Female Yorkshire Pigs Research Articles

Methane production of growing and finishing pigs in southern China

Methane from enteric fermentation and manure decomposition from livestock contributes to greenhouse gas (GHG) emissions. Research on CH4 production from enteric fermentation has mainly focused on ruminant livestock. However, CH4 production from enteric fermentation in pigs may be important due to the large global pig population. This experiment measured enteric CH4 from growing and finishing pigs in China. Five 58.9±1.15kg and five 89.4±0.85kg Duroc×Landrace×Large Yorkshire female pigs were randomly selected from a commercial farm in Shenzhen, Guangdong province (China). The pigs were transferred to individual 2.0m×1.0m×1.2m respiration chambers located on the same farm and continued to be fed the commercial diets of their respective weight group on the farm. After 3 d of adaptation to the respiration chambers, daily CH4 emissions were measured from the 10 pigs for 3 d. The CH4 production of the 60 and 90kg pigs was 1.13 and 2.01g pig/d, respectively, with that of 90kg pigs higher than the 60kg group (P<0.05). When the values are adjusted/kg DM feed intake (i.e., 0.70g/kg DM versus 0.82g/kg DM) and per kg body weight (19.01g/kg versus 22.47g/kg) for the 60 and 90kg groups, there were no differences. Using these values, estimated annual CH4 production from the 60 and 90kg pigs was 0.41kg and 0.73kg, respectively, values which are lower than the 1kg pig/year adopted by the Intergovernmental Panel on Climate Change. The high variation in CH4 emission among our pigs, and those in the literature, suggest that differences in CH4 emission exist among pigs within the same herd housed under similar nutritional and husbandry management.This article is part of the special issue entitled: Greenhouse Gases in Animal Agriculture – Finding a Balance between Food and Emissions, Guest Edited by T.A. McAllister, Section Guest Editors; K.A. Beauchemin, X. Hao, S. McGinn and Editor for Animal Feed Science and Technology, P.H. Robinson.

Abstract No. 21: Major motor nerve injury following irreversible electroporation (IRE) in a porcine model

To determine whether irreversible electroporation (IRE) may be performed safely in close proximity to the sciatic nerve in a porcine model. Single session IRE treatment adjacent to the normal sciatic nerve at the sciatic notch was performed unilaterally in 4 female Yorkshire pigs under general anesthesia and using pancuronium as a paralytic agent. Two to four 19-gauge monopolar electrodes were placed in close proximity to the sciatic nerve under CT guidance. IRE was performed using 1500–1800 V/cm. One of the animals was sacrificed acutely. The other 3 pigs were observed for up to 7 days before they were euthanized for pathologic correlation. The pigs were administered analgesics and encouraged to ambulate in the observation period. Contrast-enhanced CT was performed immediately following IRE, and contrast-enhanced CT or MRI was performed on the day of sacrifice. Gross and microscopic histology of the ablated tissue were evaluated. The closest electrode was placed within 0–5mm of the sciatic nerve in each animal. Maximal dimension of the ablation zone on immediate post-ablation contrast-enhanced CT measured 2.5 cm with 2 electrodes (n=1) and 6.9–7.2 cm (n=3, mean 7.0 cm) with 4 electrodes. Size of the ablation zone decreased to 3.6–6.8 cm (mean 5.3cm) on contrast-enhanced MRI within 2–7 days. Two of the 3 non-acute pigs were unable to ambulate over an observation period of 2–7 days, including one pig euthanized at day 2 post-IRE due to immobility. Histology confirmed injury to the sciatic nerve in the acute and 2 immobile pigs. IRE performed in one animal at 5mm from the sciatic nerve did not result in nerve injury or immobility; in this case, the zone of non-enhancement on post-ablation CT did not extend beyond the nerve. Major motor nerves less than 5 mm from the IRE electrode may be injured and become nonfunctional. Further investigation is warranted to determine the safety parameters of IRE in perineural tissue.

