Multiple Oblique Illumination and High-Definition Microscopy for Breast Fine-Needle Aspirates

ABSTRACT We wished to determine whether small-intestinal submucosa (SIS) will epithelialize when used as a ureteral replacement material. An 11-mm segment of native ureter was excised from eight New Zealand White rabbits and replaced with an 11-mm porcine SIS graft, which was circumferentially wrapped around a ureteral stent. The SIS ureteral grafts were harvested at 11 days or 35 days postimplantation and examined grossly and by standard light microscopy techniques. Partial epithelialization with the ingrowth of urothelium, smooth muscle cells, and blood vessels was observed in the grafts harvested at 11 days postimplantation. The SIS ureteral grafts examined at 35 days postimplantation showed additional restructuring of the smooth muscle cell layer and more organized epithelialization in comparison to the SIS graft examined at 11 days. After 35 days of regenerative healing, elements of all three layers of the native ureter were observed within the collagen matrix of the SIS graft. No significant complications were observed, but all subjects (8/8) demonstrated mild intra-abdominal adhesions. Mild collecting system dilatations were observed in 4/4 (100%) of the animals harvested at 35 days and in 0/4 (0%) of the animals harvested at 11 days. We have this demonstrated in this preliminary study that SIS xenografts will epithelialize when used as a ureteral replacement material. The repair mechanism of these ureteral grafts occurred through a regenerative healing process rather than by scar formation. With further studies, this material may prove to be a useful treatment option in patients with ureteral injuries.     

First prize: ureteral segmental replacement revisited

BACKGROUND AND PURPOSE: Long strictures of the proximal ureter are difficult to manage, and circumferential replacement with various natural and synthetic materials has been unsuccessful. We sought to use cultured autologous cells seeded onto graft material for proximal-ureteral replacement. Additionally, we wished to determine if urothelial cell-seeded de-epithelialized small bowel would generate adequate ureteral replacement. MATERIALS AND METHODS: Three sets of experiments were performed. First, autologous pig-bladder smooth-muscle and urothelial cells were expanded in culture on large sheets of multilayer small-intestinal submucosa (SIS). These sheets were then tubularized and used to replace a 5-cm segment of proximal ureter in pigs. Second, autologous cells harvested from the bladders of Beagle dogs were cultured and seeded on porcine ureteral acellular matrix, which was used to replace a 3-cm segment of ureter in dogs. Segments were wrapped in omentum to enhance vascularity. Third, a de-epithelialized small-bowel segment seeded with autologous bladder-epithelial cells was transversally retubularized (Monti) into a 4-cm ureteral replacement. Follow-up studies consisted of retrograde pyelography, serum chemistry assays, hematoxylin/eosin studies, and immunohistopathologic examination using antibodies against alpha-smooth-muscle actin and pancytokeratin AE1-AE3. RESULTS: Coculture of urinary-tract cells on large segments of SIS failed to create adequate ureteral replacement. All grafts were contracted and stenotic, with complete obstruction of the ipsilateral renal unit. Similar results were seen in the Beagles. Despite clinical obstruction and gross contraction of the graft, a circumferential muscular ureteral wall lined with multilayer transitional epithelium was present. Urotheliumseeded de-epithelialized Monti bowel segments resulted in patent ureteral replacement without hydroureteronephrosis and with normal renal function, serum electrolytes, and acid-base balance. However, bowel mucosa fully regenerated, with multilayer transitional epithelium growing adluminally in continuity with the proximal and distal anastomotic sites. CONCLUSIONS: Seeding of ureteral grafts with autologous bladder cells does not promote success in two largeanimal models using different xenogenic acellular matrices. However, muscle and urothelium regeneration occurs with ureteral acellular matrix in the dog. Urothelium-seeded de-epithelialized Monti bowel segments may be an acceptable substitute for long proximal ureteral segments. Further technical refinements are required to replace the bowel mucosa completely with normal urothelium.     

