OPTN/SRTR 2022 Annual Data Report: Vascularized Composite Allograft.

This year's chapter on vascularized composite allograft (VCA) encompasses reviews of data collected from 2014 (when VCA was included in the Final Rule) through 2022. The present Annual Data Report shows that the number of VCA recipients in the United States continues to be small and has remained consistent from the prior report. The data continue to be limited by sample size, with trends persistently demonstrating a predominance of White males in the young/middle-aged population as both donors and recipients for nonuterus VCA transplants, and White women younger than 35 years as the predominant recipients of uterus transplant. Similar to the 2021 report, there were only eight failed uterus grafts and one failed nonuterus VCA graft reported from 2014 through 2022. Standardization of definitions of success and failure as well as outcome measures for the different VCA types remain unmet needs in VCA transplantation.

Acute Rejection Rates in Vascularized Composite Allografts: A Systematic Review of Case Reports.

INTRODUCTION: Vascularized Composite Allografts (VCA) are usually performed in a full major histocompatibility complex mismatch setting, with a risk of acute rejection depending on factors such as the type of immunosuppression therapy and the quality of graft preservation. In this systematic review, we present the different immunosuppression protocols used in VCA and point out relationships between acute rejection rates and possible factors that might influence it. METHODS: This systematic review was performed according to the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. We systematically searched Medline (PubMed), Embase, and The Cochrane Library between November 2022 and February 2023, using following Mesh Terms: Transplant, Transplantation, Hand, Face, Uterus, Penis, Abdominal Wall, Larynx, and Composite Tissue Allografts. All VCA case reports and reviews describing multiple case reports were included. RESULTS: We discovered 211 VCA cases reported. The preferred treatment was a combination of antithymocyte globulins, mycophenolate mofetil (MMF), tacrolimus, and steroids; and a combination of MMF, tacrolimus, and steroids for induction and maintenance treatment, respectively. Burn patients showed a higher acute rejection rate (P = 0.073) and were administered higher MMF doses (P = 0.020). CONCLUSIONS: In contrast to previous statements, the field of VCA is not rapidly evolving, as it has encountered challenges in addressing immune-related concerns. This is highlighted by the absence of a standardized immunosuppression regimen. Consequently, more substantial data are required to draw more conclusive results regarding the immunogenicity of VCAs and the potential superiority of one immunosuppressive treatment over another. Future efforts should be made to report the VCA surgeries comprehensively, and muti-institutional long-term prospective follow-up studies should be performed to compare the number of acute rejections with influencing factors.

Machine Perfusion Enables 24-h Preservation of Vascularized Composite Allografts in a Swine Model of Allotransplantation.

The current gold standard for preserving vascularized composite allografts (VCA) is 4°C static cold storage (SCS), albeit muscle vulnerability to ischemia can be described as early as after 2 h of SCS. Alternatively, machine perfusion (MP) is growing in the world of organ preservation. Herein, we investigated the outcomes of oxygenated acellular subnormothermic machine perfusion (SNMP) for 24-h VCA preservation before allotransplantation in a swine model. Six partial hindlimbs were procured on adult pigs and preserved ex vivo for 24 h with either SNMP (n = 3) or SCS (n = 3) before heterotopic allotransplantation. Recipient animals received immunosuppression and were followed up for 14 days. Clinical monitoring was carried out twice daily, and graft biopsies and blood samples were regularly collected. Two blinded pathologists assessed skin and muscle samples. Overall survival was higher in the SNMP group. Early euthanasia of 2 animals in the SCS group was linked to significant graft degeneration. Analyses of the grafts showed massive muscle degeneration in the SCS group and a normal aspect in the SNMP group 2 weeks after allotransplantation. Therefore, this 24-h SNMP protocol using a modified Steen solution generated better clinical and histological outcomes in allotransplantation when compared to time-matched SCS.

Sub-zero non-freezing of vascularized composite allografts in a rodent partial hindlimb model.

Ischemia is a major limiting factor in Vascularized Composite Allotransplantation (VCA) as irreversible muscular injury can occur after as early as 4-6 h of static cold storage (SCS). Organ preservation technologies have led to the development of storage protocols extending rat liver ex vivo preservation up to 4 days. Development of such a protocol for VCAs has the added challenge of inherent ice nucleating factors of the graft, therefore, this study focused on developing a robust protocol for VCA supercooling. Rodent partial hindlimbs underwent subnormothermic machine perfusion (SNMP) with several loading solutions, followed by a storage solution with cryoprotective agents (CPA) developed for VCAs. Storage occurred in suspended animation for 24h and VCAs were recovered using SNMP with modified Steen. This study shows a robust VCA supercooling preservation protocol in a rodent model. Further optimization is expected to allow for its application in a transplantation model, which would be a breakthrough in the field of VCA preservation.