Gastrotomy closure with a new tissue anchoring device: A porcine survival study

To evaluate the feasibility, reproducibility and efficacy of a new tissue anchoring device in a porcine survival model. Gastrotomies were performed using a needle-knife and balloon dilator in 10 female Yorkshire pigs weighing 30-35 kg. Gastric closure was attempted using a new tissue anchoring device. The tightness of the closure was confirmed by means of air insufflation and the ability to maintain gastric distension with stability in peritoneal pressure measured with a Veress needle. All animals were monitored daily for signs of peritonitis and sepsis over 14 d. During necropsy, the peritoneal cavity and the gastric access site were examined. Transgastric access, closure and 14 d survival was achieved in all pigs. The mean closure time was 18.1 ± 19.2 min and a mean of 2.1 ± 1 devices were used. Supplementary clips were necessary in 2 cases. The closure time was progressively reduced (24.8 ± 13.9 min in the first 5 pigs vs 11.4 ± 5.9 min in the last 5, P = NS). At necropsy, the gastric access site was correctly closed in all cases with all brace-bars present. One device was misplaced in the mesocolon. Minimal adhesions were observed in 3 pigs and signs of mild peritonitis and adhesions in one. The use of this new tissue anchoring device in porcine stomachs is feasible, reproducible and effective and requires a short learning curve.

Antithrombin-heparin covalent complex reduces microemboli during cardiopulmonary bypass in a pig model

AbstractTranscranial Doppler-detected high-intensity transient signals (HITS) during cardiopulmonary bypass (CPB) surgery have been associated with postoperative neurocognitive dysfunction, suggesting microemboli in the brain could be a contributing factor. HITS occur despite administration of unfractionated heparin (UFH). This study was done to determine whether antithrombin-heparin covalent complex (ATH), a more potent anticoagulant than heparin, can reduce HITS during CPB. In a pig CPB model, ATH, UFH, or UFH + antithrombin (AT) was intravenously administered to female Yorkshire pigs after sternotomy. Twenty minutes later, hypothermic CPB was initiated and continued for 1.25 hours, then normothermia was re-established for 45 minutes. Protamine sulfate was given to neutralize the anticoagulants, and pigs were allowed to recover. HITS were monitored using an arterial flow probe placed over the carotid artery. Compared with UFH (300 or 1000 U/kg), ATH reduced the number of HITS during CPB in a dose-dependent manner. AT (3 mg/kg) + UFH (300 U/kg) resulted in an intermediate HITS rate between UFH and ATH (2 mg/kg in terms of AT). Examination of brain sections for emboli formation confirmed that, similar to HITS, number of thrombi decreased in direct proportion to ATH dosage. These results support the hypotheses that the majority of HITS represent thromboemboli and that ATH reduces emboli formation during CPB.

Irreversible electroporation (IRE): a novel method for renal tissue ablation

• To evaluate the effects of irreversible electroporation (IRE) on renal parenchyma and the renal collecting system in a porcine model. • Eight female Yorkshire pigs underwent a series of laparoscopic ablations using either monopolar or bipolar IRE (Angiodynamics, Queensbury, NY, USA). • The pigs were killed between 10 min and 14 days after IRE, and the kidneys were harvested for gross and histological analysis, including NADH staining for cellular viability. • In all, 24 ablations were performed and all the pigs survived without complications. • Initial gross lesions were diffusely haemorrhagic, decreasing progressively in size (30-40%) to small white scars over the 14-day period. • Immediately after IRE, ablated tissue was characterized by diffuse tubular desquamation, eosinophilia, and nuclear pyknosis, with absence of cellular viability by NADH. • At 7 days after IRE, there was diffuse cellular necrosis with early peripheral granulation changes, and by 14 days there was marked tissue granulation, chronic inflammation, and dystrophic calcification with early fibrosis and cellular contraction. • Initial patchy urothelial injury and ulceration showed signs of repair and viability by 14 days after IRE. • Renal IRE in the porcine kidney leads to predictable histological changes characteristic of cellular death within 1 h of ablation, with relative urothelial sparing. • Further animal studies are warranted to determine safety and efficacy of this novel ablation technology.