Ureteral replacement using collagen sponge tube grafts

The use of a collagen sponge tube graft as a material for segmental ureteral replacement was investigated. The structural design of the collagen sponge graft was achieved by cell culture on the matrix. MGH-U1 cells, derived from bladder cell carcinoma, were grown in vitro on the collagen sponge matrix with excellent biocompatibility and without evidence of cytotoxicity. The collagen sponge demonstrated biodegradability when implanted subcutaneously in dogs. However, a urine exposure test of collagen sponge in rat bladders revealed extensive salt deposits on its surface in some rats, as observed by crystallographic examination. Segmental ureteral replacements by collagen sponge tube grafts, accompanied by ureteral splint catheters, were performed in dogs. There was extensive uro-epithelial cell regeneration on the inner surface of the collagen grafts, without evidence of severe hydronephrosis, 5 to 12 weeks following the procedure. The results indicate the potential for ureteral replacement by collagen sponge tube grafts, which would act as non-toxic, biodegradable scaffolds inducing the regenerative activity of the ureter.     

Reconstruction of ureteral defects with microvascular vein grafts in a rat model

This study was undertaken to test the performance of an autologous vein graft as a ureteral replacement in the rat model. Twenty-six rats were divided into three groups. In Group 1 (n = 10), the animals had a 3-mm segment of the ipsilateral ureter excised and the ureteral defect repaired, using a superficial epigastric vein graft. In Group 2 (n = 10), the same ureteral defect was created and again repaired, using a superficial epigastric vein graft, with the addition of a Silastic stent. The control, Group 3 (n = 6), had the ureter transected and repaired solely by means of primary anastomosis. Animals from each group underwent urography and were sacrificed for histology at three different postoperative intervals: 1, 4, and 12 weeks. Postoperative urography results showed normal renal function in the animals with ureteral reconstruction using vein grafting aided by a stent, as well as in those with primary ureteral anastomosis. No renal function return was seen in the animals with ureteral reconstruction by vein grafting alone, without stent support. Histologically, a progressive loss of the vascular endothelium, and replacement with the urothelium typical of the ureter, was seen in the stented vein grafts. Severe ureteral obstruction at the proximal site of the graft and hydronephrosis were seen in the vein-graft group without stenting. This study demonstrates that autologous vein grafts can be used successfully to correct a ureteral deficit, contingent on accurate microsurgical technique and immediate stenting.     

Use of single layer small intestinal submucosa for long segment ureteral replacement: a pilot study

PURPOSE: Previous studies have demonstrated successful use of small intestinal submucosa (SIS) as a tube for replacing short segment (11 mm) proximal ureteral defects. However, such small segment ureteral defects could be managed by resection re-anastomosis. We evaluated the use of 1-layer SIS as a tube for the replacement of long segment ureteral defects. MATERIALS AND METHODS: The ureters of 5 female mongrel dogs were accessed through a median laparotomy incision. A 4 cm segment of mid ureter was resected on the right side. The right ureteral segments were replaced by tubularized SIS segments using 6-zero polydioxanone interrupted sutures. Internal pigtail stents were left for 6 weeks. All animals were sacrificed at 12 weeks. Ureteral patency was assessed by excretory urography and magnetic resonance urography 7 and 12 weeks after the initial procedures. Inflammation and regeneration were assessed histologically. RESULTS: At 12 weeks all ureters on the experimental side were completely occluded with significant hydroureteronephrosis and the subsequent deterioration of kidney function. At autopsy there was failure to calibrate any of the experimental ureters with a 3Fr catheter. Although histologically urothelium and muscular cells had proliferated over the graft, they were embedded in an intense fibrotic and inflammatory process. CONCLUSIONS: Technically 1-layer SIS was easily modeled, providing the conditions for watertight anastomosis. The regeneration of urothelium and muscle was induced and supported by the graft. However, functional replacement was not successful. One-layer SIS is not a suitable material for replacing long segment (4 cm) ureteral defects.     

小肠黏膜下层修复尿道的实验研究

目的探讨小肠黏膜下层(small intestinal submucosa,SIS)在尿道修复重建中的应用价值.方法 24只日本雄性大耳白兔,随机分为A、B、C及D组(n=6).A、B组切除前尿道2.0 cm,A组,用管状SIS修复尿道缺损;B组将其断端与周围组织直接缝合作为对照.C、D组仅切除2.0 cm尿道前壁,保留一半尿道壁为底板,C组用片状SIS修复尿道缺损;D组将其残端与周围组织直接缝合作为对照.均于修复后6、12周行组织学观察;12周行尿道膀胱造影及尿动力学检查.结果术后6周,A、C组修复的尿道有单层上皮细胞覆盖,基层组织中可见SIS的微小碎片包裹,出现不规则紊乱的平滑肌细胞生长,A组较C组的炎性反应重,有白细胞及淋巴细胞浸润,C组出现新生血管.术后12周,C组的上皮组织及基层下组织与D组无明显差别,平滑肌排列规则,血管数目进一步增多,炎性反应消失,未见SIS组织;A组中仍可见少数SIS的微小碎片;B组1只、D组2只尿道自行修复,余可见尿道闭塞,大量结缔组织生长,炎性细胞浸润,无正常上皮结构.术后12周尿道膀胱造影,A、C组可见尿道完整、光滑,无尿液外渗、尿道憩室等形成;尿动力学检查示A、C组的膀胱容量、最大尿道压分别与术前比较,差异无统计学意义(P＞0.05),而B、D组不能置入测压管检测.结论 SIS可作为兔尿道修复重建的良好支架材料,片状SIS修复优于管状SIS修复.     

