C30 SURGICAL MANAGEMENT OF UNCOMMON UROGENITAL SINUS ANOMALIES

C30 SURGICAL MANAGEMENT OF UNCOMMON UROGENITAL SINUS ANOMALIES

Transplantation, Bucharest, Romania, 2Fundeni Clinical Institute, Dept. of ICU, Bucharest, Romania Introduction & Objectives: Barbagli’s dorsal onlay ...

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Transplantation, Bucharest, Romania, 2Fundeni Clinical Institute, Dept. of ICU, Bucharest, Romania Introduction & Objectives: Barbagli’s dorsal onlay graft using buccal mucosa (BMG) proved to be the most effective and reliable technique in the urethral surgeons armamentarium. Membranous or proximal bulbar urethral strictures occur after about 3-6% TURP operations. Classical treatment (repeated dilatations or urethrotomies), because of the risk of incontinence, is often ineffective and has a poor quality of life. We present the surgical technique and the results of Barbagli’s dorsal onlay BMG in the repair of “sphincter” strictures post TURP. Material & Methods: Between Jan. 2003 – Jan. 2011 we performed 357 urethral urethroplasties for urethral strictures or other urethral pathology. Barbagli’s dorsal onlay graft using prepuce or BMG was used in the majority of cases (212 pts of urethral strictures from the meatus to the external sphincter). From these, 58 pts were strictures after TURP, 21 involving proximal bulbar and membranous urethra, close to the external sphincter (“sphincter” strictures). The diagnosis protocol includes anamnesis, clinical examination, bipolar urethrography. In 16 pts we performed urethroscopy and internal urethrotomy 3 mo. prior to the urethroplasty, in order to visualize the level of the stricture and the integrity of the external sphincter. We exclude from the operation patients with history of urinary incontinence after TURP with cervico-prostatic sclerosis and with external sphincter damages. Our surgical technique involves a “Y” inverted perineal incision and complete dissection of the bulbar urethra until we reach the urogenital (UG) diaphragm. We performed dorsal incision of the stricture. The buccal mucosa graft is quilted on the inferior fascia of the UG diaphragm. The margins of the urethral incision are sutured to the graft. A 20 Fr Foley catheter is inseted for 14 days. Results: We register no major intra or postoperative complications (3 perineal hematomas treated conservatively). After median 20 mo. follow-up interval (range 10-34 mo.) we register moderate incontinence in 4 pts. (19%) and recurrence of the stricture in 5 pts. (24%). The medium term success rate was 76% (pts. voiding complete with more than 12 ml/s peak flow rate and a good quality of life). Conclusions: Similar to the corpora, inferior fascia of the UG diaphragm is a good grafting bed for BMG. Because of low rate of complications, especially incontinence, Barbagli’s dorsal onlay BMG represents a good treatment choice of the majority of recurred post TURP “sphincter” strictures, instead of repeated dilatations or multiple urethrotomies.

C29

Potential of amniotic fluid-derived stem cells for urological tissue engineering

Olkowska J.1, Gurtowska N.1, Bajek A.1, Walentowicz-Sadlecka M.2, Sadlecki P.2, Kloskowski T.1, Tworkiewicz J.1, Grabiec M.2, Drewa T.1 1 Nicolaus Copernicus University, Collegium Medicum, Dept. of Tissue Engineering, Bydgoszcz, Poland, 2Nicolaus Copernicus University, Collegium Medicum, Dept. of Obstetrics, Gynecology and Gynecologic Oncology, Bydgoszcz, Poland Introduction & Objectives: Cells present in amniotic fluid have been used in prenatal diagnosis for more than decades. Recent reports suggest that amniotic fluid contains stem cells with a high proliferation and differentiation potential, which can be used in clinical practice. Tissue engineering techniques and stem cells are a promising tool in overcoming problems of damaged urinary tract tissues and complications of current procedures. The aim of this study was to isolate and characterize the human amniotic fluid-derived stem cells (AFSCs). Material & Methods: The human amniotic fluid (2-3 ml) was obtained during planned amniopunctions and centrifuged at 350g for 10 min. After washing with PBS, cells were gently suspended and plated. When reached confluence the colony forming assay, differentiation potential and aging process were performed. Briefly, cells after 14 days culture in 6-well plates were stained with 2% rhodamine B, and then colonies were counted to evaluate the clonogenicity potential. For the osteogenic differentiation potential cells were plated at a density of 12 x 103 cells/well on 6-well plates, and cultured in 2 ml/well of osteogenic medium at 37°C in a humidified atmosphere (5% CO2) for 14 days. Osteogenic differentiation of AFSCs was assessed using von Kossa assay. Aging of these cells was evaluated by morphological changes in subsequent passages observation in an inverted optics microscope. Results: AFSCs were allowed to adhere to plastic culture dishes and were grown till 6 passage. These cells formed an average of 8,2 ± 1,7 colonies. The clonogenicity that correlates with number of stem cells was 0,16%. Cells cultured in medium supplemented with differentiation factors toward osteogenic lineage showed change of morphology, from fibroblast-like to polygonal, after 5 days. In subsequent passages, the size of cells has been increasing and the density of growth in culture has been diminishing. Cells were grown till 6 passages, then stopped to proliferate. Conclusions: In conclusion, even if further investigations are required, the results obtained in this study support the finding that amniotic fluid contains cells with high proliferation and differentiation potential. These stem cells features make them good candidates for further in vitro and in vivo studies for urological disorders treatment.

