Precise treatment of pelvic fracture urethral injury associated with urethrorectal fistula.

OBJECTIVE: To evaluate the effect of a new strategy of transperineal anastomotic urethroplasty (TAU) with proximal transection in treating pelvic fracture urethral injury (PFUI) associated with urethrorectal fistula (URF). PATIENTS AND METHODS: A retrospective review of all patients treated by TAU with proximal transection and fistula repair for PFUI associated with URF was performed between August 2013 and July 2022. Information on demographics, peri-operative variables, and postoperative follow-up outcomes was collected. Successful surgery was defined as restoration of a uniform urethral calibre using flexible cystoscopy (third postoperative month) without strictures or leakage, with no further interventions required. Functional outcomes, including erectile function (assessed using the five-item International Index of Erectile Function) and urinary continence, were assessed. RESULTS: Forty patients diagnosed with PFUI associated with URF and treated by TAU with proximal transection and rectal fistula repair were enrolled. Six patients (15.0%) had a history of failed urethral reconstruction. The mean stenosis length and fistula diameter were 2.9 cm and 1.2 cm, respectively. All patients underwent faecal diversion before urethroplasty. After a median (range) follow-up of 45 (3-115) months, the final success rate was 90.0% (36/40). Postoperative complications included haematoma in three patients, epididymo-orchitis in three, wound infection in one, wound bleeding in one, delayed wound healing in three, and wound numbness in three. The overall incidence of postoperative erectile dysfunction reached 75.0%, with a median (range) score of 9 (0-19). Normal continence was achieved in 31 patients (77.5%). Occasional incontinence without the need for urinal pads occurred in eight patients, whereas one patient required urinal pads. CONCLUSIONS: Transperineal anastomotic urethroplasty with proximal transection is a precise and effective surgical strategy for treating PFUI associated with URF. This strategy ensures a high success rate and improves surgical efficiency.

Bacterial cellulose-based hydrogel with regulated rehydration and enhanced antibacterial activity for wound healing.

Bacterial cellulose (BC) hydrogels are promising medical biomaterials that have been widely used for tissue repair, wound healing and cartilage engineering. However, the high water content of BC hydrogels increases the difficulty of storage and transportation. Moreover, they will lose their original hydrogel structure after dehydration, which severely limits their practical applications. Introducing the bio-based polyelectrolytes is expected to solve this problem. Here, we modified BC and combined it with quaternized chitosan (QCS) via a chemical reaction to obtain a dehydrated dialdehyde bacterial cellulose/quaternized chitosan (DBC/QCS) hydrogel with repeated swelling behavior and good antibacterial properties. The hydrogel can recover the initial state on the macro scale with a swelling ratio over 1000 % and possesses excellent antimicrobial properties against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) with a killing rate of 80.8 % and 81.3 %, respectively. In addition, the hydrogel has excellent biocompatibility, which is conducive to the stretching of L929 cells. After 14 d of in vivo wound modeling in rats, it was found that the hydrogel loaded with pirfenidone (PFD) could promote collagen deposition and accelerate wound healing with scar prevention. This rehydratable hydrogel can be stored and transported under dry conditions, which is promising for practical applications.

Glycated serum albumin decreases connexin 43 phosphorylation in the corpus cavernosum.

