Orthotopic Small Bowel Transplantation Research Articles

Surgical technique for orthotopic small bowel transplantation in a rat

Objectives: Presently, the results of small bowel transplantation (SBT) procedures are far from ideal. Problems such as rejection, sepsis and graft-versus host disease are still encountered after SBT. Here, we describe a refined rat model of SBT for researchers who wish to investigate these complications. Materials and Methods: Sixty male outbred Sprague Dawley (SD) rats were used for the study. Orthotopic small bowel transplantations were performed. An immunosuppressive regimen was not implemented and the recipients were monitored for seven days. If the recipient survived until the experimental end point, the transplantation was deemed to be successful. Recipient survival shorter than seven days was considered to be a technical failure. Results: The mean operation times for donor and recipients were 40 minutes (40 ± 5 min) and 65 minutes (61 ± 10), respectively. Cold and warm ischemia times were 32.5 ± 5 min and 25 ± 7 min, respectively. End-to side venous and arterial anastomosis times were approximately 20 minutes (20 ± 7 min). Four recipients died within the first three days after transplantation due to technical failure [bleeding (n:3), microvascular anastomosis thrombosis ( n :1)]. The rest of the recipients ( n : 26) reached the experimental end point in a healthy condition (success rate was 86%). Conclusion: This model is easy to perform and offers various advantages. The principle advantages of our surgical model are that it minimizes the mechanical injury of the graft and provides a high quality intestinal graft. It also allows us to make rapid and safe anastomosis in the recipient operation.

Microsurgical skills and perioperative management of orthotopic small bowel transplantation in mice

Objective To establish a stable murine models of orthotopic small bowel transplantation. Methods Syngeneic orthotopic small bowel transplantation was performed between C57BL/6 mice. Donor intestinal graft was harvested along with vascular pedicles (including graft's portal vein and the superior mesenteric artery with a Carrel patch of aorta) by microsurgery. The donor intestine was revascularized by end-to-side arterial and venous anastomoses. The native intestine was cut off. The enteric continuity was reestablished with end-to-end intestinal anastomoses. Microsurgical skills and perioperative management were modified through study. Results Orthotopic small bowel transplantation in mice was performed in 70 cases, the success rate of transplantation reached up to 73.5% as technology enters its maturity, although it was only 38.1% in the start-up stage. In maturing stage, the operation time of donor was (46.3±3.8) min, the cold ischemic time of graft was (35.2±4.2) min, the operation time of recipient was (98.0±5.5) min, the vascular anastomotic time was (30.6±3.2) min and the intestinal anastomotic time was (28.6±2.8) min. Conclusion The reliable model of orthotopic small bowel transplantation in mice has been established by modified microsurgical skills and perioperative, which could lay a foundation for further research. Key words: Small bowel transplantation; Orthotopic; Mice; Microsurgical skills; Perioperative management

A CD52 antibody impairs mouse-transplanted intestinal tight junctions

BackgroundInduction immunosuppression strategies using a CD52 monoclonal antibody (mAb) have been introduced for small bowel transplantation, resulting in improved outcomes. However, little information is known about the effects of the antibody on the microstructure of the intestinal barrier, which functions to prevent bacterial translocation. In this study, we used a murine orthotopic small bowel transplantation model to investigate the impact of a CD52 mAb on tight junctions (TJs), which are considered a central part of the intestinal barrier, of the transplanted intestine. Materials and methodsC57BL/6 mice were used as recipients. The grafts were harvested from BALB/c mice in allogeneic groups and C57BL/6 mice in syngeneic groups. The anti-mouse CD52 mAb was applied as a surrogate antibody. Transmission electron microscopy was used to evaluate the TJ ultrastructure. The expression of the TJ proteins occludin and ZO-1 was analyzed by Western blot. The distribution of TJ proteins was observed by immunofluorescence, and the permeability of the transplanted intestine was assessed in vivo using FITC-dextran. ResultsAfter CD52 mAb application, a compromised TJ ultrastructure was observed. In addition, TJ protein expression (occludin and ZO-1) decreased and the intestinal permeability increased. ConclusionsThe anti-mouse CD52 mAb impaired the transplanted intestinal TJ and barrier.

Normothermic Extracorporeal Membrane Oxygenation Support: Improves Donor Intestinal Graft Function After Cardiac Death.

