Urethral replacement: a comparison between small intestinal submucosa grafts and spontaneous regeneration

OBJECTIVE: To evaluate the use of commercially available single-layer small intestinal submucosa (SIS) for urethral replacement, both as an onlay and as a tube, in a rabbit model. MATERIALS AND METHODS: Thirty-six male rabbits were assigned to four experimental groups. Group 1 had the ventral wall of the penile urethra excised for 15 mm; in group 2 this created defect was patched with a SIS onlay graft; group 3 had complete excision of a 15 mm segment of the penile urethra; and in group 4, this created defect was replaced with a SIS tube graft. In all rabbits the urethra was stented for 2 weeks. A retrograde urethrogram was taken in all rabbits before death at 3, 6 and 12 weeks after surgery. The urethra was then exposed, examined carefully and excised for histopathological examination. RESULTS: In groups 1 and 2 the retrograde urethrograms were normal in 13 rabbits and there was relative narrowing in two rabbits in group 1 and three in group 2. In groups 3 and 4 all rabbits developed urethral fistulae or strictures. Histological examination of the urethra showed epithelial regeneration supported by smooth muscle backing in all rabbits in group 1, while rabbits in group 2 showed no regeneration of smooth muscle. By contrast, rabbits in groups 3 and 4 showed incomplete regeneration and progressive fibrosis. CONCLUSIONS: Single-layer SIS is not a suitable urethral substitute in this animal model. When used as an onlay, healing is inferior to spontaneous urethral regeneration, as SIS impedes smooth muscle cell regeneration. When used as a tube, there is complete scarring and urethral luminal occlusion.     

Evaluation of small intestinal submucosa as scaffolds for intestinal tissue engineering

BACKGROUND: Biodegradable scaffolds have been used for regenerating the small intestine. The aim of this study was to evaluate the feasibility of small intestinal submucosa (SIS) as scaffolds for intestinal regeneration in a rat model. MATERIALS AND METHODS: A tubular one-ply or four-ply SIS was interposed between isolated jejunal segments in rats. The scaffolds were harvested at 2, 4, and 8 weeks after implantation, and the specimens were examined grossly and histologically. RESULTS: Significant contractions were observed in SIS scaffolds after implantation. The one-ply SIS contracted to 44% of its initial length at 2 weeks and continued to contract to 6% of its initial length at 8 weeks. The contraction of four-ply SIS scaffolds was less than that of the one-ply SIS, reaching 29% of its initial length at 8 weeks. Minimal epithelial and smooth muscular regeneration was observed in the SIS scaffolds after implantation. CONCLUSIONS: A significant shrinkage was observed in the SIS scaffolds after implantation. Although the four-ply SIS contracted less than the one-ply SIS, neither scaffold supported significant amount of intestinal regeneration.     

Challenges in a larger bladder replacement with cell-seeded and unseeded small intestinal submucosa grafts in a subtotal cystectomy model

OBJECTIVE: To evaluate small intestinal submucosa (SIS), unseeded or seeded, as a possible augmentation material in a canine model of subtotal cystectomy. MATERIALS AND METHODS: In all, 22 male dogs had a 90% partial cystectomy and were then divided into three groups. At 1 month after the initial cystectomy, dogs in group 1 (unseeded, six) and group 2 (seeded, six) received a bladder augmentation with a corresponding SIS graft. The dogs in group 3 (ten) received no further surgery and were considered the surgical control group. All dogs were evaluated before and after surgery with blood chemistry, urine culture, intravenous urography, cystograms and cystometrograms. After surgery (at 1, 5 and 9 months), the bladders were examined using routine histology and immunohistochemistry. RESULTS: All 22 dogs survived the subtotal cystectomy, and 18 survived their intended survival period. One dog, in group 2 (seeded), was killed at 1 month after augmentation due to bladder perforation caused by a large piece of incompletely absorbed SIS. Three other dogs (group 1, two; and group 2, one) were killed within 2 months after augmentation due to bladder obstruction by stones. Group 1 and group 2 SIS grafts had moderate to heavy adhesion, graft shrinkage, and some had bone and calcification at the graft site. Histologically, there was limited bladder regeneration in both groups. Interestingly, dogs in group 3 at 1 month after cystectomy (when group 1 and 2 received their augmentations) had severely shrunken bladders and histologically had severe inflammation, fibroblast infiltration and muscle hypertrophy. These results verify the subtotal cystectomy model. CONCLUSIONS: The use of seeded or unseeded SIS in a subtotal cystectomy model does not induce the same quality and quantity of bladder regeneration that is seen in the 40% non-inflammatory cystectomy model. This study provides important insights into the process of regeneration in a severely damaged bladder. The results led us to re-evaluate the critical elements required for a complete bladder replacement using tissue-engineering techniques.     

