Small Bowel Transplantation In Mice Research Articles

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

IDO expressing dendritic cells suppress allograft rejection of small bowel transplantation in mice by expansion of Foxp3+ regulatory T cells

BackgroundIndoleamine 2,3-dioxygenase (IDO), the enzyme that catalyzes the first and rate-limiting step of tryptophan catabolism, suppresses T-cell responses by tryptophan depletion and accumulation of kynurenine metabolites. IDO prevents allograft rejection in various transplantations. MethodsDendritic cells (DC) highly expressing IDO (IDO+ DC) were cultured through transduction of adenovirus vectors carrying the IDO sequence. IDO+ DC were incubated with CD4+ CD25− T cells to detect T cell proliferation. The effects of IDO+ DC and 3-Hydroxyanthranilic acid (3-HAA) were verified in an allogeneic murine small bowel transplantation (SBT) model. Foxp3+ Treg cells of recipient mice were detected by flow cytometry and cytokines in plasma were determined by ELISA. ResultsIDO+ DC effectively suppressed proliferation of CD4+ CD25− T cells in vitro, and this effect could be enhanced by adding 3-HAA. In the SBT transplantation model, both 3-HAA (P<0.05) and IDO+ DC (P<0.01) prolonged the survival time of transplanted mice. Mice treated with IDO+ DC achieved longer mean survival time than 3-HAA administrated mice (11.5d vs. 18.5d). Grafts from IDO+ DC, 3-HAA and combination treatment group showed reduced inflammation and minimal architectural distortion. IFN-γ production was significantly inhibited by IDO+ DC and 3-HAA (P<0.05). The expression of IL-2 was slightly lower with 3-HAA or IDO+ DC treatment. However, IL-10 was higher in 3-HAA, IDO+ DC and combination treatment groups, while TGF-β was elevated in all non-control groups. ConclusionsIDO+ DC plus 3-HAA has an immunoprotective role and represents a potential strategy to suppress acute rejection and prolong survival of grafts in SBT.

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.

A Novel Microsurgical Model for Orthotopic Small Bowel Transplantation in Mice

A Novel Microsurgical Model for Orthotopic Small Bowel Transplantation in Mice

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.

Development of a Simple Model of Extrinsic Denervation of the Small Bowel in Mouse

Small bowel transplantation (SBT) is associated with poorly understood enteric dysfunction. The study of SBT in mice is hindered by the technical difficulty of orthotopic SBT in the mouse. Our aim was to develop an easy preparation of extrinsic denervation of the entire jejunoileum in mice as a model of orthotopic SBT. All neurolymphatic tissues accompanying the superior mesenteric artery (SMA) and vein (SMV) were ligated just distal to the middle colic vessels. The SMA and SMV were then stripped of investing adventitia, and the mesentery to jejunum and colon were transected radially. Jejunum and colon were not transected and reanastomosed. To confirm extrinsic denervation 1, 3, and 6 months later, segments of small bowel were stained for protein gene product 9.5 (PGP9.5) and tyrosine hydroxylase (TH). Tyrosine hydroxylase immunoreactive intensity was then quantified using a semiquantitative analysis. Immunohistochemical fluorescence showed persistence of PGP9.5 immunoreactivity confirming enteric nerves in jejunoileum; however, there was no TH immunoreactivity in jejunoileum in denervated mice despite the expected preservation of TH immunoreactivity in the still-innervated duodenum at 1 month. At 3 months, sparse immunoreactivity for TH was present, and by 6 months, reinnervation of TH-containing nerves appeared similar to controls. Quantification of intensity at each time-point further confirmed this trend. This technique in the mouse accomplishes a complete extrinsic denervation of jejunoileum early postoperatively (1 and 3 months); reinnervation occurs by 6 months. This is an easily learned murine model of orthotopic SBT.

Orthotopic Small Bowel Transplantation in Mice

On the basis of our experience with rat small bowel transplantation (SBT), we established a model of orthotopic SBT in mice. The immediate success rate was approximately 50%. Indefinite survival was obtained in the syngeneic group. The effect of different immunosuppressive treatments was also investigated: mean survival was 10.2+/-1.2 days in untreated mice receiving allogeneic grafts, 14.2+/-1.6 days with cyclosporine, 24+/-5.1 days in the rapamycin group, and 73.6+/-29.1 days with tacrolimus-treated animals. From this study, we conclude that although the model is technically difficult, it may provide an excellent tool to further investigate the physiology and immunology of small-intestinal transplantation.