Background:Organs from cortrolled donors after cardiac death (CDCD) are particularly susceptible to the effects of warm ischaemic injury. Perfuison by normothermic extracorporeal membrane oxygenation (NECMO) is increasingly advocated as a useful remedy to the effects of ischaemia-reperfusion injury and is reported to enable transplantation of organs from donors previously deemed unsuitable. To our knowledge, no studies are available for small bowel transplantation using DCD supported by NECMO. In an experimental setting of small bowel transplantation, we studied the feasibility of using intestinal grafts retrieved from DCD supported by NECMO. Method: Thirty White piglets underwent segmental orthotopic small bowel transplantation using FK506 (0.1 mg/kg per day, POD 0-7) immunosuppression and were randomly divided as follow: Living donor (LD) group (n =6) received grafts from HBD; DCD group (n=6) received grafts from DCD; E1 group (n =6) received grafts from DCD supported by NECMO for 1 hour; E3 group (n =6) received grafts from DCD supported by NECMO for 3 hours; E5 group (n =6) received grafts from DCD supported by NECMO for 5 hours. The DCD pigs were sacrificed inducing the cardiac death by a lethal atracurium injection. After 10 minutes (no touch period time), they received NECMO support for 1, 3 or 5 hours. Histopathologic and ultrastructural analysis, mucosal tight junction protein and adenosine triphosphate concentrations were determined immediately after NECMO support. At day 7 after the transplantation, maltose absorption test and histopathologic analysis were performed. FK506 was as immunosuppression. Result: NECMO support can aggravate intestinal mucosa injury and reduce the level of mucosal tight junction protein. Energy charge could be partly increased at one hour but decrease rapidly in the subsequent stages of NECMO support. Ischemical reperfusion injury of E1 group is the lighest in all the groups after transplantation. At day 7 after the transplantation, the intestinal absorptive function of E1 group recovered quickly, which is closest to the LD group, while the DCD and E5 group recovered slowest. Conclusion: Normothermic extracorporeal membrane oxygenation support for short term could improve donor intestinal graft function after cardiac death. This is the first study in a reliable large animal transplant model demonstrating the efficacy of NECMO to rescue ischemic injured DCD intestinal grafts.

Postoperative Enteral Nutrition Improves Survival After Orthotopic Small Bowel Transplantation in Mice

BackgroundThe aim of this study was to assess the effect postoperative enteral nutrition support on the survival after orthotopic small bowel transplantation in mice. MethodsA model of orthotopic small bowel transplantation used C57BL/6 mice as both donors and recipients. ResultsWe demonstrated that postoperative ileus was the deciding complication after transplantation. Postoperative enteral nutritional support resolved this problem and significantly improved survival and growth rates after transplantation. ConclusionThese data suggest the importance of routine nutritional support after orthotopic small bowel transplantation in mice.

Long-Term Comparison of Rat Model of Chronic Allograft Rejection of Orthotopic Small Bowel Transplantation Induced by Cyclosporine versus Tacrolimus

ObjectiveTo establish 2 models of long-term chronic allograft rejection (CR) of orthotopic small bowel transplantation (OSBT) using cyclosporine (CsA) or tacrolimus (FK506). MethodsF344 and Lewis rats served as donors and recipients respectively for OSBT. In the study 1, the rat was administered CsA (5 mg/kg/d) intramuscularly from postoperative day (POD) 0 to 13. In study 2, intramuscular FK 506 (0.3, 0.5, or 1.0 mg/kg/d) was delivered on POD 0–13, 20 and 27. We observed body weight gain, survival rate, and histology. ResultsIn study 1, all allografts demonstrated one or more histological features of CR but no serious villous blunting. In addition, there was no significant difference in the features of chronic allograft rejection between POD 60 and POD 90. In study 2, the rats that received FK506 at doses of 0.3 or 0.5 mg/kg/day survived to POD 126 but all died before OD 156. Recipients that were administered FK506 at a dose of 1.0 mg/kg/d survived beyond POD 180 showing the same body weight gain as the isogeneic group. Histopathologic analysis revealed distinctive features of CR in the rat models of OSBT induced by CsA or FK506. ConclusionUsing CsA or FK506, we developed a rat CR model of OSBT; recipients administered FK506 survived longer and showed more classic characteristics of CR compared with those treated with CsA.

