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.

Vascularized osteomyocutaneous allografts are permissive to tolerance by induction-based immunomodulatory therapy.

Vascularized composite allografts (VCAs) are unique among transplanted organs in that they are composed of multiple tissues with disparate antigenic and immunologic properties. As the predominant indications for VCAs are non-life-threatening conditions, there is an immediate need to develop tolerance induction strategies and to elucidate the mechanisms of VCA rejection and tolerance using VCA-specific animal models. In this study, we explore the effects of in vitro induced donor antigen-specific CD4(-) CD8(-) double negative (DN) Treg-based therapy, in a fully MHC mismatched mouse VCA such as a vascularized osteomyocutaneous as compared to a non-VCA such as a full thickness skin (FTS) transplantation model to elucidate the unique features of VCA rejection and tolerance. We demonstrate that combined therapy with antigen-induced CD4 derived DN Tregs and a short course of anti-lymphocyte serum, rapamycin and IL-2/Fc fusion protein results in donor-specific tolerance to VCA, but not FTS allografts. Macrochimerism was detected in VCA but not FTS allograft recipients up to >60 days after transplantation. Moreover, a significant increase of CD4(+) Foxp3(+) Tregs was found in the peripheral blood of tolerant VCA recipients. These data suggest that VCA are permissive to tolerance induced by DN Treg-based induction therapy.

Composite tissue vasculopathy and degeneration following multiple episodes of acute rejection in reconstructive transplantation.

Transplant vasculopathy has not been systematically investigated in composite tissue allotransplantation (CTA). The impact of multiple acute rejections (ARs) on long-term graft outcomes in reconstructive transplantation remains unknown. This study in a rat hind-limb allotransplantation model systematically analyzes vasculopathy and tissue-specific pathological changes secondary to multiple AR episodes. LEW rats were transplanted with BN rat hind limbs and treated as follows: Group 1 (Iso): isografts. Group 2 (CsA): Cyclosporine (CsA) qd; Group 3 (mult AR): CsA and dexamethasone only when AR was observed. No AR was observed in Groups 1 and 2. Multiple AR were observed in Group 3, and each episode was completely reversed (clinically) with pulsed CsA + dexamethasone treatment. Group 3 animals demonstrated significant vascular lesions along with skin and muscle atrophy, upregulation of profibrotic gene expression and fibrosis when compared to Groups 1 and 2. In addition, allograft bone was sclerotic, weak and prone to malunion and nonunion. Interestingly, vasculopathy was a late finding, whereas muscle atrophy with macrophage infiltration was seen early, after only a few AR episodes. Taken together, multiple AR episodes lead to vasculopathy and tissue-specific pathology in CTA. This is the first evidence of 'composite tissue vasculopathy and degeneration (CTVD)' in CTA.

Immunosuppression and rejection in human hand transplantation.

Avoidance or at least minimization of maintenance immunosuppression represents the key step for promoting wider applicability of reconstructive transplantation. Understanding the mechanisms of composite tissue allograft rejection is essential in working toward that goal. We herein review the current knowledge on acute rejection in reconstructive transplantation and discuss findings in the light of novel immunosuppressive and immunomodulatory strategies.

Alemtuzumab: key for minimization of maintenance immunosuppression in reconstructive transplantation?

Minimization of immunosuppression has become the key effort in solid organ transplantation. Alemtuzumab, the humanized CD-52 monoclonal antibody, is an effective depleting agent increasingly used in transplantation trials. In this article, we summarize the current experience with alemtuzumab use in hand transplantation and discuss its role in current and future approaches toward minimization of maintenance immunosuppression in reconstructive transplantation.

Atypical acute rejection after hand transplantation.

Skin rejection after hand transplantation is characterized by a maculopapular erythematous rash that may be diffuse, patchy or focal, and distributed over forearms and dorsum of the hands. This 'classical' pattern of rejection usually spares the skin of the palm and does not affect the nails. Herein, we report the experience on four cases presenting with an 'atypical' pattern of rejection that is novel in involving the palmar skin and the nails. All patients were young and exposed to repetitive and persistent mechanical stress of the palm. Characteristic features of rejection included a desquamative rash associated with dry skin, red papules, scaling and lichenification localized to the palm. Skin lesions were associated with nail dystrophy, degeneration, deformation or loss. Histology of the skin and nail bed revealed a lymphocytic infiltrate with predominance of T cells (CD3+, CD4+ and CD8+), with small numbers of B cells (CD20+ and CD79a+) and a low number of Forkhead transcription factor 3 (FOXP3)-positive cells in one patient. The lesions persisted over weeks to months, responded poorly to steroid treatment and were managed with antithymocyte globulin (ATG; Thymoglobulin, Genzyme, Cambridge, MA), alemtuzumab and/or intensified maintenance immunosuppression.

Sensitization and desensitization of burn patients as potential candidates for vascularized composite allotransplantation.