Laparoscopic replacement of urinary tract segments using biodegradable materials in a large-animal model.

OBJECTIVE: We elected to evaluate laparoscopic segmental bladder and ureteral replacement with free biodegradable graft materials in a large-animal model. MATERIALS AND METHODS: In 18 Yucatan minipigs, a 1.5- to 2.8-cm segment of the upper ureter was excised. In 15 study animals, the ureter was laparoscopically replaced: by a stinted (6F double-J stent) tube graft made of acellular matrix (AMX) prepared from minipig ureters (MUMX) in 6 animals, acellular matrix prepared from domestic pig ureters (DUMX) in 3, and small-intestinal submucosa (SIS) in 6. In 3 control animals, the ureteral gap was bridged only by an indwelling stent. The stent was removed at 6 weeks, and retrograde ureteropyelography was performed preoperatively and at 8 and 12 weeks postoperatively, when animals were sacrificed. In 18 Yucatan minipigs, 3 x 3-cm bladder dome segments were laparoscopically replaced: by acellular matrix prepared from minipig small bowel (MBMX) in 6 animals, and SIS in 6 animals. The bladder was closed primarily in 6 control animals. Bladder capacity was assessed preoperatively and at 6 and at 12 weeks, when the animals were sacrificed. RESULTS: The average operating time for ureteral replacement was 187 (range 105-360) minutes. At 12 weeks, all animals had complete obstruction at the level of the replacement, with fibrosis +/- bone formation at the level of the stricture. For the bladder replacement groups, the average operating time was 147 (range 85-200) minutes. At 12 weeks, the bladder capacity was 60% of the preoperative capacity in the control group, 118% in the MBMX group, and 142% in the SIS group. Histologic examination showed regeneration of urothelium and some muscle with both MBMX and SIS. CONCLUSIONS: We were able to develop a reliable laparoscopic technique for both segmental ureteral and partial bladder replacement in a porcine model. With AMX and SIS replacement, regeneration of urothelium occurred in both ureter and bladder. However, functional replacement was successful only in the bladder.     

Laparoscopic augmentation cystoplasty with different biodegradable grafts in an animal model

PURPOSE: Recently a variety of biodegradable organic materials have been used for bladder wall replacement. We sought to study the effectiveness of 4 different types of biodegradable materials for bladder augmentation using laparoscopic techniques. MATERIALS AND METHODS: Thirty one minipigs underwent successful transperitoneal laparoscopic partial cystectomy and subsequent closure (6 control) or patch augmentation (25): porcine bowel acellular tissue matrix (ATM) (6), bovine pericardium (BPC) (6), human placental membranes (HPM) (6) or porcine small intestinal submucosa (SIS) (7). An intracorporeal suturing technique with the EndoStitch device (U.S. Surgical, Norwalk, CT) and Lapra-Ty clips (Ethicon, Enodsurgery Inc. Cincinnati, OH) was used to anastomose the graft to the bladder wall. Postoperatively, a urethral catheter was left for one week. Bladders were evaluated by cystoscopy at 6 and 12 weeks and harvested at 12 weeks. RESULTS: Grafts remained in place in all groups except for the BPC group, where all grafts failed to incorporate. For the ATM and SIS groups, at 6 weeks, there was mucosal coverage of the grafts without evidence of encrustation. In the control group, at 12 weeks, the bladder capacity was 23% less than preoperatively. In the ATM, HPM and SIS groups, at 12 weeks, the bladder capacities were larger than preoperatively by 16%, 51% and 43% respectively; also the grafts had contracted to 70%, 65%, and 60% of their original sizes, respectively. Histologically, there was patchy epithelialization of ATM and SIS grafts with a mixture of squamoid and transitional cell epithelia. The graft persisted as a well-vascularized fibrous band in HPM, ATM, and SIS without evidence of significant inflammatory response. CONCLUSION: A laparoscopic technique for partial bladder wall replacement using a free graft is feasible. The biodegradable grafts of ATM, HPM and SIS are tolerated by host bladder and are associated with predominantly only mucosal regeneration at 12 weeks post-operatively.     

