International consensus recommendations on face transplantation: A 2-step Delphi study.

Face transplantation is a viable reconstructive approach for severe craniofacial defects. Despite the evolution witnessed in the field, ethical aspects, clinical and psychosocial implications, public perception, and economic sustainability remain the subject of debate and unanswered questions. Furthermore, poor data reporting and sharing, the absence of standardized metrics for outcome evaluation, and the lack of consensus definitions of success and failure have hampered the development of a "transplantation culture" on a global scale. We completed a 2-round online modified Delphi process with 35 international face transplant stakeholders, including surgeons, clinicians, psychologists, psychiatrists, ethicists, policymakers, and researchers, with a representation of 10 of the 19 face transplant teams that had already performed the procedure and 73% of face transplants. Themes addressed included patient assessment and selection, indications, social support networks, clinical framework, surgical considerations, data on patient progress and outcomes, definitions of success and failure, public image and perception, and financial sustainability. The presented recommendations are the product of a shared commitment of face transplant teams to foster the development of face transplantation and are aimed at providing a gold standard of practice and policy.

A World Update of Progress in Lower Extremity Transplantation: What's Hot and What's Not.

ABSTRACT: The field of vascularized composite allotransplantation (VCA) is the new frontier of solid organ transplantation (SOT). VCA spans life-enhancing/life-changing procedures such as upper extremity, craniofacial (including eye), laryngeal, tracheal, abdominal wall, penis, and lower extremity transplants. VCAs such as uterus transplants are life giving unlike any other SOT. Of all VCAs that have shown successful intermediate- to long-term graft survival with functional and immunologic outcomes, lower extremity VCAs have remained largely underexplored. Lower extremity transplantation (LET) can offer patients with improved function compared to the use of conventional prostheses, reducing concerns of phantom limb pain and stump complications, and offer an option for eligible amputees that either fail prosthetic rehabilitation or do not adapt to prosthetics. Nevertheless, these benefits must be carefully weighed against the risks of VCA, which are not trivial, including the adverse effects of lifelong immunosuppression, extremely challenging perioperative care, and delayed nerve regeneration. There have been 5 lower extremity transplants to date, ranging from unilateral or bilateral to quadrimembral, progressively increasing in risk that resulted in fatalities in 3 of the 5 cases, emphasizing the inherent risks. The advantages of LET over prosthetics must be carefully weighed, demanding rigorous candidate selection for optimal outcomes.

Composite tissue xenopreservation: Preliminary results of staged VCA in rat to mouse model.

BACKGROUND: The time between procurement and transplantation of composite tissues, especially regarding the limited donor pool, is a challenge effecting the outcomes of the transplantation. Current preservation techniques mainly include either cold preservation with a solution or machine perfusion using blood or certain oxygen-carrying solutions. However, none enables preservation beyond 24 h. Increasing this time to several days will provide better usage of the donor pool, safer transplantation of VCA with significant muscle content, and gives time to stabilize a patient before long surgical procedures. Herein, we described a novel strategy of xenopreservation (preservation via xenotransplantation) to preserve composite tissues for 7 days, followed by staged transplantation. MATERIALS AND METHODS: We used two concordant species, female Sprague Dawley rats (n = 10) and female CF-1 mice (n = 10) in this study. Four of pair of animals are used for anatomical study. The groin flap of the rat was used as a xenograft and xenotransplanted to the neck area of the carrier mouse. Cyclosporine (CsA) was administered used as immunosuppressant. After 7 days of preservation on the mouse neck, xenotransplanted groin flap (called xenopreserved flap) was re-harvested, skin and vessels samples were collected for histopathological evaluation, and the xenopreserved flap was transplanted to the donor rat's opposite groin area. Anastomoses were performed between the flap's pedicle and the femoral vessels. Clinical observation regarding inflammation and tissue perfusion of the xenopreserved flap was monitored daily. Fifteen days after the second surgical procedure, the rats were euthanized, and skin and vessel samples were collected. Histologic evaluation, including inflammatory cell numbers, was performed. Wilcoxon test was used to compare the changes in inflammation severity and p < .05 was set for statistical significance. RESULTS: All xenopreserved groin flaps except one survived. Mean lymphocyte count before the second operation (at the end of the xenopreservation procedure) was 20,22 ± 0.44 and reduced to 13,14 ± 0.47 at the end of 15 days, and the difference was statistically significant (p < .05). CONCLUSION: This proof-of-concept study with preliminary results showed that xenotransplantation might be a novel strategy for preservation of VCA for a certain period of time. However, additional translational studies are needed to modulate the tissue changes following xenopreservation.

