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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.
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Fascia Lata , Antígenos HLA , Engenharia Tecidual , Humanos , Animais , Engenharia Tecidual/métodos , Antígenos HLA/imunologia , Ratos , Alicerces Teciduais/química , Materiais Biocompatíveis/química , Masculino , Matriz Extracelular Descelularizada/química , Matriz Extracelular/química , Matriz Extracelular/metabolismoRESUMO
The fascial system has gained recognition for its integral role in connecting skin, superficial and deep fasciae, and underlying muscles. However, consensus on its microstructure depending on its topography remains elusive as well as its implications in clinical practices, such as reconstructive surgery and physiotherapy techniques. This study focuses on the iliotibial tract (ITT) implicated in the iliotibial band syndrome. The goal is to describe microstructural characteristics using classical 2D histology and cryogenic contrast-enhanced microcomputed tomography (cryo-CECT) such as the total thickness, number of layers, layer thickness, fibre orientation and tortuosity, according to the specific topography. The total thickness of the ITT varied across topographic regions, with the superior part being on average thicker but non-significantly different from the other regions. The inferior part showed heterogeneity, with the anterior region (AI) being the thinnest and the posterior one (PI) the thickest. The ITT exhibited 1-3 layers, with no significant differences among regions. Most commonly, it consisted of two layers, except for the antero-superior (AS) and antero-middle (AM) regions, which sometimes had only one layer. The posterior regions frequently had 2 or 3 layers, with the PI region having the highest mean (2.7 layers). The intermediate layer was the thickest one, varying from the AI region (0.368 mm ± 0.114) to the PI region (0.640 mm ± 0.305). The superficial layer showed regional variability, with the AS region being the thinnest. The deep layer appeared thinner than the superficial one. Fibre orientation analysis indicated that the intermediate layer mainly consisted of oblique longitudinal fibres, orientated downward and forward, while the superficial and deep layers had transversal or oblique transversal fibres. Cryo-CECT 3D observations confirmed these findings, revealing distinct orientations for different layers. Fibre tortuosity exhibited differences based on orientation. Transversal fibres (>65°) were significantly less tortuous than longitudinal fibres (<25°) and oblique intermediate fibres (25°-65°), aligning with 3D plot observations. This quantitative study highlights various microstructural characteristics of the ITT, offering insights into its regional variations. The analysis accuracy is increased due to the novel technology of cryo-CECT which emerges as a valuable tool for precise assessment of 3D fibre orientation and tortuosity. These findings contribute to a deeper understanding of the ITT structure, useful in clinical practices, such as reconstructive surgery and physiotherapy, and future research endeavours.
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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.
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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.
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Aloenxertos , Transplante Ósseo , Animais , Transplante Ósseo/métodos , Projetos Piloto , Suínos , Fêmur/patologia , Fêmur/diagnóstico por imagem , Remodelação Óssea/fisiologia , Transplante Homólogo/métodos , FemininoRESUMO
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.
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Aloenxertos , Fêmur , Hidróxido de Sódio , Animais , Suínos , Hidróxido de Sódio/farmacologia , Fêmur/patologia , Transplante Ósseo , Perfusão , Estresse Mecânico , Fenômenos BiomecânicosRESUMO
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.
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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.
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Transplante de Face , Alotransplante de Tecidos Compostos Vascularizados , Humanos , Transplante de Face/métodos , Consenso , Técnica Delphi , Projetos de PesquisaRESUMO
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.