Bladder regeneration in a canine model using hyaluronic acid-poly(lactic-co-glycolic-acid) nanoparticle modified porcine small intestinal submucosa

What's known on the subject? and What does the study add? Urinary bladder tissue can be produced by several regenerative techniques although consistent regeneration remains elusive and no single protocol is superior to the others. Small intestinal submucosa has been used as substrate for regeneration in several models and allows for full‐thickness bladder regeneration. Nanotechnology has been applied to impart specific physical and biological properties onto biological substrates. Nanoparticles composed of PLGA can be modified by the addition of hyaluronic acid and can be added to SIS. This modified SIS demonstrates a potential improvement in biological activity as suggested by more consistent smooth muscle regeneration in a canine model of bladder regeneration. This study demonstrates both the feasibilty and potential of synthetically‐modified natural biomaterials.

OBJECTIVE

? To determine if hyaluronic acid (HA) can be incorporated into porcine small intestinal submucosa (SIS) through poly (lactide‐co‐glycolide‐acid) (PLGA) nanoparticles to improve the consistency of the naturally derived biomaterial and promote bladder tissue regeneration.

METHODS

? Beagle dogs were subjected to 40% partial cystectomy followed by bladder augmentation with commercial SIS or HA‐PLGA‐modified SIS. ? Urodynamic testing was performed before and after augmentation to assess bladder volume. ? A scoring system was created to evaluate gross and histological presentations of regenerative bladders.

RESULTS

? All dogs showed full‐thickness bladder regeneration. ? Histological assessment showed improved smooth muscle regeneration in the HA‐PLGA‐modified SIS group. ? For both groups of dogs, urodynamics and graft measurements showed an approximate 40% reduction in bladder capacity and graft size from pre‐augmentation to post‐regeneration measurements. ? Application of the scoring system and statistical analysis failed to show a significant difference between the groups.

CONCLUSIONS

? SIS can be modified through the addition of HA‐PLGA nanoparticles. The modified grafts showed evidence of improved smooth muscle regeneration on histological assessment, although this difference was not evident on a novel grading scale. ? The volume loss and graft shrinkage experienced are consistent with previous models of SIS bladder regeneration at the 10‐week time point. ? Additional research into the delivery of HA and the long‐term benefits of HA on bladder regeneration is needed to determine the full benefit of HA‐PLGA‐modified SIS. In addition, a more objective biochemical characterization will be needed to evaluate the quality of regeneration.     

Regeneration of uterine horn using porcine small intestinal submucosa grafts in rabbits.

Tubal factor infertility may be reversed using porcine small-intestinal submucosa (SIS). The method uses as a model the New Zealand White rabbit uerine horn. In surgery, SIS grafts were prepared from porcine jejunum; the uterine horn segment was resected and a graft was placed; then the contralateral adnexa was resected. Fecundability was tested with natural mating. Three out of six rabbits became pregnant. Gross and microscopic examination confirmed regeneration of all tissue layers. Thus, this study determined that SIS facilitates successful regeneration of uterine horn morphology in a manner similar to that observed in other tissues and species.     