Vascularized Composite Allograft Versus Prosthetic for Reconstruction After Facial and Hand Trauma: Comparing Cost, Complications, and Long-term Outcome.

ABSTRACT: In the past decade, vascularized composite allotransplantation (VCA) has become clinical reality for reconstruction after face and hand trauma. It offers patients the unique opportunity to regain form and function in a way that had only been achieved with traditional reconstruction or with the use of prostheses. On the other hand, prostheses for facial and hand reconstruction have continued to evolve over the years and, in many cases, represent the primary option for patients after hand and face trauma. We compared the cost, associated complications, and long-term outcomes of VCA with prostheses for reconstruction of the face and hand/upper extremity. Ultimately, VCA and prostheses represent 2 different reconstructive options with distinct benefit profiles and associated limitations and should ideally not be perceived as competing choices. Our work adds a valuable component to the general framework guiding the decision to offer VCA or prostheses for reconstruction after face and upper extremity trauma.

Significantly Improved Cold Preservation of Rat Hind Limb Vascularized Composite Allografts Using the New PrC-210 Free Radical Scavenger.

Vascularized composite allotransplantation (VCA) represents a promising reconstructive solution primarily conducted to improve quality of life. However, tissue damage caused by cold-ischemia (CI) storage prior to transplant represents a major factor limiting widespread application. This study investigates the addition of the novel free radical scavenger PrC-210 to UW Organ Preservation Solution (UW Solution) to suppress CI-induced skeletal muscle injury in a rat hind limb amputation model. Lewis rats received systemic perfusion of UW solution +/- PrC-210 (0 mM control, 10 mM, 20 mM, 30 mM, or 40 mM), followed by bilateral transfemoral amputation. Limbs were stored in 40 mL of the same perfusate at 4 °C for 48 h. Muscle punch biopsies were taken at set times over the 48 h cold-storage period and analyzed for caspase-3,7 activity, cytochrome C levels, and qualitative histology. A single 15 s perfusion of PrC-210-containing UW Solution conferred a dose-dependent reduction in CI-induced muscle cell death over 48 h. In the presence of PrC-210, muscle cell mitochondrial cytochrome C release was equivalent to 0 h controls, with profound reductions in the caspase-3,7 apoptotic marker that correlated with limb histology. PrC-210 conferred complete prevention of ROS-induced mitochondrial lysis in vitro, as measured by cytochrome C release. We conclude that the addition of 30 mM PrC210 to UW Solution conferred the most consistent reduction in CI limb damage, and it warrants further investigation for clinical application in the VCA setting.

OPTN/SRTR 2021 Annual Data Report: Vascularized Composite Allograft.

Year 2020 marked the first OPTN/SRTR Annual Data Report that included a chapter on vascularized composite allograft (VCA), which encompassed reviews of data collected between 2014 (when VCA was included in the Final Rule) and 2020. The present Annual Data Report shows that the number of VCA recipients in the United States continues to be small and trended downward in 2021. While data continue to be limited by sample size, trends continue to show a predominance in White, young/middle-aged, male recipients. Similar to the 2020 report, eight uterus and one non-uterus VCA graft failures were reported from 2014 through 2021. Critical to advancement of VCA transplantation will be the standardization of definitions, protocols, and outcome measures for the different VCA types. Like intestinal transplants, it is likely that VCA transplants will be concentrated and performed at referral transplant centers.

Insights from immunoproteomic profiling of a rejected full-face transplant.

Vascularized composite allografts (VCAs) of faces and extremities are subject to chronic rejection that is incompletely understood. Here we report on immunoproteomic evaluation of a full facial VCA removed 88 months after transplantation due to chronic rejection. CD8-positive T cells of donor (graft) origin infiltrate deep intragraft arteries in apposition to degenerating endothelium of chimeric recipient origin in association with arteriosclerotic alterations. Digital spatial proteomic profiling highlighted proteins expressed by activated cytotoxic T cells and macrophages as well as pathway components involved in atherogenic responses, including Indoleamine 2,3-Dioxygenase 1 (IDO1) and Stimulator of Interferon Response CGAMP Interactor (STING). Chronic facial VCA rejection thus involves T cell/macrophage-mediated accelerated arteriosclerosis not normally represented in punch biopsies and potentially driven by persistent graft-resident effector T cells and recipient target endothelium that chimerically repopulates graft arteries.

Methods of ex vivo analysis of tissue status in vascularized composite allografts.