Sensitization describes the acquired ability of the immune system to react to foreign human leukocyte antigens (HLA) by producing antibodies and developing memory cells. In the field of transplantation, recipient preformed HLA antibodies due to previous sensitization have been identified - beneath ABO incompatibility - as a major factor for acute graft rejection. Several reasons for sensitization have largely been studied, such as previous blood transfusions, pregnancies or former transplants. Recent studies indicate that the use of assist devices (e.g. ECMO) or cadaveric skin allotransplantation providing temporary coverage in burn patients may lead to additional sensitization. As vascularized composite allotransplantation (VCA) has become a rapidly advancing therapeutic option for reconstruction of complex tissue defects in burns, it seems even more important to become familiar with immunological principles and to be cautiously aware of both sources of sensitization and therapeutic concepts in burns avoiding sensitization. This may also include emergency VCAs in burn patients as potential strategy for early definitive reconstruction avoiding procedures triggering HLA antibody formation. We hereby provide an overview on current evidence in the field of pre- and peritransplant sensitization, followed by posttransplant strategies of desensitization and their potential impact on future treatments of burn patients.

Biologics and donor bone marrow cells for targeted immunomodulation in vascularized composite allotransplantation: a translational trial in swine.

INTRODUCTION: Bone marrow (BM) infusion following organ transplantation is a prerequisite for potential donor-antigen-specific tolerance induction. We developed a preclinical swine model to determine the optimal dose of BM cells to achieve microchimerism. Furthermore, induction therapy was optimized by augmenting the BM infusion with biologics in the form of costimulatory blockade: cytotoxic T-lymphocyte antigen 4 immunoglobulin (CTLA4-Ig). MATERIALS AND METHODS: Yucatan miniature swine (n = 12) underwent total body and thymic irradiation for cytodepletion. Animal groups received 15, 30, or 60 million cells per kilogram of whole unmodified BM. The optimal dose of BM cell infusion (BMT) was then applied to subsequent experiments evaluating the addition of CTLA4lg. Group 1 (control) received no treatment. Group 2 received FK506 only; group 3 received irradiation, BMT, and FK506; group 4 received FK506 and CTLA4-lg. RESULTS: Microchimerism was established in all animals after BM cell infusion; at postoperative day 9, it was significantly increased for 60 million cells per kilogram (P = .0001). Transplanted animals in group 1 rejected the allograft 5 to 8 days after transplantation. Group 2 rejected the allograft (skin and muscle) 30 to 32 days after transplantation (2 days after cessation of immunosuppression). Group 3 rejected the skin portion of the allograft at 50, 52, and 53 days posttransplant. Remaining allograft components (muscle, bone, nerve, vessel) survived indefinitely. Group 4 animals demonstrated significantly prolonged muscle survival beyond 150 days posttransplant; the skin component survived past 150 days in two of three animals. Skin and muscle histology in all long-term surviving animals were normal. CONCLUSIONS: BM cell infusion with 60 million cells per kilogram results in stable levels of microchimerism. The addition of costimulatory blockade (CTLA4lg) prolonged allograft skin survival and overall graft survival. Such targeted immunomodulatory protocols might facilitate immune tolerance and eliminate the need for multidrug immunosuppression to maintain graft survival after vascularized composite allotransplantation.

Immunologic aspects and rejection in solid organ versus reconstructive transplantation.

The immunosuppressive medications developed over the past 3 decades have paved the way for solid organ transplantation to become the treatment of choice for end-stage organ failure. At the end of the century, composite tissue transplantation in humans was performed with success using the same immunosuppressive medications and therapeutic principles. A decade later, experience from >100 cases of reconstructive transplantation have increased the knowledge, changed the view, and affected the therapeutic principles in this novel field. We herein portray the evolution of this novel type of transplant with particular reference to immunologic aspects, particularly differences between reconstructive and solid organ transplantation.

Immunosuppressive agents in transplantation: mechanisms of action and current anti-rejection strategies.

Over the past century, the concept of interfering with the immune response at various sites by blocking the formation, stimulation, proliferation, and differentiation of lymphocytes has led to relentless development of new immunosuppressive drugs. These agents are associated with reduced risk of short- and long-term toxicity and have dramatically improved allograft and patient survival, especially in recipients of solid organ transplants. Current protocols in such patients are nearly all calcineurin-inhibitor based, using cyclosporine or tacrolimus, as part of dual, triple, or sequential therapy. This review focuses on agents currently in clinical use at transplant centers in United States. The drugs are described in terms of their basic mechanisms of action, therapeutic uses, clinical studies, and adverse effects. In addition, the efficacy and toxicity of a few promising new therapeutic approaches are examined. Finally, important challenges regarding pharmacological immunosuppression as it relates to solid organ and composite tissue allotransplantation are discussed.

Mixed allogeneic chimerism and tolerance to composite tissue allografts.

The development of effective immunosuppressive drugs has made solid organ allotransplantation the preferred approach for treatment of end-organ failure. The benefits of these immunosuppressants outweigh their risks in preventing rejection of lifesaving solid-organ allografts. On the contrary, composite tissue allotransplants are non-lifesaving and whether the risks of immunosuppressants justify their benefits is a subject of debate. Hence, composite tissue allografts (CTA) have not enjoyed widespread clinical application for reconstruction of large tissue defects. Therefore, a method of preventing rejection that would eliminate the need for toxic immunosuppressants is of particular importance in CTA. Bone marrow transplantation (BMT) to establish mixed chimerism induces tolerance to a variety of allografts in animal models. This article reviews mixed chimerism-based tolerance protocols. Their limitations and their relevance to CTA are discussed, highlighting some unique characteristics (high antigenicity and the presence of active bone marrow) that make CTAs different from solid organ allografts.