Endoscopic urethroplasty with unseeded small intestinal submucosa collagen matrix grafts: a pilot study

PURPOSE: We evaluated small intestinal submucosa (SIS) as a substitute for skin in endoscopic urethroplasty performed as treatment for inflammatory and iatrogenic strictures of the male bulbar urethra, and in the early treatment of bulbomembranous urethral injuries associated with recent pelvic fractures. Tissue integration and epithelialization of SIS in endoscopic urethroplasty were assessed, as was the long-term maintenance of urethral patency following this treatment. MATERIALS AND METHODS: Nine patients with bulbar urethral strictures defined by urethrography were enrolled in the study. Following optical urethrotomy the SIS grafts were tubularized over a purpose specific graft carrying balloon device and secured into the opened urethra as described for endoscopic urethroplasty. Patients were followed with urethroscopy and urethrography at regular intervals as per protocol or when symptoms arose. Failure was defined as the need for any further intervention. RESULTS: Two patients with short inflammatory strictures maintained urethral patency without any intervention at 1 and 2 years, respectively. Stricture recurrence developed in 6 patients within 3 months of surgery. Of these, 3 have undergone subsequent open urethroplasty, 2 are currently awaiting urethroplasty and 1 is maintaining urethral patency with regular self-dilatation. One patient was lost to followup. CONCLUSIONS: Endoscopic urethroplasty with unseeded SIS grafts was unsuccessful in this study.     

Experimental assessment of small intestinal submucosa as a small bowel graft in a rat model

Background/Purpose:

Small intestinal submucosa (SIS) is an extracellular matrix used in tissue engineering. The purpose of this study is to evaluate the feasibility of using SIS as a scafford for small bowel regeneration in a rat model.

Methods:

A 2-cm length tubular SIS graft from donor Sprague Dawley rats was interposed with bilateral anastomosis in the median tract of an isolated ileal loop of Lewis rats used to construct an ileostomy. The grafts were harvested and analyzed at each of the time-points ranging from 2 weeks to 24 weeks after operation using histology and immunohistochemistry.

Results:

Macroscopic examination found no adhesion in the surrounding area of neointestine by 24 weeks, and no stenosis was visible. The shrinkage of neointestine was indicated from 20% to 40%. Histologic and immunohistochemical evaluation showed that SIS grafts were colonized by numerous inflammation cells by 2 weeks. Neovascularization was evident, but the luminal surface was not epithelized. By 4 weeks, transitional mucosal epithelial layer began to line the luminal surface of the graft, and nearly 70% luminal surface of the graft had been covered by mucosal epithelium at 8 weeks. By 12 weeks, the luminal surface was covered completely by a mucosal layer with distinct bundles of smooth muscle cells in the neointestine. At 24 weeks, the neointestine wall showed 3 layers of mucosa, smooth muscle, and serosa.

Conclusions:

The preliminary study suggested that SIS allow rapid regeneration of mucosa and smooth muscle and might be a viable material for the creation of neointestine.     

Porcine small intestine submucosa for repair of goat meniscal defects

This study evaluated the effectiveness of small intestine submucosa used as a graft to fill surgically created meniscal defects in a large animal model. Four goats underwent unilateral interior subtotal meniscectomies (approximately 70%) within the avascular portion of the medial meniscus. The contralateral meniscus acted as a control. Grafts of porcine small intestine submucosa were trimmed to fill the resected defects and sutured into place. After surgery the operated knees were casted in partial flexion to limit weight bearing on the affected limb. All of the animals were sacrificed at 12 weeks at which point meniscal regeneration and articular cartilage degradation were evaluated by gross and histologic examination. Grossly, the defects in the small intestine submucosa-grafted goats were partially filled with meniscal-appearing connective tissue. Histologically, the replacement tissue was typified by the presence of dense, cellular, irregularly organized connective tissue. Evaluation of the articular cartilage displayed increased degeneration in the grafted compartment of the operative knees. Each of the operative menisci partially regenerated. The grafts were conducive to repopulation with host meniscal elements. Despite partial meniscal regeneration, comparatively more articular cartilage degeneration in the treated knees was observed than in the untreated contralateral controls.     