Yes, we can: reliable colonic closure with the Padlock-G clip in a survival porcine study (with video)

Secure closure of the colonic access site is one of the most important issues for the development of natural orifice transluminal endoscopic surgery. To evaluate the feasibility, reproducibility, and efficacy of a new over-the-scope clip. Descriptive study, in vivo porcine colon. In 10 female Yorkshire pigs weighing 30 to 35 kg, a 10-mm colotomy was performed by using a needle-knife and advancing the endoscope to the peritoneal cavity. Colonic closure was performed by using the Padlock-G clip (Aponos Medical, Kingston, NH) delivered with the Lock-It system (Aponos). Animals were monitored daily for signs of peritonitis and sepsis over a period of 14 days. During necropsy, the peritoneal cavity was examined, and the colon segment containing the incision was excised for pathological study. Closure was achieved in all cases. Nine of 10 pigs survived 14 days without complications, but 1 pig was killed immediately after the procedure because of severe bleeding during the colonic incision. The median closure time was 8 minutes (range 1-30 minutes). At necropsy, adhesions were observed in 5 cases. The incision was hardly visible at the serosa side in 3 cases and was not identified in 6 cases. In 6 cases, the clip was still slightly attached to the mucosa, and it was not found in 3 cases. The pathological study revealed a complete remodeling of the colonic wall in all cases. Animal model, noncomparative study. The Padlock-G clip procedure is feasible, reproducible, effective, and easy to perform, and it provides a reliable colonic closure.

Delay of 2 or 6 Weeks Adversely Affects the Functional Outcome of Augmented Primary Repair of the Porcine Anterior Cruciate Ligament

Background Enhanced primary anterior cruciate ligament repair, in which suture repair is performed in conjunction with a collagen-platelet composite to stimulate healing, is a potential new treatment option for anterior cruciate ligament injuries. Previous studies have evaluated this approach at the time of anterior cruciate ligament disruption. Hypothesis Delaying surgery by 2 or 6 weeks would have a significant effect on the functional outcome of the repair. Study Design Controlled laboratory study. Methods Sixteen female Yorkshire pigs underwent staged, bilateral surgical anterior cruciate ligament transections. Anterior cruciate ligament transection was initially performed on 1 knee and the knee closed. Two or 6 weeks later, enhanced primary repair was performed in that knee while the contralateral knee had an anterior cruciate ligament transection and immediate repair. Biomechanical parameters were measured after 15 weeks in vivo to determine the effect of delay time relative to immediate repair on the healing response. Results Yield load of the repairs at 15 weeks was decreased by 40% and 60% in the groups where repair was delayed for 2 and 6 weeks, respectively (P = .01). Maximum load showed similar results (55% and 60% decrease in the 2- and 6-week delay groups, respectively; P = .011). Linear stiffness also was adversely affected by delay (50% decrease compared with immediate repair after either a 2- or 6-week delay, P = .011). Anterior-posterior laxity after 15 weeks of healing was 40% higher in knees repaired after a 2-week delay and 10% higher in those repaired after a 6-week delay (P = .012) when tested at 30° of flexion, but was not significantly affected by delay when tested at 60° or 90° (P = .21). Conclusion A delay between anterior cruciate ligament injury and enhanced primary repair has a significant negative effect on the functional performance of the repair. Clinical Relevance As future investigations assess new techniques of anterior cruciate ligament repair, the timing of the repair should be considered in the design and the interpretation of experimental studies.

Bone Marrow Mesenchymal Stem Cells Stimulate Cardiac Stem Cell Proliferation and Differentiation

The regenerative potential of the heart is insufficient to fully restore functioning myocardium after injury, motivating the quest for a cell-based replacement strategy. Bone marrow-derived mesenchymal stem cells (MSCs) have the capacity for cardiac repair that appears to exceed their capacity for differentiation into cardiac myocytes. Here, we test the hypothesis that bone marrow derived MSCs stimulate the proliferation and differentiation of endogenous cardiac stem cells (CSCs) as part of their regenerative repertoire. Female Yorkshire pigs (n=31) underwent experimental myocardial infarction (MI), and 3 days later, received transendocardial injections of allogeneic male bone marrow-derived MSCs, MSC concentrated conditioned medium (CCM), or placebo (Plasmalyte). A no-injection control group was also studied. MSCs engrafted and differentiated into cardiomyocytes and vascular structures. In addition, endogenous c-kit(+) CSCs increased 20-fold in MSC-treated animals versus controls (P<0.001), there was a 6-fold increase in GATA-4(+) CSCs in MSC versus control (P<0.001), and mitotic myocytes increased 4-fold (P=0.005). Porcine endomyocardial biopsies were harvested and plated as organotypic cultures in the presence or absence of MSC feeder layers. In vitro, MSCs stimulated c-kit(+) CSCs proliferation into enriched populations of adult cardioblasts that expressed Nkx2-5 and troponin I. MSCs stimulate host CSCs, a new mechanism of action underlying successful cell-based therapeutics.