Background: Glycated serum albumin (GSA) is an early glycosylation product that participates in diabetic vascular complications. This study examined the role of GSA in early damage to the corpus cavernosum and the involved mechanisms. Methods: Nine 8-week-old male Sprague-Dawley (SD) rats (250-300 g) were divided into the control (saline vehicle, n=3) and GSA (200 µg/kg, n=6) groups. The corpus cavernosum tissues were harvested. Phosphorylated and non-phosphorylated connexin 43 (Cx43), endothelial nitric oxide synthase (eNOS), phosphatidylinositol 3-kinase (PI3K), and serine-threonine kinase (Akt) were tested by immunohistochemistry and western blotting. Human umbilical vein endothelial cells (HUVECs) overexpressing Cx43 were used to analyze the Cx43 phosphorylation sites (S368, S262, Y265, S255, and S279/282) using western blotting. Results: The expression of phosphorylated Cx43 in the penis was significantly lower in GSA-treated rats than in controls. The expression levels of p-Cx43, p-eNOS, p-PI3K, and p-Akt were significantly decreased in HUVECs exposed to GSA in dose- and time-dependent manners. The most significant impact on all four proteins was observed with 1 µg/mL of GSA for 12 h. Phosphorylation at the S368, S262, Y265, S255, and S279/282 sites of Cx43 was downregulated by GSA, and S368 was the most significantly suppressed phosphorylation site compared with the other sites. Conclusions: GSA decreases the expression of p-Cx43 in the corpus cavernosum of rats. This effect might be also related to the decreased phosphorylation of p-eNOS, p-PI3K, and p-Akt, as well as by the downregulation of phosphorylation at the S368 site.

Comparison of the therapeutic effects of human umbilical cord blood-derived mesenchymal stem cells and adipose-derived stem cells on erectile dysfunction in a rat model of bilateral cavernous nerve injury.

Background: Cavernous nerve injury (CNI) is the leading cause of erectile dysfunction (ED) after radical prostatectomy and pelvic fracture. Transplantation of human adipose-derived stem cells (ASCs) has been widely used to restore erectile function in CNI-ED rats and patients. Umbilical cord blood-derived MSCs (CBMSCs) are similarly low immunogenic but much primitive compared to ASCs and more promising in large-scale commercial applications due to the extensive establishment of cord blood banks. However, whether CBMSCs and ASCs have differential therapeutic efficacy on CNI-ED and the underlying mechanisms are still not clear. Materials and methods: A bilateral cavernous nerve injury (BCNI) rat model was established by crushing the bilateral cavernous nerves. After crushing, ASCs and CBMSCs were intracavernously injected immediately. Erectile function, Masson staining, and immunofluorescence analyses of penile tissues were assessed at 4 and 12 weeks. PKH-26-labeled ASCs or CBMSCs were intracavernously injected to determine the presence and differentiation of ASCs or CBMSCs in the penis 3 days after injection. In vitro experiments including intracellular ROS detection, mitochondrial membrane potential assay, EdU cell proliferation staining, cell apoptosis assay, and protein chip assay were conducted to explore the underlying mechanism of CBMSC treatment compared with ASC treatment. Results: CBMSC injection significantly restored erectile function, rescued the loss of cavernous corporal smooth muscles, and increased the ratio of smooth muscle to collagen. PKH-26-labeled CBMSCs or ASCs did not colocalize with endothelial cells or smooth muscle cells in the corpus cavernosum. Moreover, the conditioned medium (CM) of CBMSCs could significantly inhibit the oxidative stress and elevate the mitochondria membrane potential and proliferation of Schwann cells. Better therapeutic effects were observed in the CBMSC group than the ASC group both in vivo and in vitro. In addition, the content of neurotrophic factors and matrix metalloproteinases in CBMSC-CM, especially NT4, VEGF, MMP1, and MMP3 was significantly higher than that of ASC-CM. Conclusion: Intracavernous injection of CBMSCs exhibited a better erectile function restoration than that of ASCs in CNI-ED rats owing to richer secretory factors, which can promote nerve regeneration and reduce extracellular matrix deposition. CBMSC transplantation would be a promising therapeutic strategy for CNI-ED regeneration in the future.

A prospective randomized controlled study on scheduled PDE5i and vacuum erectile devices in the treatment of erectile dysfunction after nerve sparing prostatectomy.