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Matriz Extracelular , Engenharia Tecidual , Suínos , Animais , Humanos , Engenharia Tecidual/métodos , Matriz Extracelular/química , Perfusão , Transplante Ósseo , Aloenxertos , Alicerces Teciduais/químicaRESUMO
Background: Calcific aortic stenosis (AS) is the most prevalent heart valve disease in developed countries. The aortic valve cusps progressively thicken and the valve does not open fully due to the presence of calcifications. In vivo imaging, usually used for diagnosis, does not allow the visualization of the microstructural changes associated with AS. Methods: Ex vivo high-resolution microfocus computed tomography (microCT) was used to quantitatively describe the microstructure of calcified aortic valve cusps in full 3D. As case study in our work, this quantitative analysis was applied to normal-flow low-gradient severe AS (NF-LG-SAS), for which the medical prognostic is still highly debated in the current literature, and high-gradient severe AS (HG-SAS). Results: The volume proportion of calcification, the size and number of calcified particles and their density composition was quantified. A new size-based classification considering small-sized particles that are not detected with in vivo imaging was defined for macro-, meso- and microscale calcifications. Volume and thickness of aortic valve cusps, including the complete thickness distribution, were also determined. Moreover, changes in the cusp soft tissues were also visualized with microCT and confirmed by scanning electron microscopy images of the same sample. NF-LG-SAS cusps contained lower relative amount of calcifications than HG-SAS. Moreover, the number and size of calcified objects and the volume and thickness of the cusps were also lower in NF-LG-SAS cusps than in HG-SAS. Conclusions: The application of high-resolution ex vivo microCT to stenotic aortic valve cusps provided a quantitative description of the general structure of the cusps and of the calcifications present in the cusp soft tissues. This detailed description could help in the future to better understand the mechanisms of AS.
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The lack of viability of massive bone allografts for critical-size bone defect treatment remains a challenge in orthopedic surgery. The literature has reviewed the advantages of a multi-combined treatment with the synergy of an osteoconductive extracellular matrix (ECM), osteogenic stem cells, and growth factors (GFs). Questions are still open about the need for ECM components, the influence of the decellularization process on the latter, the related potential loss of function, and the necessity of using pre-differentiated cells. In order to fill in this gap, a bone allograft surrounded by an osteogenic membrane made of a decellularized collagen matrix from human fascia lata and seeded with periosteal mesenchymal stem cells (PMSCs) was analyzed in terms of de-/recellularization, osteogenic properties, PMSC self-differentiation, and angiogenic potential. While the decellularization processes altered the ECM content differently, the main GF content was decreased in soft tissues but relatively increased in hard bone tissues. The spontaneous osteogenic differentiation was necessarily obtained through contact with a mineralized bone matrix. Trying to deepen the knowledge on the complex matrix-cell interplay could further propel these tissue engineering concepts and lead us to provide the biological elements that allow bone integration in vivo.
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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.
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Vértebras Lombares , Medula Espinal , Humanos , Masculino , Feminino , Medula Espinal/anatomia & histologia , Imageamento por Ressonância Magnética , Região Lombossacral , CadáverRESUMO
INTRODUCTION: Large bone defects are challenging for surgeons. Available reimplanted bone substitutes can't properly restore optimal function along and long term osteointegration of the bone graft. Bone substitute based on the perfusion-decellularization technique seem to be interesting in order to overcome these limitations. We present here an evaluation of the biomechanics of the bones thus obtained. MATERIAL AND METHODS: Two decellularization protocols were chosen for this study. One using Sodium Dodecyl Sulfate (SDS) (D1) and one using NaOH and H2O2 (D2). The decellularization was performed on porcine forearms. We then carried out compression, three-point bending, indentation and screw pull-out tests on each sample. Once these tests were completed, we compared the results obtained between the different decellularization protocols and with samples left native. RESULTS: The difference in the means was similar between the tests performed on bones decellularized with the SDS protocol and native bones for pull-out test: +1.4% (CI95% [-10.5%- 12.4%]) of mean differences when comparing Native vs D1, compression -14.9% (CI95% [-42.7%- 12.5%]), 3-point bending -5.7% (CI95% [-22.5%- 11.1%]) and indentation -10.8% (CI95% [-19.5%- 4.6%]). Bones decellularized with the NaOH protocol showed different results from those obtained with the SDS protocol or native bones during the pull-out screw +40.7% (CI95% [24.3%- 57%]) for Native vs D2 protocol and 3-point bending tests +39.2% (CI95% [13.7%- 64.6%]) for Native vs D2 protocol. The other tests, compression and indentation, gave similar results for all our samples. CONCLUSION: Vascularized decellularized grafts seem to be an interesting means for bone reconstruction. Our study shows that the decellularization method affects the mechanical results of our specimens. Some methods seem to limit these alterations and could be used in the future for bone decellularization.
