RESUMO
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.
Assuntos
Transplante de Face , Alotransplante de Tecidos Compostos Vascularizados , Humanos , Transplante de Face/métodos , Consenso , Técnica Delphi , Projetos de PesquisaRESUMO
INTRODUCTION: To mitigate the post-operative complication rates associated with massive bone allografts, tissue engineering techniques have been employed to decellularize entire bones through perfusion with a sequence of solvents. Mechanical assessment was performed in order to compare conventional massive bone allografts and perfusion/decellularized massive bone allografts. MATERIAL AND METHODS: Ten porcine femurs were included. Five were decellularized by perfusion. The remaining 5 were left untreated as the "control" group. Biomechanical testing was conducted on each bone, encompassing five different assessments: screw pull-out, 3-points bending, torsion, compression and Vickers indentation. RESULTS: Under the experimental conditions of this study, all five destructive tested variables (maximum force until screw pull-out, maximum elongation until screw pull-out, energy to pull out the screw, fracture resistance in flexion and maximum constrain of compression) were statistically significantly superior in the control group. All seven nondestructive variables (Young's modulus in flexion, Young's modulus in shear stress, Young's modulus in compression, Elastic conventional limit in compression, lengthening to rupture in compression, resilience in compression and Vickers Hardness) showed no significant difference. DISCUSSION: Descriptive statistical results suggest a tendency for the biomechanical characteristics of decellularized bone to decrease compared with the control group. However, statistical inferences demonstrated a slight significant superiority of the control group with destructive mechanical stresses. Nondestructive mechanical tests (within the elastic phase of Young's modulus) were not significantly different.
Assuntos
Aloenxertos , Fêmur , Hidróxido de Sódio , Animais , Suínos , Hidróxido de Sódio/farmacologia , Fêmur/patologia , Transplante Ósseo , Perfusão , Estresse Mecânico , Fenômenos BiomecânicosRESUMO
The topographical neuroanatomy of the human spinal cord (SC) is currently based on the adjacent vertebrae. This morphometric study sought to develop a dataset allowing for statistical analysis of human SC segment characteristics. Overall, 32 human SCs were dissected (18 female and 14 male donors), and individual SC segments were identified. Anterior and posterior lengths, thicknesses and widths were measured by two examiners. Statistical analyses included t-tests, as well as intraclass and Pearson's correlation coefficients. The SC length was significantly shorter in females than males. The cranial (C4) and caudal (T1/T2) limits of the cervical enlargement, along with its maximal width (C6-C7), were identified by combining widths and thicknesses of the segments. The thoracic region, T2 to T12, could be identified using segments widths and thicknesses values. The length of the lumbosacral region, from segments L2 to S5, was particularly stable, independently of SC length and sex. The lumbar enlargement was characterized by a thickness increase between the segments L2 and S1 which reached its maximum at the level of L3, L4, and L5, whereas the width was not significantly increased. From the S2 to S5 segments, width and thickness were equal, with both decreasing of 1 mm per segment. The morphometrical analysis of 32 human SCs provided a dataset allowing for statistical analysis of segmental measures with significant results. A combined approach mostly using widths and thicknesses provided landmarks of potential interest for the localization of SC segments in a clinical MRI setting.