Tacrolimus-Eluting Disk within the Allograft Enables Vascularized Composite Allograft Survival with Site-Specific Immunosuppression without Systemic Toxicity.

AIM: Widespread clinical application of vascularized composite allotransplantation (VCA) has been limited by the need for lifelong systemic immunosuppression to prevent rejection. Our goal was to develop a site-specific immunosuppressive strategy that promotes VCA allograft survival and minimizes the risk of systemic side effects. METHODS: Tacrolimus loaded polycaprolactone (TAC-PCL) disks were prepared and tested for their efficacy in sustaining VCA allograft survival via site-specific immunosuppression. Brown Norway-to-Lewis rat hind limb transplantations were performed; animals received one TAC disk either in the transplanted (DTx) or in the contralateral non-transplanted (DnonTx) limbs. In another group, animals received DTx and lymphadenectomy on Tx side. Blood and allograft levels of TAC were measured using LC-MS/MS. Systemic toxicity was evaluated. RESULTS: Animals that received DTx achieved long-term allograft survival (> 200 days) without signs of metabolic and infectious complications. In these animals, TAC blood levels were low but stable between 2 to 5 ng/mL for nearly 100 days. High concentrations of TAC were achieved in the allografts and the draining lymph nodes (DLN). Animals that underwent lymphadenectomy rejected their allograft by 175 days. Animals that received DnonTx rejected their allografts by day 70. CONCLUSION: Controlled delivery of TAC directly within the allograft (with a single TAC disk) effectively inhibits rejection and prolongs VCA allograft survival, while mitigating the complications of systemic immunosuppression. There was a survival benefit of delivering TAC within the allograft as compared to a remote site. We believe this approach of local drug delivery has significant implications for drug administration in transplantation.

Treg-inducing microparticles promote donor-specific tolerance in experimental vascularized composite allotransplantation.

For individuals who sustain devastating composite tissue loss, vascularized composite allotransplantation (VCA; e.g., hand and face transplantation) has the potential to restore appearance and function of the damaged tissues. As with solid organ transplantation, however, rejection must be controlled by multidrug systemic immunosuppression with substantial side effects. As an alternative therapeutic approach inspired by natural mechanisms the body uses to control inflammation, we developed a system to enrich regulatory T cells (Tregs) in an allograft. Microparticles were engineered to sustainably release TGF-ß1, IL-2, and rapamycin, to induce Treg differentiation from naïve T cells. In a rat hindlimb VCA model, local administration of this Treg-inducing system, referred to as TRI-MP, prolonged allograft survival indefinitely without long-term systemic immunosuppression. TRI-MP treatment reduced expression of inflammatory mediators and enhanced expression of Treg-associated cytokines in allograft tissue. TRI-MP also enriched Treg and reduced inflammatory Th1 populations in allograft draining lymph nodes. This local immunotherapy imparted systemic donor-specific tolerance in otherwise immunocompetent rats, as evidenced by acceptance of secondary skin grafts from the hindlimb donor strain and rejection of skin grafts from a third-party donor strain. Ultimately, this therapeutic approach may reduce, or even eliminate, the need for systemic immunosuppression in VCA or solid organ transplantation.