Minced Urothelium to Create Epithelialized Subcutaneous Conduits

We used in vivo cell expansion to create 3-dimensional subcutaneous conduits lined with an inner layer of autologous urothelial mucosa. Laparotomy and excision of a fifth of the bladder were done in 5 female Yorkshire pigs (Parsons Farm, Westhampton, Massachusetts) under general anesthesia. After mechanical removal of the detrusor muscle the bladder mucosa was minced to obtain 0.2 x 0.8 x 0.8 mm particles, which were attached to the outer surface of latex tubes using a thin layer of fibrin glue. Seven to 10 tubes were placed in the abdominal wall subcutaneous tissue in each original donor pig with tubes lacking particles serving as controls. Biopsy was done 1 to 4 weeks after transplantation for histological evaluation. One week after transplantation particles were still present in the granulation tissue. At 2 weeks the epithelium was differentiated with transitional uroepithelium facing the lumen, ie toward the tube. No epithelium was detected around control tubes. After autologous transplantation of bladder mucosal particles organized in 3-dimensional fashion in pig subcutaneous tissue the transplanted cells proliferated, migrated and reorganized to form a continuous epithelial lining facing the lumen. This novel approach to urothelial transplantation may allow successful formation of a conduit to the bladder or of a neourethra.

Pre-clinical evaluation of liposomal gene transfer to improve dermal and epidermal regeneration

Liposomal gene transfer effectively enhances dermal and epidermal regeneration in burned rodents. To advance this treatment to clinical studies, we investigated the efficacy of liposomal gene transfer in a clinically relevant porcine wound model. Mimicking the clinical scenario, six female Yorkshire pigs (40-50 kg) received up to 12 burns of 50 cm(2) area that were fully excised and covered with skin autograft meshed at 4:1 ratio 24 h post-burn. Animals received control injections (empty liposomes), liposomes (DMRIE-C) containing 1 mg LacZ-cDNA, or liposomes (DMRIE-C) with 1 mg of platelet-derived growth factor (PDGF)-cDNA, or the naked PDGF gene. Serial biopsies were taken from different wound sites at multiple time points up to 12 days post-wounding. Transfection efficacy and transfection rate of LacZ and localization of beta-gal were determined by immunohistochemical and immunofluorescent techniques. RT-PCR and multiplex protein analysis (ELISA) were used to measure levels of growth factor mRNA transcribed and growth factor protein translated. Wound re-epithelialization and graft adhesion was evaluated using planimetric analysis and clinical scores. We found that peak transfection of liposomal beta-galactosidase occurred on day 2, with a fluorescence increase of 154% to baseline (P<0.001). Transfection intensity dropped to 115% above baseline on day 4 (P<0.001) and 109% on day 7. Immunohistochemistry showed a maximum transfection rate of 34% of cells in wound tissue. Gene transfer of liposomal PDGF-cDNA resulted in increased PDGF-mRNA and protein expression on days 2 and 4, and accelerated wound re-epithlialization as well as graft adhesion on day 9 (P<0.05). In this study, we showed that liposomal cDNA gene transfer is possible in a porcine wound model, and by using PDGF-cDNA we further showed that dermal and epidermal regeneration can be improved. These data indicate that liposomal gene transfer can be a new therapeutic approach to improve wound healing in humans.

