RESUMEN
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.
Asunto(s)
Aloinjertos Compuestos , Diaminas , Soluciones Preservantes de Órganos , Daño por Reperfusión , Compuestos de Sulfhidrilo , Ratas , Animales , Depuradores de Radicales Libres , Caspasa 3 , Aloinjertos Compuestos/patología , Citocromos c , Calidad de Vida , Ratas Endogámicas Lew , Glutatión/farmacología , Alopurinol/farmacología , Insulina/farmacología , Isquemia , Preservación de Órganos , Frío , Daño por Reperfusión/patología , Rafinosa , AdenosinaRESUMEN
SUMMARY: Skin is one of the target tissues of rejection in face transplants and, because of its easy accessibility, has become the gold standard in the diagnosis of rejection. The allograft contains deeper tissues where rejection can occur, but samples cannot be obtained because of difficult access. Deep tissue changes were monitored on computed tomographic scans of the midface in six face transplant recipients with the help of image segmentation. The maxillary sinus was identified as a dynamic anatomical compartment. Observed changes in volume of the aeration relative to the opacification (aeration coefficient) of the maxillary sinus were quantified with the help of image segmentation. Changes in the aeration coefficient as a surrogate of mucosal swelling were quantified and related to time, treatment, and skin rejection grade. Lower aeration coefficients were found only in patients with transplanted maxillary sinus mucosa. Pathologic changes were not observed in face transplant recipients with a native maxillary sinus. The data show that the aeration coefficient was significantly lower at the time of biopsy-proven allograft rejection. Neither mechanical, nor infectious, nor medication side effects sufficiently explain the findings presented herein. The authors' findings are important to consider for clinical management of face transplant patients who receive parts of the sinonasal tract. The authors identify a potential radiologic biomarker of deep tissue allograft rejection. In the future, the proposed methodology might prove useful in monitoring deeper dynamic tissue changes in vascularized composite allografts and might help in designing patient-specific, individualized treatment strategies.
Asunto(s)
Aloinjertos Compuestos/diagnóstico por imagen , Trasplante Facial/efectos adversos , Rechazo de Injerto/diagnóstico , Seno Maxilar/diagnóstico por imagen , Mucosa Respiratoria/trasplante , Adulto , Aloinjertos Compuestos/patología , Femenino , Rechazo de Injerto/etiología , Rechazo de Injerto/patología , Rechazo de Injerto/prevención & control , Humanos , Masculino , Seno Maxilar/patología , Persona de Mediana Edad , Mucosa Respiratoria/patología , Estudios Retrospectivos , Tomografía Computarizada por Rayos X , Resultado del Tratamiento , Adulto JovenRESUMEN
BACKGROUND: Cutaneous changes of facial vascularized composite allotransplants (fVCAs) are extensively described in the literature. Parts of the nose, nasal, and oral cavities are included in most fVCAs. Distinctively, the nose and mouth are lined by mucosa. Little is known about the histopathology and complications of the mucosa involved in fVCA patients. METHODS: The study constitutes a retrospective cohort study of nine fVCA patients. Medical records were reviewed for information about changes of oral and nasal mucous membranes. Types of mucosal lesions were recorded and analyzed. Uni- and multivariate generalized estimating equation (GEE) models were used to assess the odds of developing mucosal inflammation in the presence of clinico-pathologic variables. RESULTS: A total of 186 clinical encounters with examination of oral and nasal mucous membranes were included. Membranes were devoid of clinical pathology in 101 instances (53% of all clinical assessments). Ulcerations/erosions (27%), edema (18%), and erythema (14%) were the most common lesions. Oral lesions affected the lips (58%), buccal mucosa (38%), and palate (5%). Sinonasal processes predominantly affected nasal vestibules and septae. In univariate analysis, sirolimus, skin rejection, and skin Banff grade were associated with the presence of an acute inflammatory mucosal lesion (p<0.05). In multivariate analysis, skin Banff grade and sirolimus were independent predictors of mucosal inflammation. CONCLUSION: Pathologies of fVCA mucous membranes are more common than previously reported. Mucosal assessment plays an important role in the pleomorphic allograft rejection process evaluation rather than diagnosis and treatment based on cutaneous pathology. A closer look at the pathophysiology of fVCA mucosal rejection and inflammation is warranted.
