Peter D. Furness

Laparoscopic Palomo Varicocele Ligation in Children and Adolescents

PURPOSEnWe evaluate our experience using the laparoscopic Palomo varicocele ligation (LPV) technique in male children and adolescents with varicoceles.nnnMATERIALS AND METHODSnBetween September 1994 and September 2002, 122 varicoceles were treated using LPV with either mass cord vascular clip application (68) or LigaSure (Valleylab, Boulder, Colorado) vascular sealing (54). All procedures were performed as day surgery cases and patients were allowed to return to normal activities as soon as they could tolerate them. Followup was scheduled for 6 to 12 weeks and 1 year postoperatively. Testicular size was evaluated using a Prader orchidometer.nnnRESULTSnOf the 122 patients 103 were evaluable at the initial postoperative visit and 96 at 1-year followup. Patient ages ranged from 9 to 19 years (mean 14.6). Operating time was 9 to 52 minutes (mean 28), although in the last 2 years (28 cases) mean operative time has decreased to 14.6 minutes. Indications for surgery included ipsilateral testicular hypotrophy in 84 cases, symptoms in 3 and parental choice in 16. At followup only 1 recurrent varicocele (vascular sealant group) was identified which was subsequently re-treated successfully using vascular sealant LPV. At 1 year 64 of 78 testes (82%) demonstrated catch-up growth and there was no evidence of testicular loss or persistent hypotrophy in the others. Reactive hydroceles were identified in 6 patients equally distributed between the 2 LPV techniques. Surgery was required on 2 of these hydroceles due to size, and the other 4 are small and are being observed. Temporary scrotal emphysema occurred in the vast majority of patients and 1 patient required laparoscopic closure of a small sigmoid serosal tear. No patients required narcotic medications for greater than 48 hours.nnnCONCLUSIONSnLPV is a highly successful method to correct varicoceles in young males with catch-up growth similar to series using other accepted standard techniques. LPV can be performed safely and rapidly as an outpatient and allows early return to activity. As with the open Palomo technique, hydrocele is a bothersome complication that may require a second surgical procedure if correction is warranted.

COCULTURE OF BLADDER UROTHELIAL AND SMOOTH MUSCLE CELLS ON SMALL INTESTINAL SUBMUCOSA: POTENTIAL APPLICATIONS FOR TISSUE ENGINEERING TECHNOLOGY

PURPOSEnSmall intestinal submucosa is a xenogenic, acellular, collagen rich membrane with inherent growth factors that has previously been shown to promote in vivo bladder regeneration. We evaluate in vitro use of small intestinal submucosa to support the individual and combined growth of bladder urothelial cells and smooth muscle cells for potential use in tissue engineering techniques, and in vitro study of the cellular mechanisms involved in bladder regeneration.nnnMATERIALS AND METHODSnPrimary cultures of human bladder urothelial cells and smooth muscle cells were established using standard enzymatic digestion or explant techniques. Cultured cells were then seeded on small intestinal submucosa at a density of 1 x 105 cells per cm.2, incubated and harvested at 3, 7, 14 and 28 days. The 5 separate culture methods evaluated were urothelial cells seeded alone on the mucosal surface of small intestinal submucosa, smooth muscle cells seeded alone on the mucosal surface, layered coculture of smooth muscle cells seeded on the mucosal surface followed by urothelial cells 1 hour later, sandwich coculture of smooth muscle cells seeded on the serosal surface followed by seeding of urothelial cells on the mucosal surface 24 hours later, and mixed coculture of urothelial cells and smooth muscle cells mixed and seeded together on the mucosal surface. Following harvesting at the designated time points small intestinal submucosa cell constructs were formalin fixed and processed for routine histology including Masson trichrome staining. Specific cell growth characteristics were studied with particular attention to cell morphology, cell proliferation and layering, cell sorting, presence of a pseudostratified urothelium and matrix penetrance. To aid in the identification of smooth muscle cells and urothelial cells in the coculture groups, immunohistochemical analysis was performed with antibodies to alpha-smooth muscle actin and cytokeratins AE1/AE3.nnnRESULTSnProgressive 3-dimensional growth of urothelial cells and smooth muscle cells occurred in vitro on small intestinal submucosa. When seeded alone urothelial cells and smooth muscle cells grew in several layers with minimal to no matrix penetration. In contrast, layered, mixed and sandwich coculture methods demonstrated significant enhancement of smooth muscle cell penetration of the membrane. The layered and sandwich coculture techniques resulted in organized cell sorting, formation of a well-defined pseudostratified urothelium and multilayered smooth muscle cells with enhanced matrix penetration. With the mixed coculture technique there was no evidence of cell sorting although matrix penetrance by the smooth muscle cells was evident. Immunohistochemical studies demonstrated that urothelial cells and smooth muscle cells maintain the expression of the phenotypic markers of differentiation alpha-smooth muscle actin and cytokeratins AE1/AE3.nnnCONCLUSIONSnSmall intestinal submucosa supports the 3-dimensional growth of human bladder cells in vitro. Successful combined growth of bladder cells on small intestinal submucosa with different seeding techniques has important future clinical implications with respect to tissue engineering technology. The results of our study demonstrate that there are important smooth muscle cell-epithelial cell interactions involved in determining the type of in vitro cell growth that occurs on small intestinal submucosa. Small intestinal submucosa is a valuable tool for in vitro study of the cell-cell and cell-matrix interactions that are involved in regeneration and various disease processes of the bladder.

