Peter Rubenwolf

Expression and Localisation of Aquaporin Water Channels in Human Urothelium In Situ and In Vitro

BACKGROUND Urothelium is generally considered to be impermeable to water and constituents of urine. The possibility that human urothelium expresses aquaporin (AQP) water channels as the basis for water and solute transport has not previously been investigated. OBJECTIVE To investigate the expression of AQP water channels by human urothelium in situ, in proliferating urothelial cell cultures and in differentiated tissue constructs. DESIGN, SETTING, AND PARTICIPANTS AQP expression by human urothelium in situ and cultured urothelial cells was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunolabelling. Expression screening was carried out on samples of freshly isolated urothelia from multiple surgical (bladder and ureteric) specimens and on proliferating and differentiated normal human urothelial (NHU) cells in culture. Urothelial tissue constructs were established and investigated for expression of urothelial differentiation markers and AQPs. MEASUREMENTS Qualitative study. RESULTS AND LIMITATIONS Transcripts for AQP3, AQP4, AQP7, AQP9, and AQP11 were expressed consistently by freshly isolated urothelia as well as by cultured NHU cells. AQP0, AQP1, AQP2, AQP5, AQP6, AQP8, AQP10, and AQP12 were not expressed. Immunochemistry confirmed expression of AQP3, AQP4, AQP7, and AQP9 at the protein level. AQP3 was shown to be intensely expressed at cell borders in the basal and intermediate layers in both urothelium in situ and differentiated tissue constructs in vitro. CONCLUSIONS This is the first study to demonstrate that AQPs are expressed by human urothelium, suggesting a potential role in transurothelial water and solute transport. Our findings challenge the traditional concept of the urinary tract as an impermeable transit and storage unit and provide a versatile platform for further investigations into the biological and clinical relevance of AQPs in human urothelium.

Delayed-onset Ureteral Obstruction After Endoscopic Dextranomer/Hyaluronic Acid Copolymer (Deflux) Injection for Treatment of Vesicoureteral Reflux in Children: A Case Series

We report 4 patients with upper urinary tract (UUT) obstruction requiring ureteric reimplantation at 1, 7, 28, and 63 months after dextranomer/hyaluronic acid copolymer (Dx/HA) injection for vesicoureteric reflux. Histopathologic evaluation of ureteric segments revealed extensive foreign body formation in all cases. We conclude that UUT obstruction is a rare but serious complication after Dx/HA injection that can occur even years after surgery. The incidence of delayed-onset UUT obstruction may be higher than previously noted. Long-term follow-up and a critical reappraisal of the method are needed to assess the late sequelae of Dx/HA injection therapy for vesicoureteric reflux.

Psoas hitch and Boari flap ureteroneocystostomy.

Introduction In the middle of the last century, Dolff [1], Paquin [2] and Zimmermann et al. [3] developed principles for ureteroneocystostomy after gynaecological ureter injuries. Turner-Warwick and Worth [4] adopted these techniques, named it the ‘Psoas Bladder-Hitch Procedure’ and applied this technique of ureteroneocystostomy for the treatment of distal ureteric obstruction, ureteric fistulas and ‘distended duplication’ of the upper urinary tract.

Loss of aquaporin 3 protein expression constitutes an independent prognostic factor for progression-free survival: an immunohistochemical study on stage pT1 urothelial bladder cancer