Eur Urol Suppl 2011;10(9):620

C30

Surgical management of uncommon urogenital sinus anomalies

Ardelean M.A.1, Ionescu S.2, Tica C.3, Varik K.4, Schimpl G.1 1 Paracelsus Medical University, Dept. of Pediatric Surgery, Salzburg, Austria, 2 Emergency Children Hospital “Marie Curie”, Dept. of Pediatric Surgery, Bucharest, Romania, 3Emergency Hospital Constanta, Dept. of Pediatric Surgery, Constanta, Romania, 4University Hospital, Dept. of Pediatric Surgery, Tartu, Estonia Introduction & Objectives: The aim of the study is to present the challenging surgical treatment of rare forms of urogenital sinus (UGS) anomalies. Material & Methods: Many studies published in the last 15 years contribute to the improvement of treatment of children with UGS associated with CAH, cloacal persistence, or with “simple forms” of UGS. Five patients with rare forms of UGS, their surgical correction and outcomes, are reviewed. Results: A 7-year-old girl with posterior cloaca, accessory urethra, hydrocolpos and urine incontinence. Correction was achieved by urogenital mobilization (UM). The child is continent and has a normal calibrated vagina. An 8-monthold girl with posterior cloaca, accessory urethra and hydronephrosis. Surgical treatment consisted of an UM with a transanorectal approach. She is continent. A 10-year-old girl with urogenital sinus, agenesis of the bladder and vagina. An ileal bladder replacement, ureteral reimplantation, and continent urinary diversion were performed. She is socially continent. A 2-year-old girl with urethral agenesis, megabladder, narrow vesicovaginal fistula and introitus stenosis. To date the patient has a continent urinary diversion. She is scheduled for introitus plasty at adolescence. A 9-year-old girl with UGS persistence after anorectoplasty in infancy (at birth: cloacal persistence, pseudoexstrophy). The correction of UGS by UM and the correction of outer genitalia were undertaken. The patient is continent. Conclusions: Patients with UGS and normal anorectum should be continent after repair. In most patients with complex abnormalities social continence can be achieved. Vaginal dilatations before puberty are questionable. Vaginal agenesis or hypoplasia requires a vaginoplasty performed in adolescence when the patient has reached psychological maturity.

C31

Urine as a potential source of stem cells for urological tissue reconstruction

Tworkiewicz J.1, Adamowicz J.1, Bajek A.1, Kloskowski T.1, Drewa T.2 1 Nicolaus Copernicus University, Dept. of Tissue Engineering, Bydgoszcz, Poland, 2Oncology Centre Hospital, Dept. of Urology, Bydgoszcz, Poland Introduction & Objectives: Urine is investigated to be potential source of stem cells for urinary tract regeneration. Many research teams make an effort to isolate the stem cells from the urine. Zhang et al. were the first who performed succesful isolation of urine derived stem cells. A lot of different cell sources were evaluated for application in urologic regenerative medicine. Mesenchymal stem cells, hair follicle stem cells, adipose-derived stem cells were demonstrated to differentiate into urological structures. The isolation of these cells is invasive procedure for the patients and expensive to carry out. The method of isolation and culture of urinederived stem cells is still under development. Constitution of urine to be stem cell source for experimental therapies of regenerative medicine would be a milestone in urogenital repair. There is a need to focus on improving the method of urine derived stem cells isolation in order to increase its effectiveness and repeatability. Harvesting stem cells from urine is a perfect idea for wide introducing stem cellsbase therapies for clinical practice due to low costs of cell isolation, easy availability for patient and a non-invasive procedure. Material & Methods: 20 urine samples (300ml) were collected from 4 healthy male individuals. They included fresh urine, morning urine, urine voided by urethral catheter under sterile conditions and urine voided one day after catheterization. Each urine sample was centrifuged at 500g for 5min. The supernatant was discarded and the cell palet was resuispended in complete medium containing DMEM/Ham F12 (PAA, Austria) supplemented with 10% FBS (PAA, Austria), 10ng/ml bFGF (Sigma, Germany), 5g/ml amfotericin B and 100g/ml penicillin/ streptomycin (PAA, Austria). Cells were then counted with the use of trypan blue test and seeded on twelve-well culture plates (BD Bioscience, USA) and cultivated in complete medium in 37C. 10 primary cultures were established. Results: The average number of living cells isolated from the diluted (fresh) urine (s.g. 1.010) was 4x104 cells. The average number of living cells isolated from the morning urine (s.g. 1.032) approximately 9x104. Approximately 10x104 cells were obtained from urine voided by catheter. The highest number 2x106 cells were obtained from the urine collected one day after catheterization. Most of living cells did not attach to culture plates and were removed during medium changing in second day of in vitro cultivation. 5-10 cells attached to culture plates in each primary culture. The cells proliferated and formed colonies. The cell number obtained after one week of cultivation was approximately 1x104. Morphologically all cells were similar to urine derived stem cells. Conclusions: We report that it is possible to culture urine derived stem cells in medium supplemented only with FBS. Urine can be an alternative source of cells for therapeutic applications in urological tissue engineering.