Asunto(s)
Aloinjertos Compuestos/patología , Cara/cirugía , Rechazo de Injerto/patología , Mucosa Bucal/patología , Mucosa Nasal/patología , Alotrasplante Compuesto Vascularizado/efectos adversos , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios RetrospectivosRESUMEN
BACKGROUND: Histologic criteria for diagnosing acute rejection in vascularized composite tissue allograft (VCA) have been established by the Banff 2007 Working Classification of Skin-Containing Composite Tissue Allograft, but the role of early vascular lesions in graft rejection warrants additional analysis. METHODS: We performed a retrospective study of 34 skin biopsies performed over 430 d for rejection surveillance, in Canada's first face allotransplant recipient. Three observers reviewed all biopsies to assess the nature and intensity of the inflammatory skin infiltrate. A complete histological and immunohistochemical review of the vascular components was performed with a focus on lymphocytic vasculitis, intravascular fibrin, vessel caliber, extent of injury, C4d positivity, and inflammatory cell phenotyping. We then correlated these data points to clinical and immunosuppression parameters. RESULTS: Acute vascular damage in biopsies that would be classified as mild acute rejection correlates with troughs in immunosuppression and subsides when immunosuppressive tacrolimus doses are increased. Grade 0 Banff rejection and Grade I without lymphocytic vasculitis were almost indistinguishable, whereas Grade I with lymphocytic vasculitis was an easy and reproducible histologic finding. CONCLUSIONS: Our results highlight the possible relevance of vascular injury in the context of VCA, as its presence might underlie a more aggressive form of immune rejection. If these findings are validated in other VCA patients, vascular injury in mild rejection might warrant a different clinical approach.
Asunto(s)
Trasplante Facial/efectos adversos , Rechazo de Injerto/diagnóstico , Inmunosupresores/administración & dosificación , Tacrolimus/administración & dosificación , Vasculitis/complicaciones , Anciano , Biopsia , Canadá , Aloinjertos Compuestos/irrigación sanguínea , Aloinjertos Compuestos/patología , Relación Dosis-Respuesta a Droga , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Rechazo de Injerto/prevención & control , Supervivencia de Injerto/efectos de los fármacos , Supervivencia de Injerto/inmunología , Humanos , Inmunosupresores/farmacocinética , Masculino , Estudios Retrospectivos , Índice de Severidad de la Enfermedad , Piel/irrigación sanguínea , Piel/patología , Tacrolimus/farmacocinética , Trasplante Homólogo/efectos adversos , Resultado del Tratamiento , Vasculitis/diagnóstico , Vasculitis/tratamiento farmacológico , Vasculitis/inmunologíaRESUMEN
BACKGROUND: Transplantation of vascularized composite allografts is limited mainly by the need for life-long immunosuppression. The consequent side effects and looming specter of chronic rejection portend eventual allograft loss. Development of tolerogenic protocols is thus of utmost importance to the field of vascularized composite allograft transplantation. METHODS: With a modified delayed tolerance induction protocol, 10 cynomolgus macaques received hand (n = 2) or face vascularized composite allografts across both full and haploidentical major histocompatibility complex barriers before donor bone marrow transplantation at a later date. Protocol and for-cause allograft skin biopsies were performed for immunohistochemical analysis and analysis of donor-recipient leukocyte contribution; mixed chimerism in peripheral blood and in vitro immune responses were assessed serially. RESULTS: Before bone marrow transplantation, maintenance immunosuppression for 4 months led to lethal complications, including posttransplant lymphoproliferative disorder (in two of four recipients), which necessitated early study termination. Shortening the maintenance period to 2 months was clinically relevant and allowed all subsequent subjects (n = 6) to complete the delayed tolerance induction protocol. Acute rejection developed within the first 2 to 4 weeks after transplantation, with corresponding near-complete turnover of allograft leukocytes from donor to recipient origin, but donor-specific antibodies remained negative. After bone marrow transplantation, mixed chimerism failed to develop, although carboxyfluorescein succinimidyl ester mixed lymphocyte reaction demonstrated generalized unresponsiveness. However, the accrual of subsequent rejection episodes eventually culminated in graft vasculopathy and irreversible allograft loss. CONCLUSIONS: Despite the various advantages of the delayed tolerance induction protocol, it failed to reliably induce mixed chimerism and thus immunologic tolerance to vascularized composite allografts, given currently available immunosuppression treatment options. Ongoing work shows promise in overcoming these limitations.