BackgroundTreatment of patients with stage pT1 urothelial bladder cancer (UBC) continues to be a challenge due to its unpredictable clinical course. Reliable molecular markers that help to determine appropriate individual treatment are still lacking. Loss of aquaporin (AQP) 3 protein expression has previously been shown in muscle-invasive UBC. The aim of the present study was to investigate the prognostic value of AQP3 protein expression with regard to the prognosis of stage pT1 UBC.MethodAQP 3 protein expression was investigated by immunohistochemistry in specimens of 87 stage T1 UBC patients, who were diagnosed by transurethral resection of the bladder (TURB) and subsequent second resection at a high-volume urological centre between 2002 and 2009. Patients underwent adjuvant instillation therapy with Bacillus Calmette-Guérin (BCG). Loss of AQP3 protein expression was defined as complete absence of the protein within the whole tumour. Expression status was correlated retrospectively with clinicopathological and follow-up data (median: 31 months). Multivariate Cox regression analysis was used to assess the value of AQP3 tumour expression with regard to recurrence-free (RFS), progression-free (PFS) and cancer-specific survival (CSS). RFS, PFS and CSS were calculated by Kaplan-Meier analysis and Log rank test.Results59% of patients were shown to exhibit AQP3-positive tumours, whereas 41% of tumours did not express the marker. Loss of AQP3 protein expression was associated with a statistically significantly worse PFS (20% vs. 72%, p=0.020). This finding was confirmed by multivariate Cox regression analysis (HR 7.58, CI 1.29 – 44.68; p=0.025).ConclusionsLoss of AQP3 protein expression in pT1 UBC appears to play a key role in disease progression and is associated with worse PFS. Considering its potential prognostic value, assessment of AQP3 protein expression could be used to help stratify the behavior of patients with pT1 UBC.

Aquaporin Expression Contributes to Human Transurothelial Permeability In Vitro and Is Modulated by NaCl

It is generally considered that the bladder is impervious and stores urine in unmodified form on account of the barrier imposed by the highly-specialised uro-epithelial lining. However, recent evidence, including demonstration of aquaporin (AQP) expression by human urothelium, suggests that urothelium may be able to modify urine content. Here we have we applied functional assays to an in vitro-differentiated normal human urothelial cell culture system and examined both whether AQP expression was responsive to changes in osmolality, and the effects of blocking AQP channels on water and urea transport. AQP3 expression was up-regulated by increased osmolality, but only in response to NaCl. A small but similar effect was seen with AQP9, but not AQP4 or AQP7. Differentiated urothelium revealed significant barrier function (mean TER 3862 Ω.cm2), with mean diffusive water and urea permeability coefficients of 6.33×10−5 and 2.45×10−5 cm/s, respectively. AQP blockade with mercuric chloride resulted in decreased water and urea flux. The diffusive permeability of urothelial cell sheets remained constant following conditioning in hyperosmotic NaCl, but there was a significant increase in water and urea flux across an osmotic gradient. Taken collectively with evidence emerging from studies in other species, our results support an active role for human urothelium in sensing and responding to hypertonic salt concentrations through alterations in AQP protein expression, with AQP channels providing a mechanism for modifying urine composition. These observations challenge the traditional concept of an impermeable bladder epithelium and suggest that the urothelium may play a modulatory role in water and salt homeostasis.

15 years of continent urinary diversion and enterocystoplasty in children and adolescents: the Würzburg experience

Study Type – Therapy (case series)
Level of Evidence 4

Intraoperative Monitoring of Bladder and Internal Anal Sphincter Innervation: A Predictor of Erectile Function following Low Anterior Rectal Resection for Rectal Cancer? Results of a Prospective Clinical Study

Background: The objective was to investigate whether two-dimensional intraoperative neuromonitoring (IONM) of pelvic autonomic nerves has the potential to predict erectile function (EF) following surgery for rectal cancer. Methods: A consecutive series of 17 sexually active male rectal cancer patients undergoing IONM-based nerve-sparing low anterior rectal resection were evaluated prospectively. IONM was performed by electric stimulation of the pelvic splanchnic nerves with concomitant electromyography of the internal anal sphincter and cystomanometry. Sexual function was assessed using a validated questionnaire. Results: The degree of agreement between electromyography-based and cystomanometry-based IONM with postoperative EF was moderate and good (κ = 0.43 and κ = 0.66). Combined assessment yielded the best agreement (κ = 0.76) with sensitivity of 90%, specificity of 86%, positive predictive value of 90%, negative predictive value of 86%, and overall accuracy of 88%, respectively, in terms of prediction of postoperative EF. Conclusion: The method may be suitable to predict male EF following rectal resection.