Adipose-derived stromal cell therapy combined with a short course nonmyeloablative conditioning promotes long-term graft tolerance in vascularized composite allotransplantation.

The risks of chronic immunosuppression limit the utility of vascularized composite allotransplantation (VCA) as a reconstructive option in complex tissue defects. We evaluated a novel, clinically translatable, radiation-free conditioning protocol that combines anti-lymphocyte serum (ALS), tacrolimus, and cytotoxic T-lymphocyte-associated protein 4 immunoglobulin (CTLA4-Ig) with adipose-derived stromal cells (ASCs) to allow VCA survival without long-term systemic immunosuppression. Full-mismatched rat hind-limb-transplant recipients received tacrolimus (0.5 mg/kg) for 14 days and were assigned to 4 groups: controls (CTRL) received no conditioning; ASC-group received CTLA4-Ig (10 mg/kg body weight i.p. postoperative day [POD] 2, 4, 7) and donor ASCs (1 × 106 iv, POD 2, 4, 7, 15, 28); the ASC-cyclophosphamide (CYP)-group received CTLA4-Ig, ASC plus cyclophosphamide (50 mg/kg ip, POD 3); the ASC-ALS-group received CTLA4-Ig, ASCs plus ALS (500 µL ip, POD 1, 5). Banff grade III or 120 days were endpoints. ASCs suppressed alloresponse in vitro. Median rejection-free VCA survival was 28 days in CTRL (n = 7), 34 in ASC (n = 6), and 27.5 in ASC-CYP (n = 4). In contrast, ASC-ALS achieved significantly longer, rejection-free VCA survival in 6/7 animals (86%), with persistent mixed donor-cell chimerism, and elevated systemic and allograft skin Tregs , with no signs of acute cellular rejection. Taken together, a regimen comprised of short-course tacrolimus, repeated CTLA4-Ig and ASC administration, combined with ALS, promotes long-term VCA survival without chronic immunosuppression.

Heterotopic Transplantation of Allogeneic Vertical Rectus Abdominis Myocutaneous Flaps in Miniature Swine.

BACKGROUND: Vascularized composite tissue allotransplantation (VCA) opens new possibilities for reconstruction of complex tissue defects, including upper extremity and facial transplantation. The main challenges in VCA transplantation are the side effects of long-term immunosuppression and chronic graft rejection. Translational preclinical animal models are crucial for VCA research to improve clinical outcomes and to study underlying immunologic mechanisms. Herein, we describe a novel, large animal, non-bone-bearing VCA model in inbred, swine leukocyte antigen-typed miniature swine. METHODS: Transplantation of vertical rectus abdominis myocutaneous (VRAM) flaps was performed between fully swine leukocyte antigen-mismatched miniature swine. The flaps were transferred to the posterolateral aspect of the neck of recipients and anastomosed to the common carotid artery and internal jugular vein. Different immunosuppressive drug regimens were used. Clinical graft evaluation was performed daily, and punch biopsies were taken for histology. RESULTS: Ten VRAM transplants were performed. The mean ischemia time was 89.4 min (SD ± 47), mean pedicle length 7.5 cm (SD ± 2), mean venous diameter 2.5 mm (SD ± 0.4), and mean arterial diameter 2.2 mm (SD ± 0.3). Follow-up demonstrated good correlation between clinical appearance and progression of graft rejection confirmed by histologic assessment. Complications were intraoperative cardiac arrest in one recipient and one flap loss due to venous compromise. CONCLUSIONS: VRAM transplantation in miniature swine is an appropriate preclinical VCA model, with the advantage of good clinical and histologic correlation during the course of rejection, as well as easy access to the graft. The availability of inbred, haplotyped animals allows studies across different major histocompatibility complex barriers in a non-bone-bearing VCA.

Pharmacokinetics and Biodistribution of Tacrolimus after Topical Administration: Implications for Vascularized Composite Allotransplantation.

