Minkai Xie

Evaluation of stretched electrospun silk fibroin matrices seeded with urothelial cells for urethra reconstruction

BACKGROUND We investigated the feasibility of urethral reconstruction using stretched electrospun silk fibroin matrices. MATERIALS AND METHODS A novel electrospun silk fibroin matrix was prepared. The structure of the material was assessed by scanning electron microscopy and a porosity test. Canine urothelial cells were isolated, expanded, and seeded onto the material for 1 wk to obtain a tissue-engineered graft. The tissue-engineered graft was assessed using hematoxylin and eosin staining and scanning electron microscopy. A dorsal urethral mucosal defect was created in nine female beagle dogs. In the experimental group, tissue-engineered mucosa was used to repair urethra mucosa defects in six dogs. No substitute was used in the three dogs of the control group. Retrograde urethrography was performed at 1, 2, and 6 mo after grafting. The urethral grafts were analyzed grossly and histologically. RESULTS Scanning electron microscope and a porosity test revealed that the material had a three-dimensional porous structure. Urothelial cells grew on the material and showed good biocompatibility with the stretched silk fibroin matrices. Canines implanted with tissue-engineered mucosa voided without difficulty. Retrograde urethrography revealed no signs of stricture. Histologic staining showed gradual epithelial cell development and stratified epithelial layers at 1, 2, and 6 mo. The canines in the control group showed difficulty in voiding. Retrograde urethrography showed urethra stricture. Histologic staining showed that no or only one layer of epithelial cells developed. A severe inflammatory reaction was also observed in the control group. CONCLUSIONS Stretched electrospun silk fibroin matrices have good biocompatibility with urothelial cells, which could prove to be a potential material for use in urethra reconstruction.

Skeletal myogenic differentiation of human urine-derived cells as a potential source for skeletal muscle regeneration.

Stem cells are regarded as possible cell therapy candidates for skeletal muscle regeneration. However, invasive harvesting of those cells can cause potential harvest‐site morbidity. The goal of this study was to assess whether human urine‐derived stem cells (USCs), obtained through non‐invasive procedures, can differentiate into skeletal muscle linage cells (Sk‐MCs) and potentially be used for skeletal muscle regeneration. In this study, USCs were harvested from six healthy individuals aged 25–55. Expression profiles of cell‐surface markers were assessed by flow cytometry. To optimize the myogenic differentiation medium, we selected two from four different types of myogenic differentiation media to induce the USCs. Differentiated USCs were identified with myogenic markers by gene and protein expression. USCs were implanted into the tibialis anterior muscles of nude mice for 1 month. The results showed that USCs displayed surface markers with positive staining for CD24, CD29, CD44, CD73, CD90, CD105, CD117, CD133, CD146, SSEA‐4 and STRO‐1, and negative staining for CD14, CD31, CD34 and CD45. After myogenic differentiation, a change in morphology was observed from ‘rice‐grain’‐like cells to spindle‐shaped cells. The USCs expressed specific Sk‐MC transcripts and protein markers (myf5, myoD, myosin, and desmin) after being induced with different myogenic culture media. Implanted cells expressed Sk‐MC markers stably in vivo. Our findings suggest that USCs are able to differentiate into the Sk‐MC lineage in vitro and after being implanted in vivo. Thus, they might be a potential source for cell injection therapy in the use of skeletal muscle regeneration. Copyright

Changing trends in the causes and management of male urethral stricture disease in China: an observational descriptive study from 13 centres.

To determine whether there have been any changes in the causes and management of urethral strictures in China.

Tunable structures and properties of electrospun regenerated silk fibroin mats annealed in water vapor at different times and temperatures

Regenerated silk fibroin (SF) mats were fabricated using electrospinning technique, followed by mild water vapor annealing to effectively tune the structures and improve the mechanical properties of the mats at different annealing times and temperatures. The breaking strength and the breaking energy of the mats treated with water vapor at 65°C for 12 h reached 6.0MPa and 171.7 J/kg, respectively. The conformational transition of the SF mats was significantly influenced by the treating temperature, while the influence of time was comparatively limited. The influence is consistent with the time-temperature equivalent principle and would be helpful for the preparation of water-vapor-annealed silk-based biomaterials for various applications.

Urethral reconstruction with a 3D porous bacterial cellulose scaffold seeded with lingual keratinocytes in a rabbit model.

The goal of this study was to evaluate the effects of urethral reconstruction with a three-dimensional (3D) porous bacterial cellulose (BC) scaffold seeded with lingual keratinocytes in a rabbit model. A novel 3D porous BC scaffold was prepared by gelatin sponge interfering in the BC fermentation process. Rabbit lingual keratinocytes were isolated, expanded, and seeded onto 3D porous BC. BC alone (group 1, N  =  10), 3D porous BC alone (group 2, N  =  10), and 3D porous BC seeded with lingual keratinocytes (group 3, N  =  10) were used to repair rabbit ventral urethral defects (2.0   ×   0.8 cm). Scanning electron microscopy revealed that BC consisted of a compact laminate while 3D porous BC was composed of a porous sheet buttressed by a dense outer layer. The average pore diameter and porosity of the 3D porous BC were 4.23   ±   1.14 μm and 67.00   ±   6.80%, respectively. At 3 months postoperatively, macroscopic examinations and retrograde urethrograms of urethras revealed that all urethras maintained wide calibers in group 3. Strictures were found in all rabbits in groups 1 and 2. Histologically, at 1 month postoperatively, intact epithelium occurred in group 3, and discontinued epithelium was found in groups 1 and 2. However, groups 2 and 3 exhibited similar epithelial regeneration, which was superior to that of group 1 at 3 months (p  <  0.05). Comparisons of smooth muscle content and endothelia density among the three groups revealed a significant increase at each time point (p  <  0.05). Our results demonstrated that 3D porous BC seeded with lingual keratinocytes enhanced urethral tissue regeneration. 3D porous BC could potentially be used as an optimized scaffold for urethral reconstruction.

