PURPOSE: Tissue engineering has been used for bladder augmentations with small intestinal submucosa (SIS). Although favorable short-term outcomes have been reported, long-term followup has been poor. We investigate whether tissue engineering with stem cells improves the morphological and genetic composition. MATERIALS AND METHODS: A total of 33 Lewis rats (Harlan Laboratories, Indianapolis, Indiana) were used to investigate bladder augmentations with 4-layer SIS in certain groups, including the control group (sham operation), partial cystectomy with oversewn defect group (OG), augmentation with unseeded SIS group (USG) and augmentation with stem cell seeded SIS group (SSG). Bladders from 4 rats per group were harvested 1 and 3 months after surgery. Morphological analyses were performed using Masson's trichrome and immunohistochemical staining with cytokeratin AE1/AE3, smooth muscle alpha-actin and S100. Gene expression was evaluated using quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) for collagen I (CI), collagen III (CIII), cytokeratins 8 and 19, and smooth muscle myosin heavy chain (MHC). RESULTS: At 1 month trichrome staining revealed collagen admixed with indiscrete cells and morphology similar to that in controls in USG and SSG, respectively. Discrete smooth muscles fascicles and S100 staining were found in all groups except USG. Organized urothelium with increased basal cell layer staining was present in controls and SSG only. At 3 months increased collagen formation was present in OG and USG. Immunostaining showed hyperplasia of the urothelium with increased staining of the basal cell layer, discrete muscle fascicles and positive nerve staining in all groups. Using quantitative RT-PCR expression levels in SSG were more improved than in USG, especially for CI, CIII and MHC. This was further evident at 3 months when CI and CIII were over expressed in OG and USG but not in the control group or SSG. Furthermore, RT-PCR showed that cytokeratins 8 and 19, and MHC had greater expression levels in SSG than in USG. CONCLUSIONS: Bladder reconstitution occurs more rapidly using stem cell seeded SIS. Although in USG and SSG all 3 cellular constituents appear to develop by 3 months, only SSG had gene expression levels similar to those in controls. The results suggest an explanation for the fibrosis noted in unseeded SIS bladder augmentations and the possible solution using stem cells.
PURPOSE: Tissue engineering has been used for bladder augmentations with small intestinal submucosa (SIS). Although favorable short-term outcomes have been reported, long-term followup has been poor. We investigate whether tissue engineering with stem cells improves the morphological and genetic composition. MATERIALS AND METHODS: A total of 33 Lewis rats (Harlan Laboratories, Indianapolis, Indiana) were used to investigate bladder augmentations with 4-layer SIS in certain groups, including the control group (sham operation), partial cystectomy with oversewn defect group (OG), augmentation with unseeded SIS group (USG) and augmentation with stem cell seeded SIS group (SSG). Bladders from 4 rats per group were harvested 1 and 3 months after surgery. Morphological analyses were performed using Masson's trichrome and immunohistochemical staining with cytokeratin AE1/AE3, smooth muscle alpha-actin and S100. Gene expression was evaluated using quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) for collagen I (CI), collagen III (CIII), cytokeratins 8 and 19, and smooth muscle myosin heavy chain (MHC). RESULTS: At 1 month trichrome staining revealed collagen admixed with indiscrete cells and morphology similar to that in controls in USG and SSG, respectively. Discrete smooth muscles fascicles and S100 staining were found in all groups except USG. Organized urothelium with increased basal cell layer staining was present in controls and SSG only. At 3 months increased collagen formation was present in OG and USG. Immunostaining showed hyperplasia of the urothelium with increased staining of the basal cell layer, discrete muscle fascicles and positive nerve staining in all groups. Using quantitative RT-PCR expression levels in SSG were more improved than in USG, especially for CI, CIII and MHC. This was further evident at 3 months when CI and CIII were over expressed in OG and USG but not in the control group or SSG. Furthermore, RT-PCR showed that cytokeratins 8 and 19, and MHC had greater expression levels in SSG than in USG. CONCLUSIONS: Bladder reconstitution occurs more rapidly using stem cell seeded SIS. Although in USG and SSG all 3 cellular constituents appear to develop by 3 months, only SSG had gene expression levels similar to those in controls. The results suggest an explanation for the fibrosis noted in unseeded SIS bladder augmentations and the possible solution using stem cells.
Authors: Mila Torii Corrêa Leite; Luiz G Freitas-Filho; Andréia Silva Oliveira; Patrícia Semedo-Kuriki; Marcus Laks; Victor Eduardo Arrua Arias; Pedro S Peixoto Journal: Pediatr Surg Int Date: 2014-04 Impact factor: 1.827