Asunto(s)
Aloinjertos Compuestos/inmunología , Rechazo de Injerto/prevención & control , Tolerancia Inmunológica , Acondicionamiento Pretrasplante/métodos , Alotrasplante Compuesto Vascularizado/efectos adversos , Animales , Biopsia , Trasplante de Médula Ósea/métodos , Aloinjertos Compuestos/patología , Aloinjertos Compuestos/trasplante , Modelos Animales de Enfermedad , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Supervivencia de Injerto/inmunología , Humanos , Terapia de Inmunosupresión/efectos adversos , Terapia de Inmunosupresión/métodos , Inmunosupresores/administración & dosificación , Inmunosupresores/efectos adversos , Leucocitos/inmunología , Prueba de Cultivo Mixto de Linfocitos , Trastornos Linfoproliferativos/epidemiología , Trastornos Linfoproliferativos/etiología , Macaca fascicularis , Complicaciones Posoperatorias/epidemiología , Complicaciones Posoperatorias/etiología , Piel/irrigación sanguínea , Piel/inmunología , Piel/patología , Quimera por Trasplante/inmunología , Acondicionamiento Pretrasplante/efectos adversos , Trasplante Homólogo/efectos adversos , Insuficiencia del Tratamiento , Alotrasplante Compuesto Vascularizado/métodosRESUMEN
BACKGROUND: Facial vascularized composite allotransplantation (fVCA) presents an established approach to restore form and function of patients with catastrophic facial defects. Skin is one of the target tissues of the rejection process, and due to its easy accessibility has become the gold standard in the diagnosis of rejection. Mucosal rejection frequently occurs; however, the added value of mucosal rejection assessment for patient management is unknown. METHODS: We conducted a systematic review of manuscripts listed in the MEDLINE/PubMed and GoogleScholar databases to identify articles that provide data on mucosal rejection following fVCA. For inclusion, papers had to be available as full-text and written in English. Non-VCA studies and animal studies were excluded. We followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. RESULTS: We included 17 articles that described changes in allotransplanted mucosa of fVCAs. These articles yielded data on 168 BANFF graded biopsies of corresponding skin and mucosa biopsies. Rejection grades were consistently higher in mucosal biopsies. Concordance between allograft skin and mucosa biopsy grades increased with an increasing skin-BANFF grade. Mucosa rejection grades were on average lower in the early stages of the posttransplant period (
Asunto(s)
Aloinjertos Compuestos/trasplante , Trasplante Facial/efectos adversos , Rechazo de Injerto/inmunología , Membrana Mucosa/trasplante , Trasplante de Piel/efectos adversos , Piel/inmunología , Alotrasplante Compuesto Vascularizado/efectos adversos , Adulto , Biopsia , Aloinjertos Compuestos/inmunología , Aloinjertos Compuestos/patología , Femenino , Rechazo de Injerto/patología , Rechazo de Injerto/terapia , Supervivencia de Injerto , Humanos , Masculino , Persona de Mediana Edad , Membrana Mucosa/inmunología , Membrana Mucosa/patología , Piel/patología , Resultado del TratamientoRESUMEN
Vascularized composite allotransplantation (VCA), such as hand and face transplantation, is emerging as a potential solution in patients that suffered severe injuries. However, adverse effects of chronic high-dose immunosuppression regimens strongly limit the access to these procedures. In this study, we developed an in situ forming implant (ISFI) loaded with rapamycin to promote VCA acceptance. We hypothesized that the sustained delivery of low-dose rapamycin in proximity to the graft may promote graft survival and induce an immunoregulatory microenvironment, boosting the expansion of T regulatory cells (Treg). In vitro and in vivo analysis of rapamycin-loaded ISFI (Rapa-ISFI) showed sustained drug release with subtherapeutic systemic levels and persistent tissue levels. A single injection of Rapa-ISFI in the groin on the same side as a transplanted limb significantly prolonged VCA survival. Moreover, treatment with Rapa-ISFI increased the levels of multilineage mixed chimerism and the frequency of Treg both in the circulation and VCA-skin. Our study shows that Rapa-ISFI therapy represents a promising approach for minimizing immunosuppression, decreasing toxicity and increasing patient compliance. Importantly, the use of such a delivery system may favor the reprogramming of allogeneic responses towards a regulatory function in VCA and, potentially, in other transplants and inflammatory conditions.
Asunto(s)
Aloinjertos Compuestos/efectos de los fármacos , Rechazo de Injerto/prevención & control , Supervivencia de Injerto/efectos de los fármacos , Miembro Posterior/trasplante , Sirolimus/farmacología , Linfocitos T Reguladores/inmunología , Alotrasplante Compuesto Vascularizado/efectos adversos , Animales , Aloinjertos Compuestos/inmunología , Aloinjertos Compuestos/patología , Sistemas de Liberación de Medicamentos , Inmunosupresores/farmacología , Masculino , Ratas , Ratas Endogámicas Lew , Quimera por Trasplante , Tolerancia al Trasplante/inmunologíaRESUMEN
BACKGROUND: Currently, patients receiving vascularized composite allotransplantation (VCA) grafts must take long-term systemic immunosuppressive therapy to prevent immunologic rejection. The morbidity and mortality associated with these medications is the single greatest barrier to more patients being able to receive these life-enhancing transplants. In contrast to solid organs, VCA, exemplified by hand or face transplants, allow visual diagnosis of clinical acute rejection (AR), directed biopsy and targeted graft therapies. Local immunosuppression in VCA could reduce systemic drug exposure and limit adverse effects. This proof of concept study evaluated, in a large animal forelimb VCA model, the efficacy and tolerability of a novel graft-implanted enzyme-responsive, tacrolimus (TAC)-eluting hydrogel platform, in achieving long-term graft survival. METHODS: Orthotopic forelimb VCA were performed in single haplotype mismatched mini-swine. Controls (n = 2) received no treatment. Two groups received TAC hydrogel: high dose (n = 4, 91 mg TAC) and low dose (n = 4, 49 mg TAC). The goal was to find a dose that was tolerable and resulted in long-term graft survival. Limbs were evaluated for clinical and histopathological signs of AR. TAC levels were measured in serial blood and skin tissue samples. Tolerability of the dose was evaluated by monitoring animal feeding behavior and weight. RESULTS: Control limbs underwent Banff Grade IV AR by post-operative day six. Low dose TAC hydrogel treatment resulted in long-term graft survival time to onset of Grade IV AR ranging from 56 days to 93 days. High dose TAC hydrogel also resulted in long-term graft survival (24 to 42 days), but was not well tolerated. CONCLUSION: Graft-implanted TAC-loaded hydrogel delays the onset of Grade IV AR of mismatched porcine forelimb VCA grafts, resulting in long term graft survival and demonstrates dose-dependent tolerability.