Small Intestinal Submucosa as a Vascular Graft: A Review

Continuing investigations of vascular graft materials suggest that unacceptable graft complications continue and that the ideal graft material has not yet been found. We have developed and tested a biologic vascular graft material, small intestine submucosa (SIS), in normal dogs. This material, when used as an autograft, allograft, or xenograft has demonstrated biocompatibility and high patency rates in aorta, carotid and femoral arteries, and superior vena cava locations. The grafts are completely endothelialized at 28 days post-implantation. At 90 days, the grafts are histologically similar to normal arteries and veins and contain a smooth muscle media and a dense fibrous connective tissue adventita. Follow-up periods of up to 5 years found no evidence of infection, intimal hyperplasia, or aneurysmal dilation. One infection-challenge study suggested that SIS may be infection resistant, possibly because of early capillary penetration of the SIS (2 to 4 days after implantation) and delivery of body defenses to the local site. We conclude that SIS is a suitable blood interface material and is worthy of continued investigation. It may serve as a structural framework for the application of tissue engineering technologies in the development of the elusive ideal vascular graft material.     

Small intestinal submucosa as a small-caliber venous graft: a novel model for hepatocyte transplantation on synthetic biodegradable polymer scaffolds with direct access to the portal venous system

BACKGROUND/PURPOSE: Hepatotrophic factors in the portal blood are critically important for the survival of heterotopically transplanted hepatocytes. Currently, no model exists for the implantation of hepatocytes on biodegradable polymer scaffolds with direct access to the portal blood. This study investigates the use of small intestinal submucosa (SIS) as a small-caliber venous conduit that may be used for the implantation of tissue-engineered liver. METHODS: SIS was prepared from segments of rat jejunum and implanted as a venous conduit between the portal vein and inferior vena cava in 26 heparinized Lewis rats. Venograms were performed periodically, and the grafts were harvested at various time-points and examined by scanning electron microscopy (SEM) and histology. Von Willebrand Factor (vWF) staining was performed to assess endothelialization. RESULTS: Five rats died of technical complications. Seventeen of 21 rats (81%) maintained patent grafts at the time of death up to 8 weeks. Venograms demonstrated patent grafts at 3 and 8 weeks. SEM results showed a smooth luminal surface with endothelial-like cells by 3 weeks. Histology demonstrated a confluent luminal endothelial monolayer, absence of thrombus, and neovascularization in the SIS graft. VWF staining results were positive, confirming the growth of endothelial cells on the luminal surface. In preliminary studies, implantation of hepatocytes seeded on biodegradable polymer tubes into the SIS graft demonstrated clusters of viable cells after 2 days. CONCLUSIONS: Rat SIS can be prepared readily, maintains high patency as a small-caliber venous graft, and may be a useful model for the transplantation of tissue-engineered liver with access to the portal circulation.     

Grafts of porcine intestinal submucosa for repair of cervical and abdominal esophageal defects in the rat.

Porcine small intestinal submucosa (SIS) is a cell-free collagen matrix that has demonstrated its ability as scaffold material for constructive remodeling of damaged or missing tissue. The purpose of this study was to evaluate the morphology and function of esophagoplasty in rat using a porcine SIS scaffold for the repair of a semi-circumferential defect in the cervical or in the abdominal esophagus. Sixty-seven rats underwent surgical excision of the anterior wall either of the cervical or of the abdominal esophagus and subsequent repair of the defect with an SIS patch graft. Outcomes of weight gain, signs of dysphagia, hematological and serum chemistry parameters, and barium swallow studies were used to assess the progress of healing and function over a 150-day time period. The grafts were studied for gross changes and histology at predetermined time points. Ninety-four percent of the SIS-treated rats survived, showing no significant differences in survival rate between groups. The grafted animals did well, without signs of dysphagia, and gaining weight. Barium swallow studies showed no evidence of fistula, significant stenosis, or diverticula. No hematological or serum biochemistry abnormalities were found. By 150 days, the SIS graft was replaced with esophageal-derived tissues. Specimens were completely lined by keratinized stratified squamous epithelium and showed complete regeneration of muscle fibers and scarce immunoreactivity for nerve. In the rat model, a patch graft technique using porcine SIS appears to induce esophageal regrowth either in cervical and abdominal esophagus. The repair mechanism occurred through a regenerative healing process.     

