1.4 SILK FIBROIN IN TISSUE ENGINEERING
1.4.12 Bladder tissue engineering
The urinary bladder is a hollow muscular and distensible (or elastic) organ which sits on the pelvic floor and collects urine excreted by the kidneys before disposal by urination. A variety of conditions encountered in urology result in bladder dysfunction, and many of these require reconstructive procedures. Currently, gastrointestinal segments are considered the gold standard for bladder reconstructive procedures (Atala, 2011).
However, significant complications including chronic urinary tract infection, metabolic abnormalities, urinary stone formation, bowel dysfunction and secondary malignancies are associated with this approach. Replacement of bladder tissues with in vitro engineered functionally equivalent tissues composed of biomaterial-based, bladder-shaped scaffolds seeded with autologous urothelial and smooth muscle cells, could improve the outcome of reconstructive surgery (Atala, 2011). Biomaterials derived from SF have been previously utilized in vascular and orthopedic applications and thus possesses robust physical and biological properties which would be well suited for urologic applications.
The in vitro cytocompatibility of SF film with the transitional epithelial cells from the urinary bladders of New Zealand rabbits was evaluated by Liu et al (2008a). The SF film was compatible with the transitional cells and there was no significant difference between the control and experimental groups as evident from MTT assay. In a similar study, Liu et al (2007) revealed that all the 12 rabbits in the experimental group, which had a urethral defect of 1.5 cm, did not show signs of urethral stricture following the surgery. Also, the implanted SF film was degraded completely at 16 weeks and the defect was repaired by inducing the growth of the smooth muscle cells and urethral epithelial cells. Later, Liu et al (2008b) studied the effect of SF film for repairing urethral defect in 14 adult male dogs. It was found that the implanted SF film was degraded completely at week 12 and the defect was repaired by mucous membrane of urethra and smooth muscle cells regularly in the experimental group I (1.5-cm defect, n=6). While, pale, dense and rigid appearance of the urethra with a narrow cavity was found in the experimental group II (3.0-cm defect, n=6). Thus the SF film can repair a short-length defect within 1.5 cm, while it is unpredictable to repair defect longer than 3.0 cm only by the materials.
Zhang et al (2010) demonstrated the repairing of urethral defect with adipose- derived MSCs seeded on a porous SF scaffold in 39 New Zealand rabbits. The incidence rate of post-operative urethral stricture and fistula was found to be around 76.92% in
group C (control), which was significantly higher than group A (23.07%, SF implant) and group B (15.38%, SF implant seeded with MSCs). Also, histological examination showed that blood vessel formation and urethral epithelium, smooth muscle regeneration were much better in groups of A and B than those in group C, which were better in group B than those in group A. Thus, it was suggested that compared to SF alone, the MSCs seeded SF significantly enhances urethral defect repairing in rabbits. The fabrication of SF based woven patches using gel spinning technique and its use in bladder augmentation was described by Cannon et al (2010) in mice models and by Mauney et al (2011) in murine models. Both these studies revealed that, in contrast to PGA- and SIS- patches, by 10 weeks, the silk matrix significantly increases the bladder capacity and voided volume while maintaining similar degrees of compliance relative to the control group, as evident from histological, voided stain on paper and renal / bladder ultrasound analyses (Figure 1.13).
Stress urinary incontinence remains a worldwide problem affecting patients of all ages. Implantation of sub-urethral sling is the cornerstone treatment, but, the current slings have inherent disadvantages. Zou et al (2010) developed a tissue engineered sling with bone marrow derived mesenchymal stem cells (bmMSCs) (derived from Sprague–
Dawley rats) seeded silk scaffold. Forty stress urinary incontinence female rat models were divided into four groups. Group A (n=5) had sham operation, Group B (n=5) had no sling placed, Group C (n=15) was treated with a silk-sling, and Group D (n=15) was treated with tissue engineered sling. It was found that the Group B had a significantly lower leak-point pressure (24.0 ± 4.2 cmH2O) at 4 weeks, while Group C (38.0 ± 3.3 cmH2O) and Group D (36.3 ± 3.1 cmH2O) almost reached to the normal level shown by Group A (41.6 ± 3.8 cmH2O) (Zou et al, 2010). Thus, the MSCs seeded sling showed convincing functional effects for the treatment of stress urinary incontinence in a rat model and better ligament-like tissue formation suggested potential long-term function.
Figure 1.13. Murine bladder augmentation model: (a) abdominal incision and extrusion of bladder, hashed line denotes defect site; (b) gel spun silk matrix tailored to area of the defect site; Hematoxylin and eosin analysis of regeneration bladder defect augmented with silk scaffold scaffold after 21 d (c) and 70 d (d) post-implantation period. (*) denotes scaffold fragments, brackets denote area of tissue regeneration. (Reproduced with permission, Mauney et al 2011).
Figure 1.14. Images of (a) flat and (b) aligned grooved patterned silk fibroin films of 2 μm in thickness. Fluorescent images of 10 days old GFP expressing rat corneal fibroblast cells growing on (a’) flat and (b’) patterned silk fibroin film surfaces (red indicates stained actin filaments and green represents GFP fluorescence). (Reproduced with permission, Lawrence et al, 2009).