Impact of Antimouse CD52 Monoclonal Antibody on Graft’s γδ Intraepithelial Lymphocytes After Orthotopic Small Bowel Transplantation in Mice

Alemtuzumab, a humanized CD52 monoclonal antibody (mAb), has been used for immune induction in clinical small bowel transplantation (SBT). However, the local impact of CD52 mAb on the transplanted intestine is not known. In this study, we investigated the effects of CD52 mAb on the intestinal intraepithelial lymphocytes (iIELs) in grafts using a murine orthotopic SBT model. The antimouse CD52 mAb was used as a surrogate antibody. Graft rejection was assessed by histopathologic analysis, iIELs were isolated for flow cytometric analysis, the mucosal keratinocyte growth factor (KGF) expression was evaluated by real-time polymerase chain reaction, and epithelial cell (EC) proliferation (percent bromodeoxyuridine incorporation) was measured by immunohistochemistry. No mice showed evidence of rejection. Seven days after SBT, when the CD52 mAb was used, the numbers of iIELs and both T-cell receptor (TCR) γδ and TCRαβ subpopulations were decreased, and the number of TCRγδ iIELs of donor origin was prominently reduced in the allografts. Furthermore, TCRγδ iIEL-derived KGF messenger RNA expression and EC proliferation decreased significantly after CD52 mAb application. These data demonstrate that the antimouse CD52 mAb could deplete iIELs in the transplanted intestine, especially the TCRγδ subset of donor origin. The reduction of TCRγδ iIELs accompanied the decrease in mucosal KGF expression and EC proliferation, which may slow down the epithelial turnover and retard the repair of the damaged epithelium.

Short-Term TNF-Alpha Inhibition Reduces Neutrophilic Infiltration of Recipient Lung and Leads to Accumulation of Graft Cytoprotective Markers in a Rat Intestinal Transplantation

Aim: Ischemia/reperfusion (I/R) injury is one of the major problems in intestinal transplantation (ITx). Polimorphnuclear neutrophils (PMNs) mediate lung injury as a consequence of the systemic inflammatory response induced after IRI of the small bowel graft. The aim of the present study was to evaluate the effects of different TNF-α inhibition concepts on PMN infiltration of recipient lung and expression level of graft cytoprotective markers as well. Methods: Orthotopic small bowel transplantation was performed in an isogenic model in male Lewis (LEW) rats. Grafts were flushed with UW solution and stored for 6hrs at 4°C before implantation. The TNF-α inhibitor groups received either infliximab (I) (8 mg/kg b.w. i.v. immediately after reperfusion), etanercept (E) (1 mg/kg b.w. s.c.; pre reperfusion, PODs 1,3,5,7) or pentoxifylline (P) (50 mg/kg b.w. i.p pre reperfusion; 25 mg/kg b.w. i.p PODs 1-5). Controls (C) received no further treatment. Tissue samples of proximal and distal graft were taken 20 min, 12h, 7d and 6 mths after reperfusion (each n=6 per time point and group) for immunohistology (HSP70). Lung sections (12h, 7d and 6 mths after reperfusion) were stained for Myeloperoxidase (MPO, specific staining of neutrophil granulocytes). Significant differences were evaluated by multifactorial analysis using two-way ANOVA and Bonferroni post test. Results: Seven day survival was significantly improved following TNF-α inhibition with infliximab (I 100% vs. C 60%: p< 0.05). Beyond POD 7 survival was 100% in all groups. TNF-α inhibition with infliximab correlated with significantly enhanced accumulation of cytoprotective markers (HSP-70) in small bowel grafts. Moreover, MPO stain revealed a significant reduction of neutrophil infiltration of the lung following TNF-α inhibition with all three treatment modalities. Discussion: PMN infiltration of recipient lung was significantly reduced after short-term TNF-α inhibition. Single shot treatment with infliximab correlated with a significant increase in HSP-70 expression in intestinal grafts. These findings may help to: 1. improve injury of distant organs after ITx and 2. ameliorate initial graft injury after I/R, which is believed to exert long-term effects on graft outcome by activating innate immunity, thus contributing to chronic allograft enteropathy.

