Local tissue reaction and histopathological characteristics of three different bulking agents: a rabbit model

ABSTRACT Purpose: We assessed the efficacy and safety of a single injection of three bulking agents over the short- and long-term follow-ups in rabbits. Dermal and preputial matrices were compared with Deflux (DxHA) injection. Material and methods: Twenty-four rabbits were divided into three groups. Group I (n=8) underwent the injection of a lyophilized dermal matrix (LDM) beneath the seromuscular layer of the bladder wall. Rabbits in group II (n=8) were injected with lyophilized preputial matrix (LPM). Rabbits of group III (n=8) were injected with DxHA as the control group. They were followed up for 1 and 6 months after the injection. Subcutaneous injection of all bulking agents was also performed in nude mice. Biopsies were stained with LCA (leukocyte common antibody), CD68, CD31, and CD34. Scanning electron microscopy (SEM) and MTT assay were also performed. Results: Immunohistochemistry staining with CD68 and LCA revealed higher inflammation grade in LDM as compared with LPM and DxHA. Fibrosis grade was also higher in LDM both in short- and long-term follow-ups. However, no significant difference was detected in CD31 and CD34 staining between control and experimental groups. SEM analysis showed that the particle size of LPM was more similar to DxHA. MTT assay revealed that cell proliferation was similar in DxHA, LDM, and LPM. In-vivo assay in nude mice model showed more promising results in LPM as compared with LDM. Conclusion: The long-term results demonstrated that LPM was more similar to Deflux with the least local tissue reaction, inflammation, and fibrosis grade.

Local tissue reaction and histopathological characteristics of three different bulking agents: a rabbit model.

PURPOSE: We assessed the efficacy and safety of a single injection of three bulking agents over the short- and long-term follow-ups in rabbits. Dermal and preputial matrices were compared with Deflux (DxHA) injection. MATERIAL AND METHODS: Twenty-four rabbits were divided into three groups. Group I (n=8) underwent the injection of a lyophilized dermal matrix (LDM) beneath the seromuscular layer of the bladder wall. Rabbits in group II (n=8) were injected with lyophilized preputial matrix (LPM). Rabbits of group III (n=8) were injected with DxHA as the control group. They were followed up for 1 and 6 months after the injection. Subcutaneous injection of all bulking agents was also performed in nude mice. Biopsies were stained with LCA (leukocyte common antibody), CD68, CD31, and CD34. Scanning electron microscopy (SEM) and MTT assay were also performed. RESULTS: Immunohistochemistry staining with CD68 and LCA revealed higher inflammation grade in LDM as compared with LPM and DxHA. Fibrosis grade was also higher in LDM both in short- and long-term follow-ups. However, no significant difference was detected in CD31 and CD34 staining between control and experimental groups. SEM analysis showed that the particle size of LPM was more similar to DxHA. MTT assay revealed that cell proliferation was similar in DxHA, LDM, and LPM. In-vivo assay in nude mice model showed more promising results in LPM as compared with LDM. CONCLUSION: The long-term results demonstrated that LPM was more similar to Deflux with the least local tissue reaction, inflammation, and fibrosis grade.

Prevention of Renal Scarring in Acute Pyelonephritis by Probiotic Therapy: an Experimental Study.

We evaluated the protective effects of probiotic administration as a prophylaxis treatment and immediately after fever onset in increasing the immune response and decreasing the renal scarring in a rat model of acute pyelonephritis. Twenty-four rats were apportioned to three groups. In GI (n = 8), the rats were injected with direct inoculation of Escherichia coli into the right kidney. In GII (n = 8), the rats received a probiotic regimen 1 month before E. coli injection and the probiotic regimen was continued for the next 2 months. In GIII (n = 8), the probiotic regimen was started just after E. coli injection and was continued for 2 months. Technetium-99m-DMSA renal scan, histopathological evaluations, concentrations of CA19-9, IgA, blood urea nitrogen (BUN), and creatinine were assessed 1 and 2 months post-injection. It took an average of 4.2 ± 1.1 h between the injection and onset of fever in GI and GII. In GIII, this period was longer (7.5 ± 1.4). Probiotic administration resulted in reduction of interstitial fibrosis and tubular and glomerular atrophy in GII in all follow-ups. Technetium-99m-DMSA renal scan showed that the right kidney reached near the normal cortical integrity (47%) in GII compared to GI (32%) after 2 months of injection. However, the renal integrity did not improve significantly in GIII (41%). In GII, CA19-9 was lower (p < 0.05), while the levels of serum and fecal IgA were higher (p < 0.05). Administration of the probiotic regimen in the rat model may decrease renal damage in pyelonephritis. In spite of better results in the prophylactic group compared to the treatment group, no strong evidence was found to prove the advantage of its prophylactic application over the treatment administration.

