Orthotopic Small Bowel Transplantation in Rats

Summary

Small bowel transplantation has become an accepted treatment option for patients with irreversible intestinal failure. Our experimental model of orthotopic small bowel transplantation in rats serves as a reliable tool to address underlying immunologic and inflammatory processes that complicate intestinal transplantation.

Abstract

Small bowel transplantation has become an accepted clinical option for patients with short gut syndrome and failure of parenteral nutrition (irreversible intestinal failure). In specialized centers improved operative and managing strategies have led to excellent short- and intermediate term patient and graft survival while providing high quality of life ^1,3. Unlike in the more common transplantation of other solid organs (i.e. heart, liver) many underlying mechanisms of graft function and immunologic alterations induced by intestinal transplantation are not entirely known^6,7. Episodes of acute rejection, sepsis and chronic graft failure are the main obstacles still contributing to less favorable long term outcome and hindering a more widespread employment of the procedure despite a growing number of patients on home parenteral nutrition who would potentially benefit from such a transplant. The small intestine contains a large number of passenger leucocytes commonly referred to as part of the gut associated lymphoid system (GALT) this being part of the reason for the high immunogenity of the intestinal graft. The presence and close proximity of many commensals and pathogens in the gut explains the severity of sepsis episodes once graft mucosal integrity is compromised (for example by rejection). To advance the field of intestinal- and multiorgan transplantation more data generated from reliable and feasible animal models is needed. The model provided herein combines both reliability and feasibility once established in a standardized manner and can provide valuable insight in the underlying complex molecular, cellular and functional mechanisms that are triggered by intestinal transplantation. We have successfully used and refined the described procedure over more than 5 years in our laboratory ^8-11. The JoVE video-based format is especially useful to demonstrate the complex procedure and avoid initial pitfalls for groups planning to establish an orthotopic rodent model investigating intestinal transplantation.

Protocol

1. Donor Operation

1. The donor rat should be kept fasting for 24 hr (free access to water/glucose solution).
2. To induce Isoflurane inhalation anesthesia, start with 2% on standard atomizer, and then reduce to 1% after performing laparotomy. Perform a toe pinch to check sedation.
3. Shave the abdomen and clean with skin prep 3 times (Kodan). Then perform a median incision after subcutaneous administration of the analgesic.
4. After the graft is wrapped in saline soaked gauze, separate the physiological adhesions between the pancreas and the ascending colon gently with a Q-tip (under the surgical microscope with 6x magnification).
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Results

Normal postoperative course

The transplanted animals should recover quickly from the procedure under a heat lamp for approximately 1 hr. Hypothermia is a major cause of postoperative mortality and should be carefully avoided intra- and postoperatively. Intraoperative fluid losses must be replaced by s.c. injection of 2.5 ml normal saline plus 2.5 ml Glucose 5% every 8 hr for the first 24 hr. The rats should also have free access to glucose solution (or gel diet) and water p.o. for the first 24 hr.......

Discussion

While intestinal transplant models in rats have been described as early as in the 1970ies ⁵ most of the recently employed models involve heterotopic intestinal transplantation using different techniques ¹³. While the heterotopic procedures in general have the advantage of easier microsurgical techniques and easier accessibility of the graft for assessment, heterotopic intestinal transplantation has the big disadvantage of not taking into account the multiple interactions of the transplanted s.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
University of Wisconsin (UW) solution (ViaSpan)	Bristol-Myers Squibb
Uro-Nebacetin N solution	Nycomed	6967855
Ampicillin	Ratiopharm
Carprofen (Rimadyl)	Pfizer
Prolene 10-0 unresorbable suture	Ethicon
Monocryl 6-0 resorbable suture	Ethicon
Vicryl 3-0 resorbable suture	Ethicon
i.v. Catheter G 20 1.1x33 mm	Braun
i.v. Catheter G 22 0.9x25 mm	Braun
Kodan Skin Prep	Schülke
NaCl 0.9% Infusion solution	Braun
Curved forceps small	FineScienceTools	11009-13
Micro forceps curved	AESCULAP	BD 333
Micro forceps curved	AESCULAP	FD281R
Micro forceps straight 1	WPI	5
Micro forceps straight 2	WPI	2
Micro needle holder	WPI	14081
Micro scissors	FineScienceTools	15006-09
Micro scalpel	MANI	Ophthalmic knife
Micro clamps	AESCULAP	FB329R