Metabolic Consequences after Urinary Diversion

Metabolic disturbances are well-known, but sometimes neglected immediate consequences or late sequelae following urinary diversion (UD) using bowel segments. Whereas subclinical disturbances appear to be quite common, clinically relevant metabolic complications, however, are rare. Exclusion of bowel segments for UD results in loss of absorptive surface for its physiological function. Previous studies demonstrated that at least some of the absorptive and secreting properties of the bowel are preserved when exposed to urine. For each bowel segment typical consequences and complications have been reported. The use of ileal and/or colonic segments may result in hyperchloremic metabolic acidosis, which can be prevented if prophylactic treatment with alkali supplementation is started early. The resection of ileal segments may be responsible for malabsorption of vitamin B12 and bile acids with subsequent neurological and hematological late sequelae as well as potential worsening of the patient’s bowel habits. Hence, careful patient and procedure selection, meticulous long-term follow-up, and prophylactic treatment of subclinical acidosis is of paramount importance in the prevention of true metabolic complications.

Bladder augmentation using bowel segments (enterocystoplasty).

Gastric segments, small and large bowel segments as well as the ureter are used for bladder augmentation [ 1 ] . Since the late 19th century, ileal segments have been used for bladder augmentation (ileocystoplasty) [ 2,3 ] . Later, these segments were detubularised and reconfi gured to create, together with the remnant bladder, a spherical reservoir [ 4 ] . In the middle of the past century, the use of caecum for bladder augmentation was reported [ 5,6,7,8 ] . In ileocaecocystoplasty, the presence of the appendix is advantageous for patients who are unable to perform clean intermittent self-catheterisation (CISC) through the urethra. In these cases, the submucosally embedded appendix can be used as an additional continent cutaneous stoma to evacuate the augmented bladder, as in patients with heterotopic continent cutaneous diversion [ 9 ] . If ureteric re-implantation is necessary, the ileocaecal valve can be used as anti-refl ux mechanism for severely dilated ureters [ 10,11 ] . Nondilated ureters can be implanted into the large bowel by the submucosal tunnel technique [ 12 ] . At the beginning of the 20th century, the use of sigmoid colon was reported for bladder augmentation (colocystoplasty) [ 13,14 ] . Gastrocystoplasty was reported in 1978 by Leong and Ong [ 15,16 ] . However, complications of bladder augmentation by gastric segments, e.g. hyponatraemic, hypochloraemic alkalosis, haematuria-dysuria syndrome [ 17,18 ] and secondary malignancies after the 10th postoperative year [ 19,20,21,22 ] made this type of bladder augmentation obsolete. ILLUSTRATIONS by STEPHAN SPITZER, www.spitzer-illustration.com

Persistent Histological Changes in the Exstrophic Bladder After Primary Closure—A Cause for Concern?

PURPOSE We investigated bladder biopsies from patients with classic bladder exstrophy for the histological features and discuss the potential clinical significance of the findings. MATERIALS AND METHODS Bladder tissues were collected from patients with bladder exstrophy between 2004 and 2011. These specimens were obtained at primary bladder closure (group 1, 29 patients), during secondary reconstructive procedures (group 2, 27) or during cystectomy for failed reconstruction (group 3, 15). All tissue specimens were investigated for inflammatory, proliferative, metaplastic and dysplastic changes. Expression of urothelial differentiation markers CK13 and CK20 was determined by immunohistochemical analysis. RESULTS Inflammatory, proliferative and metaplastic changes were found in bladder specimens of all subgroups. Neither dysplasia nor neoplasia was present. Severe epithelial changes such as cystitis glandularis and intestinal metaplasia were observed in up to 62% of bladders several years after primary closure. Aberrant expression patterns of CK13 and CK20 suggesting abnormal urothelial differentiation were shown to be present in the urothelium of all subgroups. CONCLUSIONS Our findings provide prima facie evidence that the epithelial changes observed in the unclosed bladder template persist or even progress in a subset of bladders after primary closure. Although the malignant potential of cystitis glandularis and intestinal metaplasia is controversial, some patients may be at increased risk for dysplasia/neoplasia in the long term. Since the natural history of these lesions in the exstrophic bladder is unknown, these patients require lifelong surveillance.