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Antebraço , Peróxido de Hidrogênio , Suínos , Animais , Hidróxido de Sódio , Fenômenos Biomecânicos , Artrodese , Alicerces Teciduais , Engenharia Tecidual , Matriz ExtracelularRESUMO
Sirolimus, by targeting the mammalian target of rapamycin (mTOR) pathway, has demonstrated efficacy on lymphatic malformations (LMs) in adults and neonates. The current hypothesis is that the earlier the lesion is treated, the better it responds. This has prompted the idea that sirolimus administration might be efficacious to treat fetal LMs as well. Here we report a successful management of a cervicofacial fetal LM with sirolimus taken orally by the mother from the 22nd week of pregnancy until 2 weeks before planned delivery. Repeated cordocentesis recorded a 30% transplacental crossing of sirolimus. Continuation of sirolimus after birth allowed resection of the residual mass. We have followed the physical and neurological evolution of the child for 6 years since the fetal administration of sirolimus. We conclude that early administration of sirolimus during pregnancy with maternal serum monitoring may be proposed to high-risk fetal LMs in selected cases.
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Sirolimo , Humanos , Sirolimo/uso terapêutico , Sirolimo/administração & dosagem , Feminino , Gravidez , Resultado do Tratamento , Anormalidades Linfáticas/tratamento farmacológico , Adulto , Administração Oral , Recém-Nascido , Ultrassonografia Pré-Natal , Cordocentese , Inibidores de MTOR/uso terapêutico , Idade Gestacional , Troca Materno-FetalRESUMO
Introduction: Nipple-areolar complex (NAC) reconstruction after breast cancer surgery is challenging and does not always provide optimal long-term esthetic results. Therefore, generating a NAC using tissue engineering techniques, such as a decellularization-recellularization process, is an alternative option to recreate a specific 3D NAC morphological unit, which is then covered with an in vitro regenerated epidermis and, thereafter, skin-grafted on the reconstructed breast. Materials and methods: Human NACs were harvested from cadaveric donors and decellularized using sequential detergent baths. Cellular clearance and extracellular matrix (ECM) preservation were analyzed by histology, as well as by DNA, ECM proteins, growth factors, and residual sodium dodecyl sulfate (SDS) quantification. In vivo biocompatibility was evaluated 30 days after the subcutaneous implantation of native and decellularized human NACs in rats. In vitro scaffold cytocompatibility was assessed by static seeding of human fibroblasts on their hypodermal side for 7 days, while human keratinocytes were seeded on the scaffold epidermal side for 10 days by using the reconstructed human epidermis (RHE) technique to investigate the regeneration of a new epidermis. Results: The decellularized NAC showed a preserved 3D morphology and appeared white. After decellularization, a DNA reduction of 98.3% and the absence of nuclear and HLA staining in histological sections confirmed complete cellular clearance. The ECM architecture and main ECM proteins were preserved, associated with the detection and decrease in growth factors, while a very low amount of residual SDS was detected after decellularization. The decellularized scaffolds were in vivo biocompatible, fully revascularized, and did not induce the production of rat anti-human antibodies after 30 days of subcutaneous implantation. Scaffold in vitro cytocompatibility was confirmed by the increasing proliferation of seeded human fibroblasts during 7 days of culture, associated with a high number of living cells and a similar viability compared to the control cells after 7 days of static culture. Moreover, the RHE technique allowed us to recreate a keratinized pluristratified epithelium after 10 days of culture. Conclusion: Tissue engineering allowed us to create an acellular and biocompatible NAC with a preserved morphology, microarchitecture, and matrix proteins while maintaining their cell growth potential and ability to regenerate the skin epidermis. Thus, tissue engineering could provide a novel alternative to personalized and natural NAC reconstruction.