Snodgrass hypospadias repair with vascularized dartos flap: the perfect repair for virgin cases of hypospadias?

PURPOSEnSince its introduction, the Snodgrass hypospadias repair has been applied to virtually all forms of hypospadias repair. However, fistula rates have still been reported to be as high as 5% from large center, multiple surgeon studies and 16% from smaller center studies. We report on the use of the Snodgrass repair in conjunction with routine use of a vascularized dartos flap and 2-layer closure of the neourethra from multiple institutions.nnnMATERIALS AND METHODSnRecords of patients who underwent a primary 1-stage hypospadias repair with the Snodgrass technique in conjunction with vascularized dartos flap coverage were reviewed. Nearly identical surgical technique was used by all 6 surgeons in each case, which included a 2-layer closure of the neourethra, preservation of the well vascularized periurethral tissue and routine use of vascularized dartos flap coverage. A total of 514 cases were identified, including 414 with distal and 100 with midshaft or proximal hypospadias. Stents were used in 292 of the 514 repairs.nnnRESULTSnOf the 414 distal cases there were no fistulas and 1 case of meatal stenosis. Of the 100 proximal cases there were 3 fistulas and 1 case of meatal stenosis. The overall complication rate was less than 1% for all cases combined.nnnCONCLUSIONSnThis series represents the largest reported multi-institutional experience with the Snodgrass technique. When used in conjunction with vascularized dartos flap coverage, 2-layer closure of the neourethra and special attention to preservation of the periurethral vascular supply, this repair can be performed with a near 0 complication rate. We believe that this is the optimal repair for routine cases of hypospadias.

MULTI-INSTITUTIONAL STUDY OF TESTICULAR, MICROLITHIASIS IN CHILDHOOD: A BENIGN OR PREMALIGNANT CONDITION?

PURPOSEnTesticular microlithiasis, a rare ultrasonographic diagnosis in children, has been shown to coexist in benign and malignant conditions. The natural history of incidentally discovered testicular microlithiasis has not been well defined in the pediatric population. The concern that testicular microlithiasis may be a premalignant condition has been raised. Reports indicate as much as a 45% association of germ cell tumors with testicular microlithiasis at the time of tumor diagnosis and there have been 4 cases of interval testicular tumor development associated with preexisting testicular microlithiasis. To address this issue we performed a multi-institutional study to evaluate children with incidentally diagnosed testicular microlithiasis.nnnMATERIALS AND METHODSnData on 26 patients with a mean age of 12.3 years at presentation with incidentally discovered testicular microlithiasis were collected from 7 institutions. Presenting scrotal conditions were reviewed. Two children with a previous testicular malignancy were excluded from study.nnnRESULTSnFollowup ranged from 1 month to 7 years (mean 27.6 months). Testicular biopsy and tumor marker (alpha-fetoprotein and beta-human chorionic gonadotropin) determinations were performed in 9 and 15 patients, respectively. To date no testicular tumor has developed during clinical followup.nnnCONCLUSIONSnOur multi-institutional study has not yet shown a trend toward the malignant degeneration of incidentally diagnosed testicular microlithiasis in children. However, we still advocate close surveillance of patients with testicular microlithiasis, such as yearly testicular ultrasound, physical examination, and judicious tumor marker determinations. We propose that a registry be started to follow prospectively patients with testicular microlithiasis to define its significance better.

