[Show abstract][Hide abstract] ABSTRACT: To study the outcome of composite cystoplasty using cultured urothelial cells combined with de-epithelialized colon or uterus in a porcine surgical model, using appropriate controls, and to characterize the neo-epithelium created by composite cystoplasty.
Urothelial cells were isolated and propagated in vitro from open bladder biopsies taken from nine female minipigs. Cohesive sheets of confluent urothelial cells were transferred to polyglactin carrier meshes and sutured to de-epithelialized autologous colon in four animals and de-epithelialized autologous uterus in five. These composite segments were then used for augmentation cystoplasty. Conventional colocystoplasty, de-epithelialized colocystoplasty and sham operations were carried out in six control animals. After killing the animals at approximately 90 days the bladders were removed for examination and immunohistochemical analysis, using a panel of antibodies against cytokeratins and urothelial differentiation-associated antigens.
Macroscopically, the bladders augmented with composite segments derived from uterine muscle had no evidence of shrinkage or contracture. Histological analysis showed that in four of five composite uterocystoplasties, the neo-urothelium was stratified and had a transitional morphology, although in some areas coverage was incomplete. Immunohistochemical analysis showed evidence of squamous differentiation in both native and augmented segments. All composite and de-epithelialized colonic segments showed significant contraction with poor urothelial coverage, reflecting the unsuitability of the thin-walled porcine colon for de-epithelialization.
The functional and macroscopic outcome of bladder augmentation with a composite derived from cultured urothelium and de-epithelialized smooth muscle of uterine origin endorses the feasibility of composite cystoplasty.
BJU International 04/2004; 93(4):609-16. · 3.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Reconstruction of the urinary bladder with bowel to restore storage capacity is associated with significant complications arising from substituting an absorptive, mucus-producing intestinal epithelium for the barrier urothelium of the bladder. To overcome these problems, we are developing a "composite enterocystoplasty" procedure to replace the epithelium of the bowel with autologous in vitro-propagated normal urothelial cells. The aims of this study were to evaluate synthetic biomaterials as delivery vehicles for the cultured urothelial cells and provide support during transfer and cell adherence to the de-epithelialized bowel wall. The surgical compliance of 12 biomaterials was evaluated, along with their ability to support urothelial cell attachment. Transfer of urothelial cells onto biomaterials as single cell suspensions or intact cell sheets was investigated. Seeding of a single cell suspension on to non-woven mesh resulted in poor cell attachment. Seeding onto woven mesh was more efficient, but the most effective transfer method involved producing an intact cell sheet that could be combined with woven, knitted and non-woven biomaterials. Transfer of the cell sheet : mesh complexes onto a de-epithelialized bladder stroma produced a stratified epithelium incorporating the strands of the mesh and expressing urothelial-associated antigens after 48 h in organ culture. Thus, we have developed and evaluated a suitable transfer method for in vitro propagated urothelial cells to be used in "composite enterocystoplasty".
Journal of Materials Science Materials in Medicine 01/2001; 12(10-12):991-6. · 2.38 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To determine if the indications for cutaneous vesicostomy have changed with the advent of prenatal diagnosis and to assess if temporary vesicostomy remains an effective measure in preventing urinary infection and stabilising renal function in children with severe uropathy.
Retrospective case note review.
Nineteen patients underwent temporary cutaneous vesicostomy over a 12-year period. Of these, 10 had a prenatally detected uropathy. Children with prenatal findings were diverted at a significantly younger age than cases presenting clinically (33 (range 10-240) vs. 182 days (range 28-425), p = 0.02). In addition, there were significantly more boys diverted in the prenatally detected group (p = 0.02). The majority of clinically presenting infants were diverted because of severe or recurrent urosepsis whereas most prenatally detected cases underwent surgery for progressive renal impairment. In both groups vesicostomy was successful in preventing further urosepsis and resulted in improved renal function. Follow-up data, however, revealed 9 (50%) of 18 patients with chronic renal failure. One patient died prior to stoma closure from septicaemia and renal disease.