Intragraft distribution of lymphocytes expressing β7 integrins after small bowel transplantation in mice

Lymphocytes with activated β7 integrins (αEβ7 and α4β7) contribute to inflammatory reactions in the small bowel. Since the selective recruitment of lymphocytes to the lymphoid compartments of the small bowel is controlled by distinct adhesion molecule interactions, a compartment-dependent use of β7 integrins may influence the rejection response within intestinal transplants. To further delineate the nature of β7 integrin-mediated graft infiltration, we analysed their expression on T lymphocytes in the heterotopically transplanted small bowel of BALB/c and C57BL/6 mice. Lymphocytes isolated from the epithelium, lamina propria (LP), Peyer's patches (PP), and mesenteric lymph nodes (MLN) were analysed by three-color fluorescence flow cytometry using monoclonal antibodies (mAb) to integrin the subunits, lymphocyte markers, and MHC I of the donor and recipient strains. On postoperative day 5 (POD) after allogeneic small bowel transplantation (SBT), 43% of intraepithelial lymphocytes (IEL) and 63% of LP, 93% of MLN, and 93% of PP lymphocytes were of host origin. In the MLN and PP of allografts, a major infiltrating lymphocyte population consisted of CD8 + cells with increased expression of α4β7 and decreased expression of L-selectin, an adhesion molecule profile characteristic of intestinal effector cell phenotypes. An increase in α4β7 levels was also found on CD8 + host lymphocytes in the LP. The integrin profile of a number of host IEL suggests an ongoing transition from the phenotype of graft infiltrating lymphocytes with high levels of α4β7 and low levels of αεβ7 to that of resident IEL with high levels of αεβ7 and low levels of α4β7. The importance of β7-mediated lymphocyte trafficking to the graft is attested by the significant reduction in the host lymphocyte population in the LP, PP, and epithelium following the administration of a β7-blocking mAb to allograft recipients. In conclusion, while the infiltration patterns of lymphocytes may vary between the lymphoid compartments of intestinal allografts, host CD8 + lymphocytes with high levels of α4β7 constitute a major effector cell population that affects the entire graft.

Modified Surgical Model of Paratopic Small Bowel Transplantation in Mice

A murine paratopic model of intestinal transplantation was developed and used to study the immune response to allografts in normal C57BL/6 recipients (H2b) and C57BL/6 recipients treated with tacrolimus (1 mg/kg, Days 0–7). B6C3F1 mice (C57BL/6 × C3H/HeJ, H2bxk) were used as donors. The paratopic model differed from the standard heterotopic model in that continuity of the intestine graft with the recipient intestine was established by anastomosing the graft ileum to the side of the recipient jejunum. The success rate of this modified procedure was 82% (94/114). Major complications included hypovolemic shock and/or anesthetic-related deaths (8%), infection (8%), and vascular thrombosis (2%). Rejection was assessed using both clinical features (i.e., edema and closure of the stoma, mucous discharge, and a palpable mass) and histologic features (apoptosis of crypt cells, crypt destruction, lymphocytic infiltrate, and mucosal ulceration). Syngeneic grafts appeared clinically and histologically normal throughout the study period. Untreated recipients of allografts developed clinical and histologic evidence of rejection by Day 4 which progressed to severe rejection and graft destruction by Day 18. After initially developing evidence of mild to moderate graft rejection on Day 8, the severity of allograft rejection decreased significantly by Days 10 to 14 in tacrolimus-treated mice. By Day 28 evidence of moderate rejection had recurred in mice treated with an 8-day course of tacrolimus. Consistent with these observations, graft function, as assessed by maltose absorption, was impaired in rejecting allografts when compared with syngeneic grafts. Allografts in mice treated with tacrolimus demonstrated improved maltose absorption relative to allografts from untreated mice. These studies describe a modified technique for intestinal transplantation in mice and provide a detailed analysis of complications as well as an assessment of rejection and its functional consequences. Based on these results, we conclude that the paratopic model of intestinal transplantation in mice is a useful tool for studying the host immune response to intestinal allografts.

Prévention du rejet d'allogreffe intestinale par des anticorps antimolécules d'adhésion dans un modèle murin

Study aim Small bowel transplantation is still hampered by a high morbidity and mortality linked to the heavy non specific immunosuppression which is required by the transplantation of this lymphoid organ. Adhesion molecules appear to be potential targets for specific immunosuppression. The aim of the study was to investigate the effect of a transitory administration of anti-LFA-1 or anti-α4 monoclonal antibodies (mAb) in the prevention of rejection in a model of fetal small-bowel transplantation in mice. Materials and methods The small bowel of C57BL/6 (H-2 b) fetus (16 to 20 days of gestation) was transplanted into adult C3H/He mice (H-2 k) or C57BL/6 recipient mice. Recipients were treated with a short course of either antiLFA-1 mAb alone, either with anti-α4 mAb alone, or with both mAb. Biopsies with histological study of the grafts were performed between post-operative day 5 and 60. A score of development and rejection was assigned to each sample. Results Normal intestinal development with no sign of rejection was observed in 24/28 syngenic grafts till postoperative day 45. In the absence of treatment, intense rejection was observed as soon as day 5 and all allogenic grafts were rejected ( n=22). In contrast, in anti-LFA-1 mAb treated mice, 18/20 allogenic grafts developed normally with minimal signs of rejection. In anti-α4 treated mice, a transient protective effect on small bowel allograft survival was observed on day 7 but thereafter, all grafts were massively rejected within a few days ( n = 18). The combination of both mAb didn't improve the survival of the grafts when compared to anti-LFA-1 mAb treated grafts ( n = 10). Conclusion These results demonstrate that a transitory administration of anti-LFA-1 mAb, but not of anti-α4 mAb, is able to prolong significantly the survival of non vascularized small bowel fetal grafts in mice. Our results are promising for the possible use of the anti-LFA-1 mAb in clinical intestinal transplantation.