Cavernous nerve injury is an important cause of erectile dysfunction (ED). Although protective nerve technology has been widely used in nerve-sparing radical prostatectomy (nsRP), the incidence of ED is still very high after surgery. The purpose of our study was to evaluate erectile function (EF) and penile length in the non-erectile state (PLNES) following scheduled phosphodiesterase 5 inhibitor (PDE5i), vacuum erectile device (VED) treatment, and combination therapy after nsRP. One hundred patients with localized prostate cancer and normal EF were randomized to scheduled PDE5i group, VED treatment group, a combined treatment group, and the control group without any intervention. The International Index of Erectile Function-5 (IIEF-5) scores and PLNES were evaluated after 6 months and 12 months of treatment. Sexual Encounter Profile (SEP-Question 2 and SEP-Question 3) were evaluated after 12 months of treatment. Ninety-one of the 100 randomized patients completed the study. We found that the 5 mg tadalafil once a day (OaD) combined with VED can help improve IIEF-5 scores in nsRP patients after both 6 months and 12 months. VED alone or combined with tadalafil OaD can help patients maintain PLNES. VED combined with tadalafil OaD can improve the rate of successful penetration (SEP-Question 2) after 12 months. There were no significant differences in the return to target EF after 12 months among the groups. No significant correlation was noted between the variables and return to target EF (IIEF ≥ 17), and between the variables and effective shortening of the patient's penis (shortening ≥ 1 cm) after 12 months of intervention.

Urethra-inspired biomimetic scaffold: A therapeutic strategy to promote angiogenesis for urethral regeneration in a rabbit model.

Limited angiogenesis and epithelialization make urethral regeneration using conventional tissue-engineered grafts a great challenge. Consequently, inspired from the native urethra, bacterial cellulose (BC) and bladder acellular matrix (BAM) were combined to design a three dimensional (3D) biomimetic scaffold. The developed BC/BAM scaffold was engineered for accelerating urethral regeneration by enhancing angiogenesis and epithelialization. The BC/BAM scaffold reveals the closest mimic of native urethra in terms of the 3D porous nanofibrous structure and component including collagen, glycosaminoglycans, and intrinsic vascular endothelial growth factor (VEGF). In vitro studies showed that the bioinspired BC/BAM scaffold promoted in vitro angiogenesis by facilitating human umbilical vein endothelial cells (HUVECs) growth, expression of endothelial function related proteins and capillary-like tube formation. Effect of the BC/BAM scaffold on angiogenesis and epithelialization was studied by its implantation in a rabbit urethral defect model for 1 and 3 months. Results demonstrated that the improved blood vessels formation in the urethra-inspired BC/BAM scaffold significantly promoted epithelialization and accelerated urethral regeneration. The urethra-inspired BC/BAM scaffold provides us a new design approach to construct grafts for urethral regeneration. STATEMENT OF SIGNIFICANCE: Findings in urethral regeneration demonstrate that an ideal tissue-engineered urethra should have adequate angiogenesis to support epithelialization for urethral regeneration in vivo. In this study, inspired from the native urethra, a bioinspired bacterial cellulose/bladder acellular matrix (BC/BAM) scaffold was developed to promote angiogenesis and epithelialization. The designed scaffold showed the closest physical structure and component to natural urethra, which is beneficial to angiogenesis and regeneration of urethral epithelium. This is the first time to utilize BC and dissolved BAM to develop biomimetic scaffold in urethral tissue engineering. Our biomimetic strategy on urethra graft design provided enhanced angiogenesis and epithelialization to achieve an accelerated and successful rabbit urethral repair. We believe that our urethra-inspired biomimetic scaffold would provide new insights into the design of urethral tissue engineering grafts.

Erratum: A smart bilayered scaffold supporting keratinocytes and muscle cells in micro/nano-scale for urethral reconstruction.

[This corrects the article DOI: 10.7150/thno.22080.].

A smart bilayered scaffold supporting keratinocytes and muscle cells in micro/nano-scale for urethral reconstruction.