Assuntos
Vértebras Lombares , Medula Espinal , Humanos , Masculino , Feminino , Medula Espinal/anatomia & histologia , Imageamento por Ressonância Magnética , Região Lombossacral , CadáverRESUMO
In Hashimoto's thyroiditis (HT), oxidative stress (OS) is driven by Th1 cytokines' response interfering with the normal function of thyrocytes. OS results from an imbalance between an excessive production of reactive oxygen species (ROS) and a lowering of antioxidant production. Moreover, OS has been shown to inhibit Sirtuin 1 (SIRT1), which is able to prevent hypoxia-inducible factor (HIF)-1α stabilization. The aims of this study were to determine the involvement of NADPH-oxidases (NOX), SIRT1, and HIF-1α in HT pathophysiology as well as the status of antioxidant proteins such as peroxiredoxin 1 (PRDX1), catalase, and superoxide dismutase 1 (SOD1). The protein expressions of NOX2, NOX4, antioxidant enzymes, SIRT1, and HIF-1α, as well as glucose transporter-1 (GLUT-1) and vascular endothelial growth factor A (VEGF-A), were analyzed by Western blot in primary cultures of human thyrocytes that were or were not incubated with Th1 cytokines. The same proteins were also analyzed by immunohistochemistry in thyroid samples from control and HT patients. In human thyrocytes incubated with Th1 cytokines, NOX4 expression was increased whereas antioxidants, such as PRDX1, catalase, and SOD1, were reduced. Th1 cytokines also induced a significant decrease of SIRT1 protein expression associated with an upregulation of HIF-1α, GLUT-1, and VEGF-A proteins. With the exception of PRDX1 and SOD1, similar results were obtained in HT thyroids. OS due to an increase of ROS produced by NOX4 and a loss of antioxidant defenses (PRDX1, catalase, SOD1) correlates to a reduction of SIRT1 and an upregulation of HIF 1α, GLUT-1, and VEGF-A. Our study placed SIRT1 as a key regulator of OS and we, therefore, believe it could be considered as a potential therapeutic target in HT.
Assuntos
Doença de Hashimoto/etiologia , Doença de Hashimoto/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Estresse Oxidativo , Sirtuína 1/genética , Células Th1/imunologia , Células Th1/metabolismo , Fator A de Crescimento do Endotélio Vascular/genética , Adulto , Doenças Autoimunes/etiologia , Doenças Autoimunes/metabolismo , Autoimunidade/genética , Biomarcadores , Citocinas/genética , Citocinas/metabolismo , Feminino , Imunofluorescência , Regulação da Expressão Gênica , Doença de Hashimoto/diagnóstico , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Modelos Biológicos , NADPH Oxidase 2/genética , NADPH Oxidase 2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Sirtuína 1/metabolismo , Superóxido Dismutase-1/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Timócitos/imunologia , Timócitos/metabolismo , Testes de Função Tireóidea , Glândula Tireoide/imunologia , Glândula Tireoide/metabolismo , Glândula Tireoide/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Adulto JovemRESUMO
Graves' disease (GD) is an autoimmune thyroiditis often associated with Graves' orbitopathy (GO). GD thyroid and GO orbital fat share high oxidative stress (OS) and hypervascularization. We investigated the metabolic pathways leading to OS and angiogenesis, aiming to further decipher the link between local and systemic GD manifestations. Plasma and thyroid samples were obtained from patients operated on for multinodular goiters (controls) or GD. Orbital fats were from GO or control patients. The NADPH-oxidase-4 (NOX4)/HIF-1α/VEGF-A signaling pathway was investigated by Western blotting and immunostaining. miR-199a family expression was evaluated following quantitative real-time PCR and/or in situ hybridization. In GD thyroids and GO orbital fats, NOX4 was upregulated and correlated with HIF-1α stabilization and VEGF-A overexpression. The biotin assay identified NOX4, HIF-1α and VEGF-A as direct targets of miR-199a-5p in cultured thyrocytes. Interestingly, GD thyroids, GD plasmas and GO orbital fats showed a downregulation of miR-199a-3p/-5p. Our results also highlighted an activation of STAT-3 signaling in GD thyroids and GO orbital fats, a transcription factor known to negatively regulate miR-199a expression. We identified NOX4/HIF-1α/VEGF-A as critical actors in GD and GO. STAT-3-dependent regulation of miR-199a is proposed as a common driver leading to these events in GD thyroids and GO orbital fats.