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Introduction: The human fascia lata (HFL) is used widely in reconstructive surgery in indications other than fracture repair. The goal of this study was to compare microscopic, molecular, and mechanical properties of HFL and periosteum (HP) from a bone tissue engineering perspective. Material and Methods: Cadaveric HP and HFL (N = 4 each) microscopic morphology was characterized using histology and immunohistochemistry (IHC), and the extracellular matrix (ECM) ultrastructure assessed by means of scanning electron microscopy (SEM). DNA, collagen, elastin, glycosaminoglycans, major histocompatibility complex Type 1, and bone morphogenetic protein (BMP) contents were quantified. HP (N = 6) and HFL (N = 11) were submitted to stretch tests. Results: Histology and IHC highlighted similarities (Type I collagen fibers and two-layer organization) but also differences (fiber thickness and compaction and cell type) between both tissues, as confirmed using SEM. The collagen content was statistically higher in HFL than HP (735 vs. 160.2 µg/mg dry weight, respectively, p < 0.0001). On the contrary, DNA content was lower in HFL than HP (404.75 vs. 1,102.2 µg/mg dry weight, respectively, p = 0.0032), as was the immunogenic potential (p = 0.0033). BMP-2 and BMP-7 contents did not differ between both tissues (p = 0.132 and p = 0.699, respectively). HFL supported a significantly higher tension stress than HP. Conclusion: HP and HFL display morphological differences, despite their similar molecular ECM components. The stronger stretching resistance of HFL can specifically be explained by its higher collagen content. However, HFL contains many fewer cells and is less immunogenic than HP, as latter is rich in periosteal stem cells. In conclusion, HFL is likely suitable to replace HP architecture to confer a guide for bone consolidation, with an absence of osteogenicity. This study could pave the way to a bio-engineered periosteum built from HFL.
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Introduction: Durable reconstruction of critical size bone defects is still a surgical challenge despite the availability of numerous autologous and substitute bone options. In this paper, we have investigated the possibility of creating a living bone allograft, using the perfusion/decellularization/recellularization (PDR) technique, which was applied to an original model of vascularized porcine bone graft. Materials and Methods: 11 porcine bone forelimbs, including radius and ulna, were harvested along with their vasculature including the interosseous artery and then decellularized using a sequential detergent perfusion protocol. Cellular clearance, vasculature, extracellular matrix (ECM), and preservation of biomechanical properties were evaluated. The cytocompatibility and in vitro osteoinductive potential of acellular extracellular matrix were studied by static seeding of NIH-3T3 cells and porcine adipose mesenchymal stem cells (pAMSC), respectively. Results: The vascularized bone grafts were successfully decellularized, with an excellent preservation of the 3D morphology and ECM microarchitecture. Measurements of DNA and ECM components revealed complete cellular clearance and preservation of ECM's major proteins. Bone mineral density (BMD) acquisitions revealed a slight, yet non-significant, decrease after decellularization, while biomechanical testing was unmodified. Cone beam computed tomography (CBCT) acquisitions after vascular injection of barium sulphate confirmed the preservation of the vascular network throughout the whole graft. The non-toxicity of the scaffold was proven by the very low amount of residual sodium dodecyl sulfate (SDS) in the ECM and confirmed by the high live/dead ratio of fibroblasts seeded on periosteum and bone ECM-grafts after 3, 7, and 16 days of culture. Moreover, cell proliferation tests showed a significant multiplication of seeded cell populations at the same endpoints. Lastly, the differentiation study using pAMSC confirmed the ECM graft's potential to promote osteogenic differentiation. An osteoid-like deposition occurred when pAMSC were cultured on bone ECM in both proliferative and osteogenic differentiation media. Conclusion: Fully decellularized bone grafts can be obtained by perfusion decellularization, thereby preserving ECM architecture and their vascular network, while promoting cell growth and differentiation. These vascularized decellularized bone shaft allografts thus present a true potential for future in vivo reimplantation. Therefore, they may offer new perspectives for repairing large bone defects and for bone tissue engineering.
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Gestational gigantomastia (GGM) is a rare condition characterized by a massive overgrowth of breast tissue during pregnancy. Surgical sanction may be required when conservative measures fail. In this study, we report the case of a 29-year-old woman who presented with an evolutive GGM responsible for physical and emotional distress, despite medical treatment. A multidisciplinary decision was made to induce delivery at 32 weeks. In the postdelivery period, the patient developed breast wounds, complicated with septic cardiomyopathy. An emergency bilateral mastectomy was then carried out, together with banking of both nipple-areola complexes. Thereafter, delayed bilateral 2-stage breast reconstruction was started at 12 months with subcutaneous tissue expanders, later on followed by implants removal and autologous reconstruction with bilateral deep inferior epigastric artery perforator flaps and bilateral nipple replantation.