Real-time intra-operative near-infrared fluorescence identification of the extrahepatic bile ducts using clinically available contrast agents

Iatrogenic bile duct injuries are serious complications with patient morbidity. We hypothesized that the invisible near-infrared (NIR) fluorescence properties of methylene blue (MB) and indocyanine green (ICG) could be exploited for real-time, intraoperative imaging of the extrahepatic bile ducts during open and laparoscopic surgeries. In all, 2.0 mg/kg of MB and 0.05 mg/kg of ICG were injected intravenously into 35-kg female Yorkshire pigs and the extrahepatic bile ducts were imaged over time using either the Fluorescence-Assisted Resection and Exploration (FLARE) image-guided surgery system (open surgery) or a custom NIR fluorescence laparoscopy system. Surgical anatomy was confirmed using x-ray cholangiography. The contrast-to-background ratio (CBR), contrast-to-liver ratio (CLR), and chemical concentrations in the cystic duct (CD) and common bile duct (CBD) were measured, and the performance of each agent was quantified. Using NIR fluorescence of MB, the CD and CBD could be identified with good sensitivity (CBR and CLR > or =4), during both open and laparoscopic surgeries, from 10 to 120 min postinjection. Functional impairment of the ducts, including constriction and injury were immediately identifiable. Using NIR fluorescence of ICG, extrahepatic bile ducts did not become visible until 90 min postinjection because of strong residual liver retention; however, between 90 and 240 min, ICG provided exquisitely high sensitivity for both CD and CBD, with CBR > or =8 and CLR > or =4. We demonstrate that 2 clinically available NIR fluorophores, MB fluorescing at 700 nm and ICG fluorescing at 800 nm, provide sensitive, prolonged identification of the extrahepatic bile ducts and assessment of their functional status.

Hemodynamic Changes and Fluid Shifts After Large-Volume Fluid Infiltration

While certain parameters such as blood loss and serum lidocaine levels following liposuction have been well studied, fluid shifts between the intravascular and extravascular space have not. With the advent of large volume liposuction, prudent fluid management has become obligatory. Hence, the reason for our study.To test the impact of large volume infiltration on intercompartmental fluid shifts, we measured urine output and hemodynamic changes in 10 anesthetized female Yorkshire pigs weighing between 50 and 85 kg. Eight pigs were infused with 5 to 10 L of tumescent fluid. Two pigs were anesthetized, received no wetting solution, and served as controls. Hemodynamic variables were recorded before infusion and hourly for 48 hours. Animals were extubated after 4 hours of anesthesia. Plasma volume was measured using Evan's Blue Dye, and intravascular fluid shifts were calculated using Foldager's method.Total fluid shift into the intravascular space ranged between 511 and 1036 mL per animal with a mean of 767 mL in the first 3 hours. Higher volumes of fluid infiltration did not lead to fluid overload in the experimental group. Hemodynamic changes were characterized by significant increases in central venous pressure, cardiac output, pulmonary artery pressure, and heart rate consistent with the increase in intravascular volume. Hemodynamic parameters returned to baseline 20 hours following tumescent fluid infiltration.In this porcine model, animals were able to tolerate large fluid challenges delivered by clysis with statistically significant but only modest increases in hemodynamic parameters which gradually returned to baseline within 20 hours.

NOTES TRANSGASTRIC PARTIAL CYSTECTOMY

Background & Aims: The evolution of flexible endoscopes and endoscopic devices has recently enabled per-oral transgastric abdominal exploration in animal models. This study was undertaken to assess the ability to systematically identify abdominopelvic organs and to determine the feasibility of organ resection via the transgastric route. Methods: Female Yorkshire pigs were used for the study. Under general anesthesia, a gastroscope was used to place a sterile overtube into the esophagus. After antibacterial lavage, the gastric wall was incised, and a sterile dual-channel endoscope was advanced into the peritoneal cavity. Endoscopic abdominal exploration was then methodically performed in 9 animals with oophorectomy and partial hysterectomy in 6 animals. The gastric incision was closed with endoclips. Of the animals that underwent organ resection, 3 were sacrificed immediately after surgery, and the subsequent 3 survived. Results: The peritoneal cavity was accessed uneventfully, and the stomach, liver, small bowel, colon, urinary bladder, uterus, fallopian tubes, and ovaries were able to be evaluated in all cases. The gallbladder could be visualized in only 5 of the 9 animals, and retroperitoneal structures could not be consistently identified. Oophorectomy with partial hysterectomy was performed in 6 animals. The 3 animals in the survival group did well for 24 hours without sequelae. Conclusions: This study shows the ability of endoscopic transgastric peritoneal exploration to successfully identify most abdominopelvic organs and demonstrates the technical feasibility of transgastric organ resection.