Application of Human Acellular Breast Dermal Matrix (ABDM) in Implant-Based Breast Reconstruction: An Experimental Study.

BACKGROUND: The use of acellular dermal matrices (ABDM) has become more common for breast reconstruction to improve postoperative outcomes. We evaluated the efficacy of breast reconstruction by the application of human ABDM in a sheep model. METHODS: The sheep in group I (GI) (N = 4) underwent the following procedures on the right side: (1) breast reconstruction using human ABDM after total mastectomy, (2) human ABDM under the skin, near the breast area and on the left side, (3) fat injection and human ABDM after partial mastectomy, and (4) replacement of ABDM in the abdominal wall far from the breast. Sheep in group II (GII) (N = 4) underwent the following procedures. On the right side: (1) breast reconstruction using ABDM after total mastectomy, (2) replacement of ABDM under the skin, near the breast area, and on the left side, (3) application of vicryl synthetic mesh after partial mastectomy and (4) replacement of mesh under the skin, near the breast area. RESULTS: Histological evaluations of decellularized skin scaffolds demonstrated a collagen-based matrix with preserved ECM and complete nuclear removal. Histological evaluations of implanted ABDM demonstrated a viable matrix with fibroblast infiltration and revascularization in all follow-ups. The overall surgical complication rate was significantly lower in the ABDM implant under the skin and near the breast in both short- and long-term follow-ups. CONCLUSION: The results of this study demonstrated that the application of novel prepared ABDMs has promising outcomes for breast reconstruction to provide total coverage without the need for breast expansion before implant placement. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors http://www.springer.com/00266.

Aortic valve conduit implantation in the descending thoracic aorta in a sheep model: The outcomes of pre-seeded scaffold.

BACKGROUND: We evaluated the outcomes of implanting pre-seeded decellularized aortic valve conduit (AVC) with bone marrow-derived mesenchymal stem cells (MSCs) in a sheep model. METHODS: Eight sheep AVCs were obtained under sterile conditions and decellularized by using detergent-based methods. Decellularized AVCs were seeded with autologous bone marrow-derived MSCs in a dynamic bioreactor system. Pre-seeded AVCs were implanted in the descending thoracic aorta in a sheep model. In all sheep, a decellularized pericardial patch was also anastomosed to the proximal part in order to reduce the incidence of rupture. Pathological evaluations, echocardiography, multislice computed tomography (CT), and CT angiography were performed for the evaluation of implanted AVCs. RESULTS: The longest survival period was 19 months in pre-seeded animals with complete recellularization at the long-term follow-up. Immunohistochemical staining for desmin, smooth muscle actin, and cytokeratin was significantly positive in the pre-seeded samples and reached near normal ranges. CT angiography revealed no intimal tearing after 18 months of follow-up. CONCLUSION: Pre-seeded AVCs with bone marrow-derived MSCs may have satisfactory results in postoperative cell seeding capabilities with promising functional potentiality. This modality may be beneficial and may provide a new era of biological grafts in cardiovascular surgery.

In-vivo trachea regeneration: fabrication of a tissue-engineered trachea in nude mice using the body as a natural bioreactor.

PURPOSE: To investigate the outcomes of implanting rat decellularized trachea scaffold (DTS) between the paravertebral muscles of nude mice using the body as a bioreactor for total graft recellularization. METHODS: The tracheas of four rats were aseptically resected and decellularized. To assess the efficiency of the decellularization procedure, all decellularized scaffolds and native control tissues were evaluated with scanning electron microscopy (SEM), DAPI staining, DNA quantification, biomechanical analyses and hydroxyproline measurement. They were then implanted between the paravertebral muscles of four nude mice. The biopsies were precisely evaluated at 1, 3, 6 and 12 months postoperatively for tracheal cartilage and soft tissue recellularization by staining for TTF1, CD34, S100 and leukocyte common antibody. RESULTS: Hematoxylin and eosin (H&E) staining, SEM and the tensile test confirmed the preservation of the tissue structure and the biophysical and biochemical properties of the DTS. The present study clearly demonstrated that the hydroxyproline content of the DTS was similar to that of the native tissue. On the other hand, in biopsy samples obtained after 12 months, histological evaluation showed superior organization and cell seeding in both the cartilage and connective tissues. CONCLUSION: This study demonstrated the feasibility of using a natural bioreactor for recellularizing DTS; this may have the potential to facilitate homologous transplantation for repairing segmental trachea defects.

The application of tissue-engineered preputial matrix and fibrin sealant for urethral reconstruction in rabbit model.