Short-Term TNF-Alpha Inhibition Reduces Neutrophilic Infiltration of Recipient Lung and Leads to Accumulation of Graft Cytoprotective Markers in a Rat Intestinal Transplantation

Aim: Ischemia/reperfusion (I/R) injury is one of the major problems in intestinal transplantation (ITx). Polimorphnuclear neutrophils (PMNs) mediate lung injury as a consequence of the systemic inflammatory response induced after IRI of the small bowel graft. The aim of the present study was to evaluate the effects of different TNF-α inhibition concepts on PMN infiltration of recipient lung and expression level of graft cytoprotective markers as well. Methods: Orthotopic small bowel transplantation was performed in an isogenic model in male Lewis (LEW) rats. Grafts were flushed with UW solution and stored for 6hrs at 4°C before implantation. The TNF-α inhibitor groups received either infliximab (I) (8 mg/kg b.w. i.v. immediately after reperfusion), etanercept (E) (1 mg/kg b.w. s.c.; pre reperfusion, PODs 1,3,5,7) or pentoxifylline (P) (50 mg/kg b.w. i.p pre reperfusion; 25 mg/kg b.w. i.p PODs 1-5). Controls (C) received no further treatment. Tissue samples of proximal and distal graft were taken 20 min, 12h, 7d and 6 mths after reperfusion (each n=6 per time point and group) for immunohistology (HSP70). Lung sections (12h, 7d and 6 mths after reperfusion) were stained for Myeloperoxidase (MPO, specific staining of neutrophil granulocytes). Significant differences were evaluated by multifactorial analysis using two-way ANOVA and Bonferroni post test. Results: Seven day survival was significantly improved following TNF-α inhibition with infliximab (I 100% vs. C 60%: p< 0.05). Beyond POD 7 survival was 100% in all groups. TNF-α inhibition with infliximab correlated with significantly enhanced accumulation of cytoprotective markers (HSP-70) in small bowel grafts. Moreover, MPO stain revealed a significant reduction of neutrophil infiltration of the lung following TNF-α inhibition with all three treatment modalities. Discussion: PMN infiltration of recipient lung was significantly reduced after short-term TNF-α inhibition. Single shot treatment with infliximab correlated with a significant increase in HSP-70 expression in intestinal grafts. These findings may help to: 1. improve injury of distant organs after ITx and 2. ameliorate initial graft injury after I/R, which is believed to exert long-term effects on graft outcome by activating innate immunity, thus contributing to chronic allograft enteropathy.

Reduced Expression of Mucosal Antimicrobial Peptides in the Small Intestine During Acute Rejection after Orthotopic Small Bowel Transplantation

Background: After small bowel transplantation, episodes of acute rejection are frequent and often associated with severe systemic inflammatory response and sepsis. One potential mechanism is bacterial overgrowth and subsequent translocation of microorganisms through the mucosal barrier. Expression of antimicrobial peptides constitutes a well established defense mechanism of intact intestinal mucosa against such microbial assault, alteration of these antimicrobial peptides could facilitate translocation and prove deleterious to graft and host. Methods: We analyzed the impact of small bowel transplantation and acute rejection on the expression of antimicrobial peptides in intestinal iso- and allografts of rats. For allogenic transplantation, Brown Norway intestinal grafts were transplanted into Lewis hosts (n=9) and for isogenic transplantation Lewis rats served likewise as donor and host (n=9). Graft and host mucosa was sampled at 7 d posttransplant for RT-PCR expression analysis of known antimicrobial peptides. Only laparotomy and mucosa sampling was performed in both BN and Lew rats for sham groups (n=4, respectively). Statistical significance of mRNA expression levels was determined by non-parametric testing (Kruskal-Wallis test with Bonferroni correction and Dunn's multiple comparison test on p< 0,05 level). For histologic grading of acute rejection, the Wu rejection score was applied on HE stained graft sections by two independent reviewers. Results: Histology at 7d posttransplant displayed acute rejection in allogenic grafts and residual inflammatory changes in isogenic grafts as observed by two independent reviewers (interrater reliability analysis showed substantial agreement; Cohen's κ = 0,66, p< 0,001). In RT-PCR expression analysis of host and graft mucosa normalized to β-Actin, Cryptdin 7 and neutrophil defensin 3 (NP 3) were significantly reduced in allogenic (BN to LEW) graft mucosa versus isogenic (Lew to Lew) graft mucosa (Cryptdin7: 11fold, p< 0,05/NP3: 14fold, p< 0,05 reduction respectively). Compared to sham control (laparotomy only), the Cryptdins 5+7 and NP3 were significantly reduced in allogenic graft mucosa (Cryptdin5: 18fold, p< 0,001/Cryptdin7: 17 fold, p< 0,001/NP3: 21fold, p< 0,001 reduction respectively), but not in isogenic graft mucosa. Of the other antimicrobial peptides tested (BD1+2, HIP/PAP3, CRAMP, RELM, Lysozyme), some showed a tendency towards reduced expression in allogenic grafts which did not reach statistical significance. Conclusion: A variety of important antimicrobial peptides, both of Paneth cell- (Cryptdin 5+7), and non-Paneth cell origin (NP3) were found to be significantly reduced in the setting of acute rejection after allogenic intestinal transplantation. The operative trauma as induced by isogenic intestinal transplantation failed to negatively affect the expression of antimicrobial peptides in a similar fashion. If the reduced expression and secretion of antimicrobial peptides is a causative mechanism of bacterial translocation and sepsis during acute rejection of intestinal grafts remains to be determined.