Asunto(s)
Aloinjertos Compuestos , Tacrolimus/administración & dosificación , Alotrasplante Compuesto Vascularizado/métodos , Animales , Aloinjertos Compuestos/efectos de los fármacos , Aloinjertos Compuestos/inmunología , Aloinjertos Compuestos/patología , Implantes de Medicamentos , Miembro Anterior/trasplante , Rechazo de Injerto/inmunología , Rechazo de Injerto/prevención & control , Supervivencia de Injerto/efectos de los fármacos , Supervivencia de Injerto/inmunología , Hidrogeles , Inmunosupresores/administración & dosificación , Inmunosupresores/farmacocinética , Modelos Animales , Prueba de Estudio Conceptual , Porcinos , Porcinos Enanos , Tacrolimus/farmacocinéticaRESUMEN
BACKGROUND: Routine application of vascularized composite allotransplantation is hampered by immunosuppression-related health comorbidities. To mitigate these, we developed an inflammation-responsive hydrogel for local immunosuppression. Here, we report on its long-term effect on graft survival, immunological, and toxicological impact. METHODS: Brown Norway-to-Lewis rat hindlimb transplantations were treated either systemically with daily injections of 1 mg/kg tacrolimus (TAC) or with subcutaneous intragraft injections of hydrogel containing 7 mg TAC, every 70 days. Animals were monitored for rejection or other pathology for 280 days. Systemic and graft TAC levels, regulatory T cells, and donor cell chimerism were measured periodically. At endpoint, markers for kidney, liver, and metabolic state were compared to naive age-matched rats. RESULTS: Both daily systemic TAC and subcutaneous intragraft TAC hydrogel at 70-day intervals were able to sustain graft survival longer than 280 days in 5 of 6 recipients. In the hydrogel group, 1 graft progressed to grade 3 rejection at postoperative day 149. In systemic TAC group, 1 animal was euthanized due to lymphoma on postoperative day 275. Hydrogel treatment provided stable graft and reduced systemic TAC levels, and a 4 times smaller total TAC dose compared with systemic immunosuppression. Hydrogel-treated animals showed preserved kidney function, absence of malignancies or opportunistic infections and increased hematopoietic chimerism compared with systemic immunosuppression. CONCLUSIONS: Our findings demonstrate that localized immunosuppression with TAC hydrogel is a long-term safe and reliable treatment. It may reduce the burden of systemic immunosuppression in vascularized composite allotransplantation, potentially boosting the clinical application of this surgical intervention.
Asunto(s)
Inhibidores de la Calcineurina/administración & dosificación , Portadores de Fármacos/química , Rechazo de Injerto/prevención & control , Terapia de Inmunosupresión/efectos adversos , Tacrolimus/administración & dosificación , Alotrasplante Compuesto Vascularizado/efectos adversos , Animales , Aloinjertos Compuestos/efectos de los fármacos , Aloinjertos Compuestos/inmunología , Aloinjertos Compuestos/patología , Aloinjertos Compuestos/trasplante , Modelos Animales de Enfermedad , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Supervivencia de Injerto/efectos de los fármacos , Supervivencia de Injerto/inmunología , Miembro Posterior/trasplante , Humanos , Hidrogeles/química , Terapia de Inmunosupresión/métodos , Inyecciones Intralesiones , Inyecciones Subcutáneas , Masculino , Ratas , Ratas Endogámicas BN , Ratas Endogámicas LewRESUMEN
The emergence of skin-containing vascularized composite allografts (VCAs) has provided impetus to understand factors affecting rejection and tolerance of skin. VCA tolerance can be established in miniature swine across haploidentical MHC barriers using mixed chimerism. Because the deceased donor pool for VCAs does not permit MHC antigen matching, clinical VCAs are transplanted across varying MHC disparities. We investigated whether sharing of MHC class I or II antigens between donors and recipients influences VCA skin tolerance. Miniature swine were conditioned nonmyeloablatively and received hematopoietic stem cell transplants and VCAs across MHC class I (n = 3) or class II (n = 3) barriers. In vitro immune responsiveness was assessed, and VCA skin-resident leukocytes were characterized by flow cytometry. Stable mixed chimerism was established in all animals. MHC class II-mismatched chimeras were tolerant of VCAs. MHC class I-mismatched animals, however, rejected VCA skin, characterized by infiltration of recipient-type CD8+ lymphocytes. Systemic donor-specific nonresponsiveness was maintained, including after VCA rejection. This study shows that MHC antigen matching influences VCA skin rejection and suggests that local regulation of immune tolerance is critical in long-term acceptance of all VCA components. These results help elucidate novel mechanisms underlying skin tolerance and identify clinically relevant VCA tolerance strategies.
