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      Towards a Novel Patch Material for Cardiac Applications: Tissue-Specific Extracellular Matrix Introduces Essential Key Features to Decellularized Amniotic Membrane

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          Abstract

          There is a growing need for scaffold material with tissue-specific bioactivity for use in regenerative medicine, tissue engineering, and for surgical repair of structural defects. We developed a novel composite biomaterial by processing human cardiac extracellular matrix (ECM) into a hydrogel and combining it with cell-free amniotic membrane via a dry-coating procedure. Cardiac biocompatibility and immunogenicity were tested in vitro using human cardiac fibroblasts, epicardial progenitor cells, murine HL-1 cells, and human immune cells derived from buffy coat. Processing of the ECM preserved important matrix proteins as demonstrated by mass spectrometry. ECM coating did not alter the mechanical characteristics of decellularized amniotic membrane but did cause a clear increase in adhesion capacity, cell proliferation and viability. Activated monocytes secreted less pro-inflammatory cytokines, and both macrophage polarization towards the pro-inflammatory M1 type and T cell proliferation were prevented. We conclude that the incorporation of human cardiac ECM hydrogel shifts and enhances the bioactivity of decellularized amniotic membrane, facilitating its use in future cardiac applications.

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          Most cited references38

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          HL-1 cells: a cardiac muscle cell line that contracts and retains phenotypic characteristics of the adult cardiomyocyte.

          We have derived a cardiac muscle cell line, designated HL-1, from the AT-1 mouse atrial cardiomyocyte tumor lineage. HL-1 cells can be serially passaged, yet they maintain the ability to contract and retain differentiated cardiac morphological, biochemical, and electrophysiological properties. Ultrastructural characteristics typical of embryonic atrial cardiac muscle cells were found consistently in the cultured HL-1 cells. Reverse transcriptase-PCR-based analyses confirmed a pattern of gene expression similar to that of adult atrial myocytes, including expression of alpha-cardiac myosin heavy chain, alpha-cardiac actin, and connexin43. They also express the gene for atrial natriuretic factor. Immunohistochemical staining of the HL-1 cells indicated that the distribution of the cardiac-specific markers desmin, sarcomeric myosin, and atrial natriuretic factor was similar to that of cultured atrial cardiomyocytes. A delayed rectifier potassium current (IKr) was the most prominent outward current in HL-1 cells. The activating currents displayed inward rectification and deactivating current tails were voltage-dependent, saturated at >+20 mV, and were highly sensitive to dofetilide (IC50 of 46.9 nM). Specific binding of [3H]dofetilide was saturable and fit a one-site binding isotherm with a Kd of 140 +/- 60 nM and a Bmax of 118 fmol per 10(5) cells. HL-1 cells represent a cardiac myocyte cell line that can be repeatedly passaged and yet maintain a cardiac-specific phenotype.
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            Preparation and rheological characterization of a gel form of the porcine urinary bladder matrix.

            Biologic scaffolds composed of extracellular matrix (ECM) have been used to facilitate the repair and reconstruction of a variety of tissues in clinical and pre-clinical studies. The clinical utility of such scaffolds can be limited by the geometric and mechanical properties of the tissue or organ from which the ECM is harvested. An injectable gel form of ECM could potentially conform to any three-dimensional shape and could be delivered to sites of interest by minimally invasive techniques. The objectives of the present study were to prepare a gel form of ECM harvested from the urinary bladder (urinary bladder matrix or UBM), to characterize the rheological properties of the gel, and finally to evaluate the ability of the gel to support in vitro growth of smooth muscle cells. Following enzymatic solubilization with pepsin, UBM was induced to self-assemble into a gel when brought to physiological conditions. The UBM gel supported the adhesion and growth of rat aortic smooth muscle cells when cultured under static in vitro conditions. The present study showed that an intact form of UBM can be successfully solubilized without purification steps and induced to repolymerize into a gel form of the UBM biologic scaffold material.
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              Characterisation of a soft elastomer poly(glycerol sebacate) designed to match the mechanical properties of myocardial tissue.

              The myocardial tissue lacks significant intrinsic regenerative capability to replace the lost cells. Therefore, the heart is a major target of research within the field of tissue engineering, which aims to replace infarcted myocardium and enhance cardiac function. The primary objective of this work was to develop a biocompatible, degradable and superelastic heart patch from poly(glycerol sebacate) (PGS). PGS was synthesised at 110, 120 and 130 degrees C by polycondensation of glycerol and sebacic acid with a mole ratio of 1:1. The investigation was focused on the mechanical and biodegrading behaviours of the developed PGS. PGS materials synthesised at 110, 120 and 130 degrees C have Young's moduli of 0.056, 0.22 and 1.2 MPa, respectively, which satisfy the mechanical requirements on the materials applied for the heart patch and 3D myocardial tissue engineering construction. Degradation assessment in phosphate buffered saline and Knockout DMEM culture medium has demonstrated that the PGS has a wide range of degradability, from being degradable in a couple of weeks to being nearly inert. The matching of physical characteristics to those of the heart, the ability to fine tune degradation rates in biologically relevant media and initial data showing biocompatibility indicate that this material has promise for cardiac tissue engineering applications.
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                Author and article information

                Journal
                Int J Mol Sci
                Int J Mol Sci
                ijms
                International Journal of Molecular Sciences
                MDPI
                1422-0067
                29 March 2018
                April 2018
                : 19
                : 4
                : 1032
                Affiliations
                [1 ]Charité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, 13353 Berlin, Germany; matthias.becker@ 123456charite.de (M.B.); josemaring@ 123456hotmail.com (J.A.M.); maria.schneider@ 123456charite.de (M.Sc.); aaron.herrera@ 123456charite.de (A.X.H.M.) martina.seifert@ 123456charite.de (M.Se.); oliver.klein@ 123456charite.de (O.K.)
                [2 ]Berlin-Brandenburg Center for Regenerative Therapies (BCRT), 13353 Berlin, Germany; falk@ 123456dhzb.de
                [3 ]Julius Wolff Institute for Biomechanics and Musculoskeletal Regeneration, 13353 Berlin, Germany
                [4 ]Department of Obstetrics and Gynecology, Charite Medical University, 13353 Berlin, Germany; thorsten.braun@ 123456charite.de
                [5 ]German Centre for Cardiovascular Research (DZHK), Partner Site Berlin, 13316 Berlin, Germany
                [6 ]Deutsches Herzzentrum Berlin (DHZB), Augustenburger Platz 1, 13353 Berlin, Germany
                Author notes
                [* ]Correspondence: stamm@ 123456dhzb.de ; Tel.: +49-30-453-321-09
                Article
                ijms-19-01032
                10.3390/ijms19041032
                5979550
                29596384
                5df96e4b-7ad3-4f8d-aa94-a8c8748b677c
                © 2018 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 09 March 2018
                : 27 March 2018
                Categories
                Article

                Molecular biology
                extracellular matrix,hydrogel,cardioprotection,patch,epicardium,amnion,immunocompatibility

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