Postnatal ontogeny of intestinal GCPII and the RFC in pig

In humans and pigs, hydrolysis of dietary polyglutamyl folates is carried out by intestinal brush border folate hydrolase [glutamate carboxypeptidase II (GCPII)], whereas the transport of the monoglutamyl folate derivatives occurs via the intestinal brush border reduced folate carrier (RFC). The study objective was to measure the expression of intestinal GCPII and RFC during postnatal development of pigs and their effects on plasma and liver folate concentrations. Duodenum, jejunum, ileum, liver, and plasma samples were collected from female Yorkshire pigs at birth, 24 h, 1 wk, 3 wk, and 6 mo (n=6 at each time point). GCPII mRNA transcripts and protein (normalized using beta-actin), and enzyme activity (normalized per mg mucosal protein) were highest in all segments of small intestine at birth and were undetectable in ileum after 1 wk, whereas jejunal protein and activity predominated at 6 mo. RFC mRNA transcripts were present in all segments of small intestine at birth and declined significantly throughout development to 6 mo. Conversely, RFC protein increased twofold during the first 24 h and remained constant throughout development in all segments of small intestine. Liver RFC mRNA transcripts were detected at birth but were reduced by 6 mo. Liver folate concentration increased throughout postnatal development, whereas plasma folate levels increased during the first 24 h but decreased over time, reflecting the pattern of RFC expression in small intestine. These findings show that intestinal GCPII and intestinal and hepatic RFC all exhibit ontogenic changes in the pig that are reflected in postnatal folate status.

The development of a novel intracolonic occlusion balloon for transcolonic natural orifice transluminal endoscopic surgery: description of the technique and early experience in a porcine model (with videos)

Transgastric and transvaginal approaches in natural orifice transluminal endoscopic surgery (NOTES) are the most commonly used, although the transcolonic approach may have some advantages. To develop a workable technique for transcolonic NOTES. A nonsurvival study followed by a survival study in a porcine model. Transcolonic peritoneoscopy was performed with the use of a novel intracolonic occlusion balloon. The colotomy was closed with endoclips. A necropsy and histologic evaluation were performed 2 weeks after surgery. Academic hospital, health science research center. Fifteen female Yorkshire pigs (5 nonsurvival, 10 survival). A balloon-tipped catheter was placed proximal to the colotomy site. The balloon was inflated to occlude the colonic lumen. An endoscope was inserted through the anus. Colonic incision was created with an endoscopic needle-knife at 15 to 20 cm from the anal verge. Peritoneoscopy was performed. The colotomy was closed with endoclips. Rates of complications, survival, healing, and adhesions. Two initial experiments were complicated by bowel distension and contamination of the incision area by colonic content. In the remaining 13 pigs, the experiments were performed with the use of the intracolonic occlusion balloon. No complications were documented. Necropsies were performed 2 weeks after surgery. Gross and histologic evaluations demonstrated near complete healing. Minimal adhesions were identified in 4 of 10 pigs. Imperfection of the prototype balloon. Excessive bowel distension and fecal contamination because of spillage from the proximal bowel may be barriers to performing transcolonic NOTES. Isolation of the operative area by splitting the bowel and sealing the colonic lumen with the balloon above the colonic incision may overcome these problems and optimize the technique.