Assuntos
Tecido Adiposo/metabolismo , Regulação para Baixo/genética , Doença de Graves/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , MicroRNAs/genética , NADPH Oxidase 4/genética , Glândula Tireoide/metabolismo , Fator A de Crescimento do Endotélio Vascular/genética , Adulto , Feminino , Oftalmopatia de Graves/genética , Oftalmopatia de Graves/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Transdução de Sinais/genéticaRESUMO
PURPOSE: In the mandible, the condylar neck vascularization is commonly described as mainly periosteal; while the endosteal contribution is still debated, with very limited anatomical studies. Previous works have shown the contribution of nutrient vessels through accessory foramina and their contribution in the blood supply of other parts of the mandible. Our aim was to study the condylar neck's blood supply from nutrient foramina. METHODS: Six latex-injected heads were dissected and two hundred mandibular condyles were observed on dry mandibles searching for accessory bone foramina. RESULTS: Latex-injected dissections showed a direct condylar medular arterial supply through foramina. On dry mandibles, these foramina were most frequently observed in the pterygoid fovea in 91% of cases. However, two other accessory foramina areas were identified on the lateral and medial sides of the mandibular condylar process, confirming the vascular contribution of transverse facial and maxillary arteries. CONCLUSIONS: The maxillary artery indeed provided both endosteal and periosteal blood supply to the condylar neck, with three different branches: an intramedullary ascending artery (arising from the inferior alveolar artery), a direct nutrient branch and some pterygoid osteomuscular branches.
Assuntos
Côndilo Mandibular/irrigação sanguínea , Artéria Maxilar/anatomia & histologia , Cadáver , Dissecação , Feminino , Fixadores , Humanos , Látex , Masculino , Mandíbula/anatomia & histologia , Mandíbula/irrigação sanguínea , Mandíbula/cirurgia , Côndilo Mandibular/anatomia & histologia , Côndilo Mandibular/cirurgia , Artéria Maxilar/cirurgia , Fixação de Tecidos/métodosRESUMO
BACKGROUND: Liver transplantation is the treatment for end-stage liver diseases and well-selected malignancies. The allograft shortage may be alleviated with living donation. The initial UCLouvain experience of adult living-donor liver transplantation (LDLT) is presented. METHODS: A retrospective analysis of 64 adult-to-adult LDLTs performed at our institution between 1998 and 2016 was conducted. The median age of 29 (45.3%) females and 35 (54.7%) males was 50.2 years (interquartile range, IQR 32.9-57.5). Twenty-two (34.4%) recipients had no portal hypertension. Three (4.7%) patients had a benign and 33 (51.6%) a malignant tumor [19 (29.7%) hepatocellular cancer, 11 (17.2%) secondary cancer and one (1.6%) each hemangioendothelioma, hepatoblastoma and embryonal liver sarcoma]. Median donor and recipient follow-ups were 93 months (IQR 41-159) and 39 months (22-91), respectively. RESULTS: Right and left hemi-livers were implanted in 39 (60.9%) and 25 (39.1%) cases, respectively. Median weights of right- and left-liver were 810â¯g (IQR 730-940) and 454â¯g (IQR 394-534), respectively. Graft-to-recipient weight ratios (GRWRs) were 1.17% (right, IQR 0.98%-1.4%) and 0.77% (left, 0.59%-0.95%). One- and five-year patient survivals were 85% and 71% (right) vs. 84% and 58% (left), respectively. One- and five-year graft survivals were 74% and 61% (right) vs. 76% and 53% (left), respectively. The patient and graft survival of right and left grafts and of very small (<0.6%), small (0.6%-0.79%) and large (≥0.8%) GRWR were similar. Survival of very small grafts was 86% and 86% at 3- and 12-month. No donor died while five (7.8%) developed a Clavien-Dindo complication IIIa, IIIb or IV. Recipient morbidity consisted mainly of biliary and vascular complications; three (4.7%) recipients developed a small-for-size syndrome according to the Kyushu criteria. CONCLUSIONS: Adult-to-adult LDLT is a demanding procedure that widens therapeutic possibilities of many hepatobiliary diseases. The donor procedure can be done safely with low morbidity. The recipient operation carries a major morbidity indicating an important learning curve. Shifting the risk from the donor to the recipient, by moving from the larger right-liver to the smaller left-liver grafts, should be further explored as this policy makes donor hepatectomy safer and may stimulate the development of transplant oncology.