AIM: The high doses of oral tacrolimus (TAC) (1,2) necessary to prevent acute rejection (AR) after vascularized composite allotransplantation (VCA) are associated with systemic adverse effects. The skin is the most antigenic tissue in VCA and the primary target of AR. However, the short-term use of topical TAC (Protopic®), as an off-label adjunct to oral TAC, to treat AR episodes pro re nata (PRN), has yielded inconsistent results. There is lack of data on the pharmacokinetics and tissue distribution of topical TAC in VCA, that hampers our understanding of the reasons for unreliable efficacy. Toward this goal, we evaluated the ability of topical TAC to achieve high local tissue concentrations at the site of application with low systemic concentrations. MATERIALS AND METHODS: We assessed the pharmacokinetics and tissue distribution of topical TAC (Protopic®, 0.03%) after single or repeated topical application in comparison to those after systemic delivery in rats. Animals received a single topical application of TAC ointment (Group 1) or an intravenous (IV) injection of TAC (Group 2) at a dose of 0.5 mg/kg. In another experiment, animals received daily topical application of TAC ointment (Group 3), or daily intraperitoneal (IP) injection of TAC (Group 4) at a dose of 0.5 mg/kg for 7 days. TAC concentrations in blood and tissues were analyzed by Liquid Chromatography-Mass Spectrometry (LC/MS-MS). RESULTS: Following single topical administration, TAC was absorbed slowly with a Tmax of 4 h and an absolute bioavailability of 11%. The concentrations of TAC in skin and muscle were several folds higher than whole blood concentrations. Systemic levels remained subtherapeutic (< 3 ng/ml) with repeated once daily applications. CONCLUSION: Topical application of TAC ointment (Protopic®, 0.03%) at a dose of 0.5 mg/kg/day provided high concentrations in the local tissues with low systemic exposure. Repeated topical administration of TAC is well tolerated with no local or systemic adverse effects. This study confirms the feasibility of topical application of TAC for site specific graft immunosuppression and enables future applications in VCA.

Vascularized composite allotransplantation in the United States: A descriptive analysis of the Organ Procurement and Transplantation Network Data.

On July 3, 2014, the Organ Procurement and Transplantation Network/United Network for Organ Sharing was charged with the oversight of vascularized composite allograft (VCA) procurement and transplantation in the United States. As of December 31, 2017, 61 VCA programs at 27 centers were approved in the United States. Fifty candidates have been added to the waiting list at 15 centers. Twenty-eight VCA transplants have been performed at 14 programs (10 upper limb, 10 uterus, 5 craniofacial, 1 scalp, 1 abdominal wall, and 1 penile). Twenty-two VCAs were procured from 21 deceased donors, resulting in 109 non-VCA organs transplanted (15 hearts, 3 intestine, 40 kidney, 20 livers, 24 lungs, and 7 pancreata). Six uterus transplants were performed from living donors. Fourteen candidates were still waiting at 9 centers on December 31, 2017. Two of the 10 uterus recipients had live births and 3 still had viable grafts. Seventeen of 18 nonuterus recipients had functioning grafts. At present, VCA is an emerging field with a small number of patients transplanted. Data on posttransplant survival and functional outcomes continue to be collected to further the understanding of this complex and evolving field. Further systematic data are important for policy refinement and assurance of patient safety.

Composite Graft Pretreatment With Hydrogen Sulfide Delays the Onset of Acute Rejection.

