RESUMO
Regardless of etiology, peripheral nerve injuries (PNI) result in disruption/loss of neuromuscular junctions, target muscle denervation, and poor sensorimotor outcomes with associated pain and disability. Adipose-derived stem cells (ASCs) have shown promise in neuroregeneration. However, there is a paucity of objective assessments reflective of functional neuroregeneration in experimental PNI. Here, we use a multimodal, static, and dynamic approach to evaluate functional outcomes after ASC therapy in a rodent PNI model. METHODS: Lewis rats were divided into 3 groups: 10 mm sciatic nerve resection ("CUT" group; n = 10), transection and repair ("REP" group; n = 10), transection and repair plus single-dose ASCs ("ASC" group; n = 12). Allogeneic (Brown Norway rat) ASCs (1 × 106) were administered intravenously on postoperative day 1. Functional outcome was assessed by static sciatic index, toe spread factor, and a dynamic swim test on a weekly basis for 6 weeks. Sciatic nerves and gastrocnemius muscles were harvested at endpoint (6 weeks) for histological analysis. RESULTS: The ASC group showed accelerated functional recovery on the swim test at 2 weeks postoperatively, with continued improvement over 4 weeks, culminating in superior overall outcomes at 6 weeks compared with the REP group. The CUT group showed no significant improvement from baseline. Nerve histomorphometry correlated well with the swim test results in the ASC group. Gastrocnemius muscle weights showed no difference between the REP and the ASC groups. CONCLUSION: Our study confirms that early, single dose, systemic administration of ASC after PNI accelerates and enhances overall motor recovery on static and dynamic functional tests as evidenced by improvements in voluntary as well as involuntary motions.
RESUMO
Early results of hand and face transplants and other grafts such as those of uterus, penis, trachea, larynx, or abdominal wall have confirmed the potential for vascularized composite allotransplantation (VCA) to restore appearance, anatomy, function, independence, and social integration in patients suffering from devastating tissue deficits untreatable by conventional treatment options. Despite such promise, these novel and complex procedures face challenges and controversies that remain open to discussion and debate. Indeed, many barriers to clinical advancement and negative stakeholder perceptions still exist. The bioethical challenges surrounding VCA include but are not limited to justice and vulnerability of subjects, and their experiences with risks, benefits and outcomes, provider economy of fame, public awareness and attitudes toward transplantation, and policy and regulatory issues shaping progress of the field. The First International Workshop on Bioethical Challenges in Reconstructive Transplantation was organized by the Brocher Foundation in Hermance, Switzerland. VCA professionals representing teams from across the world examined bioethical issues in VCA related to standards for safety, efficacy, feasibility, privacy, confidentiality, and equitability. Key discussion topics from the workshop were included in a survey questionnaire implemented across VCA professionals attending the 13th Congress of International Society of VCA held in Salzburg, Austria. The insights from the Brocher workshop and International Society of VCA survey as presented here could help inform the future development of clinical practice and policy strategies in VCA to ensure value, accessibility, and acceptance of these procedures by potential donors, potential or actual recipients and their families, and providers and payers.
