Design and Assembly Procedures for Large-Sized Biohybrid Scaffolds as Patches for Myocardial Infarct

[EN] Objective: To assemble a biohybrid cardiac patch consisting of a large (5x5 cm) elastomer scaffold whose pores are filled with a self-assembling peptide (SAP) gel entrapping adipose stem cells, to be used as a novel implant in a big animal model (sheep) of myocardial infarction. The study focuses on the way to determine optimal procedures for incorporating the SAP solution and the cells in the patch to ensure cell colonization and a homogeneous cell distribution in the construct before implantation. The problems associated with the scale-up of the different procedures raised by the large size of the construct are discussed. Materials and Methods: Experiments were performed to choose between different assembling alternatives: incorporation of the SAP gel before cell seeding or simultaneous SAP and cell loading of the scaffold; surface seeding of cells or cell injection into the scaffold pores; dissemination of the cells throughout the scaffold before incubation by gentle shaking or by centrifugation. Immunocytochemistry techniques and confocal and scanning electron microscopies were employed to assess and quantify cell colonization of the material and early cell distribution. Cell concentrations and the uniformity of cellular distribution throughout the scaffold were taken as the main criteria to decide between the different alternative procedures. Results: The combination of peptide preloading, cell injection, and shaking before incubation yielded the best results in terms of greater cell density and the most uniform distribution after 24 h of culture compared with the other methods. These techniques could be scaled-up to obtain large biohybrid cardiac patches with success. Conclusions: The results obtained after the different seeding methods allowed us to establish an effective protocol for the assembly of large biohybrid patches for their subsequent implantation in the heart of a big animal model of myocardial infarct in the context of a preclinical study.The authors acknowledge the financing from the European Commission through the ‘‘Regeneration of cardiac tissue assisted by bioactive implants’’ (RECATABI) FP7 NMP3-SL-2009-229239 project. MMP acknowledges support of Instituto de Salud Carlos III with assistance from the European Regional Development Fund through CIBER-BBN initiative.Martínez Ramos, C.; Rodríguez Pérez, E.; Perez Garnes, M.; Chachques, JC.; Moratal Pérez, D.; Vallés Lluch, A.; Monleón Pradas, M. (2014). Design and Assembly Procedures for Large-Sized Biohybrid Scaffolds as Patches for Myocardial Infarct. Tissue Engineering Part C Methods. 20(10):817-827. https://doi.org/10.1089/ten.TEC.2013.0489S817827201

Endothelial progenitor cells and integrins: adhesive needs

In the last decade there have been multiple studies concerning the contribution of endothelial progenitor cells (EPCs) to new vessel formation in different physiological and pathological settings. The process by which EPCs contribute to new vessel formation in adults is termed postnatal vasculogenesis and occurs via four inter-related steps. They must respond to chemoattractant signals and mobilize from the bone marrow to the peripheral blood; home in on sites of new vessel formation; invade and migrate at the same sites; and differentiate into mature endothelial cells (ECs) and/or regulate pre-existing ECs via paracrine or juxtacrine signals. During these four steps, EPCs interact with different physiological compartments, namely bone marrow, peripheral blood, blood vessels and homing tissues. The success of each step depends on the ability of EPCs to interact, adapt and respond to multiple molecular cues. The present review summarizes the interactions between integrins expressed by EPCs and their ligands: extracellular matrix components and cell surface proteins present at sites of postnatal vasculogenesis. The data summarized here indicate that integrins represent a major molecular determinant of EPC function, with different integrin subunits regulating different steps of EPC biology. Specifically, integrin α4β1 is a key regulator of EPC retention and/or mobilization from the bone marrow, while integrins α5β1, α6β1, αvβ3 and αvβ5 are major determinants of EPC homing, invasion, differentiation and paracrine factor production. β2 integrins are the major regulators of EPC transendothelial migration. The relevance of integrins in EPC biology is also demonstrated by many studies that use extracellular matrix-based scaffolds as a clinical tool to improve the vasculogenic functions of EPCs. We propose that targeted and tissue-specific manipulation of EPC integrin-mediated interactions may be crucial to further improve the usage of this cell population as a relevant clinical agent

HCQ Practice Habits and Impact by COVID Pandemic

Supplental Tables 1 & 2

Parking Fees for Phototherapy Patients

Supplemental methods for assessment of parking fees experienced by phototherapy patients study.THIS DATASET IS ARCHIVED AT DANS/EASY, BUT NOT ACCESSIBLE HERE. TO VIEW A LIST OF FILES AND ACCESS THE FILES IN THIS DATASET CLICK ON THE DOI-LINK ABOV

A Necrotic Stimulus is Required to Maximize Matrix-Mediated Myogenesis.