Vascularized composite allotransplantation can improve quality of life and restore functionality. However, the complex tissue composition of vascularized composite allografts (VCAs) presents unique clinical challenges that increase the likelihood of transplant rejection. Under prolonged static cold storage, highly damage-susceptible tissues such as muscle and nerve undergo irreversible degradation that may render allografts non-functional. Skin-containing VCA elicits an immunogenic response that increases the risk of recipient allograft rejection. The development of quantitative metrics to evaluate VCAs prior to and following transplantation are key to mitigating allograft rejection. Correspondingly, a broad range of bioanalytical methods have emerged to assess the progression of VCA rejection and characterize transplantation outcomes. To consolidate the current range of relevant technologies and expand on potential for development, methods to evaluate ex vivo VCA status are herein reviewed and comparatively assessed. The use of implantable physiological status monitoring biochips, non-invasive bioimpedance monitoring to assess edema, and deep learning algorithms to fuse disparate inputs to stratify VCAs are identified.

Continuous versus Pulsatile Flow in 24-Hour Vascularized Composite Allograft Machine Perfusion in Swine: A Pilot Study.

INTRODUCTION: Multiple perfusion systems have been investigated on vascularized composite allografts, with various temperatures and different preservation solutions, most using continuous flow (CF). However, physiological flow is pulsatile and provides better outcomes in kidney and lung ex vivo perfusions. The objective of this pilot study is to compare pulsatile flow (PF) with CF in our 24-h subnormothermic machine perfusion protocol for swine hindlimbs. METHODS: Partial hindlimbs were harvested from Yorkshire pigs and perfused with a modified Steen solution at 21°C for 24 h either with CF (n = 3) or with pulsatile flow (PF) at 60 beats/min (n = 3). Perfusion parameters, endothelial markers, and muscle biopsies were assessed at different timepoints. RESULTS: Overall, lactate levels were significantly lower in the PF group (P = 0.001). Glucose uptake and potassium concentration were similar in both groups throughout perfusion. Total nitric oxide levels were significantly higher in the PF group throughout perfusion (P = 0.032). Nitric oxide/endothelin-1 ratio also tends to be higher in the PF group, reflecting a potentially better vasoconductivity with PF, although not reaching statistical significance (P = 0.095). Arterial resistances were higher in the PF group (P < 0.001). Histological assessment did not show significant difference in muscular injury between the two groups. Weight increased quicker in the CF group but reached similar values with the PF after 24 h. CONCLUSIONS: This pilot study suggests that PF may provide superior preservation of vascularized composite allografts when perfused for 24 h at subnormothermic temperatures, with potential improvement in endothelial function and decreased ischemic injury.

Tolerance Induction in Vascularized Composite Allotransplantation-A Brief Review of Preclinical Models.

Pre-clinical studies are an obligatory tool to develop and translate novel therapeutic strategies into clinical practice. Acute and chronic rejection mediated by the recipient's immune system remains an important limiting factor for the (long-term) survival of vascularized composite allografts (VCA). Furthermore, high intensity immunosuppressive (IS) protocols are needed to mitigate the immediate and long-term effects of rejection. These IS regiments can have significant side-effects such as predisposing transplant recipients to infections, organ dysfunction and malignancies. To overcome these problems, tolerance induction has been proposed as one strategy to reduce the intensity of IS protocols and to thereby mitigate long-term effects of allograft rejection. In this review article, we provide an overview about animal models and strategies that have been used to induce tolerance. The induction of donor-specific tolerance was achieved in preclinical animal models and clinical translation may help improve short and long-term outcomes in VCAs in the future.

Current opinion: advances in machine perfusion and preservation of vascularized composite allografts - will time still matter?

PURPOSE OF REVIEW: A major hurdle hindering more widespread application of reconstructive transplantation is the very limited cold ischemia time (CIT) of vascularized composite allografts (VCAs). In this review, we discuss cutting edge machine perfusion protocols and preservation strategies to overcome this limitation. RECENT FINDINGS: Several preclinical machine perfusion studies have demonstrated the multifactorial utility of this technology to extend preservation windows, assess graft viability prior to transplantation and salvage damaged tissue, yet there are currently no clinically approved machine perfusion protocols for reconstructive transplantation. Thus, machine perfusion remains an open challenge in VCA due to the complexity of the various tissue types. In addition, multiple other promising avenues to prolong preservation of composite allografts have emerged. These include cryopreservation, high subzero preservation, vitrification and nanowarming. Despite several studies demonstrating extended preservation windows, there are several limitations that must be overcome prior to clinical translation. As both machine perfusion and subzero preservation protocols have rapidly advanced in the past few years, special consideration should be given to their potential complementary utilization. SUMMARY: Current and emerging machine perfusion and preservation technologies in VCA have great promise to transform the field of reconstructive transplantation, as every extra hour of CIT helps ease the complexities of the peri-transplant workflow. Amongst the many advantages, longer preservation windows may allow for elective procedures, improved matching, establishment of novel immunomodulatory protocols and global transport of grafts, ultimately enabling us the ability to offer this life changing procedure to more patients.