Long-Term Comparison of Rat Model of Chronic Rejection Orthotopic Small Bowel Transplantation Induced by Cyclosporine Versus Tacrolimus

Long-Term Comparison of Rat Model of Chronic Rejection Orthotopic Small Bowel Transplantation Induced by Cyclosporine Versus Tacrolimus

A Novel Microsurgical Model for Orthotopic Small Bowel Transplantation in Mice

A Novel Microsurgical Model for Orthotopic Small Bowel Transplantation in Mice

A Natural Tetrahydropyrimidine, Ectoine, Ameliorates Ischemia Reperfusion Injury after Intestinal Transplantation in Rats

Background/Aims: Ischemia reperfusion (I/R) injury after small bowel transplantation leads to inflammatory reactions and loss of structural integrity with subsequent graft contractile dysfunction in the early postoperative phase. The natural tetrahydropyrimidine ectoine (1-,4-,5-,6-tetrahydro-2-methyl-4-pyrimidine carboxylic acid; THP) protects the ileal mucosa and muscularis against effects of I/R injury in an experimental model of isolated graft reperfusion. The effects of THP treatment were evaluated in an established experimental intestinal transplant model. Methods: Isogenic, orthotopic small bowel transplantation was performed in Lewis rats (6 h cold ischemia time). Perioperative THP treatment (intraluminal/intravascular) groups were compared to vehicle-treated animals (after 3 and 24 h) and non-transplanted controls (n = 5/group). Park’s score defined the effects of I/R injury. The infiltration of neutrophils, monocytes and macrophages, mRNA expression of IL-6 and TNF-α, serum levels of IL-6 and NO and smooth muscle contractility were evaluated. Results: Improved graft outcome after intraluminal and intravascular THP treatment was defined by considerably ameliorated neutrophil infiltration and less histological signs of I/R injury (p ≤ 0.05). In the presence of THP, mRNA expression of IL-6 and TNF-α and IL-6 and NO serum levels were reduced and smooth muscle function was improved. Conclusion: THP treatment offers protection against the effects of I/R injury in intestinal transplantation in vivo, however, only as supplementary treatment option.

Mouse model of orthotopic small bowel transplantation.

To investigate the surgical procedures of orthotopic small bowel transplantation (SBT) model in mice to study the function and rejection of SBT. We established a mouse SBT allograft model as follows: the donor portal vein was anastomosed end by side with the recipient inferior vena cava; the donor superior mesenteric artery with aorta patch was anastomosed end by side with recipient abdominal aorta. After an appropriate length of the recipient's small bowel was removed, the donor's small bowel and the recipient's small bowel were end-to-end anastomosed discontinuously. The mice were fasted for 4 d after the operation, free access to water and subcutaneously injection of 2 mL of 5% glucose saline twice daily. Operation success was regarded as survival for more than 5 d. There is no antibiotic and immunosuppressor. A total of 30 transplantations were done, the 5 d survival rate was 60% (18/30), and 12 died within 5 d. Among the dead recipients, 5 died of arterial anastomotic stenosis and anastomotic thrombosis, 2 of hemorrhagic shock caused by anastomotic bleeding, and the other 5 of intra-abdominal infection caused by postoperative intestinal fistula. The donors' operative time was (40 ± 4.5) min, warm ischemia time was about 0.5 min, donor preparation time was about 3 min, and cold preservation time was (30 ± 7.5) min. The recipients' operative time was (95 ± 8.0) min, among which, the abdominal aorta and inferior vena cava clamping time was (38 ± 3.5) min, the venous anastomotic time was (10 ± 2.0) min and the arterial anastomotic time was (15 ± 3.0) min. The mean intra-operative blood loss of the surviving recipient mice was about 0.2 mL. High quality vascular anastomosis, and rehydration of donors and recipients are crucial factors for improving the success rate of SBT.