The timing and indications for vesicostomy may be changing in the light of prenatal diagnosis. Temporary vesicostomy diversion remains a simple, effective and easily reversible method of preventing urosepsis and stabilising upper tract function. The poor renal outcome on follow-up may reflect the fact vesicostomy is reserved for infants and children with severe disease.
European Urology 02/1998; 33(4):405-11. · 12.48 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We established a 3-dimensional organ culture model of urinary tract tissue in which to study the effects of seeding cultured urothelial cells onto de-epithelialized urothelial stroma.
Normal human urinary tract tissues were placed in organ culture or used to establish urothelial cell cultures. At passage 2 cell cultures were harvested and used to reconstitute autologous organ cultures by seeding onto de-epithelialized stroma. Organ cultures were harvested at intervals and analyzed by immunohistology with a panel of antibodies against differentiation associated antigens, cytokeratins, cell adhesion molecules, extracellular matrix components and proliferation associated antigens.
Human urothelial tissues were maintained in organ culture for at least 18 weeks and they retained a transitional epithelial morphology with expression of normal in situ antigenic characteristics. Within 2 weeks of reconstitution recombined organ cultures formed a stratified, polarized, transitional-like neo-epithelium that expressed many of the phenotypic and differentiated characteristics of normal tissue. Basement membrane formed at sites of direct contact between urothelial cells and stroma. After an initial stabilization period the proliferation rate of the urothelium of intact and reconstituted organ cultures decreased to the low turnover rate characteristic of normal urothelium in situ, indicating that the cells were responsive to normal growth regulatory controls.
Normal human urothelial cells, which express a proliferative nondifferentiated phenotype in monolayer culture, retain the capacity to differentiate and reform a slow turnover, stratified transitional epithelium.
The Journal of Urology 10/1997; 158(3 Pt 2):1147-52. · 3.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We attempted to determine if the degree of second trimester dilatation and/or other qualitative sonographic features of the fetal urinary tract are predictive of postnatal outcome in male neonates with posterior urethral valves.
We reviewed ultrasound reports and/or hard copy imaging in 17 cases of posterior urethral valves initially detected on second trimester scans (median gestation 18 weeks, range 15 to 23.5). Ultrasound appearance was categorized according to the pattern and severity of dilatation, cystic change or echogenicity of the renal cortex and presence of oligohydramnios. Outcome was reviewed at a median followup of 5.7 years (range 4.4 to 10).
Of the 17 cases there was a poor outcome in 10, including death in 4 and chronic renal failure in 6. Seven patients were alive and well with normal renal function at followup. The prognosis in cases of moderate or severe upper tract dilatation was poor. Of the 9 patients with marked prenatal hydroureteronephrosis 8 (89%) were dead or had chronic renal failure at followup. In contrast, only 2 of the 8 patients (25%) with mild upper tract dilatation or dilatation limited to the bladder had chronic renal failure at review (p = 0.05). Three cases of prenatal renal parenchymal change and 3 of subsequent oligohydramnios had a poor outcome postnatally.
The prognosis for boys with prenatally detected posterior urethral valves is closely associated with qualitative aspects of second trimester scan findings. This information may be of clinical value in the prenatal counseling of parents.
The Journal of Urology 10/1997; 158(3 Pt 2):1022-5. · 3.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To examine the relationship between pre-natal ultrasonographic findings typical of pelvi-ureteric junction (PUJ) obstruction and post-natal renal function in the affected kidney.
This retrospective study comprised 35 infants in whom pre-natal ultrasonography had identified a unilateral pattern of dilatation of the renal pelvis and calices typical of PUJ obstruction. In each case, the presence of unilateral hydronephrosis had been confirmed post-natally by ultrasonography, and differential function and drainage evaluated by isotope renography. The results of isotope renography were compared with the severity and timing of onset of the fetal hydronephrosis.