ELEVATED BLADDER URINE CONCENTRATION OF TRANSFORMING GROWTH FACTOR-beta 1 CORRELATES WITH UPPER URINARY TRACT OBSTRUCTION IN CHILDREN

PURPOSEnWe evaluated urinary transforming growth factor-beta1 (TGF-beta1) concentration in children with upper urinary tract obstruction as a potential tool for supporting the diagnosis of clinically significant obstruction.nnnMATERIALS AND METHODSnRenal pelvic and bladder urine samples were obtained for analysis from 30 patients a median of 5 months old who underwent surgery for obstruction at the ureteropelvic (29) and ureterovesical (1)junctions. Urinary TGF-beta1 concentration was measured using a quantitative sandwich enzyme-linked immunoassay technique. Bladder urine TGF-beta1 in patients with obstruction was compared with that in controls. In addition, we compared renal pelvic and bladder urine TGF-beta1 in patients with obstruction.nnnRESULTSnMean bladder urine TGF-beta1 plus or minus standard error of mean was 4-fold higher in patients with upper tract obstruction than in controls (195 +/- 29 versus 47 +/- 7 pg./mg. creatinine, p <0.001). In the obstructed group mean TGF-beta1 in the renal pelvic urine was 378 +/-86 pg./mg. creatinine, or twice that of the bladder urine (p = 0.02).nnnCONCLUSIONSnBladder urine TGF-beta1 in patients with upper urinary tract obstruction is significantly elevated compared with that in controls. To our knowledge our study is the first to identify a bladder urinary marker that correlates with upper urinary tract obstruction with greater than 90% sensitivity. Measuring TGF-beta1 in a voided bladder urine sample may provide an objective and noninvasive test for assisting in the diagnosis of upper urinary tract obstruction.

CHARACTERIZATION OF CULTURED BLADDER SMOOTH MUSCLE CELLS: ASSESSMENT OF IN VITRO CONTRACTILITY

PURPOSEnThe contractile properties of in vitro cultured bladder smooth muscle cells (SMC) are unknown. This study characterized the in vitro contractile response of human and rat bladder SMC to several pharmacological agonists known to induce in vivo contraction of intact bladder muscle.nnnMATERIALS AND METHODSnHuman and rat bladder SMC were seeded separately within attached collagen lattices. Contractility of SMC was analyzed by measuring alterations in lattice diameter after exposure and release to the following contractile agonists: carbachol (10(-7)-10(-3) microM), calcium-ionophore (10 microM), lysophosphatidic acid (LPA) (1 microM), endothelin (0.1 microM), KCl (3.33 mmicroM) angiotensin II (10 microM), and serotonin (100 microM). Results were recorded as a mean reduction of the lattice diameter. In addition, immunohistochemical analysis for phenotypic markers of smooth muscle cell differentiation was performed on bladder SMC cultured within collagen lattices. Human palmar fascia fibroblasts, which have been previously well characterized by in vitro contractility and immunohistochemistry, were tested in parallel and used as controls for all the above experiments.nnnRESULTSnHuman SMC had significant contractile responses to calcium-ionophore (31% +/- 4 relative percent contraction, p <0.05), LPA (34% +/- 4, p <0.05), and endothelin (37 +/- 5%, p <05). There was no significant contraction in response to carbachol, angiotensin II, KCl, or serotonin. Rat bladder SMC had a similar contractile response but did not contract in response to endothelin. In contrast to human and rat bladder SMC, fibroblasts did not contract to calcium-ionophore.nnnCONCLUSIONSnIn vitro cultured bladder SMC demonstrate loss of contractile response to normal in vivo pharmacologic agonists. Both human and rat bladder SMC can be distinguished in vitro from fibroblasts based upon their lack of contractile response to calcium- ionophore. These results demonstrate the ability to further characterize cultured bladder SMC with in vitro contractility. Further characterization is essential if we are to advance our understanding of the clinical applicability of in vitro studies utilizing cultured bladder SMC.

Lessons learned from stomal complications in children with cutaneous catheterizable continent stomas

To evaluate the impact of various factors that might ultimately influence the stoma complication rate associated with the construction of a continent catheterizable urinary (CCU) and Malone antegrade colonic enema (MACE) stoma in children.