Small intestinal submucosa as a bioscaffold for biliary tract regeneration

BACKGROUND: Porcine small intestinal submucosa (SIS) biograft is used as a bioscaffold for regeneration of a variety of tissues. To date, SIS has not been used as a biliary tract graft. The purpose of this study was to evaluate the feasibility of using SIS as a scaffold for bile duct tissue regeneration in a canine model. METHODS: Fifteen, 25- to 35-kg mongrel dogs underwent midline laparotomy and exposure of the common bile duct. Nine dogs had a longitudinal choledochotomy and a 2- x 1-cm elliptical patch of 4-ply SIS placed using 6-0 polypropylene suture. Six dogs had the anterior two thirds of the bile duct resected and a 2- to 3-cm tubularized 4-ply SIS interposition graft placed. Dogs were killed at intervals ranging from 2 weeks to 5 months. Before killing, liver function tests (alkaline phosphatase [U/L] and total bilirubin [mg/dL]) were evaluated, cholangiograms were performed, and the bile duct was examined histologically. RESULTS: Fourteen out of 15 dogs survived and were healthy at the time of killing. The one failure was a result of a bile leak in a patched animal. The SIS showed signs of incorporation with infiltration of native fibroblasts, blood vessels, and biliary mucosa within 2 weeks. Within 3 months the SIS graft was replaced with native collagen covered with a biliary epithelium. No changes occurred at 5-month follow-up. One animal with an interposition graft developed a stricture at the proximal anastomosis within 2 months. In the remaining dogs, liver enzymes were normal, and the caliber of the common bile duct remained normal. CONCLUSIONS: SIS can be used for regeneration of bile duct tissue in a canine model. In 13 of 15 dogs SIS resulted in regeneration of canine common bile duct when used as a patch or as an interposition graft. The potential for the use of SIS as a patch for biliary stricturoplasty, or as an interposition graft for repair of complex biliary injuries is encouraging.     

小肠黏膜下层移植修复海绵体神经损伤的实验研究

目的:研究小肠黏膜下层(SIS)移植替代损伤的双侧海绵体神经(CN)恢复大鼠的勃起功能。方法:制备SIS,建立动物模型,分为CN损伤组、假手术组、SIS移植组,分别给予切断双侧的CN、仅游离CN以及SIS移植修复损伤的CN。术后3个月进行阿朴吗啡试验,了解阴茎勃起情况。取中、后段阴茎海绵体组织,进行nNOS免疫组化染色,记录nNOS阳性神经纤维的数目。结果:阿朴吗啡试验:30 min内SIS移植组72.73%(8/11)的大鼠出现阴茎勃起,平均勃起(1.07±0.89)次;CN损伤组勃起率和勃起次数均为0;假手术组则为90.91%(10/11)和(2.19±1.17)次。无论是勃起率还是勃起次数,SIS移植组均显著高于CN损伤组(P<0.01),但仍然比假手术组低(P<0.05)。nNOS神经纤维数目:SIS移植组为(70.36±10.09)条,CN损伤组为(22.09±4.76)条,差异有统计学意义(P<0.01),但二者均低于假手术组[(90.81±5.69)条,P<0.01]。结论:SIS作为移植物修复损伤的大鼠CN损伤,有利于恢复CN损伤所致的勃起功能障碍。     

Penile enhancement using a porcine small intestinal submucosa graft in a rat model

Several biodegradable materials have been experimented for penile enhancement, but none show the potential for clinical use. This study was designed to use porcine small intestinal submucosa (SIS) augmenting the normal tunica albuginea to increase the functional girth of the rat penis. In all, 20 adult male Sprague-Dawley rats constituted the study population. The animals were divided into two groups: group 1 consisted of the control (n=10) and group 2 (n=10) consisted of rats that underwent penile enhancement by a longitudinal I-shaped incision of the tunica albuginea on both sides, and the dissection of the plane between tunica albuginea and cavernosal tissue was carried out (n=10). The incision was then patched with a 3 x 10 mm2 piece of SIS, using a 6/0 nylon suture material. The penile length and mid-circumference were then measured using a Vernier Caliper before and 2 months after surgery. All rat penises underwent histological examination using Masson's trichome and Verhoff's van Giesen's stain for collagen and elastic fibers. The penile length, mid-circumference and degree of fibrosis score were expressed as mean+/-s.e. (standard error) and analyzed using a Wilcoxon rank-sum test. A statistical significance was accepted at P-value < or =0.05. Our results showed similar preoperative penile length and circumference in both groups. However, 2 months after the surgery, the mean penile circumference of the SIS group has grown significantly larger than the control group, while the mean penile length remained unchanged. The histological study of the rat penises revealed minimal amounts of fibrosis under the graft, and the elastic fibers of the graft showed orientation in a circular manner. In conclusion, SIS appears promising for material use in a penile enhancement.     