Rationale: In urethral tissue engineering, the currently available reconstructive procedures are insufficient due to a lack of appropriate scaffolds that would support the needs of various cell types. To address this problem, we developed a bilayer scaffold comprising a microporous network of silk fibroin (SF) and a nanoporous bacterial cellulose (BC) scaffold and evaluated its feasibility and potential for long-segment urethral regeneration in a dog model. Methods: The freeze-drying and self-assembling method was used to fabricate the bilayer scaffold by stationary cultivation G. xylinus using SF scaffold as a template. The surface morphology, porosity and mechanical properties of all prepared SF-BC scaffolds were characterized using Scanning electron microscopy (SEM), microcomputed tomography and universal testing machine. To further investigate the suitability of the bilayer scaffolds for tissue engineering applications, biocompatibility was assessed using an MTT assay. The cell distribution, viability and morphology were evaluated by seeding epithelial cells and muscle cells on the scaffolds, using the 3D laser scanning confocal microscopy, and SEM. The effects of urethral reconstruction with SF-BC bilayer scaffold was evaluated in dog urethral defect models. Results: Scanning electron microscopy revealed that SF-BC scaffold had a clear bilayer structure. The SF-BC bilayer scaffold is highly porous with a porosity of 85%. The average pore diameter of the porous layer in the bilayer SF-BC composites was 210.2±117.8 µm. Cultures established with lingual keratinocytes and lingual muscle cells confirmed the suitability of the SF-BC structures to support cell adhesion and proliferation. In addition, SEM demonstrated the ability of cells to attach to scaffold surfaces and the biocompatibility of the matrices with cells. At 3 months after implantation, urethra reconstructed with the SF-BC scaffold seeded with keratinocytes and muscle cells displayed superior structure compared to those with only SF-BC scaffold. Principal Conclusion: These results demonstrate that the bilayer SF-BC scaffold may be a promising biomaterial with good biocompatibility for urethral regeneration and could be used for numerous other types of hollow-organ tissue engineering grafts, including vascular, bladder, ureteral, bowel, and intestinal.

Posterior Urethroplasty Complexity and Prognosis Can be Described by a Novel Method: Posterior Urethral Stenosis Score.

OBJECTIVE: To develop a standardized PU-score (posterior urethral stenosis score), with the goal of using this scoring system as a preliminary predictor of surgical complexity and prognosis of posterior urethral stenosis. PATIENTS AND METHODS: We retrospectively reviewed records of all patients who underwent posterior urethral surgery at our institution from 2013 to 2015. The PU-score is based on 5 components, namely etiology (1 or 2 points), location (1-3 points), length (1-3 points), urethral fistula (1 or 2 points), and posterior urethral false passage (1 point). We calculated the score of all patients and analyzed its association with surgical complexity, stenosis recurrence, intraoperative blood loss, erectile dysfunction, and urinary incontinence. RESULTS: There were 144 patients who underwent low complexity urethral surgery (direct vision internal urethrotomy, anastomosis with or without crural separation) with a mean score of 5.1 points, whereas 143 underwent high complexity urethroplasty (anastomosis with inferior pubectomy or urethrorectal fistula repair, perineal or scrotum skin flap urethroplasty, bladder flap urethroplasty) with a mean score of 6.9 points. The increase of PU-score was predictive of higher surgical complexity (P = .000), higher recurrence (P = .002), more intraoperative blood loss (P = .000), and decrease of preoperative (P = .037) or postoperative erectile function (P = .047). However, no association was observed between PU-score and urinary incontinence (P = .213). CONCLUSION: The PU-score is a novel and meaningful scoring system that describes the essential factors in determining the complexity and prognosis for posterior urethral stenosis.

Three-Dimensional Computerized Model Based on the Sonourethrogram: A Novel Technique to Evaluate Anterior Urethral Stricture.