Post‐conditioning reduces infarct size in an open‐chest porcine acute ischemia–reperfusion model

Timely reperfusion is a prerequisite for myocardial salvage; however, re-oxygenation of the ischemic myocardium initiates reperfusion injury. Post-conditioning diminishes the detrimental aftermath of an acute myocardial infarction through alleviation of reperfusion injury. Ischemic post-conditioning consists of a series of brief interruptions in the coronary blood supply that has to be applied within the first minutes after re-establishing the coronary flow. Sixteen female mixed Danish Landrace and Yorkshire pigs weighing 20 kg were included. The heart was exposed through a midline sternotomy. A snare was positioned around the left anterior descending coronary artery downstream of the second diagonal branch. After randomization to either no treatment (control group) or treatment by ischemic post-conditioning (post-conditioning group), the pigs underwent 45 min of ischemia and 180 min of reperfusion. The post-conditioning group had a post-conditioning algorithm applied consisting of 15 s of reperfusion alternating with 15 s of re-occlusion repeated 10 times. The groups were comparable with regard to body weight, hemodynamics and the size of the area at risk. The post-conditioning group had an absolute reduction in infarct size of 18.1% [confidence interval (CI): 6.2: 30.0%] compared with the control group (P=0.0056). In the post-conditioning group, infarction developed in 39.6+/-12.0% (1 SD) of the area at risk compared with 57.8+/-10.2% (1 SD) in the control group. When ischemic post-conditioning was applied at reperfusion, we found an absolute reduction in infarct size of 18.1% presumably attributable to a diminished reperfusion injury. The model we have developed is suitable for further studies of this promising intervention.

025 The Role of Genetics on Wound Contraction in a Porcine Model

Pigs have been accepted as good models for skin wound healing research due to their anatomical similarities to humans. The healing response of excisional wounds in juvenile female Yorkshire (Y) pigs closely resembles normal healing in humans. In contrast, identical wounds in female red Duroc (RD) pigs contracted significantly more, resulting in an 80–90% reduction in wound area. This hypercontraction was associated with marked hyperpigmentation at the wound margins, and some features of the RD healing phenotype, such as increased collagen deposition, are similar to hypertrophic scar formation in humans. Recent investigations have begun to elucidate the genetic transmissibility of the RD healing phenotype. Skin wounds were created on Y × RD F1 female animals (N = 8), and while hyperpigmentation was not observed, the hypercontraction prevalent in the RD persisted in 100% of animals studied. Further, the degree and kinetics of contraction observed in the F1 animals was significantly greater than in the RD. All F1 animals were subsequently bred to a single Yorkshire boar, producing a cohort of 20 female backcross animals. Healing in the backcross animals generally followed the Y phenotype with regard to wound contraction and pigmentation. However, between days 28 and 56, the backcross animals demonstrated significantly less contraction than the Yorkshire animals. These results suggest that the regulation of wound contraction is controlled by a restricted number of major genes and an unidentified number of other genes, which may positively or negatively modulate the influence of the major genes, as observed in the F1 and backcross animals. These studies have begun the process of elucidating the molecular basis for abnormal skin wound healing and scarring. Further ongoing studies are using genomic and proteomic approaches to further elucidate candidate genes potentially involved in the RD healing phenotype. Acknowledgments: Funding provided by CIHR and NSERC

005 The Biomechanical Behavior of Scar Tissue and Uninjured Skin in a Porcine Model

To investigate changes in the mechanical properties of skin with injury, tissue was evaluated 70 days following full thickness wounding in juvenile female Yorkshire pigs (N = 14). Samples were taken in the axial (cranial-caudal) and transverse (dorsal-ventral) directions, for both scar tissue and uninjured skin. The samples were evaluated mechanically in vitro using a protocol of stress relaxation followed by tensile failure, in an INSTRON universal test machine. Specimens were subjected to a 10% elongation and held for a 4-min isometric period, after which they were returned to the original length and failed in tension. Axial normal skin exhibited a smaller force at 10% elongation (−58%, p = 0.002), and higher failure load (p = 0.027) and displacement (p = 0.028) than transverse. Scarring reduced the failure strength on average by 60% in the axial and 65% in the transverse directions (p < 0.001), with large decreases (>50%) in displacement at failure in both directions (p < 0.001). Elongation of 10% produced a larger force in axial scars (p < 0.001) than their normal counterparts, indicating a reduction in small-displacement compliance. Axial scars exhibited greater failure strength (p = 0.009) than transverse scars, yet no significant differences were observed in elongation to failure or load at 10% elongation. These results indicate that normal skin exhibits higher compliance and failure strength in the axial direction. Following injury, the strength and compliance of the tissue are greatly reduced, and the strong compliance directionality originally observed, is lost. Therefore it appears that the healing response leads to increases in tissue stiffness, such that small displacements result in larger tissue loads. This is further complicated by the reduced ability of the scar to resist failure. Taken together, these findings illustrate the compounding negative effects of scar on tissue integrity and function. Acknowledgments: Alberta Ingenuity Fund, CIHR and NSERC

