**8. Pediatric urology: the prospect of urethral reconstruction using bovine amniotic membrane**

In the field of urology or pediatric surgery, uroepithelial reconstruction for several pathology is challenging. There is no synthetic material that is considered ideal as a substitute for the urethra and there is no research that firmly determine a good synthetic material to replace urethral defects. Strictures still found after the transplantation of acellular scaffold was also reported [73]. Therefore, the use of cell-seeded scaffold is proposed to be a better material used in urethral reconstruction. Bovine amniotic membrane has unique properties including anti-adhesive effects, bacteriostatic

### *Future of Bovine Amniotic Membrane: Bovine Membrane Application on Wound Healing… DOI: http://dx.doi.org/10.5772/intechopen.99313*

properties, wound protection, pain reduction, and epithelialization effects. Another characteristic of amniotic membrane is the lack of immunogenicity [26].

Amniotic membrane also has some advantage compared to other allographs, such as bladder mucosa, buccal mucosa, and also appendix tissue. First of all, amnion harvesting does not need extra surgery unlike other organ. Postoperative care is simpler with shorter hospital time. However, there is technical difficulty in handling the membrane. Due to the thinness of the amniotic membrane and the release of the amniotic layer to the chorion, there is a higher risk of perforation or tear or separation of the two layers during surgery [74].

Bovine amniotic membrane has been studied mostly on animal studies with promising effects. One experimental study for urinary bladder reconstruction by Bakhtiari in 2000 was using fresh and formalin based bovine amniotic membrane for canine. Urinary bladder and urethra share similar histology. Both are consisting of epithelium on the lumen surrounded by rich collagen connective tissue and muscle layer. Both are responsible for maintaining structural integrity of the organ and transporting or expelling the urine [75]. The surgical procedure was conducted by 5 cm resection from the cranial bladder. Postoperative graft using two types of bovine amniotic membrane was observed. Graft site observations on postoperative days 30 and 60 showed adhesion at the graft site (100%), the graft floating within the bladder lumen (40%), good graft adhesion to the bladder, and no evidence of leakage or fistulation. Histopathological examination revealed regeneration of uroepithelial tissue and smooth muscle at the graft site. In addition, congestion, edema, and inflammatory cell infiltration were also seen in two cases. According to this study, despite complications such as infection, release of amnion from the bladder, "less than normal" distension and adhesions of the bladder at the graft site, it can be argued that fresh and preserved bovine amnion acts as a scaffold for the repair of canine bladder defects. The regeneration of the urothelium and the presence of microscopic smooth muscle and the important complications that may occur from enterocystoplasty can encourage the use of bovine amniotic membrane for bladder reconstruction. However, long-term studies are still needed to assess other clinical and laboratory findings before measuring for clinical use [76].

In another study, Shakeri demonstrated the ability of the human amniotic membrane to induce epithelialization in experimental study on rabbits by reconstructing the urethra using the human amniotic membrane. The evaluation was conducted after 30 days post-operative. The result showed re-epithelialization of urethral without inflammation and tissue loss. The author also concluded that amniotic membrane is an inexpensive, easy, and biodegradable graft with very little antigen effect which seems to be the ideal solution for urethroplasty [74].

Amniotic membrane could be used as potential source for stem cell. Ghionzoli and Chung showed that it could be differentiated into smooth muscle and urothelial cell, both which are becoming integral parts creating urethral tissue [77, 78]. Despite using human amniotic membrane, some preclinical studies have also explored urological applications. Iijima demonstrated that amniotic membrane could successfully be used for bladder augmentation in rats. Human amniotic membrane-augmented bladder revealed regeneration of urothelium, detrusor smooth muscle, and nerve fibers within 3 months post-operatively. Bladder capacity was also found to be normal within 4 weeks post-operatively [79].

Pusateri evaluated placental membrane grafts for urethral replacement in rabbit model. The procedure consisted of mobilitation of urethra, dorsal urethrotomy and graft placement. Dorsal onlay urethroplasty was performed afterwards. Observation after 3 month showed urethral patency in all rabbits. On pathologic examination, urothelial cell replacement was observed in all rabbit without malignant transformation. Urothelium was intact and circumferentially normal in all sections of graft bed. On cystourethroscopy, there was no strictures, fistulas or massess reported [80].

Wang et al. looked further to this idea, namely using the collagen scaffolding of amniotic membrane as potential regenerative material in urethroplasty. The authors separated basement layer of amnion to retrieve denuded human amniotic scaffold. Rabbit urethral epithelial cell was inoculated on its surface and the response showed mild immune reaction with no rejection. This maximizes the biocompability of amniotic membrane making it potential biomaterial for urethral reconstruction [81]. Gunes took another approach by combining amniotic membrane and buccal mucosa for penile urethral reconstrucion in rabbit model. Both membrane were obtained from rabbit. Gunes compared whether buccal mucosa, amniotic membrane or both might be useful in urethroplasty. The best result of epithelial transformation was shown in combined group after 8 weeks with no complication regarding fistula or dehiscence observed [82].

Hariastawa et al. compared both bovine and human amniotic membrane for reconstruction of urethral defect in rabbit animal model. His study aimed to discover the difference in mucosal integrity between both groups. Epithelium layer was formed in both group with no significant ddifference of postoperative mucosal grading on day 7,14 and 28 postoperatively. Authors concluded that bovine amniotic membrane could be used as good, cheap alternative in urethral defect reconstruction [83]. In another study, Hariastawa used dried amniotic membrane scaffold with adipose derived-mesenchymal stem cell seeding for rabbit penile urethral reconstruction. Adipose cell was cultured from rabbit neck and mixed with fetal bovine serum. Viability of stem cell was tested before the surgical procedure. The urethral wall was cut transversely before the scaffold that had been seeded with stem cells was implanted as urethral graft. Urethroplasty was done afterwards. Post-operative clinical observation showed urethral integrity alongside the defect. Urethral specimen was harvested on day 28 post-operatively. The specimen was then observed using fluorescene microscope. Neovascularization and best epithelial transformation was seen in combined amniotic membrane and adipose-derived mesenchymal stem cell seeding group. The promising result showed that stem cell could be used as adjunct treatment for amniotic membrane application [84].

Although limited to human amniotic membrane, clinical researches have been accepted. The first clinical report regarding the use of human amniotic membrane for anterior urethral defect repair was reported by Razzaghi et al. This pilot study included patients with previous hypospadia repair [85]. Hypospadias repair remains a challenge due to many anatomical variation of the pathology, surgical techniques and comorbidities of patient. Complications vary from dehiscence, stenosis to fistula formation. Secondary or salvage procedure were often needed for failed primary repair [86, 87]. After reconstruction of neourethra and proper hemostatis from urehtroplasy, the allograft was used to cover the suture lines. Observation between 7 to 18 moths post-operatively showed no long term complications. Amniotic membrane graft was proposed as an applicable, low-cost, biodegradable cover for second hypospadias repair [85]. Oottamasathien et al. proposed that amniotic membrane could be used for reducing complication rate, particularly from high re-operation rate of hypospadias. The underlying premise is to provide a barrier layer with robust source of tissue, vascular growth factors and anti-inflammatory environment for soft tissue healing [88].

### **9. Summary**

Potential future application of bovine amniotic membrane could be explored widely. Broad number of biological properties found in amniotic membrane described above presents future studies. Preclinical and clinical researches could be used for basic scaffold for other applications. Combination with other biomaterials would be considered further.

*Future of Bovine Amniotic Membrane: Bovine Membrane Application on Wound Healing… DOI: http://dx.doi.org/10.5772/intechopen.99313*