INTRODUCTION: Vascularized composite allotransplantation can reconstruct devastating tissue loss by replacing like-with-like tissues, most commonly in the form of hand or face transplantation. Unresolved technical and ethical challenges have meant that such transplants remain experimental treatments. The most significant barrier to expansion of this field is the requirement for systemic immunosuppression, its toxicity and effect on longevity.Hydrogen sulfide (H2S) has been shown experimentally to ameliorate the ischemia reperfusion injury associated with composite tissue autotransplantation, which has been linked to acute rejection in solid organ transplantation. In this protocol, a large-animal model was used to evaluate the effect of H2S on acute rejection after composite tissue allotransplantation. MATERIALS AND METHODS: A musculocutaneous flap model in SLA-mismatched swine was used to evaluate acute rejection of allotransplants in 2 groups: control animals (n = 8) and a treatment group in which the allografts were pretreated with hydrogen sulfide (n = 8). Neither group was treated with systemic immunosuppression. Acute rejection was graded clinically and histopathologically by an independent, blinded pathologist. Data were analyzed by t tests with correction for multiple comparisons by the Holm-Sídák method. RESULTS: Clinically, H2S-treated tissue composites showed a delay in the onset of rejection that was statistically significant from postoperative day 6. Histopathologically, this difference between groups was also apparent, although evidence of a difference in groups disappeared beyond day 10. CONCLUSIONS: Targeted hydrogen sulfide treatment of vascularized composite allografts immediately before transplantation can delay acute rejection. This may, in turn, reduce or obviate the requirement for systemic immunosuppression.

Adipogenesis for soft tissue reconstruction.

PURPOSE OF REVIEW: It has been increasingly common to use adipose tissue for regenerative and reconstructive purposes. Applications of autologous fat transfer and different stem cell therapies have significant limitations and adipose tissue engineering may have the potential to be an important strategy in the reconstruction of large tissue defects. A better understanding of adipogenesis will help to develop strategies to make adipose tissue more effective for repairing volumetric defects. RECENT FINDINGS: We provide an overview of the current applications of adipose tissue transfer and cellular therapy methods for soft tissue reconstruction, cellular physiology, and factors influencing adipogenesis, and adipose tissue engineering. Furthermore, we discuss mechanical properties and vascularization strategies of engineered adipose tissue, and its potential applications in the clinical settings. SUMMARY: Autologous fat tissue transfer is the standard of care technique for the majority of surgeons; however, high resorption rates, poor perfusion within a large volume fat graft and widely inconsistent graft survival are the main limitations. Adipose tissue engineering is a promising field to reach the first goal of producing adipose tissue which has more predictable survival and higher graft retention rates. Advancements of scaffold and vascularization strategies will contribute to metabolically and functionally more relevant adipose tissue engineering.

Immunological considerations and concerns as pertinent to whole eye transplantation.

PURPOSE OF REVIEW: The advent of clinical vascularized composite allotransplantation (VCA), offers hope for whole eye transplantation (WET) in patients with devastating vison loss that fails or defies current treatment options. Optic nerve regeneration and reintegration remain the overarching hurdles to WET. However, the realization of WET may indeed be limited by our lack of understanding of the singular immunological features of the eye as pertinent to graft survival and functional vision restoration in the setting of transplantation. RECENT FINDINGS: Like other VCA, such as the hand or face, the eye includes multiple tissues with distinct embryonic lineage and differential antigenicity. The ultimate goal of vision restoration through WET requires optimal immune modulation of the graft for successful optic nerve regeneration. Our team is exploring barriers to our understanding of the immunology of the eye in the context of WET including the role of immune privilege and lymphatic drainage on rejection, as well as the effects ischemia, reperfusion injury and rejection on optic nerve regeneration. SUMMARY: Elucidation of the unique immunological responses in the eye and adnexa after WET will provide foundational clues that will help inform therapies that prevent immune rejection without hindering optic nerve regeneration or reintegration.

Blood product utilization in human upper-extremity transplantation: challenges, complications, considerations, and transfusion protocol conception.

