster Woman and Child, Group Biomedical Sciences; Obstetrics and Gynecology, Leuven, BELGIUM - 6) Stanford University School of Medicine, Obstetrics and Gynecoogy, Palo Alto, USA - 7) Stanford University School of Medicine, Urology, Palo Alto, USA
PURPOSE
Current fetal vesicoa-amniotic shunts (VAS) for prenatal treatment of lower urinary tract obstruction (LUTO) are limited in efficacy. The novel Vortex shunt incorporates enhanced fixation elements, a kink resistant extendible conduit, high echogenicity to facilitate ultrasound-guided deployment, and a one-way valve for bladder cycling. We aimed to evaluate the efficacy and safety of ultrasound-guided insertion, drainage, and long-term dislodgement risks of the Vortex shunt in fetal lambs with surgically induced LUTO.
MATERIAL AND METHODS
LUTO was created in 10 fetal lambs at a median of 75 days gestation. All fetuses developed severe LUTO with ultrasound features of megacystis, hydronephrosis, urinary ascites, and significant oligohydramnios/ anhydramnios. At a median of 96 days gestation, these fetuses underwent ultrasound-guided placement of the Vortex shunt. Near term, euthanasia was performed, and the fetuses were retrieved. Efficacy was assessed by the resolution of the key anatomical features of LUTO on ultrasound and necropsy. Safety parameters included dislodgement rate, patency at term, and shunt or procedural side effects.
RESULTS
The ultrasound-guided deployment of the Vortex shunt was successful for all fetuses. Bladder drainage was observed immediately after placement. There was sustained bladder emptying, normalization of hydroureteronephrosis, and amniotic fluid levels. The Vortex shunt remained in an appropriate position throughout gestation for a median of 41 days in all fetal lambs with no dislodgement. There were no shunt-related or procedure-related complications. Furthermore, shunted LUTO bladders had improved bladder contractility compared to the LUTO bladders on early with potassium chloride and carbachol testing, which were comparable to normal controls.
CONCLUSIONS
In this preclinical large animal model of LUTO, we showed that the Vortex shunt can be accurately deployed at midgestation under ultrasound guidance. We further demonstrate its long-term efficacy, function, and safety. The novel shunt system may improve the adverse outcomes of LUTO.
13:25 - 13:29
S01-7 (OP)
Anjali SRIVASTAVA 1, Sachit ANAND 1, Ajay VERMA 1, Jitendra MEENA 2 and Kalpana LUTHRA 3
1) All India Institute of Medical Sciences (AIIMS), Pediatric Surgery, New Delhi, INDIA - 2) All India Institute of Medical Sciences (AIIMS), Pediatrics, New Delhi, INDIA - 3) All India Institute of Medical Sciences (AIIMS), Biochemistry, New Delhi, INDIA
PURPOSE
Posterior urethral valves (PUV) are a major cause of congenital urinary tract obstruction, often leading to chronic kidney disease (CKD) despite successful valve ablation. Epigenetic modifications, particularly DNA methylation, have been implicated in the pathogenesis of several kidney diseases. This study aimed to explore the association of global DNA methylation levels and kidney function in PUV patients with varying kidney function.
MATERIAL AND METHODS
This cross-sectional study included 45 children with PUV post-valve ablation and 45 matched controls (ages 0-14 years). DNA was extracted from peripheral blood, and global DNA methylation levels were quantified using the 5-mC % ELISA kit. Nuclear scintigraphy studies were performed to assess the glomerular filtration rate (GFR) and the presence of kidney scars. The PUV group was categorized into different CKD-subgroups using GFR values.
RESULTS
Global DNA methylation levels were significantly higher in PUV patients (median 5-mC% = 0.4336) compared to controls (median 5-mC% = 0.3732, p < 0.001). Subgroup analysis revealed an increasing trend in 5-mC% across all CKD stages compared to controls, with significant elevation in most stages (p < 0.05). A logarithmic trend analysis demonstrated a strong correlation between advancing CKD stages and rising 5-mC% (R² = 0.8012), suggesting a potential link between DNA methylation and kidney dysfunction progression.
CONCLUSIONS
Elevated global DNA methylation levels in PUV patients, especially those with CKD, suggests a potential epigenetic marker for progressive deterioration of kidney function. Also, it appears that the quantification of global DNA methylation can guide pediatricians in early identification of at-risk PUV cases.