Small bowel transplantation in outbred rats

To investigate the clinical evolution of orthotopic small bowel transplantation in outbred rats. Seventy-two outbred Wistar rats weighting from 250 to 300g were used as donor and recipient in 36 consecutives ortothopic small intestine transplantation without immunosuppression. The graft was transplanted into the recipient using end-to-side aortic and portacaval microvascular anastomosis. Procedure duration, animal clinical course and survival were evaluated. Survival shorter than four days was considered technical failure. Recipients were sacrificed with signs of severe graft rejection or survival longer than 120 days. Necropsies were performed in all recipients to access histopathological changes in the graft. Median time for the procedure was 107 minutes. Six recipients (16.7%) presented technical failure. Twenty-seven recipients were sacrificed due to rejection, being nineteen (52.7%) between 7(th) and 15(th) postoperative day and eight (22.2%) between 34(th) and 47(th) postoperative day. Graft histology confirmed severe acute cellular rejection in those recipients. Uneventful evolution and survival longer than 120 days without rejection were observed in three recipients (8.3%). Intestinal transplantation in outbred rats without immunosuppressant regiment accomplishes variable clinical evolution.

Perioperative infliximab application has marginal effects on ischemia–reperfusion injury in experimental small bowel transplantation in rats

Ischemia-reperfusion injury leads to impaired smooth muscle function and inflammatory reactions after intestinal transplantation. In previous studies, infliximab has been shown to effectively protect allogenic intestinal grafts in the early phase after transplantation with resulting improved contractility. This study was designed to reveal protective effects of infliximab on ischemia-reperfusion injury in isogenic transplantation. Isogenic, orthotopic small bowel transplantation was performed in Lewis rats (3h cold ischemia). Five groups were defined: non-transplanted animals with no treatment (group 1), isogenic transplanted animals with vehicle treatment (groups 2/3) or with infliximab treatment (5mg/kg body weight intravenously, directly after reperfusion; groups 4/5). The treated animals were sacrificed after 3 (group 2/4) or 24h (group 3/5). Histological and immunohistochemical analysis, TUNEL staining, real-time RT-PCR, and contractility measurements in a standard organ bath were used for determination of ischemia-reperfusion injury. All transplanted animals showed reduced smooth muscle function, while no significant advantage of infliximab treatment was observed. Reduced infiltration of neutrophils was noted in the early phase in animals treated with infliximab. The structural integrity of the bowel and infiltration of ED1-positive monocytes and macrophages did not improve with infliximab treatment. At 3h after reperfusion, mRNA expression of interleukin (IL)-6, TNF-α, IL-10, and iNOS and MCP-1 displayed increased activation in the infliximab group. The protective effects of infliximab in the early phase after experimental small bowel transplantation seem to be unrelated to ischemia-reperfusion injury. The promising effects in allogenic transplantation indicate the need for further experiments with infliximab as complementary treatment under standard immunosuppressive therapy. Further experiments should focus on additional infliximab treatment in the setting of acute rejection.

2-Octylcyanoacrylate-Assisted Microvascular Anastomosis in Rat Orthotopic Small Bowel Transplantation

The purpose of this study was to investigate the potential advantages of 2-octylcyanoacrylate-assisted end-to-side arterial anastomosis over continuous suture-only anastomosis in the rat orthotopic small bowel transplantation (OSBT) model. OSBT was performed in 80 Sprague-Dawley rats, and they were divided into two groups. Group 1 was the study group (n = 40), using 2-octylcyanoacrylate-assisted end-to-side arterial anastomosis. Group 2 was the control group (n = 40), using a running 9-0 prolene suture only. The operation time and degree of leakage were observed. After sacrifice on postoperative days 90, histology and electron scanning microscope of arterial anastomosis were measured. The suture time and recovery time in group 1 were significantly shorter than that in group 2. First-degree leakage was observed mostly in group 1, and second-degree leakage was common in group 2. No difference was found between the two techniques considering the graft intestinal histological changes. Under electron scanning microscopy, the anastomoses of the two groups displayed no obvious difference. In conclusion, due to superior speed of operation and no toxicity to vessel wall, 2-octylcyanoacrylate-assisted end-to-side arterial anastomosis might be a quick and safe method that is much easier to apply in rat OSBT.