Defining chronic rejection in vascularized composite allografts - do we have reliable surrogates to look for?

PURPOSE OF REVIEW: Chronic rejection (CR) is a major threat in the field of vascularized composite tissue allografts (VCAs) as it causes graft dysfunction and usually graft loss. Unfortunately, knowledge of CR in VCA is incomplete because of the limited number of VCA recipients, the heterogeneous nature of VCAs and the short follow-up. RECENT FINDINGS: The diagnosis of CR in VCA has relied on clinical and pathological findings. Clinical changes include graft fibrosis, dyschromia and ischemic/necrotic ulcerations. Pathological changes primarily affect allograft vessels and manifest with graft vasculopathy (i.e. myo-intimal proliferation and luminal narrowing of allograft vessels, leading to graft ischemia). Attempts are made to diagnose CR with non- or minimally-invasive techniques, such as imaging studies (ultrasound biomicroscopy, functional magnetic resonance imaging) and serum biomarkers. These techniques provide interesting results and further insight into the mechanisms of CR in VCA. SUMMARY: The diagnosis of CR in VCA still relies mainly on clinicopathological graft alterations; unfortunately, these become overt rather late during the rejection process, when reversal of CR is problematic. More recent, minimally- or non-invasive techniques have provided encouraging results, but their usefulness in the diagnosis of CR requires further studies. These data highlight the paramount importance of CR prevention.

OPTN/SRTR 2020 Annual Data Report: VCA.

The first vascularized composite allograft (VCA) transplant in the United States was performed in 1998 in a 40-year-old man who received a laryn-geal transplant after experiencing severe trauma to the throat 20 years before. The following VCA was a hand transplant in 1999 in a 37-year-old man who lost his left hand 13 years before. Since then, the field of VCA transplantation has made significant strides. On July 3, 2014, the Or gan Procurement and Transplantation Network (OPTN)/United Network for Organ Sharing (UNOS) oversight of VCA procurement and transplant in the United States went into effect. In the last decade, the number of candidates listed for and transplanted with VCA has increased. While patient demographic data, whether listed candidates or patients undergoing VCA transplant, is limited by sample size, the trend is a predominance toward a young/middle-aged, White population. Overall outcomes data have been promising, with the vast majority of VCA transplants resulting in functioning grafts.

Optimization of Ex Vivo Machine Perfusion and Transplantation of Vascularized Composite Allografts.

BACKGROUND: Machine perfusion is gaining interest as an efficient method of tissue preservation of Vascularized Composite Allografts (VCA). The aim of this study was to develop a protocol for ex vivo subnormothermic oxygenated machine perfusion (SNMP) on rodent hindlimbs and to validate our protocol in a heterotopic hindlimb transplant model. METHODS: In this optimization study we compared three different solutions during 6 h of SNMP (n = 4 per group). Ten control limbs were stored in a preservation solution on Static Cold Storage [SCS]). During SNMP we monitored arterial flowrate, lactate levels, and edema. After SNMP, muscle biopsies were taken for histology examination, and energy charge analysis. We validated the best perfusion protocol in a heterotopic limb transplantation model with 30-d follow up (n = 13). As controls, we transplanted untreated limbs (n = 5) and hindlimbs preserved with either 6 or 24 h of SCS (n = 4 and n = 5). RESULTS: During SNMP, arterial outflow increased, and lactate clearance decreased in all groups. Total edema was significantly lower in the HBOC-201 group compared to the BSA group (P = 0.005), 4.9 (4.3-6.1) versus 48.8 (39.1-53.2) percentage, but not to the BSA + PEG group (P = 0.19). Energy charge levels of SCS controls decreased 4-fold compared to limbs perfused with acellular oxygen carrier HBOC-201, 0.10 (0.07-0.17) versus 0.46 (0.42-0.49) respectively (P = 0.002). CONCLUSIONS: Six hours ex vivo SNMP of rodent hindlimbs using an acellular oxygen carrier HBOC-201 results in superior tissue preservation compared to conventional SCS.

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.

Renal hemofiltration prevents metabolic acidosis and reduces inflammation during normothermic machine perfusion of the vascularized composite allograft: A preclinical study.

