Rishi Jindal

A rapid vascular anastomosis technique for hind-limb transplantation in rats

Orthotopic hind-limb transplantation in rats has been the preferred rodent model in reconstructive transplantation1; however, the surgical procedure involves extensive and time-consuming reattachment of bones, muscles, blood vessels, and nerves. Performing the vascular anastomoses is by far the most time-consuming and delicate part of this procedure and requires a high level of microsurgical skill. For investigation of ischemia-reperfusion–related injuries, such a suture technique–based model is suboptimal because prolonged tissue storage leading to more fragile vessel walls complicates completion of the vascular anastomoses. Although very experienced and technically skilled microsurgeons report an acceptable success rate with the conventional (suture) technique even with prolonged ischemia, an experimental model should not be based on the capabilities of a few but should be available to and reproducible by many.2 The objective of this study was to investigate whether the cuff technique for vascular anastomosis is advantageous with regard to mortality, morbidity, operation time, and costs when used to test for ischemia-reperfusion injury in reconstructive transplantation.

Spontaneous resolution of acute rejection and tolerance induction with IL-2 fusion protein in vascularized composite allotransplantation

Vascularized composite allotransplantation (VCA) has emerged as a treatment option for treating nonlife‐threatening conditions. Therefore, in order to make VCA a safe reconstruction option, there is a need to minimize immunosuppression, develop tolerance‐inducing strategies and elucidate the mechanisms of VCA rejection and tolerance. In this study we explored the effects of hIL‐2/Fc (a long‐lasting human IL‐2 fusion protein), in combination with antilymphocyte serum (ALS) and short‐term cyclosporine A (CsA), on graft survival, regulatory T cell (Treg) proliferation and tolerance induction in a rat hind‐limb transplant model. We demonstrate that hIL‐2/Fc therapy tips the immune balance, increasing Treg proliferation and suppressing effector T cells, and permits VCA tolerance as demonstrated by long‐term allograft survival and donor‐antigen acceptance. Moreover, we observe two distinct types of acute rejection (AR), progressive and reversible, within hIL‐2/Fc plus ALS and CsA treated recipients. Our study shows differential gene expression profiles of FoxP3 versus GzmB, Prf1 or interferon‐γ in these two types of AR, with reversible rejection demonstrating higher Treg to Teff gene expression. This correlation of gene expression profile at the first clinical sign of AR with VCA outcomes can provide the basis for further inquiry into the mechanistic aspects of VCA rejection and future drug targets.

Orthotopic hind-limb transplantation in rats.

Composite tissue allotransplantation (CTA) now represents a valid therapeutic option after the loss of a hand, forearm or digits and has become a novel therapeutic entity in reconstructive surgery. However, long term high-dose multi-drug immunosuppressive therapy is required to ensure graft survival, bearing the risk of serious side effects which halters broader application. Further progression in this field may depend on better understanding of basic immunology and ischemia reperfusion injury in composite tissue grafts. To date, orthotopic hind limb transplantation in rats has been the preferred rodent model for reconstructive transplantation (RT), however, it is an extremely demanding procedure that requires extraordinary microsurgical skills for reattachment of vasculature, bones, muscles and nerves. We have introduced the vascular cuff anastomosis technique to this model, providing a rapid and reliable approach to rat hind limb transplantation. This technique simplifies and shortens the surgical procedure and enables surgeons with basic microsurgical experience to successfully perform the operation with high survival and low complication rates. The technique seems to be well suited for immunological as well as ischemia reperfusion injury (IRI) studies.

Cadaveric Study of a Z-Plasty Modification to the Moberg Flap for Increased Advancement and Decreased Morbidity.

Background: The Moberg advancement flap is a well-established tool to provide sensate, vascularized tissue for thumb reconstruction. Modifications providing additional length have been described, but no studies have examined how much additional advancement can be achieved consistently, and at what cost. The authors hypothesized that Z-plasty modification at the base of the Moberg flap would allow additional advancement compared with the traditional technique, and maintain primary closure of the donor-site and avoid additional morbidity. Methods: Standard Moberg flaps were performed and advancement was measured on 20 cadaver specimens. Ten flaps were then modified with the O’Brien technique of incising proximally and skeletonizing the neurovascular bundles. The other 10 flaps were modified with Z-plasties at the base of the thumb. Differences in distance of advancement were compared, as was the ability to primarily close donor sites. Results: Average advancement for Moberg flaps was 7.3 ± 1.2 mm, compared with 15.0 ± 2.5 mm for the O’Brien modification (p < 0.01) and 11.3 ± 1.7 mm for the Z-plasty modification (p < 0.01). Conclusions: Although the O’Brien modification allows approximately 50 percent further advancement than the Z-plasty modification compared with the standard Moberg flaps, the increase correlates to a large area of exposed neurovascular bundles at the volar base of the thumb, which requires secondary coverage. However, all Z-plasty donor-sites could be closed primarily. Primary closure of all donor sites will decrease healing time, wound complications, digital nerve sensitivity, and cosmetic appearance. This study is the first to show a significant increase in Moberg flap advancement using Z-plasty lengthening at the thumb base while avoiding any increased morbidity.

13: A NOVEL IMMUNOMODULATORY MODEL TO PREDICT REJECTION OUTCOMES AND INDUCE TOLERANCE IN CTA

Purpose: Despite the advance of immunosuppression in composite tissue allotransplantation (CTA), tolerance remains a distant possibility and immunomodulation may prove to be the key. Induction therapy and cyclosporine A (CsA) are widely used in transplantation, yet this is not tolerogenic. Since CsA blocks interleukin-2 (IL-2) transcription and IL-2 dependent regulation, and Tregs require IL-2 for survival, we hypothesize that adding a long lasting IL-2 fusion protein (IL2/Fc) to anti-lymphocyte serum (ALS) and CsA will enhance immunoregulation and CTA tolerance.

34A: TITRATION OF BONE MARROW CELL INFUSION IN A PRECLINICAL MODEL OF COMPOSITE TISSUE ALLOTRANSPLANTATION

Introduction: Composite tissue allotransplantation (CTA) has become a clinical reality with excellent functional results and most encouraging short and intermediate outcomes. However, recipients still require lifelong multi-drug immunosuppression to prevent graft rejection. While ensuring graft survival these regimens carry a high risk for serious side effects. Current research therefore focuses on developing strategies to minimize or avoid immunosuppression for such nonlife saving procedures. In this regard we have developed a preclinical model of heterotopic hindlimb transplantation in Yucatan miniature swine using recipient conditioning, donor bone marrow infusion, and monotherapy immunosuppression. Our current study aims to determine the optimal dose of bone marrow cells to be infused to establish stable mixed chimerism.

180B: EFFECTS OF IMMUNE RESPONSE AND CYCLOSPORINE (CSA) TREATMENT ON NERVE REGENERATION IN COMPOSITE TISSUE TRANSPLANTATION (CTA)

Background: Optimization of nerve regeneration and functional return is essential to improving the viability of CTA. To achieve this goal, we must enhance our understanding of how the immune response to allogeneic tissue affects nerve regeneration in CTA. Research in nerve allografts suggests subacute Schwann cell (SC) rejection occurs even with therapeutic levels of immunosuppression. In this study, we compare nerve regeneration in allogeneic and syngeneic limb transplantion with CsA treatment.