The Buccal Mucosa Fenestrated Graft for Urethroplasty in Rabbits

ARTICLE INFO _________________________________________________________ ___________________ IBJU | THE BUCCAL MUCOSA FENESTRATED GRAFT FOR URETHROPLASTY IN RABBITS 826 In patients with previous surgery or severe hypospadias, Bracka described a two-stage repair with a free graft of buccal mucosa, reaching a success rate of approximately 87% and accounting for only 5.7% of fi stulas and 7% of stenosis (7). However, when regarding major defects, it is sometimes necessary to use many segments of buccal mucosa or the combination of buccal mucosa and skin to cover the entire surface and reconstruct the urethral plate (8). In these cases, the use of fenestrated grafting simulating mesh grafting, a technique well known in correcting major defects especially by using skin grafting in burn reconstructive plastic surgery, could be an interesting option to increase the area to be covered by grafting in situations where the availability of donor tissue is limited (9). Our proposal was to evaluate the histological integration process of fenestrated buccal mucosal grafting in the corpora cavernosa as the fi rst stage of urethral reconstruction in an experimental model in rabbits. MATERIALS AND METHODS After approval by the Research Ethics Committee of the Universidade Federal de São Paulo Hospital São Paulo, the study included a total of 16 New Zealand male rabbits, aged approximately 8 weeks and weighting 2.0 to 2.5 kg. Before the surgeries, the animals were acclimated in the Department of Experimental Surgery of the Universidade Federal de São Paulo for a period of adjustment of 72 to 96 hours. The study was performed in 4 months, including the adjustment period, 12 weeks of the longest interval between intervention and sacrifi ce and fi nally the period of histological analysis. Anesthesia was initiated by the intramuscular administration of a preanesthetic agent (Midazolam 5mg/kg) and hydration through a peripheral vein. Anesthesia was complemented with an intramuscular injection of an anesthetic solution containing ketamine hydrochloride 30 mg/kg + xylazine 5mg/kg and penile nerve block with bupivacaine 0.25% and lidocaine 1% without vasoconstriction (10,11). The surgical procedure was carried out under sterile conditions and the use of a magnifying glass of 3.5X. First the urethra was catheterized with a urethral catheter of 8 Fr and the perineal fold between the penis and the anus of the animal was sectionalized, thus allowing access to the urethra (Figures 1A and 1B). Buck’s fascia was incised at the junction of the corpus spongiosum and the corpus cavernosum on each side, thus allowing the structures to be isolated (Figure-1C). The corpus spongiosum and urethra were then exposed and completely sectioned with a scalpel, resulting in two stumps and defi ning a lengthy urethral defect, similar to a proximal hypospadia. Part of the distal urethral stump was excised to obtain complete exposure of the ventral surface of the corpus cavernosum (Figure-1D). To avoid a greater retraction or stenosis, the proximal urethral stump was fi xed at the base of the penis and perineum with a 5.0 catgut® suture. Next, buccal mucosa was extracted from the rabbit’s jugal region. The donating site was exposed with interrupted stitches using 5.0 catgut® (Figure-2A). A centesimal solution of lidocaine at 1% with vasoconstrictor (adrenaline 1:200,000) was then injected locally into the submucosa (Figure-2B), thus facilitating the withdrawal of a buccal mucosa fragment 1.0 cm long and 0.4 cm wide (Figure-2C). After review of the hemostasis the donor region was not sutured, the wound healed by itself. The buccal mucosa graft was harvested and fenestrated through two cruciform incisions created in distal portions of its longitudinal axis, approximately 1 mm from the superior and side edges. These incisions allowed for a macroscopical augmentation of 25% of the graft’s longest strip, which went from 10 mm to 12.5 mm (Figures 2C and 2D). The fenestrated buccal mucosa graft was then brought to the urethral surgical area and applied onto the defect created in the ventral region of the penis with six interrupted stitches of polydioxanone suture (PDS® II) 6-0. The submucosa area of the graft was merged with the ventral surface of the exposed corpus cavernosum, thus constituting the neourethra plate. After the main part of urethral surgery, the penile fold was then partially rebuilt with separate stitches of 5.0 catgut® suture, while the patency of the urethrostomy was maintained without the need of a catheter. IBJU | THE BUCCAL MUCOSA FENESTRATED GRAFT FOR URETHROPLASTY IN RABBITS 827 Experimental animals were divided into 4 equal groups of 4 each and sacrifi ced 2, 4, 8 and 12 weeks after surgery, respectively. The genital of each animal was examined according to its ventral surface aspect. The penises were sectioned along the base, allowing the withdrawal of surgical parts that were immediately fi xed in 10% formalin. These fi xed segments were longitudinally sectioned, from the glans to the base of the penis, to allow for a full analysis of the two fenestrated areas in relation to the graft itself. It was compared the integration and histological changes that occurred during the different times of sacrifi ce. All blocks were cut to produce segments of 5 microns in thickness and stained with hematoxylin-eosin (HE) and Masson’s trichrome. The animal group was not revealed to the pathologist for histological evaluation enabling an unbiased analysis. The infl ammatory response was classifi ed as acute, when there was infi ltration by polymorphonuclear cells, and chronic, with infi ltration by mononuclear cells. We adopted a semi-quantitative assessment of the intensity of infl ammation, graded on a 0 to 4 + scale, as follows: (0) no infl ammatory infi ltrate, (1+) minimal infl ammation, (2+) moderate infl ammation, (3+) severe infl ammation and (4+) fi nding of aggregates of leukocytes / lymphocytes with formation of micro-abscesses. The degree of sub-epithelial fi brosis was also analyzed in a semi-quantitative scale, graded from 0 to 3+, as follows: (0) no scarring, (1+) minimal scarring, (2+) moderate scarring and (3+) severe fi brotic scar.