Asunto(s)
Aloinjertos Compuestos/trasplante , Rechazo de Injerto/prevención & control , Complejo Mayor de Histocompatibilidad/inmunología , Trasplante de Piel/efectos adversos , Quimera por Trasplante/inmunología , Tolerancia al Trasplante/inmunología , Alotrasplante Compuesto Vascularizado/efectos adversos , Animales , Aloinjertos Compuestos/inmunología , Aloinjertos Compuestos/patología , Rechazo de Injerto/etiología , Supervivencia de Injerto/inmunología , Isoanticuerpos/sangre , Isoanticuerpos/inmunología , Porcinos , Porcinos EnanosRESUMEN
As reconstructive transplantation is gaining popularity as a viable alternative for upper limb amputees, it is becoming increasingly important for plastic surgeons to renew surgical skills and knowledge of this area. Forelimb allotransplantation research has been performed previously in rodent and swine models. However, preclinical canine forelimb allotransplantation studies are lacking in the literature. The purpose of this paper is to provide an overview of the surgical skills necessary to successfully perform forelimb transplantation in canines as a means to prepare for clinical application. A total of 18 transplantation operations on canines were performed. The recipient limb was shortened at the one-third proximal forearm level. The operation was performed in the following order: bones (two reconstructive plates), muscles and tendons (separately sutured), nerves (median, ulnar, and radial nerve), arteries (two), and veins (two). The total mean time of transplantation was 5 hours ± 30 minutes. All of the animals that received transplantation were treated with FK-506 (tacrolimus, 2 mg/kg) for 7 days after surgery. Most allografts survived with perfect viability without vascular problems during the early postoperative period. The canine forelimb allotransplantation model is well qualified to be a suitable training model for standard transplantation and future research work.
Asunto(s)
Aloinjertos Compuestos/patología , Aloinjertos Compuestos/trasplante , Miembro Anterior/trasplante , Supervivencia de Injerto , Tempo Operativo , Alotrasplante Compuesto Vascularizado/métodos , Animales , Perros , Modelos Animales , Técnicas de Cultivo de Órganos/métodos , Resultado del TratamientoRESUMEN
Vascularized composite allografts can undergo immune-mediated rejection, and skin biopsies are needed for monitoring of the transplant. However it is an invasive method, and requires processing time and pathological assessment. The purpose of this study is to use a new noninvasive monitoring method of the reflectance confocal microscopy (RCM) to determine severity of the allograft rejection on rats. Five groin flap allotransplantation were performed between 10 male Sprague-Dawley rats. Immunosuppressive therapy with cyclosporine A was given to the recipients during 10 days after surgery and was ended at the 10th postoperative days to allow acute transplant rejection. Following cessation of CsA, concomitant RCM evaluation and skin biopsy was performed every other day from each animal until total rejection of the allograft. Complete rejection of the allograft took nearly about 10 days and 4 or 5 RCM evaluation and skin biopsy was performed from each rat during this period. A total of 17 specimens were evaluated. A scoring system was developed based on the RCM findings. Skin biopsies were evaluated according to the Banff 2007 working classification criteria. RCM evaluation revealed epidermal irregularity and collagen destruction, however mild perivascular inflammation and degeneration of the basal epidermal layer were observed in early and late rejection period respectively with histopathologic evaluation. High correlation was found between the RCM scores and histopathologic grading. The RCM may be the useful tool to reduce the need for skin biopsy for monitoring of the skin containing vascularized composite allograft.