BACKGROUND: To introduce the role of fibrin sealant and preputial acellular matrix (PAM) as a new source of inert collagen matrix for urethral reconstruction. METHODS: A ventral urethral segmental defect was created in 24 male rabbits divided into four groups. In group 1 (G1), urethrotomy was closed in layers. In group 2 (G2), closure was followed by applying fibrin sealant. In groups 3 (G3) and 4 (G4), urethroplasty was performed with a patch graft of PAM, and in G4, fibrin sealant was also applied. Serial urethrography was performed before and after the operation. Then, the animals were euthanized, and their urethra was excised 1, 3, and 9 months postoperatively for further electron microscopic examination, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) technique, and immunohistochemical (IHC) staining with CD34, CD31, desmin, SMA, and α-actin. RESULTS: In G1 and G2, the fistula repair failed in all the time points. In G3 and G4, serial urethrography confirmed the maintenance of a wide urethral caliber without signs of strictures or extravasations. Satisfactory vascularity was observed in G3 and G4 during the whole study, which was more significant in G4 after 9 months of follow-up. The presence of a complete transitional cell layer was confirmed over the graft in G3 and G4 in all time points. IHC staining confirmed the effectiveness of fistula repair in G3 and G4, 3 months postoperatively. CONCLUSION: This rabbit model showed that PAM combined with fibrin sealant may herald a reliable option for repairing segmental urethral defects.

Neurovascular sparing vas clipping: last option for recurrent epididymo-orchitis in urethrovasal reflux due to urethral injury.

Acute scrotum is a critical clinical entity in children. This report presents a 12-year-old boy presented with recurrent epididymo-orchitis (EO) with a history of pelvic trauma and urethral disruption 10 years ago. Antegrade and retrograde studies confirmed urethrovasal reflux. The patient did not respond to prophylactic antibiotics, clean intermittent catheterization and endoscopic injection of bulking agent at the junction of the ejaculatory duct and posterior urethra. As the last option, neurovascular sparing vas clipping was performed and the patient made a full recovery. This is the first report of this technique in the treatment for recurrent EO caused by traumatic injury.

Sheep colon acellular matrix: Immunohistologic, biomechanical, scanning electron microscopic evaluation and collagen quantification.

Colon decellularization provides three-dimensional biologic scaffold without any cell elements with preservation of extracellular matrix in order to enable autologous cell seeding for tissue augmentation without any immunological response. This study was performed to investigate the safety and feasibility of sheep colon decellularization as a first step of colon tissue engineering. The process of sheep colon decellularization was done in four stages which included scaffold preparation, histologic examination and microscopic investigations to reveal the remaining cellular deposits, biomechanical evaluation and collagen quantification studies by measurement of hydroxyproline content of normal and decellularized sheep colon. Decellularized colon scaffold revealed complete cell removal under a light microscope while 4',6-diamidino-2-phenylindole, di-hydrochloride (DAPI) staining confirmed no deoxyribonucleic acid (DNA) residues. Decellularized colon displayed preserved ultrastructure, comparable biophysical properties (resistance to unidirectional stretch forces) and higher hydroxyproline content. The results of biomechanical tests proved that the decellularized matrix did not bear any unexpected damages or structural changes which would make it unable to tolerate in vivo forces and stretches. The microscopic images captured after staining the tissue with Picro-sirius red also showed that the collagen in extracellular matrix is well preserved; this was confirmed by scanning electron microscopy. This implies that the scaffold prepared by this method is suitable for tissue augmentation or transplantation.

Establishment of a patient-derived Wilms' tumor xenograft model: a promising tool for individualized cancer therapy.

OBJECTIVE: Lack of appropriate approaches that reliably predict response of Wilms' tumor (WT) to anticancer agents remains a major deficiency in clinical practice of individualized cancer therapy. The aim of this study was to establish a patient-derived tumor tissue (PDTT) xenograft model of WT for individualized chemotherapeutic regimen selection in accordance with the patient's tumor nature. MATERIAL AND METHODS: Tumor specimens of a primary WT were orthotopically implanted into three nude mice, and after 4 weeks xenografts were harvested for serial heterotopic transplantation in 20 nude mice that were divided into three experimental groups and one control group. In vitro and in vivo chemosensitivity to doxorubicin, actinomycin-D, and vincristine were evaluated. Hematoxylin and eosin (H&E) staining and immunohistochemical examination with desmin, vimentin, myogenin, and neuron-specific enolase (NSE) were also applied to determine histological stability of the xenograft during serial transplantation compared with the original tumor tissue. RESULTS: The xenograft model was successfully established. Histopathologic characteristics of the xenograft tumors were similar to the patient's tumor. Early passage of the PDTT showed a similar chemosensitivity pattern to the original tumor tissue. CONCLUSIONS: PDTT xenograft of WT provides an appropriate model for individualized cancer therapeutic regimen selection by means of its biological stability compared with original patient's tumor.