New Regenerative Vascular Grafts for Hemodialysis Access: Evaluation of a Preclinical Animal Model

The purpose of this study was to evaluate a suitable animal model for the in vivo evaluation of patency and vascular tissue regeneration in small intestinal submucosa (SIS) vascular grafts for hemodialysis access. First, a 4-mm U-shaped SIS vascular graft was implanted between the internal carotid artery (CA) and the external jugular vein (JV) in five sheep and six swine. The U-shape grafts remained functional for 53 ± 4 days in sheep and 32 ± 2 days in swine. The sheep model presented exaggerated inflammation, so the swine model was selected for the in vivo study. Based on these initial results, a 4-mm C-shape SIS vascular graft with SIS circumferential reinforcement was developed to mechanically improve the vascular graft and manage complications identified during surgery in both sheep and swine. The C-shape vascular graft was implanted in a swine model (n = 3) between the CA and JV. GORE-TEX® vascular grafts were used as controls in the contralateral side of the neck. C-shape grafts remained patent for 47 ± 4 days, whereas the GORE-TEX® grafts were patent for 30 ± 15 days. The C-shape vascular graft was easier to handle during surgery, and its circumferential reinforcement improved in vivo patency, avoiding kinks in the graft after implantation. Histological results showed neovascularization and some regeneration with the alignment of endothelial cells in the vascular wall of the grafts. The model developed may be helpful in other research involving in vivo studies of vascular grafts for hemodialysis access.     

Vein cuff interposition prevents juxta-anastomotic neointimal hyperplasia.

This study sought to minimize juxta-anastomotic neointimal hyperplasia (JNIH) following the use of polytetrafluorethylene (PTFE) conduits. PTFE anastomoses to canine carotid arteries (noncuff grafts) were compared with grafts with vein cuffs interposed proximally and distally between the graft and native artery. This technique has been suggested clinically for below-knee PTFE femoropopliteal reconstruction. Twelve dogs received aspirin for 1 week before operation, which was continued after each animal received bilateral cuff and noncuff 4-mm PTFE grafts. At sacrifice, after 3-12 weeks, graft patency was assessed and luminal diameters measured with ophthalmic calipers at three sites along the anastomoses and 1 mm proximal or distal to graft toe (A' diameter). Specimens were perfusion fixed at arterial pressure for gross and histologic study; selected arteries were additionally fixed with 4% buffered glutaraldehyde, stored at 4 C, and examined immunochemically using antimyosin antibody immunopurified for smooth muscle. Overall patency of noncuff grafts in 11 long-term surviving dogs was 4 of 11; patency of the cuff grafts was 7 of 11. Regardless of graft thrombosis, antibody positive cellular proliferation occurred mainly at noncuffed PTFE anastomoses. Luminal encroachment was predominantly due to subintimal proliferation of cells highly reactive to smooth muscle derived antibody. JNIH was most prominent 1 mm distal to the graft toe (A' distal diameter). Average A' for noncuff grafts was 1.82 mm +/- 0.97 SEM; average A' diameter for cuff grafts was 3.41 mm +/- 0.74 SEM (p less than 0.001). Vein cuff inhibition of proliferation of smooth muscle or cells derived from smooth muscle possibly relates to wider distribution of kinetic energy (less compliance mismatch) or to interposition of venous endothelium.     

Comparison of small-intestinal submucosa and expanded polytetrafluoroethylene as a vascular conduit in the presence of gram-positive contamination

