Summary
Denne roman model for ortotopisk bagben transplantation i mus, at anvende en ikke-sutur cuff teknik til super-mikrovaskulære anastomose, giver et kraftfuldt værktøj til in vivo mekanistisk immunologisk forskning relateret til vaskulariseret komposit allotransplantation (VCA).
Abstract
In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and translational research in the field of transplantation medicine. A vast array of reagents is available exclusively in this setting, including mono- and polyclonal antibodies for both diagnostic and interventional applications. In addition, a vast number of genotyped, inbred, transgenic, and knock out strains allow detailed investigation of the individual contributions of humoral and cellular components to the complex interplay of an immune response and make the mouse the gold standard for immunological research.
Vascularized Composite Allotransplantation (VCA) delineates a novel field of transplantation using allografts to replace “like with like” in patients suffering traumatic or congenital tissue loss. This surgical methodological protocol shows the use of a non-suture cuff technique for super-microvascular anastomosis in an orthotopic mouse hind limb transplantation model. The model specifically allows for comparison between established paradigms in solid organ transplantation with a novel form of transplants consisting of various different tissue components. Uniquely, this model allows for the transplantation of a viable vascularized bone marrow compartment and niche that have the potential to exert a beneficial effect on the balance of immune acceptance and rejection. This technique provides a tool to investigate alloantigen recognition and allograft rejection and acceptance, as well as enables the pursuit of functional nerve regeneration studies to further advance this novel field of transplantation.
Introduction
The late nineties heralded the pioneering days of reconstructive transplantation with the first successful hand transplant performed in France in 1998. Since then, the use of VCAs for reconstruction of devastating tissue defects has been successfully employed in a wide spectrum of patients. To date, the world counts 76 recipients of 112 upper extremities as well as 31 faces 1-3. In addition, several other types of VCAs such as abdominal wall 4, larynx 5, trachea 6, vascularized joints 7, and even penis 8 have been performed. Furthermore, the live birth of a baby was recently reported after uterus transplantation 9. This growing world experience is indicative for how reconstructive transplantation has become a valid therapeutic option for patients suffering of significant functional tissue defects not amendable to conventional reconstructive and restorative surgery and treatment.
While the idea of replacing “like with like” sparked clinical enthusiasm, initial skepticism still prevails with regards to side effects of conventional high-dose immunosuppression required to maintain allografts and their function 10,11. However, as shown by seminal work of Lee et al., these composite grafts are less likely to reject than its individual components, and furthermore, some of the tissue components such as the vascularized bone compartment have fueled optimism as they might exert unique immunological effects onto the balance of immune acceptance and rejection 12.
Our group pioneered several microsurgical animal models for solid organ transplantation, as well as vascularized composite allotransplantation 13-19. Here we describe a novel surgical procedure using a non-suture cuff technique to perform super micro-vascular anastomosis in an orthotopic mouse hind limb transplantation model. This transplant model provides a useful tool for investigating immune acceptance and rejection mechanisms, as well as the role of individual tissue components, such as the vascularized bone marrow compartment, towards tolerance induction in the immunologically versatile setting of the mouse species. Additionally, the orthotopic placement of the limb opens the possibilities for nerve regeneration and functional outcome studies, which are critically important to the setting of VCA.
Protocol
Representative Results
Discussion
Vaskulariseret Composite Allotransplantation, såsom øvre ekstremitet og ansigt transplantation til genopbygning af ødelæggende væv defekter, har udviklet sig som en brugbar løsning for patienter, der ikke kunne ændres til konventionelle rekonstruktive procedurer behandling. Tekniske fremskridt inden for rekonstruktiv mikrokirurgi samt en stor erfaring med potente immunosuppressive og immunmodulerende behandlinger i organtransplantation, nu muliggør langvarig allotransplantatoverlevelse i denne unikke patientpopu…
Disclosures
The authors have nothing to disclose.
Acknowledgements
Dette arbejde blev støttet af hæren, søværnet, NIH, Air Force, VA og sundhed anliggender at støtte AFIRM II indsats under Award No. W81XWH-13-2-0053. Den amerikanske hær Medical Research Acquisition Aktivitet, 820 Chandler Street, Fort Detrick MD 21.702-5014 er tildelingen og administration erhvervelse kontor. Udtalelser, fortolkninger, konklusioner og anbefalinger er dem af forfatteren og ikke nødvendigvis godkendt af det amerikanske forsvarsministerium.
Forfatterne vil gerne takke Jessica Izzi, DVM, Caroline Garrett, DVM og Julie Watson, DVM for deres fremragende veterinær støtte i denne undersøgelse.
Materials
| Suture, 6-0 Nylon | MWI | 31849 | |
| Suture, 6-0 Polysorb | MWI | 72667 | |
| Suture, 10-0 Nylon | Aero Surgical | TK-107038 | |
| Polyimide Tubing, Size 25 | Vention Medical | 141-0023 | |
| Polyimide Tubing, Size 27 | Vention Medical | 141-0015 | |
| Microvascular Clamps (Single) | Synovis | 00396 | |
| Microvascular Clamps (Double) | Synovis | 00414 | |
| Micro-Scissors | Synovis | SAS-18 | |
| Micro-Forceps | Synovis | FRS-15 RM-8 | |
| Micro-Dilators | Synovis | FRS-15 RM-8d.1 | |
| Micro-Needledriver | Synovis | C-14 | |
| Micro-Clamp Applicator | Synovis | CAF-4 | |
| Micro-Flushing Needle | Hamilton | N/A | 10MM, 30°, 33G |
| Lactated Ringers Solution | Fisher Scientific | NC9968051 | |
| Buprenorphine | N/A | N/A | DEA Number required; Obtained from hosptial pharmacy. |
| Enrofloxacin; Baytril | Bayer Health Care | 186599 | |
| Heparin | N/A | N/A | Obtained from hosptial pharmacy |
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