PURPOSE: The sonourethrogram is a useful alternative to the traditional retrograde urethrogram to evaluate anterior urethral strictures. With the development of 3-dimensional reconstructive techniques 3-dimensional urethral imaging can provide more accurate and useful information to enable the surgeon to make the best surgical decisions. We evaluated the accuracy and efficacy of a 3-dimensional reconstructed digital model of the urethra based on the sonourethrogram to assess anterior urethral disease. MATERIALS AND METHODS: A total of 50 patients with an anterior urethral stricture and 10 healthy volunteers were enrolled in this study from April 2014 to January 2017. All patients and volunteers underwent sonourethrogram and retrograde urethrogram. Three-dimensional urethral models were reconstructed based on the sonourethrogram. Stricture length and location on retrograde urethrogram or sonourethrogram based images were compared with those found at operation. RESULTS: The 3-dimensional digital model revealed the entire anterior urethra, including the navicular fossa, and the penile and bulbar parts. The semitransparent model clearly demonstrated the structure of the corpus spongiosum and inside the urethral lumen. Further information on spongiofibrosis could also be seen in the 3-dimensional digital model. There was no significant difference in stricture length or location in the 3-dimensional model compared with retrograde urethrogram imaging and actual surgical findings. However, the latest technique could only reconstruct the short segment of the anterior urethra due to the probe width limitation. CONCLUSIONS: The 3-dimensional computerized model based on the sonourethrogram is a novel and effective technique of evaluating anterior urethral strictures.

Use of heparinized bacterial cellulose based scaffold for improving angiogenesis in tissue regeneration.

Vascularization is a prerequisite to achieve tissue regeneration especially for long-term survival of a scaffold. During the regeneration process, the delivery of angiogenic factors is very important for developing a vascular network. In this paper, vascular endothelial growth factor (VEGF)-loaded 3D porous bacterial cellulose/gelatin (B/G) scaffolds modified with heparin were firstly prepared. The pro-angiogenic effects of scaffolds towards proliferation and migration of endothelial cells (PIECs) were evaluated as well as in vivo implantation. Results showed that the B/G scaffold modified with heparin could provide a prolonged release of VEGF for two weeks. In vitro cellular assays showed that proliferation and migration were promoted in the presence of VEGF. Subcutaneous implantation demonstrated that angiogenesis was significantly improved for the heparinized scaffolds loaded with VEGF (V-B/G/H), compared to B/G scaffold. The resulting scaffold with sustained delivery of VEGF could be potential and effective tissue engineered candidates in tissue regeneration for future clinical applications.

Adipose-derived stem cells-seeded bladder acellular matrix graft-silk fibroin enhances bladder reconstruction in a rat model.

The unfavourable clinical outcomes of host cell-seeded scaffolds for bladder augmentation warrant improved bioactive biomaterials. This study aimed to examine the feasibility of adipose-derived stem cells (ASCs)-seeded bilayer bladder acellular matrix graft (BAMG)-silk fibroin (SF) scaffold in enhancing bladder reconstruction. Sprague Dawley rats were randomly divided into three groups: the BAMG-SF-ASCs group, the acellular BAMG-SF group and the cystotomy group. The BAMG-SF-ASCs group was sampled at 2, 4 and 12 weeks, and compared with the other groups at 12 weeks. In the BAMG-SF-ASCs group, the normal bladder contour was reformed similar to that in the cystotomy group, with abundant urothelium and smooth muscle regeneration, as well as a suitable scaffold degradation speed, and trivial fibrosis and inflammation. The ASCs seeded in BAMG-SF were maintained in the regenerated region during the 12-week experimental period and significantly enhanced the vessel density, nerve regeneration and bladder function compared with acellular BAMG-SF. In addition, the BAMG-SF-ASCs group presented elevated levels of SDF-1α, VEGF and their receptors, with an obvious increase in ERK 1/2 phosphorylation. BAMG-SF is a promising biomaterial for ASCs seeding to facilitate bladder augmentation and demonstrated an enhanced angiogenic potential possibly related to the SDF-1α/CXCR4 pathway via ERK 1/2 activation.