Reconstruction of Penile Urethra With the 3-Dimensional Porous Bladder Acellular Matrix in a Rabbit Model

OBJECTIVE To evaluate the effect of reconstruction of penile urethra with the 3-dimensional (3-D) porous bladder acellular matrix (BAM) in a rabbit model. MATERIALS AND METHODS In 30 male rabbits, a ventral urethral mucosal defect (1.5 × 0.8 cm) was created. Substitution urethroplasty was performed with 5% peracetic acid (PAA)-treated BAM (3-D porous BAM; 15 rabbits, PAA-treated BAM group) and non-PAA-treated BAM (15 rabbits; non-PAA-treated BAM group) in an onlay fashion. At 1, 2, and 3 months after surgery (5 rabbits at each time point) in the 2 groups, retrograde urethrogram and histologic analysis were performed to evaluate the outcomes of urethroplasty. RESULTS In the PAA-treated BAM group, 13 rabbits maintained a wide urethral caliber without a fistula or stricture. In contrast, 10 rabbits kept a wide caliber in the non-PAA-treated BAM group. Histologically, at 1, 2, and 3 months after the surgery, the speed of urothelium regeneration in the PAA-treated BAM group was faster than that in the non-PAA-treated BAM group. The smooth muscle-to-collagen ratio and the content of smooth muscle in the PAA-treated BAM group were significantly higher than that in the non-PAA-treated BAM group at each time point (P <.05). The endothelium density between the non-PAA-treated BAM and the PAA-treated BAM groups revealed a significant increase at all 3 time points (P <.05). CONCLUSION Our results confirmed that PAA-treated BAM urethroplasty enhanced urothelium, smooth muscle regeneration and neovascularization compared with those of the non-PAA-treated BAM. The 3-D porous BAM as an optimized biological scaffold may be used for cell-based tubular and long-segmental urethral reconstruction in the future.

Imaging techniques for the diagnosis of male traumatic urethral strictures

Urethral injures, pelvic trauma, and certain congential abnormalities in male can often cause urethral strictures and scarring. Adequate and accurate imaging diagnosis of urethral strictures is critial to select a surgical approach. Although urethrography is commonly used in evaluating male urethral injury and strictures, its limitation is that the contrast offers images only at the urethral lumen but not of the periurethral structures. Sonourethrography (SUG) has the ability to define the periurethral tissues and without the radiation exposure, the degree of periurethral fibrosis can be shown with a view to guiding surgery especially in bulbar urethra. Computed tomography (CT) urethrography has the advantage of examining patients only in one position, and by generating three-dimensional images; it can accurately measure the stricture length. Magnetic resonance imaging (MRI) provide useful information in certain clinical situations, particularly posterior urethral trauma and in the evaluation of the periurethral soft tissues. However, both CT and MRI are relative high cost. The selection of imaging methods should depend on the etiology, sites, patients general condition and the type of urethral injury, using a rational combination of different imaging methods can improve the accuracy of diagnosis for traumatic urethral strictures.

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.

Tough and VEGF-releasing scaffolds composed of artificial silk fibroin mats and a natural acellular matrix

Regenerated silk fibroin (RSF) scaffolds electrospun from aqueous solutions have great potential for tissue engineering. However, the traditional RSF mats are weak and limit the applications. A bladder acellular matrix graft (BAMG), a tough natural material, was used as an electrospinning substrate to toughen the RSF scaffolds. Compared with bare RSF scaffolds, the composite scaffolds with breaking energies ranging from 458 to 970 J kg−1 show significantly improved tensile properties and suture retention strength, which may satisfy the requirements for implantation. Vascular endothelial growth factor (VEGF) was encapsulated in the RSF/BAMG composite scaffolds by means of blend and coaxially electrospinning to promote the ability of vasculogenesis and angiogenesis. Transmission electron microscope (TEM) images show that the coaxially electrospun fibers had a core–sheath structure. An ELISA assay measurement indicates that VEGF can be released for more than 16 days. The samples annealed in water vapor exhibit higher release profiles than those immersed in ethanol. An in vitro assay indicates that VEGF loaded scaffolds evidently induced the attachment and proliferation of porcine iliac endothelial cells (PIECs) compared with those without VEGF. Moreover, the VEGF remained bioactive for up to 7 days. Thus the VEGF loaded composite scaffolds could be a promising candidate for tissue engineering applications.

Electrocautery-induced cavernous nerve injury in rats that mimics radical prostatectomy in humans

To investigate the early and delayed effects of cavernous nerve electrocautery injury (CNEI) in a rat model, with the expectation that this model could be used to test rehabilitation therapies for erectile dysfunction (ED) after radical prostatectomy (RP).