Assuntos
Tomada de Decisão Clínica/ética , Aloenxertos Compostos , Confidencialidade/ética , Consentimento Livre e Esclarecido/ética , Segurança do Paciente , Obtenção de Tecidos e Órgãos/ética , Alotransplante de Tecidos Compostos Vascularizados/ética , Sobrevivência de Enxerto , Humanos , Complicações Pós-Operatórias/etiologia , Medição de Risco , Fatores de Risco , Resultado do Tratamento , Alotransplante de Tecidos Compostos Vascularizados/efeitos adversosRESUMO
BACKGROUND: In vascularized composite allotransplantation, medication nonadherence leads to increased acute rejections. Improving medication adherence would improve overall allograft survival. Regionally delivered immunosuppression, targeted to sites of allorecognition, may reduce or eliminate the need for daily systemic immunosuppression. METHODS: The authors developed biodegradable FK disks containing FK506-loaded double-walled microspheres and tested their efficacy at preventing rejection in a Brown-Norway-to-Lewis rat hindlimb transplantation model. In some experimental group animals, one FK disk was implanted subcutaneously either in native nontransplanted leg or in a transplanted allograft. Regular blood FK506 levels were measured. The endpoint was 180-day allograft survival or grade 3 rejection. At the endpoint, tissue FK506 levels were measured and mixed lymphocytic reaction was performed. RESULTS: A single FK disk maintained systemic blood FK506 levels between 5 and 15 ng/ml for 146 ± 11.1 days. After that, the levels declined to less than 5 ng/ml through the endpoint. There was significantly increased FK506 concentration in groin lymph nodes draining the implanted FK disk. Compared with other groups, animals with an FK disk in the transplanted allograft had 100 percent allograft survival to more than 180 days despite subtherapeutic levels below 5 ng/ml. In these animals, significant T-cell hyporesponsiveness was seen in groin lymph nodes draining the FK disk compared with robust splenic T-cell proliferation. CONCLUSIONS: Sustained regional immunosuppression (with a single FK506 disk) maintained the allograft by means of a high regional concentration of FK506. Notably, this was achieved at subtherapeutic blood concentrations of FK506, without any further systemic FK506 administration.
Assuntos
Portadores de Fármacos , Rejeição de Enxerto/prevenção & controle , Imunossupressores/uso terapêutico , Microesferas , Tacrolimo/uso terapêutico , Alotransplante de Tecidos Compostos Vascularizados , Animais , Masculino , Ratos , Ratos Endogâmicos BNRESUMO
BACKGROUND: Cellular therapies for immunomodulation in vascularized composite allotransplantation (VCA) have gained importance due to their potential for minimization of immunosuppression. Adipose-derived (AD) mesenchymal stem cells (MSCs) especially have shown encouraging potential. We investigated the influence of timing and frequency of AD-MSC treatment on immunologic and graft survival as well as graft vasculopathy outcomes after VCA. METHODS: Lewis rats received full-mismatched Brown Norway rat hindlimb transplants. Recipient animals were assigned to groups receiving donor-derived AD-MSCs (10 cells/animal) either on postoperative day (POD) 1, POD 4, or repeatedly on POD 4, 8, and 15, and compared to untreated controls. RESULTS: Although AD-MSC administration on POD 1 or POD 4, 8, and 15 resulted in 50% long-term graft acceptance, recipients treated on POD 4, and controls rejected before POD 50. All treated animals revealed peripheral blood chimerism (4 weeks), most pronounced after repetitive cell administration (12.92% vs 5.03% [POD 1] vs 6.31% [POD 4]; P < 0.05; all P < 0.01 vs control 1.45%). Chimerism was associated with the generation of regulatory T cells (CD4CD25FoxP3). In vitro mixed lymphocyte reactions revealed modulation of the recipient immune response after AD-MSC treatment. Graft arteries at end point revealed significant differences of arterial intimal thickness between rejecting and AD-MSC-treated animals (P < 0.01). CONCLUSIONS: Taken together, our results point to the potential for repetitive AD-MSC administration in improving outcomes after VCA. Future studies are warranted into optimization of the dosing and frequency of AD-MSC therapy, either alone or used in, combination with other cell therapies (such as hematopoietic stem cells or bone marrow-derived MSC or dendritic cells) for optimization of appropriate conditioning or maintenance regimens.