Trilayer Three-Dimensional Hydrogel Composite Scaffold Containing Encapsulated Adipose-Derived Stem Cells Promotes Bladder Reconstruction via SDF-1α/CXCR4 Pathway.

Bladder acellular matrix graft-alginate dialdehyde-gelatin hydrogel-silk mesh (BAMG-HS) encapsulated with adipose-derived stem cells (ASCs) was evaluated in a rat model of augmentation cystoplasty, including BAMG-HS-ASCs (n = 18, subgroup n = 6 for 2, 4, and 12 weeks), acellular BAMG-HS (n = 6 for 12 weeks) and cystotomy control (n = 6 for 12 weeks) groups. Equipped with good cytocompatibility and superior mechanical properties (elastic modulus: 5.33 ± 0.96 MPa, maximum load: 28.90 ± 0.69 N), BAMG-HS acted a trilayer "sandwich" scaffold with minimal interference in systemic homeostasis. ASCs in BAMG-HS promoted morphological and histological bladder restoration by accelerating scaffold degradation (p < 0.05), ameliorating fibrosis (p < 0.05) and inflammation (p < 0.01). Additionally, ASCs facilitated the recovery of bladder function by enhancing smooth muscle regeneration (p < 0.05), innervation (p < 0.01) and angiogenesis (p < 0.001). Except for a small number of endothelium-differentiated ASCs, the pro-angiogenic effects of ASCs were mainly related to ERK1/2 phosphorylation in the downstream of SDF-1α/CXCR4 pathway.

Preliminary Experience of Nontransecting Urethroplasty for Pelvic Fracture-related Urethral Injury.

OBJECTIVE: To evaluate the urinary outcomes and preservation of erectile function in patients with pelvic fracture-related urethral injury (PFUI) after nontransecting spongiosum anastomotic urethroplasty (NTSAU). MATERIALS AND METHODS: Fifty-nine male patients with PFUI following traumatic pelvic fracture underwent NTSAU. Inclusion criteria were age 18-60 years, posterior urethral stenosis <2.5 cm without previous urethroplasty, and intact erectile function. Exclusion criteria were history of open urethroplasty, long-segment posterior urethral stenosis (>2.5 cm), preoperative impotency, or age over 60 years. Pre- and postoperative outcome analyses were performed with a paired t test and chi-square test. RESULTS: Between January 2011 and August 2015, 59 patients with a mean age of 38.5 years (range, 21-59 years) and a mean stricture length of 2.0 cm (range, 1-2.5 cm) underwent simple NTSAU (group 1, n = 41) or NTSAU with inferior pubectomy (group 2, n = 18). Patients were followed for a mean 25 months (range, 12-60 months). The primary success rate was 96.6% (57 of 59), and stricture recurrence occurred in 2 (3.4%) patients. The secondary outcomes revealed no significant changes in number of events, tip rigidity, or duration of best episode between pre- and postoperative nocturnal penile tumescence test (on RigiScan) in group 1, but a slight decrease in group 2 (P <.05). The limitation was the small sample size and heterogeneous population. CONCLUSION: NTSAU is a safe, feasible, minimally invasive procedure for PFUI, optimizing erectile preservation.

The application of CT voiding urethrography in the evaluation of urethral stricture associated with fistula: a preliminary report.

PURPOSE: This study was designed to evaluate the clinical utility of CT voiding urethrography (2D/3D reformatted CT images and virtual cystourethroscopy) in terms of its ability to detect urethral stricture associated with fistula. METHODS: In the study period, 80 patients were found to have signs or symptoms of urine leakage, as detected by voiding CT scanning. 2D/3D reformatted CT images and the virtual cystourethroscopic view were generated by the software. Conventional urethrography and traditional cystourethroscopy were also used prior to operation. The accuracy of these techniques was compared to the actual findings during the operation. RESULTS: For 58 male patients suffering from urethral strictures associated with fistulas, the detection rate of the fistulas by 2D/3D reformatted CT images was superior to conventional urethrography for characterizing the site of the fistula and the length of the urethral stricture (P < 0.05). The detection rate of the fistula by 3D virtual cystourethroscopy was similar to that by traditional cystourethroscopy (P > 0.05). The duration of the examination was shorter for 3D virtual cystourethroscopy than for traditional cystourethroscopy (P < 0.05). The same results were observed in 22 cases that featured a urethrovaginal fistula. CONCLUSION: CT voiding urethrography is a useful technique for the diagnosis of urethral disease, especially in male patients suffering from urethral stricture associated with fistula.