Combination of probiotics and antibiotics in the prevention of recurrent urinary tract infection in children.

OBJECTIVE: We examined the preventive effect of probiotic and antibiotics versus antibiotics alone, in children with recurrent urinary tract infections (RUTI) in a preliminary randomized clinical trial. METHODS: Between March 2007 and April 2011, children with the history of RUTI and unilateral vesicoureteral reflux (VUR) were randomly assigned to receive concomitant probiotic and antibiotics (Lactobacillus acidophilus and bifidobacterium lactis, 10(7)/ml, as 0.25 ml/kg three times a day regimen in addition to Nitrofurantoin, 1mg/kg daily (group I). In group II, all children received conventional prophylactic antibiotics alone (Nitrofurantoin, 1 mg/kg daily). Randomization was performed via using the random numerals table in a 1:1 manner with stratification by sex, age and grade of reflux. The urine examinations were done monthly and the incidence of UTI was evaluated in these two groups. FINDINGS: Forty-one children (age: 8.3±3.1 years) in group I and 44 children (age: 8.0±3.0 years) in group II were compared. During the course of three years, 39% in group I and 50% of participants in group II experienced RUTIs (P=0.4). Incidences of UTI - febrile and afebrile - reduced in both groups without any significant differences after two years of prophylaxis. Also, incidence of afebrile UTIs did not significantly differ (0.51±1.30 and 0.81±1.41 respectively, P =0.3); however, the incidence of febrile UTIs in particular were lower in group I (0.00±0.00 versus 0.13±0.40, P =0.03) in the last year. CONCLUSION: The consumption of probiotic and antibiotics in children with RUTI is safe and more effective in reducing the incidence of febrile UTI in comparison to prophylactic antibiotics alone.

Shunt-type and stop-type varicocele in adolescents: prognostic value of these two different hemodynamic patterns.

OBJECTIVE: To investigate different varicocele hemodynamic patterns (shunt type vs. stop type) as predictors of new-onset testicular hypotrophy or recurrence after varicocelectomy. DESIGN: Prospective clinical study. SETTING: Department of pediatric urology, children's hospital medical center. PATIENT(S): Seventy-four children and adolescents with varicocele. INTERVENTION(S): Based on ultrasound findings, patients were classified into shunt-type or stop-type varicocele groups. All patients with stop-type varicocele and testicular volume discrepancy of ≥20% underwent retroperitoneal varicocelectomy and internal spermatic vein ligation. Patients with shunt-type varicocele and ≥20% testicular asymmetry were randomly divided to undergo either retroperitoneal varicocelectomy (ligation of internal spermatic vein) or inguinal varicocelectomy with ligation of both internal and external spermatic veins. Patients with testicular volume discrepancy of <20% were put on conservative management. MAIN OUTCOME MEASURE(S): Occurrence of new-onset asymmetry among untreated patients and recurrence rate among operated patients. RESULT(S): Patients with shunt-type varicocele who did not undergo surgery revealed a higher risk of developing asymmetry during follow-up (odds ratio 10.5). Ligation of both internal and external spermatic veins in shunt-type patients was associated with significantly decreased rate of recurrence (6.3%) compared with retroperitoneal approach (30.8%). CONCLUSION(S): Shunt-type varicocele is associated with a higher risk of testicular hypotrophy among untreated patients. In addition, recurrence rate in shunt-type patients who underwent inguinal varicocelectomy was lower compared with retroperitoneal approach.

Single-donor fibrin sealant for repair of urethrocutaneous fistulae following multiple hypospadias and epispadias repairs.

PURPOSE: To evaluate the efficacy of fibrin sealant for repair of urethrocutaneous fistula after multiple failed hypospadias and epispadias surgeries. MATERIALS AND METHODS: The study population comprised 11 boys (mean age 12.18 years) with history of hypospadias or epispadias and at least two failed fistula repair operations leading to recurrent urethrocutaneous fistula. During the operation, single-donor fibrin glue, either from the patient (7) or a parent (4), was applied over the suture lines and beneath the skin. A urethral catheter was kept in place for 7-10 days. Follow up ranged from 6 to 24 months (mean 12.63 months). RESULTS: Nine patients had an uneventful postoperative course. In one patient with a large fistula, partial wound dehiscence occurred. In another patient with complete hypospadias, hematoma formation caused skin dehiscence but the urethra remained intact. Both cases recovered after 6 months with no further intervention. No fistula recurrence was reported during follow up. CONCLUSION: Single-donor fibrin glue could be a useful adjunct to surgical management of patients after multiple failed attempts at hypospadias or epispadias fistula repair. Moreover, this product improves the safety margin regarding the risk of disease transmission.