BACKGROUND: Upper-extremity transplantation (UET) is a reality. Immunologic, functional, and graft survival outcomes have been encouraging. However, these complex reconstructions have unique considerations that pose distinct challenges. Transplant programs have reported morbidity and mortality due to significant intraoperative blood losses, but similar data are scant during other phases of recovery. We report experience from two centers on complete blood component demands and utilization with UET. STUDY DESIGN AND METHODS: Inpatient medical records of UET recipients from intraoperative (time from initiation of transplant surgery to exit from the operative suite) and postoperative (exit from the operative suite to discharge from the hospital) phases were retrospectively reviewed. RESULTS: Six patients received various UETs and mean (±SD) postoperative hospital stay was 46 (±14.4) days. Mean (±SD) intraoperative blood unit utilization was 14.8 (±10.2) red blood cells (RBCs), 10.5 (±11.8) plasma, 0.8 (±1.2) platelets (PLTs), and 0.3 (±0.8) cryoprecipitate units. Mean postoperative blood unit utilization was 9.3 (±10.4) RBCs, 5.3 (±6.7) plasma, 1.2 (±2.0) PLTs, and 0.7 (±1.6) cryoprecipitate units. Both intraoperative and postoperative blood utilization for unilateral versus bilateral transplant were different, but not significantly so. However, total inpatient blood use in bilateral transplants was significantly greater than in unilateral transplants. CONCLUSION: Substantial blood loss may occur in UET and require transfusion of many blood components, primarily RBCs and plasma. We propose an UET transfusion protocol and suggest that centers preparing to perform these transplants should actively engage the transfusion medicine service to ensure availability and access to appropriate blood components for the entire hospitalizations of these unique patients.

Ultra-high-field RF coil development for evaluating upper extremity imaging applications.

The purpose of this study is to develop and evaluate a custom-designed 7  T MRI coil and explore its use for upper extremity applications. An RF system composed of a transverse electromagnetic transmit coil and an eight-channel receive-only array was developed for 7  T upper extremity applications. The RF system was characterized and evaluated using scattering parameters and B1+ mapping. Finite difference time domain simulations were performed to evaluate the B1+ field distribution and specific absorption rate for the forearm region of the upper extremity. High-resolution 7  T images were acquired and compared with those at 3 T. The simulation and experimental results show very good B1+ field homogeneity across the forearm. High-resolution images of musculotendinous, osseocartilaginous, and neurovascular structures in the upper extremity are presented with T1 volumetric interpolated breath-hold examination, T2 double-echo steady state, T2 * susceptibility weighted imaging (SWI), diffusion tensor imaging, and time-of-flight sequences. Comparison between 3  T and 7  T is shown. Intricate contextual anatomy can be delineated in synovial, fibrocartilaginous, interosseous, and intraosseous trabecular structures of the forearm, as well as palmar and digital vascular anatomy (including microvascular detail in SWI). Ultra-high-field 7  T imaging holds great potential in improving the sensitivity and specificity of upper extremity imaging, especially in wrist and hand pathology secondary to bone, ligament, nerve, vascular, and other soft or hard tissue etiology.

Premise and promise of mesenchymal stem cell-based therapies in clinical vascularized composite allotransplantation.

PURPOSE OF REVIEW: Over the past decade, clinical vascularized composite allotransplantation (VCA) has enabled functional and quality of life restoration in a wide range of indications secondary to devastating tissue loss. However, the spectre of toxicity and long-term complications of chronic immunosuppression has curtailed the momentum of VCA. This study summarizes the literature evidence behind successful mesenchymal stem cell (MSC)-based cell therapies highlighting their multipronged immunomodulatory, restorative and regenerative characteristics with special emphasis towards VCA applications. RECENT FINDINGS: Experimental and clinical studies in solid organs and VCA have confirmed that MSCs facilitate immunosuppression-free allograft survival or tolerance, stimulate peripheral nerve regeneration, attenuate ischaemia-reperfusion injury, and improve tissue healing after surgery. It has been hypothesized that MSC-induced long-term operational tolerance in experimental VCA is mediated by induction of mixed donor-specific chimerism and regulatory T-cell mechanisms. All these characteristics of MSCs could thus help expand the scope and clinical feasibility of VCA. SUMMARY: Cellular therapies, especially those focusing on MSCs, are emerging in solid organ transplantation including VCA. Although some clinical trials have begun to assess the effects of MSCs in solid organ transplantation, much scientific domain remains uncharted, especially for VCA.