Assuntos
Falência Hepática/cirurgia , Neoplasias Hepáticas/cirurgia , Transplante de Fígado/mortalidade , Transplante de Fígado/métodos , Doadores Vivos , Adulto , Fatores Etários , Bélgica , Estudos de Coortes , Feminino , Rejeição de Enxerto , Sobrevivência de Enxerto , Hepatectomia/métodos , Hospitais Universitários , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Retrospectivos , Medição de Risco , Análise de Sobrevida , Transplantados , Resultado do TratamentoRESUMO
OBJECTIVE: The purpose of this study was to assess whether perfusion-decellularization technology could be applied to facial grafts. BACKGROUND: Facial allotransplantation remains an experimental procedure. Regenerative medicine techniques allow fabrication of transplantable organs from an individual's own cells, which are seeded into extracellular matrix (ECM) scaffolds from animal or human organs. Therefore, we hypothesized that ECM scaffolds also can be created from facial subunits. We explored the use of the porcine ear as a clinically relevant face subunit model to develop regenerative medicine-related platforms for facial bioengineering. METHODS: Porcine ear grafts were decellularized and histologic, immunologic, and cell culture studies done to determine whether scaffolds retained their 3D framework and molecular content; were biocompatible in vitro and in vivo, and triggered an anti-MHC immune response from the host. RESULTS: The cellular compartment of the porcine ear was completely removed except for a few cartilaginous cells, leaving behind an acellular ECM scaffold; this scaffold retained its complex 3D architecture and biochemical components. The framework of the vascular tree was intact at all hierarchical levels and sustained a physiologically relevant blood pressure when implanted in vivo. Scaffolds were biocompatible in vitro and in vivo, and elicited no MHC immune response from the host. Cells from different types remained viable and could even differentiate at the scale of a whole-ear scaffold. CONCLUSIONS: Acellular scaffolds were produced from the porcine ear, and may be a valuable platform to treat facial deformities using regenerative medicine approaches.
Assuntos
Derme Acelular , Matriz Extracelular , Transplante de Face/métodos , Engenharia Tecidual/métodos , Alicerces Teciduais , Animais , Materiais Biocompatíveis , Orelha , Projetos Piloto , SuínosRESUMO
OBJECTIVE: During the last decade, face allotransplantation has been shown to be a revolutionary reconstructive procedure for severe disfigurements. However, offer to patients remains limited due to lifelong immunosuppression. To move forward in the field, a new pathway in tissue engineering is proposed. BACKGROUND: Our previously reported technique of matrix production of a porcine auricular subunit graft has been translated to a human face model. METHODS: 5 partial and 1 total face grafts were procured from human fresh cadavers. After arterial cannulation, the specimens were perfused using a combined detergent/polar solvent decellularization protocol. Preservation of vascular patency was assessed by imaging, cell and antigen removal by DNA quantification and histology. The main extracellular matrix proteins and associated cytokines were evaluated. Lip scaffolds were cultivated with dermal, muscle progenitor and endothelial cells, either on discs or in a bioreactor. RESULTS: Decellularization was successful in all facial grafts within 12 days revealing acellular scaffolds with full preservation of innate morphology. Imaging demonstrated a preservation of the entire vascular tree patency. Removal of cells and antigens was confirmed by reduction of DNA and antigen markers negativation. Microscopic evaluation revealed preservation of tissue structures as well as of major proteins. Seeded cells were viable and well distributed within all scaffolds. CONCLUSIONS: Complex acellular facial scaffolds were obtained, preserving simultaneously a cell-friendly extracellular matrix and a perfusable vascular tree. This step will enable further engineering of postmortem facial grafts, thereby offering new perspectives in composite tissue allotransplantation.