Eleven kidneys (31%) appeared normal in the second trimester and dilatation only became apparent during the third trimester. Mean differential function in these kidneys was 48%. However, mean differential function was reduced (mean 38%) in those kidneys noted to be dilated between 16 and 24 weeks gestation. Differential function was loosely correlated with the severity of dilatation in early onset cases, i.e. the mean differential function was 42% for mild, 37% for moderate and 27% for severe dilatation. A considerable variation in differential function values was present in each group except for those with severe dilatation, which was a significant predictor of poor functional outcome when compared with mild and moderate dilatation combined (P < 0.01).
Fetal PUJ obstruction is a heterogeneous condition permitting only broad predictions of functional outcome. Severe dilatation detected on second trimester imaging predicted significant loss of function. Mild and moderate degrees of dilatation were associated with a one in three risk of functional impairment in the obstructed kidney.
[Show abstract][Hide abstract] ABSTRACT: The purpose of the work was to establish urothelium as an in vitro model for the study of proliferation, stratification, and differentiation in "complex" epithelia.
Normal human urothelial cells were cultured in a serum-free medium. The effects of epidermal growth factor (EGF), cholera toxin (CT), extracellular calcium and 13-cis-retinoic acid on cell growth, morphology, phenotype, and cytodifferentiation were studied using phase-contrast microscopy and indirect immunofluorescence. Stratification-related changes were additionally analyzed by transmission electron microscopy.
Under optimized conditions, long-term cultures were successful in 44 (74.5%) out of 59 specimens. Bacterial infection was the most common cause of failure (9 cases). Primary urothelial cells required an initial plating density of > or = 10(4) cells/cm2 for survival; passaged cells survived much lower plating densities (> or = 2.5 x 10(2) cells/cm2). CT significantly improved cell attachment, but neither CT nor EGF were essential for growth. By contrast, cells failed to proliferate without bovine pituitary extract. In media containing bovine pituitary extract, CT, and EGF, cultures had a mean population doubling time of 14.7 +/- 1.8 hours, maintained a nonstratified phenotype, and expressed the cytokeratin (CK) profile of basal/intermediate urothelium: CK7, CK8, CK17, CK18 and CK19, with variable expression of CK13. CK20 was not expressed in vitro. CK14 and CK16 were also expressed, suggestive of squamous metaplasia in culture, which could be inhibited with 13-cis-retinoic acid. Increasing extracellular calcium from 0.09 to 0.9-4.0 mM slowed cell proliferation, induced stratification and desmosome formation, and increased expression of E-cadherin. High calcium, EGF, CT, and retinoic acid did not induce markers of late/terminal urothelial cytodifferentiation.
We describe a simplified technique for the isolation and long-term culture of human urothelial cells. Urothelial cells in vitro are capable of rapid proliferation and can be induced to form integrated stratifying cell layers in high calcium medium. Stratification-related changes are not necessarily accompanied by urothelial cell maturation and differentiation.
[Show abstract][Hide abstract] ABSTRACT: Between 1982 and 1992, 67 boys with posterior urethral valves were managed at our hospital, including 32 (48%) in whom the condition was detected prenatally. We examined the relationship between gestational age at detection and outcome at a median followup of 3.9 years (range 4 months to 10 years). Detection at or before 24 weeks of gestation predicted a poor outcome with 9 of 17 patients (53%) dead or in chronic renal failure at followup. Of the cases detected later in pregnancy only 1 had a poor outcome (p = 0.01). All of the cases detected after 24 weeks of gestation had had normal second trimester scans. Growth parameters for boys in the early and late detection groups were not statistically different. However, there was a significant association between renal failure and growth with 67% versus 14% having heights less than the 3rd percentile (p = 0.05). Respiratory distress at birth predicted a poor outcome, while the presence of palpable abdominal abnormalities or vesicoureteral reflux failed to predict outcome.