Topical Tacrolimus and Mycophenolic Acid Therapy Synergizes with Low Dose Systemic Immunosuppression to Sustain Vascularized Composite Allograft Survival.

AIM: The goal of this study was to evaluate whether topical administration of tacrolimus (TAC) and mycophenolic acid (MPA) at the transplant site enables vascularized composite allograft (VCA) survival with significant minimization of the dose and adverse effects of systemic TAC (STAC) immunosuppression. MATERIALS AND METHODS: Lewis (Lew) rats received orthotopic hind limb allotransplants from fully mismatched Brown Norway (BN) donors. Group 1 (Controls) received no treatment. Other groups were treated with STAC at a dose of 1 mg/kg/day for 7 days. On post-operative day (POD) 8, the STAC dose was dropped to 0.1 mg/kg/day for Group 2 and maintained at 1 mg/kg for Group 3. Group 4 received topical application of TAC and MPA on the transplanted (Tx) limb starting POD 8 without STAC. Group 5 received topical TAC and MPA on the contralateral non-Tx limb and Group 6 received topical TAC and MPA on the Tx limb starting POD 8 along with low dose STAC (0.1 mg/kg/day). Treatment was continued until the study end point was reached, defined as either grade 3 rejection or allograft survival exceeding 100 days. .We conducted sequential LC-MS/MS measurements to assess TAC and MPA concentrations in both blood/plasma and allograft tissues. Additionally, we evaluated markers indicative of organ toxicity associated with STAC immunosuppression. RESULTS: Compared to controls, topical therapy with TAC+MPA significantly prolonged allograft survival beyond 100 daysat very low dose STAC (0.1 mg/kg/day) (Group 6). The histopathological assessment of the grafts was consistent with the clinical outcomes. .Drug levels in blood/plasma remained low or undetectable, while allograft tissues showed higher drug concentrations compared to contralateral limb tissues (P<0.05). . Urinary creatinine clearance remained within the normal range at 2.5 mL/min. CONCLUSION: Combination therapy with topical TAC and MPA synergizes with a very low dose, corticosteroid- free-STAC regimen and facilitates rejection-free, prolonged VCA survival without morbidity.

Upper-extremity transplantation using a cell-based protocol to minimize immunosuppression.

OBJECTIVE: To minimize maintenance immunosuppression in upper-extremity transplantation to favor the risk-benefit balance of this procedure. BACKGROUND: Despite favorable outcomes, broad clinical application of reconstructive transplantation is limited by the risks and side effects of multidrug immunosuppression. We present our experience with upper-extremity transplantation under a novel, donor bone marrow (BM) cell-based treatment protocol ("Pittsburgh protocol"). METHODS: Between March 2009 and September 2010, 5 patients received a bilateral hand (n = 2), a bilateral hand/forearm (n = 1), or a unilateral (n = 2) hand transplant. Patients were treated with alemtuzumab and methylprednisolone for induction, followed by tacrolimus monotherapy. On day 14, patients received an infusion of donor BM cells isolated from 9 vertebral bodies. Comprehensive follow-up included functional evaluation, imaging, and immunomonitoring. RESULTS: All patients are maintained on tacrolimus monotherapy with trough levels ranging between 4 and 12 ng/mL. Skin rejections were infrequent and reversible. Patients demonstrated sustained improvements in motor function and sensory return correlating with time after transplantation and level of amputation. Side effects included transient increase in serum creatinine, hyperglycemia managed with oral hypoglycemics, minor wound infection, and hyperuricemia but no infections. Immunomonitoring revealed transient moderate levels of donor-specific antibodies, adequate immunocompetence, and no peripheral blood chimerism. Imaging demonstrated patent vessels with only mild luminal narrowing/occlusion in 1 case. Protocol skin biopsies showed absent or minimal perivascular cellular infiltrates. CONCLUSIONS: Our data suggest that this BM cell-based treatment protocol is safe, is well tolerated, and allows upper-extremity transplantation using low-dose tacrolimus monotherapy.