OBJECTIVE: As a vascular conduit, expanded polytetrafluoroethylene (ePTFE) is susceptible to graft infection with Gram-positive organisms. Biomaterials, such as porcine small-intestinal submucosa (SIS), have been successfully used clinically as tissue substitutes outside the vascular arena. SUMMARY BACKGROUND DATA: In the present study, we compared a small-diameter conduit of SIS to ePTFE in the presence of Gram-positive contamination to evaluate infection resistance, incorporation and remodeling, morphometry, graft patency, and neointimal hyperplasia (NH) development. METHODS: Adult male mongrel pigs were randomized to receive either SIS or ePTFE (3-cm length, 6-mm diameter) and further randomized to 1 of 3 groups: Control (no graft inoculation), Staphylococcus aureus, or mucin-producing S epidermidis (each graft inoculation with 10 colonies/mL). Pressure measurements were obtained proximal and distal to the graft to create the iliac/aorta pressure ratio. Morphometric analysis of the neointima and histopathologic examinations was performed. Other outcomes included weekly WBC counts, graft incorporation, and quantitative culture of explanted grafts. RESULTS: Eighteen animals were randomized. All grafts were patent throughout the 6-week study period. Infected SIS grafts had less NH and little change in their iliac/aorta indices compared with infected ePTFE grafts. Quantitative cultures at euthanasia demonstrated no growth in either SIS group compared with 1.7 x 10(4) colonies for ePTFE S aureus and 6 x 10(2) for ePTFE S epi (each P < 0.001). All SIS grafts were incorporated. Histology demonstrated remodeling into host artery with smooth muscle and capillary ingrowth in all SIS groups. Scanning electron micrography illustrated smooth and complete endothelialization of all SIS grafts. CONCLUSIONS: Compared with ePTFE, SIS induces host tissue remodeling, exhibits a decreased neointimal response to infection, and is resistant to bacterial colonization. SIS may provide a superior alternative to ePTFE as a vascular conduit for peripheral vascular surgery.     

Laparoscopic ureteral reconstruction with small intestinal submucosa

PURPOSE: To evaluate the feasibility of laparoscopic ureteral reconstruction with small intestinal submucosa (SIS) in the pig ureter. MATERIALS AND METHODS: Eight female pigs weighing between 25 and 30 kg were enrolled. After anesthesia was administered, a double-pigtail stent was inserted, the animals were moved to a lateral decubitus position, pneumoperitoneum was established, and three 10-mm ports were positioned. The ureter was opened longitudinally for 7 cm, and two thirds of the periphery of the upper third of the left ureter was excised. The SIS was anastomosed to the upper and distal ureteral segments with chromic 4-0 sutures. The double-pigtail stent was removed 6 weeks after the initial procedure, and retrograde pyelography was performed a week later to confirm the viability of the pelvicaliceal system. RESULTS: The average duration of the procedures was 210 minutes (range 125-250 minutes). All animals survived the entire follow-up period of 7 weeks. Retrograde pyelography revealed a patent ureteral lumen, and no obstructive phenomena were observed. Histologically, the SIS-regenerated ureteral segments were remarkably similar to normal porcine ureters and were indistinguishable from neighboring tissue. CONCLUSION: Laparoscopic ureteral reconstruction with SIS proved to be effective and technically feasible. The SIS seems to be an effective biodegradable scaffold, facilitating regeneration of host tissue.     

PVP-iodine in hydrosomes and hydrogel--a novel concept in wound therapy leads to enhanced epithelialization and reduced loss of skin grafts

BACKGROUND: Moist wound treatment improves healing at a possibly increased risk of bacterial infection and many local antiseptics impair healing. A moist treatment modality with efficient antimicrobial activity would be desirable. METHODS: In this monocentric, randomized, observer blinded, phase III study, a new hydrosome polyvinyl-pyrrolidone (PVP)-iodine preparation in hydrogel containing iodine in a 3% concentration (Repithel) was investigated for its effect on epithelialization in patients receiving meshed skin grafts. Grafts of 167 patients (donor site defects, burn wounds, or chronic defects) were dressed either with Repithel (n=83) covered with a gauze (Jelonet), or Jelonet-gauze only (n=84) until healing. RESULTS: Grafts receiving Repithel healed significantly earlier (9.4 days versus 12.4 days; p<0.0001) and faster than controls as measured by neo-epithelialization of mesh holes between days 7 and 11 (91.2+/-22.8% versus 82.3%+/-28.6, p<0.0001). A subgroup analysis showed that the effects on grafted burn wounds (p=0.0042) and chronic defects (p<0.0001) was more significant than on donor sites. Also a higher take rate of grafts (p=0.0053) and a reduced loss of grafts was observed with Repithel treatment (8 grafts versus 20 grafts) (p=0.0063, respectively). Smokers had improved graft take (p=0.0069) and higher rate of epithelialization (p=0.0040) compared to smokers of the control group. CONCLUSIONS: The results demonstrate significant clinical advantages of Repithel. This new local wound healing drug combines antisepsis and wound moisture efficiently resulting in significantly enhanced epithelialization, decreased transplant losses, and significantly improved healing especially in smokers.     