The Selection of Procedures in One-stage Urethroplasty for Treatment of Coexisting Urethral Strictures in Anterior and Posterior Urethra.

OBJECTIVE: To explore selection of the procedures in one-stage urethroplasty for treatment of coexisting urethral strictures in the anterior and posterior urethra. METHODS: Between 2008 and 2014, a total of 27 patients with existing strictures simultaneously at anterior urethra and posterior urethra were treated in our hospital. Two types of procedures were selected for treatment of the anterior urethral strictures. A penile skin flap and the lingual mucosa were used for augmented urethroplasty in 20 and 7 cases, respectively. Three types of procedures, namely, non-transecting end-to-end urethral anastomosis (n = 3), traditional end-to-end urethral anastomosis (n = 17), other grafts substitution urethroplasty, including pedicle scrotal skin urethroplasty (n = 2), and lingual mucosal graft urethroplasty (n = 5), were utilized in the treatment of posterior urethral strictures. RESULTS: The patients were mean followed up 30 months with an overall success rate of 88.9%. The majority of the patients exhibited wide patent urethras on retrograde urethrography and the patients' urinary peak flow ranged from 14.2 to 37.9 ml/s. Complications developed in 3 patients (11.1%). Of the 17 patients who underwent traditional urethral end-to-end anastomosis, urethral strictures occurred in 2 patients at 4 and 6 months after the operation. These patients achieved a satisfactory voiding function after salvage pedicle scrotal skin urethroplasty. A urethral pseudodiverticulum was observed in another patient 9 months after pedicle penile flap urethroplasty; and after a salvage procedure, he regained excellent voiding function. CONCLUSION: Synchronous anterior and posterior strictures can be successfully reconstructed with a combination of substitution and anastomotic urethroplasty techniques.

Structural and functional evaluation of oxygenating keratin/silk fibroin scaffold and initial assessment of their potential for urethral tissue engineering.

In this study, we report a new type of oxygen-generating scaffold, composed of human keratin, silk, gelatin and calcium peroxide (CPO). After mixing the silk/keratin (60:40) with 2% gelatin and 20% CPO, the film demonstrated excellent mechanical properties, non-cytotoxicity and oxygen-generative ability. The detailed structure of scaffold was revealed by confocal laser and electronic scanning microscopy. The gelatin formed the network structure, which mixed with silk fibroin and keratin. The CPOs were embedded into scaffold. A shell-core structure was formed in the CPO particles, in which the CPO was located in the core and the gelatin was mainly wrapped around the CPO. Furthermore, the oxygen-release test showed that scaffold was able to steadily release high level of oxygen over two weeks in vitro. In addition, the anti-bacterial function was also proved in the scaffold. Films with CPO enhanced the repair in dog urethral defect models, resulting in patent urethra. Improved organized muscle bundles and epithelial layer were observed in animals treated with CPO films compared with those treated with non-CPO films. This study suggests that this biomaterial could be suitable for tissue engineered urinary tract reconstruction.

Urethral Reconstruction with Small Intestinal Submucosa Seeded with Oral Keratinocytes and TIMP-1 siRNA Transfected Fibroblasts in a Rabbit Model.