Assuntos
Transplante de Face , Engenharia Tecidual/métodos , Biomarcadores/metabolismo , Reatores Biológicos , Citocinas/metabolismo , Matriz Extracelular/metabolismo , Humanos , Imuno-Histoquímica , Técnicas In Vitro , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Perfusão/métodos , Reperfusão/métodos , Alicerces TeciduaisRESUMO
PURPOSE: To determine the ideal implantation site for selective tongue neurostimulation in obstructive sleep apnea, anatomy of the distal branching of the hypoglossal nerve (HGN) was revisited. METHODS: The HGN distal course and intramuscular distribution to the tongue muscles were studied in 17 embalmed and 5 fresh heads (age 60-98, BMI 20-35). Medial branches supplying selectively the genioglossus (GG) muscle were identified. Then, the distinct bundles entering the oblique (GGo) and horizontal (GGh) parts of the GG were located. Morphometric data were compared to similar measurements made on MRI sections from 12 patients (age 43-71, BMI 18-47). RESULTS: The key facts relevant to optimize stimulation and electrode design are the following: the mean width of both GG muscles in embalmed and fresh cadavers was 20.7 ± 2.9 and 21.4 ± 5 mm, respectively; it is significantly (p < 0.05) superior to the MRI value of 18.26 ± 2.0 mm. Selective nervous branches for GGh and GGo were located at 52 ± 8% of hyoid bone-mandibular symphysis distance and at 5.8 ± 1.1 mm from the inferior border of the GG muscle. The surface of stimulation is a 4.4 ± 1.1 × 6.9 ± 3.8 mm ellipse. CONCLUSIONS: According to our observations, the optimal selective or supra-selective stimulation of the tongue protractor muscles can be performed on the lateral surface of the GG at roughly equal distance between the mandibular symphysis and the hyoid bone, at a depth of about 0.6 cm above the GG lower border.
Assuntos
Nervo Hipoglosso/anatomia & histologia , Músculo Esquelético/inervação , Idoso , Idoso de 80 Anos ou mais , Cadáver , Feminino , Humanos , Neuroestimuladores Implantáveis , Masculino , Pessoa de Meia-Idade , Apneia Obstrutiva do Sono/fisiopatologia , Apneia Obstrutiva do Sono/terapiaRESUMO
INTRODUCTION: Desmoid tumors are rare proliferative and invasive benign lesions. They can be sporadic, but in most instances, desmoid tumors develop in the context of Gardner's syndrome with principal localization in the abdominal cavity and abdominal wall. CASE-REPORT: We report the case of a 24-year-old female presenting Gardner's syndrome with a symptomatic abdominal wall desmoid tumor. Lack of response to medical treatment led to surgical management consisting in a complete resection and parietal reconstruction with a biologic mesh. Postoperative course was uneventful and there was no evidence of recurrence at 12 months of follow-up. DISCUSSION: Conventional treatment of abdominal wall desmoid tumors consists in a wide and radical resection. However, complete resection is not always feasible because of difficulty to differentiate the desmoid tumor from adjacent tissues. The surgical approach may require different techniques to repair the parietal defect including prosthetic material such as synthetic or biologic meshes. Biological mesh is an ideal alternative to synthetic graft, mainly in case of infection. CONCLUSION: We have encountered a case of a symptomatic growing desmoid tumor of the abdominal wall in a young patient with Gardner's syndrome, successfully treated by complete resection and reconstruction with a biologic mesh to correct the parietal defect.
Assuntos
Parede Abdominal/cirurgia , Fibromatose Abdominal/cirurgia , Síndrome de Gardner/complicações , Telas Cirúrgicas , Feminino , Fibromatose Abdominal/complicações , Fibromatose Abdominal/patologia , Síndrome de Gardner/cirurgia , Humanos , Adulto JovemRESUMO
Congenital limb anomalies occur in Europe with a prevalence of 3.81/1,000 births and can have a major impact on patients and their families. The present study concerned a female fetus aborted at 23 weeks of gestation because she was affected by non-syndromic bilateral absence of the zeugopod (leg) and autopod (foot). Autopsy of the aborted fetus, X-ray imaging, MRI, and histochemical analysis showed that the distal extremity of both femurs was continued by a cartilage-like mass, without joint cavitation. Karyotype was normal. Moreover, no damaging variant was detected by exome sequencing. The limb characteristics of the fetus, which to our knowledge have not yet been reported in humans, suggest a developmental arrest similar to anomalies described in chicks following surgical experiments on the apical ectodermal ridge of the lower limbs.