小肠黏膜下基质用于胆管重建的研究

目的 探讨在肝外胆管节段性缺失时,小肠黏膜下基质(SIS)用于重建胆管的可行性及相关机制,为胆道重建寻找新的思路.方法 SIS制成管状,实验犬6条,胆总管节段性切除12～15 mm,以SIS管桥接.分别于术后3、9周取材,通过胆道造影、病理等方法观察胆道重建过程和组织重构效果.结果存活率100％,胆瘘、胆道闭塞发生率为0.胆管收缩、狭窄现象普遍,长度收缩约(13.5±4.1)％.病理检查见胆管内膜细胞覆盖植入SIS段胆管,管壁由成纤维细胞构成,新生血管丰富.结论 SIS材料制成管状用于替代胆管节段性缺失是可行的,可以避免胆漏,无管腔塌陷、闭塞.     

Mitochondrial metabolism in the rat during bladder regeneration induced by small intestinal submucosa

OBJECTIVE: To assess mitochondrial metabolism of bladder tissue induced by small-intestinal submucosa (SIS), by comparing the mitochondrial enzyme metabolism in this tissue with that in normal bladder tissue and thus evaluate intracellular normality. MATERIAL AND METHODS: In all, 70 rats were grouped into healthy controls (10), surgical controls with a simple bladder incision (15) and rats treated by partial cystectomy with replacement by the SIS graft (45). At 1, 3 and 6 months the rats were killed, the enzymes of mitochondrial respiratory chain complexes assayed, and the respiration of permeabilized bladder fibres assessed using polarographic analysis. RESULTS: The enzyme activities of control and treated rats at 3 months were identical. The results from the polarographic analysis of respiration were also similar to that in normal tissue apart from a decrease in the number of mitochondria. Histologically, there was complete regeneration at 6 months. CONCLUSION: After a phase of inflammation the bladder regenerates after a patch is placed. The new tissue has the same enzymatic and histological features as normal bladder tissue.     

Rat epigastric pedicle model: a clinically relevant evaluation of 1-mm PTFE grafts

Twenty-eight Sprague-Dawley rats had 1.0-mm polytetrafluoroethylene (PTFE) interposition grafts placed in their femoral artery that supplied a 3.0 X 3.0 cm epigastric pedicle flap. The model is the first to evaluate 1.0-mm PTFE under rigorous, clinically simulating conditions. Anastomoses employed a new, continuous telescoping suture technique. Grafts were selected for histological and electron microscopic evaluation, which confirmed the development of a cellular neointimal lining. Viability of the flap model had a 90% correlation with graft patency. Overall graft patency was 50%. Patency was documented in some specimens harvested at 200 days. The flap model is a good indicator of graft patency. PTFE grafts of 1.0-mm internal diameter are not yet equivalent to autologous veins. Future refinements, particularly aseptic technique, are expected to result in even higher patency rates.     

Chondropathy after meniscal tear or partial meniscectomy in a canine model.

A primary goal in considering treatment for meniscal injuries is the preservation of the health of the articular cartilage. However, the chondroprotective effects of various techniques for meniscal injury treatments are unknown. We used a canine model to quantify articular cartilage degeneration in the medial compartment of the canine knee, resulting from a surgically created tear or a partial meniscectomy (PM) of the posterior region of the medial meniscus (each group, n = 10). After sacrifice at 12 weeks, the development of gross chondropathy and the changes in cartilage tensile stiffness were quantified, and correlations between these measurements were examined. Both treatment surgical treatment groups caused significantly greater gross chondropathy as compared to the unoperated contralateral controls. Cartilage tensile stiffness was significantly lower than unoperated controls by nearly 28% in both experimental groups. However, there were no significant differences observed between the gross chondropathy or the cartilage mechanical property changes between the experimental groups. Importantly, the severity of gross chondropathy was found to significantly correlate with the decrement in tensile stiffness properties of the articular cartilage. These findings indicate that significant degeneration of canine articular cartilage develops to a similar degree in the presence of a partially healed meniscus tear or a PM of the knee.     