Gonadoblastoma and Turner Syndrome

PURPOSEnThe presence of a Y chromosome in the extrascrotal gonad of patients with intersex disorders has been associated with a high risk of GB and, potentially, GCT. Recently, modern sophisticated genotyping has revealed a subgroup of TS cases with a mosaic karyotype expressing a Y chromosome. We sought to evaluate this group of patients and address their risk of gonadoblastoma.nnnMATERIALS AND METHODSnWe reviewed the records and genotyping of all females newly diagnosed with TS between 1990 and 2002 at Childrens Hospital in Denver. All patients with TS and Y chromosome mosaicism underwent gonadectomy, and the specimens were evaluated for the presence of gonadoblastoma on histological analysis and to identify Y chromosome on genotyping.nnnRESULTSnA total of 192 girls with a clinical diagnosis of TS were identified between January 1990 and December 2002. Seven records were unavailable and 19 patients did not have karyotypic analyses available in the hospital charts. Of the remaining 166 patients 67 exhibited mosaic cell lines, of whom 8 had 45,X0/46,XY mosaic pattern and 59 had mosaic patterns without Y chromosomal elements. All 8 patients with Y mosaicism underwent uneventful laparoscopic gonadectomy on an outpatient basis. One patient observed to have bilateral dysgenetic gonads after gonadectomy was excluded from the study. Gonadoblastoma (bilateral 2 patients, unilateral 1) was detected in 3 of 7 patients (43%) with Y mosaicism.nnnCONCLUSIONSnIn our series 4.8% of evaluable patients with TS carried a 45,X0/46,XY karyotype. Gonadoblastoma can be evident even at an early age in streak gonads with Y mosaicism and may be bilateral. We recommend prophylactic laparoscopic gonadectomy of streak gonads in patients with TS who carry a Y mosaic genotype, because fertility is not an issue, surgical morbidity is minor and there may be a high potential for malignant transformation of gonadoblastomas in this population.

The use of small intestinal submucosa as an off-the-shelf urethral sling material for pediatric urinary incontinence

PURPOSEnUse of autologous rectus fascia for urethral slings in the pediatric population has produced reliable and predictable results. However, the potential morbidity and complications associated with harvesting the autologous rectus fascia have driven efforts to find a reliable off-the-shelf material for urethral slings. Small intestinal submucosa is a collagen based material that has been shown to promote tissue specific regeneration in a variety of organs. We report the clinical experience at 4 institutions with small intestinal submucosa for urethral slings.nnnMATERIALS AND METHODSnA total of 20 patients 3 to 18 years old (mean age 8.7) received urethral slings using the commercially available form of small intestinal submucosa (STRATASIS, Cook Urologic Spencer, Indiana) via a sling suspension procedure from a suprapubic approach.nnnRESULTSnThe material was consistently uniform to work with and user-friendly. All 20 patients tolerated the procedure well with no intraoperative complications. Postoperative followup has ranged from 9 to 26 months (mean 13), and 14 (70%) patients are completely dry (85% in females and 43% in males). Of the 14 dry patients 13 are on intermittent catheterization and 1 female with epispadias voids spontaneously.nnnCONCLUSIONSnThis report is the largest and longest followup series using small intestinal submucosa as an off the shelf urethral sling material in children. These continence rates are equal to autologous fascia without additional morbidity of graft harvest.

Use of Small Intestinal Submucosa for Corporal Body Grafting in Cases of Severe Penile Curvature

PURPOSEnSmall intestinal submucosa is a unique biomaterial that has been found to promote tissue specific regeneration in the urinary tract. We present our experimental and clinical experience with small intestinal submucosa (SurgiSis, Cook Biotech, Spencer, Indiana) for pediatric corporal body reconstruction.nnnMATERIAL AND METHODSnA total of 20 Fischer rats underwent implantation of a 7 x 3 mm. small intestinal submucosa graft following excision of an ellipse of tunica albuginea and 14 control animals underwent tunical excision with reimplantation of this autologous segment. The animals were euthanized, and the penis was sectioned and histologically studied at intervals of 1, 2, 4, 6, 16 and 24 weeks. In 15 pediatric patients small intestinal submucosa was used for corporal body grafting. The grafting procedure was performed along the ventral (hypospadias cases) or dorsal (epispadias cases) surface of the corporal bodies. The tunica albuginea was incised full thickness at the point of maximal curvature down to the cavernosal tissue and the defect was filled with a single layer of small intestinal submucosa.nnnRESULTSnMeasurements of the animal small intestinal submucosa grafts did not reveal significant graft contraction through 6 months. There was no graft expansion or ballooning after pharmacological induction of an artificial erection. Histologically, marked inflammation at 1 week precipitously decreased to a normal appearing tunica albuginea at 3 and 6 months. In all clinical cases small intestinal submucosa was found to be technically easy to handle. Mean followup is 14 months (range 5 to 26). All patients have a straight phallus as documented by observation of spontaneous erections or artificial erection at the time of stage 2 hypospadias repair. No complications occurred.nnnCONCLUSIONSnSmall intestinal submucosa demonstrates tissue specific regeneration properties in the rat and human tunica albuginea. It is an off-the-shelf material that is safe, technically easy to use and readily available.