Benoît Lengelé

Vascularization remains a major limitation in tissue engineering, particularly for large or complex grafts requiring rapid perfusion after implantation. Perfusion-based decellularization is a promising strategy to preserve native vascular architecture while removing cellular components. In this study, human ALT flaps (n = 5) were perfusion-decellularized via the main nutrient perforator artery. Flaps were treated intact to preserve the fascia lata vascular network prior to its careful harvest for subsequent analyses. Decellularization and selected matrix components were assessed by histology, immunohistochemistry, biochemical assays, and angiogenic profiling. Complete cellular removal was achieved, with residual DNA below accepted thresholds and near-total HLA antigens signal depletion. Collagen architecture and basement membrane components were largely preserved, while glycosaminoglycan were reduced. Contrast-enhanced imaging demonstrated retention of macroscopic vascular architecture, although functional vascular properties were not assessed. Several matrix-associated angiogenic factors, including VEGF, PDGF-BB, and IGF-1, remained detectable after decellularization in an exploratory analysis. These findings demonstrate that perfusion decellularization can generate human fascia lata scaffolds combining effective cellular removal with retention of native vascular architecture and selected matrix-associated biochemical cues. Further studies are required to establish vascular functionality, angiogenic bioactivity, and recellularization potential.

Background: Numerous protocols exist concerning the decellularization of the esophagus, a potential alternative to the classical surgical approach for the reconstruction of the digestive tract after esophagectomy. This systematic literature review (SLR) aimed to provide an overview of the effectiveness of the current protocols. Methods: This SLR was conducted in PubMed, EMBASE, and Scopus until September 2025. Study selection, data extraction, and quality assessment were performed by two independent reviewers according to the inclusion/exclusion criteria. Results: A total of 2494 references were screened after removing duplicates. Among these references, 26 articles were included. The large majority of studies (24/26) used Sodium Dodecyl Sulfate (SDS) or Sodium DeoxyCholate (SDC), and the most common physical method was the cannulation of the esophagus (17/26). The animal model was very heterogenous. All protocols except one showed no residual cell nuclei, with only 5/19 papers confirming a satisfactory residual amount of DNA. The assessment of the extracellular matrix (ECM)—mostly qualitative—revealed global preservation but with a systematic loss of glycosaminoglycans (GAGs). Conclusions: The decellularization of the esophagus is feasible, but the definition of the optimal protocol to achieve this goal remains difficult because of the important heterogeneity among the different studies.

Background: In terms of large bone defects 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. Material and methods: 72 porcine bones were used to characterize the efficiency of our NaOH-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 & Immunohistochemistry) and quantitative (fluoroscopic absorbance & ELISA) evaluations were performed to assess the decellularization and the preservation of the extracellular matrix in the bone grafts. Cytotoxicity and compatibility were also tested. Results: Comparatively to non-treated 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 HLA antigenic load of the graft by more than 50%. The majority of measured growth factors are still present in the same amount into the decellularized bones compared to the non-treated bones. Histology and immunohistochemistry show that the bones were cell-compatible, non-cytotoxic and capable of inducing osteoblastic differentiation of MSCs. Conclusion: Our decellularization/perfusion protocol allowed to create decellularized long bone grafts models thanks to their inner vascular network, ready for in vivo implantation or to be further used as seeding matrices.

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.

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.

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.