Porcine small intestinal submucosa as a carrier for skin flap prefabrication

The feasibility of porcine small intestinal submucosa (SIS) as a carrier in skin flap prefabrication was examined in this study. Thirty-eight rats were randomly divided into five groups. The saphenous vascular bundle was used as the vascular carrier. In group 1 (n = 8), an arteriovenous fistula was made by anastomosis of distal saphenous artery and vein. A SIS patch (1.5 x 2 cm2) was placed underneath the vascular bundle. In group 2 (n = 8), the vascular bundle was isolated and laid over the SIS patch. The distal saphenous vessels were ligated when the flap was raised. In group 3 (n = 8), an arteriovenous fistula was made without SIS implant. In group 4 (n = 8), the flap was raised with only the vascular bundle with the distal end ligated. After 2 weeks of maturation, the flap was raised with only the vascular bundle. In group 5 (n = 6), SIS was implanted and the flap including the SIS patch was raised and replaced without the vascular pedicles. The survival of the flaps and histology were evaluated at 5 days after flap replacement. The results showed that the average survival area in group 1 was 99% +/- 3% and the survival area in group 2 was 86% +/- 16%. The mean survival areas in group 3 and 4 were 60% +/- 9% and 25% +/- 10%, respectively. No flap survival was observed in the group 5. These were significantly lower than in groups 1 and 2 (p < 0.05, p < 0.01). Histology showed that SIS patch was incorporated into the adjacent connective tissue and increased amounts of neovascularization were seen between the collagenous sheets and dermis. In conclusion, this study demonstrated that porcine SIS can incorporate into the adjacent tissue and induce angiogenesis in flap prefabrication. This biomaterial can provide a scaffold for supporting and enhancing the survival of vascular prefabricated skin flap.     

Mucosal regeneration of a duodenal defect using small intestine submucosa

Intestinal tissue engineering has the potential of developing new treatment strategies for patients with a deficit in intestinal surface area. The purpose of this study was to investigate the capacity of small intestine submucosa (SIS) to regenerate intestinal epithelia in a rodent model for a duodenal defect. A duodenotomy was created in 20 Sprague-Dawley rats and was repaired with a circular patch of SIS. The repaired sites were retrieved 1, 2, 4, and 12 weeks after implantation. The size of the residual mucosal defect was measured. The retrieved tissues were fixed in formalin and processed by standard histologic techniques. The animals tolerated the procedure well. The mean size of the mucosal defects significantly decreased with time. Complete epithelialization of the defects was noted within 4 weeks in three of five samples. Histologically, the defects were lined with crypts and villi, but the muscularis layer did not regenerate. In the rodent model, SIS can be used as a patch to repair a duodenotomy. Mucosal regeneration was observed in the area of the defect. Further studies will determine whether SIS may be used to preserve or increase mucosal surface area in patients whose bowel length is compromised.     

Ischemia/reperfusion: a clinically relevant model of intestinal injury yielding systemic inflammation

Background/Purpose

Multisystem organ failure (MSOF) is a major cause of morbidity and mortality in the critically ill patient. Animal models of endotoxin-induced sepsis were used to develop therapeutic regimens, which thus far have failed in clinical trials. Because multiple etiologies of MSOF affect the intestine, the authors hypothesized that during sepsis the gut may act as a possible trigger of the inflammatory cascade. As ischemia and reperfusion of the small intestine disrupts gut barrier function, thereby activating systemic inflammatory responses, the authors evaluated a murine model of ischemia/reperfusion to investigate these systemic responses to local mucosal and epithelial injury.

Methods

C57BL/10 and Balb/c mice underwent variable amounts of gut ischemia by superior mesenteric artery occlusion. Animals were evaluated for survival as well as gross and microscopic intestinal damage.

Results

Maximal ischemic damage occurred in the distal jejunum and proximal ileum. More severe epithelial damage and transmural inflammation were observed in C57BL/10 mice, which correlated with a higher mortality.

Conclusions

This model mimics what is observed clinically with intestinal injury resulting from a progressive ischemic insult with eventual systemic manifestations. This reproducible model of systemic inflammation elicits variable responses from genetically different animals, the results of which may lead to a better understanding of MSOF.