13:35 - 13:39
S01-8 (OP)
Barnali DAS 1, Shannon DIMARTINO 1, Emily MORRISON 1, Abby COTSONAS 1, Ali HASHEMI GHEINANI 2, John FROEHLICH 1 and Richard LEE 1
1) Boston Children's Hospital, Department of Urology, Boston, USA - 2) University of Bern, Department for BioMedical Research DBMR, Bern, SWITZERLAND
PURPOSE
Hydronephrosis is readily detectable and monitored by ultrasound, but differential renal function (DRF) can only be determined by nuclear medicine studies, which are radiating and invasive, or MRI, which may require sedation and can be cost prohibitive. A non-invasive diagnostic marker that identifies patients with reduced DRF may significantly streamline which children with hydronephrosis need additional testing and who may require surgical intervention. Utilizing a cohort of children undergoing pyeloplasty, we identified urinary biomarkers that can stratify the cohort by DRF.
MATERIAL AND METHODS
Bladder urine was collected from 101 children at the time of pyeloplasty. Patients were stratified by high DRF (>40%, n=79) and low DRF (<=40%, n=29) as measured by MAG-3. Urine samples underwent data-independent acquisition based mass spectrometry for peptide quantification. The feature selection algorithm Boruta and decision tree analysis were used to identify relevant proteins to stratify patients. Ingenuity Pathway Analysis identified the functional significance of pertinent proteins.
RESULTS
We identified 4727 proteins of which 1200 protein groups had complete quantified data across all 101 samples. Boruta identified 17 proteins with substantial impact on a model’s predictive capability of identifying a patient as high or low DRF. Decision tree analysis identified a combination of 4 proteins that completely delineated the high and low DRF groups. Functionally, significant proteins were observed to be significantly enriched in several renal function related pathways and predicted that the TLR family (z-score = 2.3, p-value = 2.4x10-16) and GHR (z-score = 2.27, p-value = 2.12x10-18) were master regulators of the changes identified.
CONCLUSIONS
We have demonstrated that patients with hydronephrosis can be stratified by DRF using a combination of urinary biomarkers. Once validated, these markers may have significant potential to change current management of hydronephrosis by focusing care towards those patients with low DRF.
13:39 - 13:43
S01-9 (OP)
Nilsun KUAS 1, Ufuk ATES 2, Firat SERTTURK 3, Ege EVIN 4, Volkan TEKIN 5, Deniz BILLUR 6, Ayşe Hande YOZGAT 6, Ergun ERGÜN 7, Gökhan Berktuğ BAHADIR 8, Gülnur GÖLLÜ BAHADIR 7 and Ahmet Murat ÇAKMAK 7
1) Ministry of Health University Van Regional Training and Research Hospital, Van , Turkey, Pediatric Surgery, Van, TÜRKIYE - 2) Ankara University Faculty of Medicine Department of Pediatric Surgery, Ankara, TÜRKIYE - 3) Etlik City Hospital Pediatric Surgery Clinic,, Pediatric Surgery, Ankara, TÜRKIYE - 4) Ankara University Faculty Medicine, Pediatric Surgery, Ankara, TÜRKIYE - 5) Ministry of Health University Gulhane Medical Faculty, Physiology Department, Ankara, TÜRKIYE - 6) Ankara University Faculty of Medicine, Department of Histology and Embryology, Ankara, TÜRKIYE - 7) Ankara University Faculty of Medicine, Department of Pediatric Surgery, Ankara, TÜRKIYE - 8) Ministry of Health University Gulhane Medical Faculty, Department of Pediatric Surgery,, Ankara, TÜRKIYE
PURPOSE
Testicular torsion causes ischemia-reperfusion injury, significantly impacting fertility. Free oxygen radicals have been implicated in tissue damage. Hyperbaric oxygen (HBO) therapy has shown potential in mitigating oxidative stress and improving tissue outcomes. This study aimed to evaluate the efficacy of HBO in reducing testicular damage in long-term torsion.
MATERIAL AND METHODS
Fifty-six rats were randomized into seven groups: Sham (Group 1), torsion followed by detorsion at 4, 8, or 16 hours (Groups 2-4), and the same torsion protocols with HBO therapy (Groups 5-7). HBO treatment involved daily sessions of 100% oxygen at 2.4 atm for seven days post-detorsion. Parameters assessed included total antioxidant (TAS) and oxidant status (TOS), seminiferous tubule diameter, and Johnsen score.
RESULTS
HBO therapy significantly reduced TOS levels in the 16-hour torsion group compared to untreated controls (p = 0.028). However, seminiferous tubule diameter and Johnsen scores showed no significant improvement with HBO in either torsioned or contralateral testes across all timepoints. While TAS/TOS ratios improved, histological parameters remained unchanged.
CONCLUSIONS
HBO therapy reduced oxidative stress in long-term testicular torsion but did not improve histological damage or spermatogenic activity. Further studies are required to determine the therapeutic potential and optimal timing for HBO application in testicular torsion.