Assuntos
Feto/anormalidades , Feto/patologia , Articulação do Joelho/anormalidades , Deformidades Congênitas dos Membros/patologia , Extremidade Inferior/patologia , Adulto , Feminino , Humanos , Articulação do Joelho/patologia , Extremidade Inferior/crescimento & desenvolvimento , Masculino , PrognósticoAssuntos
Doenças Mandibulares/cirurgia , Reconstrução Mandibular/métodos , Osteorradionecrose/cirurgia , Retalhos Cirúrgicos/irrigação sanguínea , Procedimentos Cirúrgicos Vasculares , Adulto , Idoso , Algoritmos , Feminino , Sobrevivência de Enxerto , Humanos , Masculino , Pessoa de Meia-Idade , Complicações Pós-OperatóriasRESUMO
Critical bone loss can have several origins: infections, tumors or trauma. Therefore, massive bone allograft can be a solution for limb salvage. Such a biological reconstruction should have the ideal biomechanical qualities. However, their complication rate remains too high. Perfusion-decellularization of massive allografts could promote the vitality of these grafts, thereby improving their integration and bone remodeling. Three perfusion-decellularized massive bone allografts were compared to 3 fresh frozen massive bone allografts in a preclinical in vivo porcine study using an orthopedic surgery model. Three pigs each underwent a critical diaphyseal femoral defects followed by an allogeneic intercalary femoral graft on their both femurs (one decellularized and one conventional fresh frozen as "native") to reconstruct the defect. Clinical imaging was performed over 3 months of follow-up. The grafts were then explanted and examined by non-decalcified histology, fluoroscopic microscopy and immunohistochemistry. Bone consolidation was achieved in both groups at the same time. However, the volume of bone callus appeared to be greater in the decellularized group. Histology demonstrated a superior bone remodeling in the decellularized group, with a higher number of osteoclasts (p < 0.001) and larger areas of osteoid matrix and newly formed bone as compared to the "native" group. Immunohistochemistry showed a superior vitality and remodeling in both the cortical and medullary cavities for osteocalcin (p < 0.001), Ki67 (p < 0.001), CD3 (p < 0.001) and α-SMA (p < 0.001) as compared the "native" group. Three months after implantation, the decellularized grafts were proven to be biologically more active compared to native grafts. Fluoroscopic microscopy revealed more ossification fronts in the depth of the decellularized grafts (p = 0.021). This pilot study provides the first in vivo demonstration on the enhanced biological capacities of massive bone allograft decellularized by perfusion as compared to conventional massive bone allografts.
Assuntos
Aloenxertos , Transplante Ósseo , Animais , Transplante Ósseo/métodos , Projetos Piloto , Suínos , Fêmur/patologia , Fêmur/diagnóstico por imagem , Remodelação Óssea/fisiologia , Transplante Homólogo/métodos , FemininoRESUMO
In terms of large bone defect reconstructions, massive bone allografts may sometimes be the only solution. However, they are still burdened with a high postoperative complication rate. Our hypothesis is that the immunogenicity of residual cells in the graft is involved in this issue. Decellularization by perfusion might therefore be the answer to process and create more biologically effective massive bone allografts. Seventy-two porcine bones were used to characterize the efficiency of our sodium hydroxide-based decellularization protocol. A sequence of solvent perfusion through each nutrient artery was set up to ensure the complete decellularization of whole long bones. Qualitative (histology and immunohistochemistry [IHC]) and quantitative (fluoroscopic absorbance and enzyme-linked immunosorbent assay) evaluations were performed to assess the decellularization and the preservation of the extracellular matrix in the bone grafts. Cytotoxicity and compatibility were also tested. Comparatively to nontreated bones, our experiments showed a very high decellularization quality, demonstrating that perfusion is mandatory to achieve an entire decellularization. Moreover, results showed a good preservation of the bone composition and microarchitecture, Haversian systems and vascular network included. This protocol reduces the human leukocyte antigen antigenic load of the graft by >50%. The majority of measured growth factors is still present in the same amount in the decellularized bones compared to the nontreated bones. Histology and IHC show that the bones were cell compatible, noncytotoxic, and capable of inducing osteoblastic differentiation of mesenchymal stem cells. Our decellularization/perfusion protocol allowed to create decellularized long bone graft models, thanks to their inner vascular network, ready for in vivo implantation or to be further used as seeding matrices.