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BACKGROUND: There is a renewed interest for prepectoral reconstruction. We aimed to describe the feasibility and the early complications associated with immediate one-stage direct-to-implant (DTI) reconstruction using prepectoral anatomical polyurethane (PU) foam-coated implants alone, for women with breast cancer or mutation carriers undergoing risk-reducing surgery. METHODS: We performed a single-center, retrospective review of 50 patients (mean age of 49 years), who underwent skin-sparing mastectomy (SSM) or nipple-sparing mastectomy (NSM) and immediate prepectoral PU implant-based reconstruction. All procedures were performed by the same senior operator, from July 2018 to March 2020. RESULTS: A total of 64 mastectomies (25 SSMs and 39 NSMs) with one-stage prepectoral PU foam-coated implant reconstruction were performed. Out of 50 patients, 6 required surgical revision within 30 days, because of hematoma (2), wound dehiscence (2) infection (1), and full thickness nipple-areolar complex (NAC) necrosis (1). Four patients developed a cutaneous rash with spontaneous resolution. Statistical analysis showed a significant influence of hypothyroidism and previous radiotherapy on the risk of complications. The association with prior radiotherapy (pRT) was not significant using binary logistic regression. When excluding oncological reasons and patient's wish for NAC excision, our decision to perform an NSM was influenced by breast cup size, preoperative measurements, and breast weight. CONCLUSIONS: Early experience with immediate prepectoral DTI reconstruction with PU-covered implants alone suggests that it is a reliable procedure. Prior breast irradiation does not increase postoperative complication rates in our series. NAC preservation was decided according to preoperative lower breast measurements.
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Implantes de Mama , Neoplasias da Mama/cirurgia , Mamoplastia/métodos , Estudos Retrospectivos , Materiais Revestidos Biocompatíveis , Estudos de Viabilidade , Feminino , Humanos , Mastectomia/métodos , Pessoa de Meia-Idade , Tamanho do Órgão , Músculos Peitorais , Poliuretanos , Complicações Pós-Operatórias , ReoperaçãoRESUMO
SUMMARY: Postoperative tracheoesophageal or bronchoesophageal fistulas represent a major surgical challenge. The authors report the description of an original perforator-based intercostal artery muscle flap, aiming to cover all types of intrathoracic fistulas, from any location, in difficult cases such as postoperative fistulas after esophagectomy in an irradiated field. Between June of 2016 and January of 2019, eight male patients were treated with a perforator-based intercostal artery muscle flap. All had previous surgery for esophageal cancer and developed a tracheoesophageal or bronchoesophageal fistula during the perioperative course. The mean patient age was 55.9 ± 8.8 years. All patients received neoadjuvant chemotherapy and seven received neoadjuvant radiation therapy. A perforator-based intercostal artery muscle flap, with a mean skin paddle size of 9.86 × 5 cm, was harvested. The median operative time was 426.50 minutes. The tracheoesophageal or bronchoesophageal fistula was successfully and definitively occluded in three patients; two patients experienced recurrence; and one patient underwent re operation. At 1 year, five patients were alive (62.5 percent), and among them, three (37.5 percent) were free from any intrathoracic complications. Three patients died, because of massive digestive bleeding, mesenteric ischemia, and multiorgan failure, respectively. The perforator-based intercostal artery muscle flap, like the Taylor flap in abdominoperineal reconstruction, could become a workhorse flap for all intrathoracic reconstructions, as it can always be harvested, even if a previous thoracotomy has ruined most of the options. This surgical technique, easily feasible, reliable, and reproducible, became our first option for all postoperative tracheoesophageal or bronchoesophageal fistula patients during the postoperative course following esophagectomy. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.
Assuntos
Fístula Brônquica/cirurgia , Fístula Esofágica/cirurgia , Neoplasias Esofágicas/cirurgia , Retalho Perfurante/irrigação sanguínea , Complicações Pós-Operatórias/cirurgia , Fístula Traqueoesofágica/cirurgia , Artérias , Humanos , Músculos Intercostais/irrigação sanguínea , Masculino , Pessoa de Meia-IdadeRESUMO
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.