The Journal of Urology 09/1994; 152(2 Pt 2):698-701. · 3.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To assess morbidity in children with mild prenatally detected pelvicalyceal dilatation and to document the natural history of this ultrasound finding in post-natal life.
A retrospective (on-going) study was carried out in 29 children (39 kidneys) with mild dilatation confined to the pelvis and/or calices confirmed on initial post-natal ultrasound scan. Re-evaluation was carried out at a mean age of 4.2 years (range 1.5-7.8). Clinically, each patient's history, height, weight and blood pressure were recorded. On ultrasound examination the renal length, collecting system appearances and dimensions were recorded.
Vesico-ureteric reflux was demonstrated in 1 of 14 infants who underwent neonatal micturating cystourethrography. During cumulative follow-up totaling 122 years, there were only two documented episodes of urological morbidity, i.e. one episode of unexplained haematuria and one of urinary tract infection. By a mean age of 4.2 years the ultrasound appearances had reverted to normal in 69% of kidneys. In 31% dilatation persisted and was unchanged or diminished in severity. No case of increasing dilatation was seen. Renal growth was normal in 97% of kidneys.
Mild dilatation of the fetal urinary tract is a common prenatal ultrasound finding. When confined to the renal pelvis and/or calices it is of doubtful clinical significance and is associated with a low level of morbidity in infancy and early childhood. Invasive investigation in post-natal life is not justified.
[Show abstract][Hide abstract] ABSTRACT: Posterior urethral valves are an important cause of obstructive uropathy in the newborn. The diagnosis may be suspected on prenatal ultrasonography, the consistent finding being bilateral ureterohydronephrosis together with a thick-walled, persistently full bladder in a male fetus. There may also be variable degrees of oligohydramnios. This paper reports the case of siblings in whom posterior urethral valves were recognized prenatally. The first child progressed well after appropriate surgical management, but the second died within 24 h of birth.
[Show abstract][Hide abstract] ABSTRACT: An in vitro system for the growth of normal human urothelial cells has been developed. Urothelial cells were isolated from tissue samples in 38 patients and cultured in a defined serum-free medium. Confluent cell monolayers of 25 cm.2 were produced after 7 days in 32 cases. Subsequent subcultures at a 1:20 split ratio achieved confluency within another 7 days and a consistently high rate of growth was sustained for at least 7 passages. Characterization by immunofluorescence with a panel of antibodies demonstrated that the cultured cells were exclusively epithelial and retained the characteristic antigenic profile of normal urothelium, even after extended periods in culture. The only consistent cause of failure (6 of 38 cases) was bacterial contamination secondary to an underlying urinary tract infection in these patients.
The Journal of Urology 09/1993; 150(2 Pt 2):721-5. · 3.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Static isotope imaging with 99mtechnetium-dimercaptosuccinic acid was performed at a mean age of 34 days in 32 children (50 kidneys) whose vesicoureteral reflux had been identified as a result of prenatal ultrasound scanning. Three patterns of isotope uptake were observed: 1) noninfected primary vesicoureteral reflux (15 children, 24 kidneys), 2) noninfected secondary (obstructed) vesicoureteral reflux (9 patients, 11 kidneys) and 3) infected primary reflux (8 infants, 15 kidneys). In 20 pattern 1 kidneys (83%) renal morphology and differential isotope were normal. In the 4 kidneys (17%) that showed evidence of impaired function this took the form of global parenchymal loss, that is small kidneys rather than focal scarring. In pattern 2 the combination of fetal vesicoureteral reflux and obstruction was a potent cause of renal damage with total or near total loss of function in 7 of 9 refluxing units associated with posterior urethral valves and in 2 kidneys with secondary ureteropelvic junction obstruction. Appearances of focal scarring were confined in pattern 3 and were found in 4 kidneys (27%). This overall incidence of detectable renal damage was lower than expected. Even when infection occurs, prenatal diagnosis may lessen the risk of scarring by enabling treatment to be instituted promptly. The findings suggest that uncomplicated primary vesicoureteral reflux is a relatively benign insult to the fetal kidney and that reflux nephropathy found in children presenting clinically is the result of infected vesicoureteral reflux in postnatal life. Any comparison of published studies will prove difficult until there is a more standardized approach to imaging technique and patient selection.