Site-specific immunosuppression in vascularized composite allotransplantation: prospects and potential.

Skin is the most immunogenic component of a vascularized composite allograft (VCA) and is the primary trigger and target of rejection. The skin is directly accessible for visual monitoring of acute rejection (AR) and for directed biopsy, timely therapeutic intervention, and management of AR. Logically, antirejection drugs, biologics, or other agents delivered locally to the VCA may reduce the need for systemic immunosuppression with its adverse effects. Topical FK 506 (tacrolimus) and steroids have been used in clinical VCA as an adjunct to systemic therapy with unclear beneficial effects. However, there are no commercially available topical formulations for other widely used systemic immunosuppressive drugs such as mycophenolic acid, sirolimus, and everolimus. Investigating the site-specific therapeutic effects and efficacy of systemically active agents may enable optimizing the dosing, frequency, and duration of overall immunosuppression in VCA with minimization or elimination of long-term drug-related toxicity.

Donor derived hematopoietic stem cell niche transplantation facilitates mixed chimerism mediated donor specific tolerance.

Compelling experimental evidence confirms that the robustness and longevity of mixed chimerism (MC) relies on the persistence and availability of donor-derived hematopoietic stem cell (HSC) niches in recipients. Based on our prior work in rodent vascularized composite allotransplantation (VCA) models, we hypothesize that the vascularized bone components in VCA bearing donor HSC niches, thus may provide a unique biologic opportunity to facilitate stable MC and transplant tolerance. In this study, by utilizing a series of rodent VCA models we demonstrated that donor HSC niches in the vascularized bone facilitate persistent multilineage hematopoietic chimerism in transplant recipients and promote donor-specific tolerance without harsh myeloablation. In addition, the transplanted donor HSC niches in VCA facilitated the donor HSC niches seeding to the recipient bone marrow compartment and contributed to the maintenance and homeostasis of stable MC. Moreover, this study provided evidences that chimeric thymus plays a role in MC-mediated transplant tolerance through a mechanism of thymic central deletion. Mechanistic insights from our study could lead to the use of vascularized donor bone with pre-engrafted HSC niches as a safe, complementary strategy to induce robust and stable MC-mediated tolerance in VCA or solid organ transplantation recipients.

A Reversibly Thermoresponsive, Theranostic Nanoemulgel for Tacrolimus Delivery to Activated Macrophages: Formulation and In Vitro Validation.

Despite long-term immunosuppression, organ transplant recipients face the risk of immune rejection and graft loss. Tacrolimus (TAC, FK506, Prograf®) is an FDA-approved keystone immunosuppressant for preventing transplant rejection. However, it undergoes extensive first-pass metabolism and has a narrow therapeutic window, which leads to erratic bioavailability and toxicity. Local delivery of TAC directly into the graft, instead of systemic delivery, can improve safety, efficacy, and tolerability. Macrophages have emerged as promising therapeutic targets as their increased levels correlate with an increased risk of organ rejection and a poor prognosis post-transplantation. Here, we present a locally injectable drug delivery platform for macrophages, where TAC is incorporated into a colloidally stable nanoemulsion and then formulated as a reversibly thermoresponsive, pluronic-based nanoemulgel (NEG). This novel formulation is designed to undergo a sol-to-gel transition at physiological temperature to sustain TAC release in situ at the site of local application. We also show that TAC-NEG mitigates the release of proinflammatory cytokines and nitric oxide from lipopolysaccharide (LPS)-activated macrophages. To the best of our knowledge, this is the first TAC-loaded nanoemulgel with demonstrated anti-inflammatory effects on macrophages in vitro.