Asunto(s)
Aloinjertos Compuestos/diagnóstico por imagen , Rechazo de Injerto/diagnóstico por imagen , Microscopía Confocal , Imagen Óptica/métodos , Piel/diagnóstico por imagen , Cuidados Posteriores/métodos , Animales , Biopsia , Aloinjertos Compuestos/patología , Rechazo de Injerto/patología , Ingle , Masculino , Ratas , Ratas Sprague-Dawley , Piel/patologíaRESUMEN
La transferencia de colgajos libres es una herramienta fundamental dentro del arsenal reconstructivo para el tratamiento quirúrgico del gran quemado. Durante años ha existido controversia sobre la influencia del momento de la cirugía sobre la tasa de complicaciones, así como el alto riesgo de complicaciones al realizar reconstrucciones con colgajos libres en pacientes quemados. Diseñamos un estudio retrospectivo de las reconstrucciones quirúrgicas mediante colgajos libres realizadas en nuestra Unidad de Quemados del Hospital Universitari y Politecnic La Fe, de Valencia (España) en el periodo comprendido entre 2001 y 2013, recogiendo todos los datos relativos al paciente, a la lesión, al procedimiento reconstructivo, a los resultados y relacionando todo ello con el momento en el cual se realizó la cirugía. Para el análisis de los datos y asociaciones entre distintas variables usamos tablas de contingencia y aplicamos el test de independencia de Chi-cuadrado. Exponemos además 2 casos clínicos representativos de la cirugía reconstructiva practicada en pacientes quemados. Recogimos en total datos de 21 procedimientos de reconstrucción mediante colgajo libre en pacientes quemados. En ninguno se realizó reconstrucción primaria inmediata (en los primeros 5 días), mientras que la reconstrucción primaria temprana (entre el día 5 y el 21) se realizó en el 28,57% de los casos, la primaria intermedia (entre el día 21 y las 6 semanas) en el 28,57%, la secundaria (más de 6 semanas) en el 9,5%, y la cirugía de secuelas en el 33,33% de los casos. Las tasas de complicaciones mayores y menores fueron respectivamente 19,05% y 9,5%. El 50% de complicaciones mayores ocurrieron en el periodo de reconstrucción primaria temprana, mientras que no registramos ninguna en el periodo primario intermedio. El resto de complicaciones se repartió a partes iguales entre el periodo secundario y la cirugía de secuelas (ambas con el 25%). No encontramos asociación estadísticamente significativa entre las variables en ninguna de las comparaciones realizadas. En conclusión, creemos que la reconstrucción mediante colgajos libres en pacientes quemados presenta un riesgo más elevado de complicaciones mayores cuando se realiza en el periodo primario temprano (entre los días 5 y 21 tras la quemadura) (AU)
Free flap transfer is an essential tool within the reconstructive repertoire for the surgical treatment of major burn patients. For a long time there has been controversy about the influence of the timing over the complication rate, as well as the increased complication risk when we perform reconstruction by means of free flaps in burn patients. We designed a retrospective study about free flap reconstructions performed in our Burns Unit at the Hospital Universitari y Politecnic La Fe, Valencia (Spain), in the period from 2001 to 2013, collecting data related to the patient, the lesion, the reconstructive procedure, the results, and all this data related to timing. For the data analysis and associations between the different variables we used contingency tables and applied the Chi-square independence test. In addition we present 2 representative clinical reports about burn reconstructive surgery. Data from 21 reconstructive procedures with free flaps in burn patients were collected. Immediate primary reconstruction (in the first 5 days) is not performed in any patient, while early primary reconstruction (from day 5 to 21) was performed in 28,57% of cases; the intermediate primary reconstruction (from day 21 to 6th week) was performed in 28,57%, the secondary reconstruction (more than 6 weeks) in 9,5% and sequelae surgery in 33,33% of the cases. Major and minor complication rate were 19,05% and 9,5% respectively; 50% of major complications happened in early primary reconstruction period, whereas no complication is registered in intermediate primary period, the rest of complications are equally divided between secondary reconstruction period and sequelae surgery (both of them 25%). No statistically significant association between the variables is found in any of the comparisons performed. In conclusion, we think that reconstruction with free flaps applied to burn patients presents a higher major complication rate when performed in the early primary period (from day 5 to day 21 after the burn) (AU)
Asunto(s)
Femenino , Humanos , Masculino , Quemaduras/metabolismo , Quemaduras/patología , Colgajos Tisulares Libres/clasificación , Colgajos Tisulares Libres/trasplante , Aloinjertos Compuestos/anatomía & histología , Aloinjertos Compuestos/cirugía , Aloinjertos Compuestos/trasplante , Protocolos Clínicos/clasificación , Estudios Retrospectivos , Quemaduras/rehabilitación , Quemaduras/cirugía , Colgajos Tisulares Libres/cirugía , Colgajos Tisulares Libres , Aloinjertos Compuestos/citología , Aloinjertos Compuestos/patología , Protocolos Clínicos/normas , España/etnología , Estudios de Evaluación como AsuntoRESUMEN
BACKGROUND: Antibody-mediated rejection is caused in part by increasing circulation/production of donor-specific antibody (DSA). Activation-induced cytidine deaminase (AID) is a key regulator of class switch recombination and somatic hypermutation of immunoglobulin in B cells, yet its role in antibody-mediated transplant rejection remains unclear. We show here that AID deficiency in mice enables suppression of allograft vasculopathy (AV) after aorta transplantation, a DSA-mediated process. METHODS: Splenocytes from C57BL/6 J (B6) AID(−/−) mice were used for determining in vitro proliferation responses, alloreactivity, cell surface marker expression, and antibody production. BALB/c mouse aortas were transplanted into B6 AID(−/−) mice with or without FK506 treatment. Blood and aorta grafts were harvested on day 30 after transplantation and were subjected to DSA, histological, and immunohistological analyses. RESULTS: The AID(−/−) splenocytes were comparable to wild type splenocytes in proliferation responses, alloreactivity, and expression of cell surface markers in vitro. However, they completely failed to produce immunoglobulin G, although they were not impaired in immunoglobulin M production relative to controls. Furthermore, BALB/c aorta grafts from B6 AID(−/−) recipient mice on day 30 after transplantation showed reduced signs of AV compared to the grafts from B6 wild type recipient mice which had severe vascular intimal hyperplasia, interstitial fibrosis, and inflammation. Treatment with FK506 produced a synergistic effect in the grafts from AID(−/−) recipients with further reduction of intimal hyperplasia and fibrosis scores. CONCLUSIONS: The AID deficiency inhibits DSA-mediated AV after aorta transplantation in mice. We propose that AID could be a novel molecular target for controlling antibody-mediated rejection in organ transplantation.