Assuntos
Matriz Extracelular , Engenharia Tecidual , Suínos , Animais , Humanos , Engenharia Tecidual/métodos , Matriz Extracelular/química , Perfusão , Transplante Ósseo , Aloenxertos , Alicerces Teciduais/químicaRESUMO
Large bone defect regeneration remains a major challenge for orthopedic surgeons. Tissue engineering approaches are therefore emerging in order to overcome this limitation. However, these processes can alter some of essential native tissue properties such as intermolecular crosslinks of collagen triple helices, which are known for their essential role in tissue structure and function. We assessed the persistence of extracellular matrix (ECM) properties in human fascia lata (HFL) and periosteum (HP) after tissue engineering processes such as decellularization and sterilization. Harvested from cadaveric donors (N = 3), samples from each HFL and HP were decellularized following five different chemical protocols with and without detergents (D1-D4 and D5, respectively). D1 to D4 consisted of different combinations of Triton, Sodium dodecyl sulfate and Deoxyribonuclease, while D5 is routinely used in the institutional tissue bank. Decellularized HFL tissues were further gamma-irradiated (minimum 25 kGy) in order to study the impact of sterilization on the ECM. Polarized light microscopy (PLM) was used to estimate the thickness and density of collagen fibers. Tissue hydration and content of hydroxyproline, enzymatic crosslinks, and non-enzymatic crosslinks (pentosidine) were semi-quantified with Raman spectroscopy. ELISA was also used to analyze the maintenance of the decorin (DCN), an important small leucine rich proteoglycan for fibrillogenesis. Among the decellularization protocols, detergent-free treatments tended to further disorganize HFL samples, as more thin fibers (+53.7%) and less thick ones (-32.6%) were recorded, as well as less collagen enzymatic crosslinks (-25.2%, p = 0.19) and a significant decrease of DCN (p = 0.036). GAG content was significantly reduced in both tissue types after all decellularization protocols. On the other hand, HP samples were more sensitive to the D1 detergent-based treatments, with more disrupted collagen organization and greater, though not significant loss of enzymatic crosslinks (-37.4%, p = 0.137). Irradiation of D5 HFL samples, led to a further and significant loss in the content of enzymatic crosslinks (-29.4%, p = 0.037) than what was observed with the decellularization process. Overall, the results suggest that the decellularization processes did not significantly alter the matrix. However, the addition of a gamma-irradiation is deleterious to the collagen structural integrity of the tissue.
RESUMO
INTRODUCTION: Postmortem evaluation of the human vascular system has a long history, with advancements ranging from dissections to modern imaging techniques like computed tomography (CT scan). This study designs a novel combination of Angiofil, a liquid radiopaque polymer, and latex, a flexible cast material, for cadaveric vascular analysis. MATERIAL & METHODS: The aim was to synergize the advantages of both components, providing accurate radiological images and optimal dissection conditions. Three arterial territories (lateral circumflex femoral artery, profunda brachii artery, and radial artery) were injected and assessed through CT scans and dissections. RESULTS: The Angiofil-latex mixture allowed successful visualization of the vascular networks, offering a simple, reproducible, and non-toxic approach. Quantitative assessments of the three territories, including diameters and lengths, showed comparable results between CT scan and dissection. DISCUSSION: The technique precision and versatility make it an accessible and valuable tool for anatomical studies, potentially extending its application to MRI analyses. Overall, the Angiofil-latex combination presents a cost-effective solution for researchers, offering enhanced visibility and detailed anatomical insights for various applications, including anatomical variation studies.