Rabbit urethra replacement with a defined biomatrix or small intestinal submucosa

OBJECTIVE: The evaluation of collagen-based biomatrix (SIS COOK((R))) in comparison to a biochemically reconstructed biomatrix for replacement of the urethra in a rabbit model as a preclinical model. MATERIAL AND METHODS: Rabbits underwent partial urethra replacement (resection of 0.5 to 1.0 cm segment of the urethra), which was replaced with 1 or 4 layers Small Intestinal Submucosa (SIS COOK) patch grafts or with a biochemically defined collagen biomatrix, partly sutured with unresolvable sutures for future reference. Six animals underwent a sham control operation. The grafts of regenerated urethras were harvested at 1, 3 and 9 months after implantation. Urethrography was performed pre-operatively and before sacrificing. The animals were evaluated macroscopically and by routine histology and immunohistochemistry. RESULTS: At 1 month after implantation, the biomatrices (1 layer, 4 layers and our biochemically defined biomatrix) were well distinguishable from the normal surrounding tissues and showed blood vessels at the periphery. Macroscopically, the unresolvable reference sutures were easy to find at all time points. At 3 months the graft was still distinguishable in the 4 layers SIS group. In the 1 layer and the defined biomatrix group a good regeneration of the urethra within the graft was seen with some central fibrosis. Histological and immunohistochemical evaluation showed urothelium regeneration on the 1 layer and on biochemically defined biomatrix with decreasing number of inflammatory cells from 1 month on. In the group treated with 4 layers SIS the urothelium was completely regenerated at 3 months. Histologically, the regeneration of muscle cells in the three biomatrices was comparable. The smooth muscle cells regenerated very slowly as 1 month after implantation no muscle cells were detectable within the grafts. At 3 months a few muscle cells were present in the graft, but cell density did not increase in the following 6 months. Strictures were not observed on control urethrography pre-operatively in the animals. In one case slight narrowing of the urethra on urethrography was seen, but apparently without causing voiding problems. One rabbit developed a fistula near the operation site. CONCLUSION: The biomatrices investigated are feasible scaffolds to repair urethral lesions. The results with our biochemically defined biomatrix are comparable to one layer Small Intestinal Submucosa. Almost no smooth muscle cells population was observed after nine months for the three biomatrices. We conclude that an improved molecularly defined biomatrix focussed on stimulation of smooth muscle cell growth may be necessary to obtain optimal cellular grafting results.     

Morphologic evaluation of regenerated small bowel by small intestinal submucosa

Background/Purpose

Previous studies have shown small intestinal submucosa (SIS) can be used as biodegradable scaffolds in tissue engineering small intestine. The purpose of this study is to evaluate the regeneration of neointestine and its morphology using SIS.

Methods

A 2-cm tubular SIS graft from Sprague Dawley rat donors was interposed in the middle of a 6-cm ileal Thiry-Vella loop of Lewis rats, which was used to construct an ileostomy. The grafts were harvested at each of the time points ranging from 2 weeks to half a year after implantation, and native small intestine and grafts were investigated for morphology using histology and immunohistochemistry.

Results

At the early postoperative period, SIS grafts were colonized by numerous inflammatory cells. A mucosal epithelial layer began to line the luminal surface of the graft by 4 weeks, and by 12 weeks, the luminal surface was covered completely by a layer of neomucosa. Neomucosa with typical small bowel morphology was characterized by a columnar epithelial cell layer with goblet cells, Paneth cells, absorptive enterocytes, and enteroendocrine cells. Significant differences between neomucosa by 12 weeks and 24 weeks in the measurements of mucosal thickness, villus height, and crypt depth were found. The outer walls of SIS grafts were composed of distinct bundles of well-formed smooth muscle-like cells with some fibrovascular tissue.

Conclusions

This initial study suggests that tissue engineering neointestine using SIS can develop structural features of the normal intestine. Small intestinal submucosa might be a viable material in the creation of neointestine for patients suffering short bowel syndrome.     