BACKGROUND: To evaluate the effect of tissue inhibitor of metalloproteinase-1 small interfering RNA (TIMP-1 siRNA) transfected fibroblasts (FB) for urethral reconstruction. MATERIALS AND METHODS: A ventral urethral mucosal defect was created. Substitution urethroplasty was performed with small intestinal submucosa (SIS) alone (8 rabbits, group 1), autogenic oral keratinocytes (OK)-seeded SIS (8 rabbits, group 2) or autogenic OK and TIMP-1 siRNA transfected FB-seeded SIS (8 rabbits, group 3). At 1 and 6 months after surgery (4 rabbits at each time point), retrograde urethrogram and histologic analysis were performed to evaluate the outcomes of urethroplasty. RESULTS: TIMP-1 siRNA transfected FB decreased the secretion of type I collagen. Under retrograde urethrography, 5 rabbits in group 1, 6 in group 2 and 7 in group 3 maintained a wide urethral caliber. Histologically, inflammation and fibrosis were observed at 6 months in group 1. The speed of urothelium, smooth muscle and vessel regeneration in group 3 was faster than that in group 2. Comparison of smooth muscle-to-collagen ratio, epithelial layers, smooth muscle content and microvessel density among three groups revealed a significant increase (p < 0.05). CONCLUSIONS: TIMP-1 siRNA transfected FB could be used as a source of seed cell for urethral tissue engineering and could prevent the proliferation of urethral scar tissue.

Comparative study of different seeding methods based on a multilayer SIS scaffold: Which is the optimal procedure for urethral tissue engineering?

Seeding cells efficiently and uniformly onto three-dimensional scaffolds is key for engineering urological tissue with an ideal histological structure in vitro. Using an optimized seeding technology allows cells to cooperate positively with biomaterials, resulting in successful reconstructive surgery. In this study, we used four different types of seeding methods in a scaffold of small intestinal submucosa (SIS). The efficiency of the sandwich co-culture, layered co-culture, static-agitation seeding, and centrifugation seeding methods were compared. It was demonstrated that dynamic seeding methods, such as static-agitation and centrifugation seeding, had superior cell-matrix infiltration and mechanical properties. The seeding time could be reduced by 5-10 min using the centrifugation method. Furthermore, functional assessment of the barriers revealed that this function was better in the centrifugation seeding method than in any other method. Our study suggests that both the static-agitation and centrifugation methods are suitable for cell seeding on SIS. There is no significant change in surface area of SIS with different seeding methods. These methods reinforce the physiological and mechanical properties of biomaterials and allow for the future in vivo study of tissue-engineered urethral reconstruction. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1098-1108, 2016.

Bacterial Cellulose-Based Biomimetic Nanofibrous Scaffold with Muscle Cells for Hollow Organ Tissue Engineering.

In this study, we built a bilayer nanofibrous material by utilizing the gelatinization properties of potato starch (PS) to interrupt bacterial cellulose (BC) assembly during static culture to create more free spaces within the fibrous network. Then, muscle cells were cultured on the loose surface of the BC/PS scaffolds to build biomaterials for hollow organ reconstruction. Our results showed that the BC/PS scaffolds exhibited similar mechanical characters to those in the traditional BC scaffolds. And the pore sizes and porosities of BC/PS scaffolds could be controlled by adjusting the starch content. The average nanofiber diameters of unmodified BC and BC/PS composites is approximately to that of the urethral acellular matrix. Those scaffolds permit the muscle cells infiltration into the loose layer and the BC/PS membranes with muscle cells could enhance wound healing in vivo and vitro. Our study suggested that the use of bilayer BC/PS nanofibrous scaffolds may lead to improved vessel formation. BC/PS nanofibrous scaffolds with muscle cells enhanced the repair in dog urethral defect models, resulting in patent urethra. Improved organized muscle bundles and epithelial layer were observed in animals treated with BC/PS scaffold seeded by muscle cells compared with those treated with pure BC/PS scaffold. This study suggests that this biomaterial could be suitable for tissue engineered urinary tract reconstruction and this type of composite scaffold could be used for numerous other types of hollow organ tissue engineering grafts, including vascular, bladder, ureter, esophagus, and intestine.