RESUMO
Last twenties, tissue engineering has rapidly advanced to address the shortage of organ donors. Decellularization techniques have been developed to mitigate immune rejection and alloresponse in transplantation. However, a clear definition of effective decellularization remains elusive. This study compares various decellularization protocols using the human fascia lata model. Morphological, structural and cytotoxicity/viability analyses indicated that all the five tested protocols were equivalent and met Crapo's criteria for successful decellularization. Interestingly, only the in vivo immunization test on rats revealed differences. Only one protocol exhibited Human Leucocyte Antigen (HLA) content below 1% residual threshold, the only criterion preventing rat immunization with an absence of rat anti-human IgG switch after one month (N=4 donors for each of the 7 groups, added by negative and positive controls, n=28). By respecting a refined set of criteria, i.e. lack of visible nuclear material, <50ng DNA/mg dry weight of extracellular matrix, and <1% residual HLA content, the potential for adverse host reactions can be drastically reduced. In conclusion, this study emphasizes the importance of considering not only nuclear components but also major histocompatibility complex in decellularization protocols and proposes new guidelines to promote safer clinical development and use of bioengineered scaffolds.
RESUMO
Continuous positive airway pressure (CPAP) is an effective but cumbersome treatment for obstructive sleep apnoea (OSA). Noncompliant patients need alternative therapies. We studied a tongue neurostimulation approach: targeted hypoglossal neurostimulation (THN) therapy with the aura6000™ System. A multi-contact electrode positioned around the main trunk of the twelfth nerve connected to an implanted pulse generator stimulates segments of the nerve, activating dilator muscles. The primary objective was to improve the polysomnographically determined apnoea/hypopnoea index (AHI) at 3 months, and maintain the improvement after 12 months of treatment. 13 out of 14 operated patients were successfully implanted. At 12 months, the AHI decreased from 45±18 to 21±17, a 53% reduction (p<0.001). The 4% oxygen desaturation index fell from 29±20 to 15±16 and the arousal index from 37±13 to 25±14, both p<0.001. The Epworth sleepiness scale decreased from 11±7 to 8±4 (p=0.09). THN was neither painful nor awakened patients, who all complied with therapy. There were two transient tongue paresis. The present study represents the longest study of any hypoglossal neurostimulation reported to date. We conclude that THN is safe and effective to treat OSA in patients not compliant with CPAP.
Assuntos
Terapia por Estimulação Elétrica/métodos , Apneia Obstrutiva do Sono/terapia , Língua/inervação , Adulto , Eletrodos , Endoscopia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Modelos Estatísticos , Segurança do Paciente , Projetos Piloto , Polissonografia , Apneia Obstrutiva do Sono/cirurgia , Resultado do TratamentoRESUMO
Hoxa2 is a homeotic transcription factor, which is downregulated once chondrogenic differentiation is initiated. We previously generated a transgenic mouse model, which turns Hoxa2 on in cells expressing Collagen II A1, i.e. in cells entering chondrogenesis. As a consequence, mice display a general embryonic delay of ossification and then a postnatal growth defect. Col2a1-Cre mice were crossed with an inducible ß-actin driven Hoxa2 transgene. Spines, vertebrae and limbs were measured and skeletal elements were studied by X-ray, microCT, pQCT, TEM, western-blotting, histomorphometry and immunohistochemistry. Mice expressing Hoxa2 in chondrogenic cells feature a proportionate short stature phenotype with a severe lordosis, which appeared significant from postnatal day 4. Analysis of both cartilage and bone development in affected embryos and mice from birth till P35 did not reveal any major defect in histogenesis, except a reduced number of chondrocytes in the vertebral anlage at E13.5. In conclusion, the sustained expression of Hoxa2 in the chondrocyte lineage is characterized by a proportionate short stature resulting from skeletal growth defect. The indepth analysis of cartilage and bone histogenesis points towards an initial deficit in cell mobilization to enter chondrogenesis.