Intensive Fluid Therapy Combined with Epinephrine Improves Orthotopic Small Bowel Transplantation in Mice

Mouse model of orthotopic small bowel transplantation (SBT) has great advantages in the researches of post-transplant immunology and physiology, whereas it is infrequently utilized because of low survival rate. To improve this model, we modified the techniques and the perioperative treatments. Syngeneic orthotopic SBT was performed in C57 mice. Two groups were established in accordance with the perioperative treatments: control group (only intensive fluid therapy was administered); epinephrine group (intensive fluid therapy and epinephrine were applied jointly). Only fluid enteral nutrition was allowed after surgery in all recipients. Survival and the change of body weight were observed for 2 wk after transplantation. The ischemia/reperfusion (I/R) injury of the grafts was assessed by histopathology on postoperative day (POD) 1 and 7. The overall success rate was 76.5% (26/34). In control group and epinephrine group, the success rate was 66.7% (12/18) and 87.5% (14/16), respectively. No complications of thrombosis and bowel obstruction were found. The body weight changed in the same way in both groups. No differences of I/R injury were observed between two groups. Precise perioperative treatments, especially intensive fluid therapy combined with epinephrine application, improve the mouse model of orthotopic SBT to achieve satisfying success rate.

FK506 treatment in a Long-term chronic rejection rat model of small bowel transplantation

Although acute rejection (AR) can be significantly improved by effective immunosuppressants, such as FK506, chronic rejection (CR) remains a major hurdle to long-term allograft survival after small bowel transplantation, in part because the pathogenic mechanisms of CR are, as yet, unknown. The rat orthotopic small bowel transplantation (OSBT) model has been used by a few researchers, but without long-term survival. Rats were randomly divided into five groups: Group 1 (n=20), sham-operation rats; Group 2 (n=20), Lewis to Lewis; Group 3 (n=20), F344 to Lewis treated with FK506 (0.3 mg/kg/day); Group 4 (n=20), F344 to Lewis with FK506 (0.5 mg/kg/day); Group 5 (n=20), F344 to Lewis with FK506 (1.0 mg/kg/day). FK506 was administrated intramuscularly to recipients on postoperative days (POD) 0-13, 20 and 27. Body weight, survival rate and histology were measured. Histopathological analysis revealed distinctive abnormalities of the allograft for all animals, including changes in villous architecture, interstitial fibrosis and intimal thickening; however, survival times were significantly increased with higher doses of FK506. Rats in Group 3 and Group 4 (low and moderate FK506 doses) survived 16-18 weeks, while recipients in Group 5 (high dose) survived 24-27 weeks. FK506 treatment (1.0 mg/kg/day, intramuscularly administrated to recipients on POD0-13, 20 and 27) can be used effectively to establish a rat OBST model of CR that will be useful for the study of the pathogenesis of CR and the effectiveness of various drugs.

Expression and function of intestinal hexose transporters after small intestinal denervation

The role of neural regulation in expression and function of intestinal hexose transporters is unknown. The aim of this study is to determine the role of intestinal innervation in gene expression and function of the membrane hexose transporters, SGLT1, GLUT2, and GLUT5 in the enterocyte. We hypothesize that denervation of the small intestine decreases expression of hexose transporters, which leads to decreased glucose absorption. Six groups of Lewis rats were studied (n = 6 each) as follows: control, 1 week after sham laparotomy, 1 and 8 weeks after syngeneic (no immune rejection) orthotopic small-bowel transplantation (SBT) (SBT1 and SBT8) to induce complete extrinsic denervation, and 1 and 8 weeks after selective disruption of intrinsic neural continuity to jejunoileum by gut transection and reanastomosis (T/A1 and T/A8). All tissue was harvested between 8 AM and 10 AM. In duodenum, jejunum, and ileum, mucosal messenger RNA (mRNA) levels were quantitated by real-time polymerase chain reaction (PCR), protein by Western blotting, and transporter-mediated glucose absorption using the everted sleeve technique. Across the 6 groups, the relative gene expression of hexose transporter mRNA and protein levels were unchanged, and no difference in transporter-mediated glucose uptake was evident in any region. The glucose transporter affinity (K(m)) and functional transporter levels (V(max)) calculated for duodenum and jejunum showed no difference among the 6 groups. Baseline regulation of hexose transporter function is not mediated tonically by intrinsic or extrinsic neural continuity to the jejunoileum.