Asunto(s)
Aorta/trasplante , Linfocitos B/enzimología , Aloinjertos Compuestos/trasplante , Citidina Desaminasa/deficiencia , Rechazo de Injerto/prevención & control , Inmunoglobulina G/sangre , Isoanticuerpos/sangre , Bazo/enzimología , Animales , Aorta/efectos de los fármacos , Aorta/inmunología , Aorta/metabolismo , Aorta/patología , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Proliferación Celular , Células Cultivadas , Aloinjertos Compuestos/inmunología , Aloinjertos Compuestos/patología , Citidina Desaminasa/genética , Fibrosis , Rechazo de Injerto/enzimología , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Hiperplasia , Inmunoglobulina G/inmunología , Inmunosupresores/farmacología , Isoanticuerpos/inmunología , Activación de Linfocitos , Masculino , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Neointima , Transducción de Señal , Bazo/inmunología , Factores de TiempoRESUMEN
The mechanisms of skin rejection in vascularized composite allotransplantation (VCA) remain incompletely understood. The formation of tertiary lymphoid organs (TLO) in hand transplantation has been recently described. We assess this phenomenon in experimental and clinical VCA rejection. Skin biopsies of human (n = 187), nonhuman primate (n = 11), and rat (n = 15) VCAs were analyzed for presence of TLO. A comprehensive immunohistochemical assessment (characterization of the cell infiltrate, expression of adhesion molecules) including staining for peripheral node addressin (PNAd) was performed and correlated with rejection and time post-transplantation. TLO were identified in human, nonhuman primate, and rat skin samples. Expression of PNAd was increased in the endothelium of vessels upon rejection in human skin (P = 0.003) and correlated with B- and T-lymphocyte numbers and LFA-1 expression. PNAd expression was observed at all time-points after transplantation and increased significantly after year 5. In nonhuman primate skin, PNAd expression was found during inflammatory conditions early and late after transplantation. In rat skin, PNAd expression was strongly associated with acute rejection and time post-transplantation. Lymphoid neogenesis and TLO formation can be uniformly found in experimental and human VCA. PNAd expression in vascular endothelium correlates with skin rejection and T- and B-cell infiltration.
Asunto(s)
Aloinjertos Compuestos/fisiopatología , Antebrazo/cirugía , Rechazo de Injerto/patología , Trasplante de Mano , Linfangiogénesis/fisiología , Tejido Linfoide/patología , Piel/inmunología , Alotrasplante Compuesto Vascularizado , Animales , Antígenos CD/análisis , Biomarcadores , Biopsia , Moléculas de Adhesión Celular/análisis , Aloinjertos Compuestos/inmunología , Aloinjertos Compuestos/patología , Femenino , Antebrazo/patología , Rechazo de Injerto/tratamiento farmacológico , Rechazo de Injerto/inmunología , Miembro Posterior/trasplante , Humanos , Inmunosupresores/uso terapéutico , Subgrupos Linfocitarios/inmunología , Subgrupos Linfocitarios/patología , Macaca fascicularis , Masculino , Ratas , Ratas Endogámicas , Piel/patologíaRESUMEN
Vascularized composite allograft (VCA) transplantation can restore form and function following severe craniofacial injuries, extremity amputations or massive tissue loss. The induction of transplant tolerance would eliminate the need for long-term immunosuppression, realigning the risk-benefit ratio for these life-enhancing procedures. Skin, a critical component of VCA, has consistently presented the most stringent challenge to transplant tolerance. Here, we demonstrate, in a clinically relevant miniature swine model, induction of immunologic tolerance of VCAs across MHC barriers by induction of stable hematopoietic mixed chimerism. Recipient conditioning consisted of T cell depletion with CD3-immunotoxin, and 100 cGy total body irradiation prior to hematopoietic cell transplantation (HCT) and a 45-day course of cyclosporine A. VCA transplantation was performed either simultaneously to induction of mixed chimerism or into established mixed chimeras 85-150 days later. Following withdrawal of immunosuppression both VCAs transplanted into stable chimeras (n=4), and those transplanted at the time of HCT (n=2) accepted all components, including skin, without evidence of rejection to the experimental end point 115-504 days posttransplant. These data demonstrate that tolerance across MHC mismatches can be induced in a clinically relevant VCA model, providing proof of concept for long-term immunosuppression-free survival.