Biomaterials that are similar to skeletal muscle extracellular matrix have been shown to augment regeneration in ischemic muscle. In this study, treatment with a collagen-based matrix stimulated molecular myogenesis in an mdx murine model of necrosis. Matrix-treated animals ran ≥40% further, demonstrating functional regeneration, and expressed increased levels of myogenic transcripts. In contrast, matrix treatment was unable to induce transcriptional or functional changes in an MLC/SOD1G93A atrophic mouse model. In vitro, satellite cells were cultured: 1) under standard conditions, 2) on matrix, 3) in the presence of myocyte debris (to simulate a necrotic-like environment), or 4) with both matrix and necrotic stimuli. Exposure to both stimuli induced the greatest increases in mef2c, myf5, myoD and myogenin transcripts. Furthermore, conditioned medium collected from satellite cells cultured with both stimuli contained elevated levels of factors that modulate satellite cell activation and proliferation, such as FGF-2, HGF and SDF-1. Application of the conditioned medium to C2C12 myoblasts accelerated maturation, demonstrated by increased mef2c, myf5 and myogenin transcripts and fusion indexes. In summary, the collagen matrix required a necrotic stimulus to enhance the maturation of satellite cell and their secretion of a myogenic cocktail. Considering that matrix treatment supports myogenesis only in in vivo models that exhibit necrosis, this study demonstrates that a necrotic environment is required to maximize matrix-mediated myogenesis

A Necrotic Stimulus is Required to Maximize Matrix-Mediated Myogenesis.

Biomaterials that are similar to skeletal muscle extracellular matrix have been shown to augment regeneration in ischemic muscle. In this study, treatment with a collagen-based matrix stimulated molecular myogenesis in an mdx murine model of necrosis. Matrix-treated animals ran ≥40% further, demonstrating functional regeneration, and expressed increased levels of myogenic transcripts. In contrast, matrix treatment was unable to induce transcriptional or functional changes in an MLC/SOD1G93A atrophic mouse model. In vitro, satellite cells were cultured: 1) under standard conditions, 2) on matrix, 3) in the presence of myocyte debris (to simulate a necrotic-like environment), or 4) with both matrix and necrotic stimuli. Exposure to both stimuli induced the greatest increases in mef2c, myf5, myoD and myogenin transcripts. Furthermore, conditioned medium collected from satellite cells cultured with both stimuli contained elevated levels of factors that modulate satellite cell activation and proliferation, such as FGF-2, HGF and SDF-1. Application of the conditioned medium to C2C12 myoblasts accelerated maturation, demonstrated by increased mef2c, myf5 and myogenin transcripts and fusion indexes. In summary, the collagen matrix required a necrotic stimulus to enhance the maturation of satellite cell and their secretion of a myogenic cocktail. Considering that matrix treatment supports myogenesis only in in vivo models that exhibit necrosis, this study demonstrates that a necrotic environment is required to maximize matrix-mediated myogenesis

A stromal cell-derived factor-1 releasing matrix enhances the progenitor cell response and blood vessel growth in ischaemic skeletal muscle

Although many regenerative cell therapies are being developed to replace or regenerate ischaemic muscle, the lack of vasculature and poor persistence of the therapeutic cells represent major limiting factors to successful tissue restoration. In response to ischaemia, stromal cell-derived factor-1 (SDF-1) is up-regulated by the affected tissue to stimulate stem cell-mediated regenerative responses. Therefore, we encapsulated SDF-1 into alginate microspheres and further incorporated these into an injectable collagen-based matrix in order to improve local delivery. Microsphere-matrix impregnation reduced the time for matrix thermogelation, and also increased the viscosity reached. This double-incorporation prolonged the release of SDF-1, which maintained adhesive and migratory bioactivity, attributed to chemotaxis in response to SDF-1. In vivo, treatment of ischaemic hindlimb muscle with microsphere-matrix led to increased mobilisation of bone marrow-derived progenitor cells, and also improved recruitment of angiogenic cells expressing the SDF-1 receptor (CXCR4) from bone marrow and local tissues. Both matrix and SDF-1-releasing matrix were successful at restoring perfusion, but SDF-1 treatment appeared to play an earlier role, as evidenced by arterioles that are phenotypically older and by increased angiogenic cytokine production, stimulating the generation of a qualitative microenvironment for a rapid and therefore more efficient regeneration. These results support the release of implanted SDF-1 as a promising method for enhancing progenitor cell responses and restoring perfusion to ischaemic tissues via neovascularisation

A stromal cell-derived factor-1 releasing matrix enhances the progenitor cell response and blood vessel growth in ischaemic skeletal muscle

Although many regenerative cell therapies are being developed to replace or regenerate ischaemic muscle, the lack of vasculature and poor persistence of the therapeutic cells represent major limiting factors to successful tissue restoration. In response to ischaemia, stromal cell-derived factor-1 (SDF-1) is up-regulated by the affected tissue to stimulate stem cell-mediated regenerative responses. Therefore, we encapsulated SDF-1 into alginate microspheres and further incorporated these into an injectable collagen-based matrix in order to improve local delivery. Microsphere-matrix impregnation reduced the time for matrix thermogelation, and also increased the viscosity reached. This double-incorporation prolonged the release of SDF-1, which maintained adhesive and migratory bioactivity, attributed to chemotaxis in response to SDF-1. In vivo, treatment of ischaemic hindlimb muscle with microsphere-matrix led to increased mobilisation of bone marrow-derived progenitor cells, and also improved recruitment of angiogenic cells expressing the SDF-1 receptor (CXCR4) from bone marrow and local tissues. Both matrix and SDF-1-releasing matrix were successful at restoring perfusion, but SDF-1 treatment appeared to play an earlier role, as evidenced by arterioles that are phenotypically older and by increased angiogenic cytokine production, stimulating the generation of a qualitative microenvironment for a rapid and therefore more efficient regeneration. These results support the release of implanted SDF-1 as a promising method for enhancing progenitor cell responses and restoring perfusion to ischaemic tissues via neovascularisation