The Journal of Urology 11/1992; 148(4):1229-31. · 3.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The Fowler Stephens operation and microvascular procedures were compared for the management of 23 intra-abdominal testes in 15 boys, 7 of whom had the prune belly syndrome. The Fowler Stephens operation was used for 15 testes (4 single-stage and 11 two-stage) and microvascular transfer was used for 8 testes. Eleven testes treated by the Fowler Stephens procedure (73%) and 7 treated by microvascular procedures (88%) were viable and intrascrotal after an average follow-up of 3.0 years, giving an overall success rate of 78%. Eight of 9 post-pubertal testes were biopsied. Severe maturational arrest was evident in all and carcinoma in situ was present in one.
British Journal of Urology 09/1991; 68(2):199-202.
[Show abstract][Hide abstract] ABSTRACT: This report analyses the characteristics and outcome of 25 infants with vesicoureteric reflux detected prenatally on the basis of dilatation of the fetal urinary tract. Sixteen infants had bilateral reflux--a total of 41 refluxing units. The high proportion of males (84%) contrasts with clinically presenting reflux, which is dominated by females. Prenatally diagnosed reflux is generally of a higher grade--usually grade IV. Eight children (32%) had coexistent congenital abnormalities. Chemoprophylaxis was completely effective in 17 children (68%), who remained infection-free; 3 children (12%) had a single urinary infection and were managed conservatively whilst 5 (20%) experienced 2 or more infections and required reimplantation or vesicostomy. Spontaneous cessation of reflux was observed in 6 (35%) of 17 refluxing ureters reassessed after a mean interval of 2.1 years. The significance of isotope findings was sometimes difficult to assess but results in 30 refluxing units support the concept that focal renal scarring is usually a consequence of infected reflux in postnatal life.
[Show abstract][Hide abstract] ABSTRACT: To assess the morbidity and mortality associated with a prenatal diagnosis of bilateral fetal uropathy.
Departments of radiology, paediatric surgery, obstetrics, and pathology in two teaching hospitals that serve as referral centres for the Yorkshire region.
126 Cases of fetal uropathy were referred either prenatally or postnatally from hospitals in Yorkshire between August 1982 and December 1987. The disease was bilateral in 54 cases and unilateral in 72 cases. In 14 cases bilateral fetal uropathy was associated with coexistent disease.
All cases were managed individually by an obstetrician after discussion with the radiologists and paediatric surgeons. Babies who survived were treated prophylactically with antibiotics after delivery and were operated on if appropriate.
Assessment of prognosis for long term renal function for each baby referred between August 1982 and December 1987: follow up ranged from six months to five years.
Of the 54 fetuses with bilateral fetal uropathy, 13 were terminated as the prenatal findings of ultrasonography were considered to be incompatible with long term survival. Ten of the liveborn babies died, five of renal or pulmonary insufficiency, or both, and five of associated congenital anomalies. Thirty one infants survived to follow up; four of these had serious coexistent disease and two had impaired renal function. Thus the overall mortality was 43% and the morbidity rate 19%. The renal anomaly was associated with other serious disease in 14 cases (26%) compared with two (3%) of the 72 cases of unilateral fetal uropathy. All but two of the 27 infants with isolated bilateral urinary tract disease had excellent prospects for survival.
Although bilateral fetal uropathy is associated with a high morbidity rate and mortality, careful prenatal assessment can help to identify fetuses with a poor prognosis. The outlook for a fetus with isolated renal disease if treated promptly after delivery is excellent and compares favourably with that reported after prenatal surgical intervention.
BMJ Clinical Research 06/1989; 298(6685):1419-20. · 14.09 Impact Factor