INTRODUCTION: Recent experimental evidence suggests normothermic machine perfusion of the vascularized composite allograft results in improved preservation compared to static cold storage, with less reperfusion injury in the immediate post-operative period. However, metabolic acidosis is a common feature of vascularized composite allograft perfusion, primarily due to the inability to process metabolic by-products. We evaluated the impact of combined limb-kidney perfusion on markers of metabolic acidosis and inflammation in a porcine model. METHODS: Ten paired pig forelimbs were used for this study, grouped as either limb-only (LO, n = 5) perfusion, or limb-kidney (LK, n = 5) perfusion. Infrared thermal imaging was used to determine homogeneity of perfusion. Lactate, bicarbonate, base, pH, and electrolytes, along with an inflammatory profile generated via the quantification of cytokines and cell-free DNA in the perfusate were recorded. RESULTS: The addition of a kidney to a limb perfusion circuit resulted in the rapid stabilization of lactate, bicarbonate, base, and pH. Conversely, the LO circuit became progressively acidotic, correlating in a significant increase in pro-inflammatory cytokines. Global perfusion across the limb was more homogenous with LK compared to LO. CONCLUSION: The addition of a kidney during limb perfusion results in significant improvements in perfusate biochemistry, with no evidence of metabolic acidosis.

Youngest Composite Full-Face Transplant: A Model for Vascularized Composite Allograft in Younger Populations.

BACKGROUND: The field of face transplantation continues to evolve, with more complex defects being addressed, and, at the same time, increased outcome expectations. Given our unique long-term experience in this field, we consented one of the youngest patients to undergo a full-face transplant. METHODS: An 18-year-old woman presented with complete destruction of her central face and craniofacial structures. She had coexisting major injuries, including pituitary gland, visual axis, and motor control. After extensive rehabilitation and reconstruction techniques, the patient underwent face transplant on May 4, 2017, at the age of 21 years. RESULTS: The total operative time for the recipient was 26 hours. There were no major perioperative complications. Since transplant, the patient has undergone 3 revision surgeries. She is near completely independent from a daily life activity standpoint. She has had 1 episode of rejection above grade II that was successfully treated with a short-term increased in immunosuppression. CONCLUSIONS: Contrary to data in solid organ transplantation where youth is associated with increased risk of rejection, our current algorithm in immunosuppression, combined with this patient's compliance, has led to only 1 rejection episode beyond grade II. This successful transplant can serve as a model for future vascularized composite transplants in younger populations.

Exosomes from donor-derived adipose mesenchymal stem cells prolong the survival of vascularized composite allografts.

Donor-derived adipose-derived mesenchymal stem cells (ADMSCs) dampen the alloimmune response and exosomes are reported to have biological activity similar to their parent cells. Here, we investigated the roles of exosomes from donor-derived ADMSCs (ADMSC-exo) in vascularized composite allotransplantation (VCA). Brown Norway-to-Lewis rat hindlimb transplantations were intravenously treated with either exosome from donor-derived ADMSCs or phosphate-buffered saline, combined with a short course of immunosuppression. We established that the treatment with ADMSC-exo prolongs the survival time of VCA grafts. Skin and muscle samples from ADMSC-exo-treated animals showed no histological signs of rejection, but samples from controls showed rejection of degree III. Comparing to the control group, a significant increase of donor cell chimerism, Tr1 and Treg, while a decrease of CD4+ T and Th1 cells were observed in the ADMSC-exo-treated group. Our findings imply that ADMSC-exo may be a valuable and safe treatment for extending VCA graft survival.

Cutaneous leukocyte lineages in tolerant large animal and immunosuppressed clinical vascularized composite allograft recipients.

Vascularized composite allografts (VCAs) can restore fully functional anatomic units in patients with limb amputations or severe facial tissue loss. However, acute rejection of the skin is frequently observed and underscores the importance of developing tolerance induction protocols. In this study, we have characterized the skin immune system in VCAs. We demonstrate infiltration of recipient leukocytes, regardless of rejection status, and in tolerant mixed hematopoietic chimeras, the co-existence of these cells with donor leukocytes in the absence of rejection. Here we characterize the dermal T cell and epidermal Langerhans cell components of the skin immune system in our porcine model of VCA tolerance, and the kinetics of cutaneous chimerism in both of these populations in VCAs transplanted to tolerant and nontolerant recipients, as well as in host skin. Furthermore, in biopsies from the first patient to receive a hand transplant in our program, we demonstrate the presence of recipient T cells in the skin of the transplanted limb in the absence of clinical or histological evidence of rejection.