Comparison of the resistance to infection of intestinal submucosa arterial autografts versus polytetrafluoroethylene arterial prostheses in a dog model

Purpose: Prosthetic graft infection represents a most challenging complication to the vascular surgeon. Although expanded polytetrafluoroethylene (ePTFE) grafts have an acceptable patency rate, especially in the large-diameter arterial location, bacterial contamination of this material usually requires surgical removal of the graft.Methods: We compared the resistance of large-diameter ePTFE grafts and grafts constructed of small intestinal submucosa (SIS) to deliberate infection with Staphylococcus aureus. Eighteen dogs were divided into two equal groups, and the infrarenal aorta was replaced with either ePTFE or SIS graft material. One hundred million S. aureus organisms were deposited directly on the graft at the time of surgery, and the dogs were observed for 30 days.Results: One dog with an ePTFE graft died of hemorrhage from an anastomosis site at 21 days. Of the remaining eight dogs with ePTFE grafts, four had positive culture results from the removed graft material, and all had histologic evidence for persistent infection. These dogs also had chronic fever, and the average white blood cell count at day 30 was 15,600/mm³. All nine dogs with SIS grafts had patent grafts, were afebrile after the first week, had an average white blood cell count of 11,500/mm³ at 30 days (p value = NS), had negative culture results, and had the histologic appearance of graft remodeling with collagen that was free of active inflammation.Conclusions: We conclude that large-diameter arterial SIS grafts are more resistant to persistent infection with S. aureus than ePTFE grafts in this dog model of deliberate bacterial inoculation. (J VASC SURG 1994;19:465-72.)     

Morphologic changes in the canine urinary tract after ureteral reconstruction with the tunica albuginea (experimental study)*

Objectives: The problem of ureteral reconstruction has not been fully solved, despite dramatic developments in surgical methods. Many attempts to repair a ureteral damage with autologous or homologous organ grafts, or with organic substitutes have not really been satisfactory. Also, attempts to use alloplastic and heteroplastic prostheses made of synthetic materials have been equally disappointing. Experimental studies, particularly in dogs, indicate great regenerative abilities of ureters when appropriate surgical techniques are used. Materials and methods: In our experimental study, suitability of the tunica albuginea for reconstruction of the middle part of the ureter was investigated in 20 dogs. The dogs were divided into two groups according to a type of ureteral damage: partial (a patch graft) or complete (a tube graft). The ureteral damage was 2–3 cm long in 10 dogs and 3–3.5 cm long in the subsequent 10 dogs. Each dog underwent two operations. The first operation involved a fenestrating incision in the ureter, 23.5 cm long, so as to preserve the ureteral continuity, and then the produced injury was repaired with a patch of the tunica albuginea, with the 4 Charr catheter being inserted in the ureter. The second operation involved a complete excision of ureteral segments, 2–3.5 cm long, on the contralateral side. The excised length of the ureter was substituted with a tube graft of the tunica albuginea, also using the 4 Charr catheter. Thus, in 21 cases, a partial damage of the ureteral wall was repaired with a patch graft of the tunica albuginea. In 19 cases, a complete damage of the ureter was repaired with a tube graft of the tunica albuginea. Results: Four dogs died after the tube-graft implantation due to urinary infiltration and/or purulent inflammatory changes in the kidneys. Autopsies were performed in all the dogs after they had been put to sleep. Definitely poor results were obtained in case of the complete reconstruction of the ureteral loss with a tube graft of the tunica albuginea, 3–3.5 cm long. Within the graft area, we observed inflammatory infiltrates of varied intensity, fibrosis, cicatricial strictures or ureteral obstruction. Occasionally, necrotic foci, degenerative changes or hydronephrosis were found. In case of the reconstruction with a shorter tube graft, i.e. 2–3 cm long, similar changes were observed, however, they were less intense. Satisfactory results were obtained following the partial reconstruction of the ureteral loss with a patch graft of the tunica albuginea. In this case, we observed generally normal regeneration not only of the transitional epithelium but the muscular layer and ureteral structure as well, with full patency of the ureter preserved. The findings obtained in our study are compatible with the findings of other authors. Conclusions: (1) The success of reconstructive procedures in the ureters can be achieved if the continuity of the ureter is preserved and the catheter is removed after 14 days. (2) The substitution of a complete ureteral loss with a tube-graft of the tunica albuginea results in necrosis of the graft and restenosis. *Printed in part at the postdoctoral thesis of Tyloch Ferdynand MD: “An experimental study on the use of tunica albuginea for supplying at westage of ureters” (Badania doswiadczalne nad przydatnoscia blony bialawej jadra w operacjach plastycznych moczowodu) University School of Medical Sciences Bydgoszcz 1994.