Evaluation of a novel access and closure device for NOTES applications: a transcolonic survival study in the porcine model (with video)

Natural orifice transluminal endoscopic surgery (NOTES) is a novel, potentially less invasive alternative to laparoscopic surgery. However, the problems of transluminal access and closure represent significant obstacles to its successful introduction in humans. Our purpose was to evaluate the feasibility and safety of a novel device designed for transluminal access and closure. Experimental endoscopic study of transcolonic incision and closure with use of a prototype device in a survival porcine model. Four adult female Yorkshire pigs were used in the study. While under general anesthesia, the animals were prepped with multiple tap water enemas followed by instillation of an antibiotic suspension and povidone-iodine lavage. At a distance of 15 to 20 cm from the anus, the prototype device deployed a circumscribing purse-string suture around the planned incision site and subsequently used a blade mechanism to create a 2.5-cm linear incision. The peritoneum was then accessed with a standard double-channel enodoscope. The transcolonic incision was then closed by cinching and securing the purse-string suture with a titanium knot by use of a separate hand-activated suture-locking device. All animals were allowed to eat immediately after recovering from general anesthesia. The animals were monitored daily for signs of peritonitis and sepsis and were electively killed on day 14. The peritoneal cavity was examined for peritonitis, and the colonic incision site was examined for wound dehiscence, pericolic abscess formation, and gross adhesions. Tissue samples from both incisional and random peritoneal sites were obtained for histologic examination. Transcolonic incision and closure were successful in all 4 animals. The device performed in a rapid and reproducible fashion. All animals recovered without septic complications. At necropsy, there was no evidence of peritonitis, abscesses, or wound dehiscence. Salpingocolonic and colovesicular adhesions were noted in 3 of 4 animals. Histologic examination revealed microabscesses at the incision site in all animals. The prototype incision and closure device represents a promising solution to the problems of transluminal access for NOTES. The presence of incision-related adhesions and microabscesses signal the need for further refinement in aseptic technique.

Minced Urothelium for tissue Engineering of Epithelialized Subcutaneous Tunnels

PURPOSE Reconstructive Pediatric Urology is sometimes limited by lack of tissue for the repair. We have previously shown that tissue regeneration and creation of epithelialized tunnels can be created by transplanting minced skin particles around a cylindrical mold. In this study we wanted to create subcutaneous tunnels with an urothelial lining. MATERIAL AND METHODS One fifth of the bladder was removed by open surgery on four 50-60 kg female Yorkshire pigs. The bladder mucosa was mechanically separated from the detrusor muscle. The mucosa was then minced to 0.8x0.3x0.3 mm particles and the pieces were attached in a 1:3 expansion rate to cylidrical molds with fibrin glue. Each mold was then implanted under the abdominal pig skin. Molds without minced bladder mucosa served as controls. Molds and surrounding tissue was extracted 2,3 and 4 weeks after transplantation for microscopical evaluation. RESULTS After removal of the mold cross sections of the excised tissue demonstrated hollow subcutaneous tunnels with a transitional epithelium facing the lumen. Immunoassays for cytokeratin and uroplakins demonstrated its urothelial origin and laminin demonstrated a basal membrane separating the basal cells from the granulation tissue. No epithelial lining was found in controls. CONCLUSIONS Minced particles of bladder mucosa can be transplanted to the subcutaneous fat under the pig abdominal skin. The particles expand in vivo and reorganize to form a continuous epithelium encasing the implanted mold. By removing the mold, uro-epithelialized subcutaneous tunnels were created. These could eventually be connected to the urinary system and used as conduits for catheterization or as a neourethra.