Assuntos
Tecido Adiposo/citologia , Aloenxertos Compostos/irrigação sanguínea , Aloenxertos Compostos/transplante , Sobrevivência de Enxerto , Membro Posterior/irrigação sanguínea , Membro Posterior/transplante , Imunoterapia/métodos , Transplante de Células-Tronco Mesenquimais/métodos , Células-Tronco Mesenquimais/imunologia , Alotransplante de Tecidos Compostos Vascularizados/métodos , Animais , Proliferação de Células , Células Cultivadas , Aloenxertos Compostos/imunologia , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/prevenção & controle , Membro Posterior/imunologia , Imunoterapia/efeitos adversos , Ativação Linfocitária , Masculino , Transplante de Células-Tronco Mesenquimais/efeitos adversos , Modelos Animais , Ratos Endogâmicos BN , Ratos Endogâmicos Lew , Linfócitos T Reguladores/imunologia , Fatores de Tempo , Quimeras de Transplante , Tolerância ao Transplante , Doenças Vasculares/imunologia , Doenças Vasculares/prevenção & controle , Alotransplante de Tecidos Compostos Vascularizados/efeitos adversosRESUMO
Pancreatic islet transplantation (PIT) represents a potential therapy to circumvent the need for exogenous insulin in type 1 diabetes. However, PIT remains limited by lack of donor islets and the need for long-term multidrug immunosuppression to prevent alloimmune islet rejection. Our goal was to evaluate a local immunoregulatory strategy that sustains islet allograft survival and restores glucose homeostasis in the absence of systemic immunosuppression. Nanogram quantities of murine CTLA4/Fc fusion protein were controllably delivered within human acellular dermal matrix scaffolds using an inkjet-based biopatterning technology and cotransplanted with allogeneic islets under the renal capsule to create an immunoregulatory microenvironment around the islet allograft. We achieved long-term engraftment of small loads of allogeneic islet cells with 40% of MHC-mismatched mouse recipients maintaining sustained normoglycemia following pancreatic ß-cell ablation by streptozotocin. Biopatterned CTLA4/Fc local therapy was associated with expansion of Foxp3+ regulatory T cells and shifts in cytokine production and gene expression from proinflammatory to regulatory profiles, thus substantially benefiting islet allografts survival and function. This study is a new paradigm for targeted therapies in PIT that demonstrates the favorable effects of immune alterations in the transplant milieu and suggests a unique strategy for minimizing systemic immunosuppression and promoting islet allograft survival.
Assuntos
Abatacepte/metabolismo , Glucose/metabolismo , Transplante das Ilhotas Pancreáticas , Animais , Células Cultivadas , Citocinas/metabolismo , Sobrevivência de Enxerto/imunologia , Sobrevivência de Enxerto/fisiologia , Homeostase/imunologia , Homeostase/fisiologia , Imunomodulação/imunologia , Imunomodulação/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Reguladores/metabolismoRESUMO
BACKGROUND: The restoration of complex tissue deficits with vascularized composite allotransplantation is a paradigm shift in reconstructive surgery. Clinical adoption of vascularized composite allotransplantation is limited by the need for systemic immunosuppression, with associated morbidity and mortality. Small-animal models lack the biological fidelity and preclinical relevance to enable translation of immunologic insights to humans. Large-animal models have been described; however, limitations persist, including the inability of heterotopic models to evaluate functional nerve regeneration, and the sensitivity of primates to toxicity of immunosuppressive drugs. The authors' novel orthotopic porcine limb transplant model has broad applicability and translational relevance to both immunologic and functional outcomes after vascularized composite allotransplantation. METHODS: Recipients underwent amputation at a level corresponding to the mid forearm. Replantation or transplantation of grafts was performed by plate fixation of the radio-ulna, microsurgical repair of brachial artery and median nerve, and extensor and flexor tendon repairs. Viability of replants was monitored clinically and radiologically. Transplants were monitored for clinicopathologic signs of rejection. Animals mobilized freely postoperatively. RESULTS: Replantations remained viable until the endpoint of 14 days. Transplants developed Banff grade 4 acute rejection by postoperative day 7. Doppler sonography and angiography confirmed vascular patency. Serial biopsy specimens of skin and histopathology of replants at endpoint confirmed tissue viability and bone healing. CONCLUSIONS: An orthotopic load-bearing porcine forelimb vascularized composite allotransplantation model was successfully established. Technical, procedural, and logistic considerations were optimized to allow model use for immunologic, bone healing, functional nerve regeneration, and other translational studies.