Asunto(s)
Aloinjertos Compuestos/inmunología , Rechazo de Injerto/inmunología , Supervivencia de Injerto/inmunología , Trasplante de Células Madre Hematopoyéticas , Complejo Mayor de Histocompatibilidad/inmunología , Alotrasplante Compuesto Vascularizado , Animales , Aloinjertos Compuestos/patología , Histocompatibilidad , Técnicas para Inmunoenzimas , Inmunosupresores/uso terapéutico , Prueba de Cultivo Mixto de Linfocitos , Porcinos , Porcinos Enanos , Linfocitos T Reguladores/inmunología , Quimera por Trasplante/inmunología , Tolerancia al Trasplante/inmunologíaRESUMEN
PURPOSE: Injuries involving the tendon-bone interface (TBI) are difficult to address. Standard techniques typically lead to diminished strength of the healed insertion site. We hypothesized that these injuries would benefit from being reconstructed with decellularized composite grafts replacing both tendon and bone. To test this hypothesis, decellularized grafts were compared with conventional pullout repairs in an in vivo animal model. METHODS: We harvested 48 Achilles TBI grafts from rats and decellularized them. Tendon-bone interface graft reconstruction and pullout repairs were compared using a pair-matched design. Biomechanical properties were evaluated at 2, 4, 8, and 12 weeks. We evaluated histological analysis of insertion morphology and collagen type I/III content. RESULTS: There was a significant increase in ultimate failure load (35 ± 11 vs 24 ± 7 N) and ultimate tensile stress (1.5 ± 0.3 vs 1.0 ± 0.4 N/mm(2)) of the TBI grafts compared with pullout repairs at 2 weeks. These differences remained at 4 weeks. At 12 weeks, both TBI grafts and pullout repairs were as strong as native tissue and not significantly different from each other. Histology showed a more organized extracellular matrix in the TBI graft group at the early time points. Repopulation of the decellularized grafts increased over time. At 12 weeks, the insertion points of both groups were richly populated with cells that possessed morphologies similar to those found in native TBI. CONCLUSIONS: This study showed that decellularized TBI grafts were stronger compared with conventional pullout repairs at 2 and 4 weeks but were comparable at 12 weeks. A more organized extracellular matrix and different collagen composition in the early time points may explain the observed differences in strength. CLINICAL RELEVANCE: In the future, decellularized TBI grafts may be used to reconstruct tendon-bone insertion tears in multiple areas including the flexor tendon system.
Asunto(s)
Trasplante Óseo/métodos , Aloinjertos Compuestos , Traumatismos de la Mano/cirugía , Tendones/trasplante , Tenodesis/métodos , Animales , Fenómenos Biomecánicos , Aloinjertos Compuestos/patología , Aloinjertos Compuestos/fisiopatología , Modelos Animales de Enfermedad , Humanos , Ratones , Falla de Prótesis , Ratas , Ratas Wistar , Tendones/patología , Tendones/fisiopatología , Recolección de Tejidos y ÓrganosRESUMEN
BACKGROUND: We have previously reported that Massachusetts General Hospital miniature swine, which had accepted class I-mismatched kidneys long-term after 12 days of high-dose cyclosporine A, uniformly accepted donor-major histocompatibility complex (MHC)-matched kidneys without immunosuppression but rejected donor MHC-matched split-thickness skin grafts by day 25, without changes in renal graft function or antidonor in vitro responses. We have now tested whether this "split tolerance" would also be observed for the primarily vascularized skin of vascularized composite allografts (VCAs). METHODS: Group 1 animals (n=3) received donor MHC-matched VCAs less than 70 days after primary kidney transplant (KTx). Group 2 animals (n=3) received a second donor-matched kidney transplant followed by a donor-matched VCA more than 200 days after primary KTx. RESULTS: Animals in Group 1 lost the epidermis on days 28, 30, and 40, with all other components of the VCAs remaining viable. Histology showed cellular infiltration localized to dermal-epidermal junction. One of three recipients of VCAs in Group 2, accepted all components of the VCA, including epidermis (>200 days). The other two recipients lost only the epidermis on days 45 and 85, with survival of the remainder of the VCA long-term. CONCLUSIONS: All tissues of a VCA are accepted long-term on animals tolerant of class I-mismatched kidneys, with the exception of epidermis, the survival of which is markedly prolonged compared with split-thickness skin grafts but not indefinite. Exposure of tolerant animals to second donor-matched kidneys before VCA increases the longevity of the VCA epidermis, suggesting an increase in the immunomodulatory mechanisms associated with tolerance of the kidney.