Diaphragmatic hernia repair porcine model to compare the performance of biodegradable membranes against Gore-Tex®.

BACKGROUND: Patch repair of congenital diaphragmatic hernia (CDH) using Gore-Tex® is associated with infection, adhesions, hernia recurrence, long-term musculoskeletal sequels and poor tissue regeneration. To overcome these limitations, the performance of two novel biodegradable membranes was tested to repair CDH in a growing pig model. METHODS: Twelve male pigs were randomly assigned to 3 different groups of 4 animals each, determined by the type of patch used during thoracoscopic diaphragmatic hernia repair (Gore-Tex®, polycaprolactone electrospun membrane-PCLem, and decellularized human chorion membrane-dHCM). After 7 weeks, all animals were euthanized, followed by necropsy for diaphragmatic evaluation and histological analysis. RESULTS: Thoracoscopic defect creation and diaphragmatic repair were performed without any technical difficulty in all groups. However, hernia recurrence rate was 0% in Gore-Tex®, 50% in PCLem and 100% in dHCM groups. At euthanasia, Gore-Tex® patches appeared virtually unchanged and covered with a fibrotic capsule, while PCLem and dHCM patches were replaced by either floppy connective tissue or vascularized and floppy regenerated membranous tissue, respectively. CONCLUSION: Gore-Tex® was associated with a higher survival rate and lower recurrence. Nevertheless, the proposed biodegradable membranes were associated with better tissue integration when compared with Gore-Tex®.

Analytical validation and algorithm improvement of HepatoPredict kit to assess hepatocellular carcinoma prognosis before a liver transplantation.

Objectives: To verify the analytical performance of the HepatoPredict kit, a novel tool developed to stratify Hepatocellular Carcinoma (HCC) patients according to their risk of relapse after a Liver Transplantation (LT). Methods: The HepatoPredict tool combines clinical variables and a gene expression signature in an ensemble of machine-learning algorithms to forecast the benefit of a LT in HCC patients. To ensure the accuracy and reliability of this method, extensive analytical validation was conducted to verify its specificity and robustness. The experiments were designed following the guidelines for multi-target genomic assays such as ISO201395-2019, MIQE, CLSI-MM16, CLSI-MM17, and CLSI-EP17-A. The validation process included reproducibility between operators and between RNA extractions and RT-qPCR runs, and interference of input RNA levels or varying reagent levels. A recently retrained version of the HepatoPredict algorithms was also tested. Results: The validation process demonstrated that the HepatoPredict kit met the required standards for robustness (p > 0.05), analytical specificity (inclusivity of 95 %), and sensitivity (LoB, LoD, linear range, and amplification efficiency between 90 and 110 %). The operator, equipment, input RNA, and reagents used had no significant effect on the HepatoPredict results. Additionally, the testing of a recently retrained version of the HepatoPredict algorithm, showed that this new version further improved the accuracy of the kit and performed better than existing clinical criteria in accurately identifying HCC patients who are more likely to benefit LT. Conclusions: Even with the introduced variations in molecular and clinical variables, the HepatoPredict kit's prognostic information remains consistent. It can accurately identify HCC patients who are more likely to benefit from a LT. Its robust performance also confirms that it can be easily integrated into standard diagnostic laboratories.

New Vascular Graft Using the Decellularized Human Chorion Membrane.

The increase of both arterial occlusive diseases and coronary heart diseases leads to a higher demand for small-diameter vascular grafts (<6 mm). The gold standard for small-diameter vessel replacement is the use of autologous veins. Nevertheless, up to 30% of these patients need to use vascular grafts. Although synthetic polymers have been successfully used for the replacement of large-diameter vascular grafts (>6 mm), they are associated with thrombosis, intimal hyperplasia, calcification, and chronic inflammation when used as small-diameter vascular grafts. Therefore, natural materials have been studied for this application. In this study, a decellularized human chorion membrane (dHCM) vascular graft with a 3-4 mm diameter was created. Herein, the biocompatibility of dHCM with endothelial cells was demonstrated in vitro and ex ovo. Blood biocompatibility of dHCM was also shown by studying plasma protein adsorption, platelet adhesion and activation, and its hemolytic potential. Furthermore, dHCM antibacterial properties against Staphylococcus aureus were also studied. In summary, the dHCM reticular layer side presented all the needed characteristics to be used in the inner side of a vascular graft. Additionally, the mechanical properties of the dHCM tubular construct were studied, being similar to the ones of the saphenous vein, the gold standard for autologous small-diameter vessel replacement.

Method to decellularize the human chorion membrane.

The human placenta is considered a biological waste, thus it is a great source of extracellular matrix (ECM) proteins. The human chorion membrane (HCM) is a membrane that composes the human placenta and is constituted by collagens type I, II, IV, V and VI, fibronectin and laminin. To the best of our knowledge, the potential of HCM alone is largely unexplored as a substrate to be used in tissue engineering and regenerative medicine. In this work, we describe, for the first time, the process and method to decellularize the chorion membrane alone. To verify the success of the decellularization protocol, the presence and distribution of cell nuclei and double-stranded DNA were quantified and analyzed by DAPI staining, PicoGreen and electrophoresis. After the decellularization protocol an ECM compact and handleably membrane is obtained, the decellularized human chorion membrane (dHCM).

Decellularized Human Chorion Membrane as a Novel Biomaterial for Tissue Regeneration.

Although some placenta-derived products are already used for tissue regeneration, the human chorion membrane (HCM) alone has been poorly explored. In fact, just one study uses decellularized HCM (dHCM) with native tissue architecture (i.e., without extracellular matrix (ECM) suspension creation) as a substrate for cell differentiation. The aim of this work is to fully characterize the dHCM for the presence and distribution of cell nuclei, DNA and ECM components. Moreover, mechanical properties, in vitro biological performance and in vivo biocompatibility were also studied. Our results demonstrated that the HCM was successfully decellularized and the main ECM proteins were preserved. The dHCM has two different surfaces, the reticular layer side and the trophoblast side; and is biocompatible both in vitro and in vivo. Importantly, the in vivo experiments demonstrated that on day 28 the dHCM starts to be integrated by the host tissue. Altogether, these results support the hypothesis that dHCM may be used as a biomaterial for different tissue regeneration strategies, particularly when a membrane is needed to separate tissues, organs or other biologic compartments.