Assuntos
Membro Anterior/transplante , Pesquisa Translacional Biomédica , Alotransplante de Tecidos Compostos Vascularizados/métodos , Animais , Animais Endogâmicos , Regeneração Óssea/fisiologia , Rejeição de Enxerto/patologia , Rejeição de Enxerto/fisiopatologia , Sobrevivência de Enxerto/fisiologia , Haplótipos , Teste de Histocompatibilidade , Suínos , Coleta de Tecidos e Órgãos/métodos , Suporte de Carga/fisiologiaRESUMO
BACKGROUND: Strategies aiming at minimization or elimination of systemic immunosuppression are key immediate goals for clinical expansion of vascularized composite allotransplantation (VCA). We compared the in vitro and in vivo immunomodulatory efficacy of adipose-derived mesenchymal stem cells (AD-MSCs) and bone marrow (BM)-derived MSCs in a rat VCA model. METHODS: Both cell types were tested in vitro for suppressor function using mixed lymphocyte reactivity assays. AD-MSCs or BM-MSCs were administered intravenously (1 × 10 or 5 × 10 cells/animal) to Lewis rat recipients of mismatched Brown Norway hindlimb transplants. Short course tacrolimus (FK-506) monotherapy was withdrawn at postoperative day 21. In vivo regulatory T-cell induction, peripheral blood chimerism, and microchimerism in lymphatic organs were analyzed. RESULTS: AD-MSCs and BM-MSCs exhibited strong dose-dependent suppressor function in vitro, which was significantly more pronounced for AD cells. In vivo, all animals revealed peripheral multi-lineage chimerism at four weeks (P < 0.01) independent of cell type and dosage. Regulatory T-cell levels were increased with both cell types, the most in AD-MSC groups. These immunomodulatory effects were only transient. MSC treatment resulted in long-term (>120 day) allograft survival in 47% of the animals, which correlated with durable microchimerism in BM and spleen. CONCLUSIONS: AD-MSCs and BM-MSCs exert immunomodulatory effects that prolong survival of immunogenic skin-bearing VCA grafts with short course (21 day) tacrolimus induction therapy. The in vivo findings in terms of allograft survival did not reflect superior immunomodulatory characteristics of AD-MSCs found in vitro.
Assuntos
Tecido Adiposo/citologia , Transplante de Medula Óssea , Aloenxertos Compostos/irrigação sanguínea , Aloenxertos Compostos/transplante , Sobrevivência de Enxerto , Membro Posterior/irrigação sanguínea , Membro Posterior/transplante , Terapia de Imunossupressão/métodos , Transplante de Células-Tronco Mesenquimais , Transplante de Pele , Alotransplante de Tecidos Compostos Vascularizados , Animais , Células Cultivadas , Aloenxertos Compostos/efeitos dos fármacos , Esquema de Medicação , Sobrevivência de Enxerto/efeitos dos fármacos , Membro Posterior/efeitos dos fármacos , Imunossupressores/administração & dosagem , Masculino , Ratos Endogâmicos BN , Ratos Endogâmicos Lew , Linfócitos T Reguladores/imunologia , Tacrolimo/administração & dosagem , Fatores de Tempo , Quimeras de Transplante , Tolerância ao TransplanteRESUMO
Conventional breast cancer extirpation involves resection of parts of or the whole gland, resulting in asymmetry and disfiguration. Given the unsatisfactory aesthetic outcomes, patients often desire postmastectomy reconstructive procedures. Autologous fat grafting has been proposed for reconstructive purposes for decades to restore form and anatomy after mastectomy. Fat has the inherent advantage of being autologous tissue and the most natural-appearing filler, but given its inconsistent engraftment and retention rates, it lacks reliability. Implementation of autologous fat grafts with cellular adjuncts, such as multipotent adipose-derived stem cells (ADSCs), has shown promising results. However, it is pertinent and critical to question whether these cells could promote any residual tumor cells to proliferate, differentiate, or metastasize or even induce de novo carcinogenesis. Thus far, preclinical and clinical study findings are discordant. A trend towards potential promotion of both breast cancer growth and invasion by ADSCs found in basic science studies was indeed not confirmed in clinical trials. Whether experimental findings eventually correlate with or will be predictive of clinical outcomes remains unclear. Herein, we aimed to concisely review current experimental findings on the interaction of mesenchymal stem cells and breast cancer, mainly focusing on ADSCs as a promising tool for regenerative medicine, and discuss the implications in clinical translation.