Protein extraction and 2‐DE of water‐ and lipid‐soluble proteins from bovine pericardium, a low‐cellularity tissue

Griffiths, Leigh G.; Choe, Leila H.; Lee, Kelvin H.; Reardon, Kenneth F.; Orton, E. Christopher

doi:10.1002/elps.200800108

Cited by 11 publications

(5 citation statements)

References 72 publications

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“… 16 ) and ( B ) in the native pericardium (collected by Griffiths et al . 17 ). Total protein content, classified as matrisome and other proteins, for ( C ) the decellularized myocardial and ( D ) decellularized pericardial scaffolds.…”

Section: Resultsmentioning

confidence: 99%

Head-to-head comparison of two engineered cardiac grafts for myocardial repair: From scaffold characterization to pre-clinical testing

Perea‐Gil

Gálvez‐Montón

Jorba

et al. 2018

Sci Rep

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Cardiac tissue engineering, which combines cells and supportive scaffolds, is an emerging treatment for restoring cardiac function after myocardial infarction (MI), although, the optimal construct remains a challenge. We developed two engineered cardiac grafts, based on decellularized scaffolds from myocardial and pericardial tissues and repopulated them with adipose tissue mesenchymal stem cells (ATMSCs). The structure, macromechanical and micromechanical scaffold properties were preserved upon the decellularization and recellularization processes, except for recellularized myocardium micromechanics that was ∼2-fold stiffer than native tissue and decellularized scaffolds. Proteome characterization of the two acellular matrices showed enrichment of matrisome proteins and major cardiac extracellular matrix components, considerably higher for the recellularized pericardium. Moreover, the pericardial scaffold demonstrated better cell penetrance and retention, as well as a bigger pore size. Both engineered cardiac grafts were further evaluated in pre-clinical MI swine models. Forty days after graft implantation, swine treated with the engineered cardiac grafts showed significant ventricular function recovery. Irrespective of the scaffold origin or cell recolonization, all scaffolds integrated with the underlying myocardium and showed signs of neovascularization and nerve sprouting. Collectively, engineered cardiac grafts -with pericardial or myocardial scaffolds- were effective in restoring cardiac function post-MI, and pericardial scaffolds showed better structural integrity and recolonization capability.

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Section: Resultsmentioning

confidence: 99%

Head-to-head comparison of two engineered cardiac grafts for myocardial repair: From scaffold characterization to pre-clinical testing

Perea‐Gil

Gálvez‐Montón

Jorba

et al. 2018

Sci Rep

View full text Add to dashboard Cite

show abstract

“…25 Protein extraction protocols, generally utilize protein chemistry-based sequential extraction techniques to solubilize and remove proteins in a stepwise manner, by serial application of solutions with characteristics designed to favor solubility of a particular protein type. 11,25 Failure to solubilize a particular undesirable cellular or antigenic component at any given step of the procedure may result in its precipitation and/or aggregation with other molecules, making it difficult if not impossible to solubilize and remove at later steps. In applying these principles to our AR process, we reasoned that since the AR buffer at each step was designed to favor solubility of a given protein class (hydrophilic or lipophilic), antigenic components of the tissue which are soluble in the AR solution will diffuse into the AR buffer, while insoluble or macromolecular tissue components, such as the FIGURE 8.…”

Section: Discussionmentioning

confidence: 99%

Cardiac Extracellular Matrix Scaffold Generated Using Sarcomeric Disassembly and Antigen Removal

Papalamprou

Griffiths

2015

Ann Biomed Eng

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Xenogeneic cardiac extracellular matrix (cECM) scaffolds for reconstructive cardiac surgery applications have potential to overcome the limitations of current clinically utilized patch materials. A potentially ideal cECM scaffold would be immunologically acceptable while preserving the native cECM niche. Production of such a scaffold necessitates removal of cellular and antigenic components from cardiac tissue while preserving cECM structure/function properties. Existing decellularization methodologies predominantly utilize denaturing detergents which might irreversibly alter cECM material properties. To overcome potential deficiencies of current approaches, the effect of sarcomere relaxation and disassembly on resultant cECM scaffold cellularity was investigated. Additionally, the ability of sequential differential protein solubilization (antigen removal-AR) to reduce cECM scaffold antigenicity was examined. Sarcomeric relaxation and disassembly were necessary to achieve scaffold acellularity. All groups in which AR was employed displayed statistically significant decreases in residual antigenicity regardless of their degree of acellularity. AR combined with sarcomeric disassembly preserved structural, biochemical, mechanical and recellularization properties of the cECM scaffold. However, sodium dodecyl sulfate significantly altered cECM properties. This study demonstrates the importance of solubilizing cellular elements and antigenic components in a stepwise manner for production of a potentially ideal cECM scaffold and may have implications for future tissue engineering and regenerative medicine applications.

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“…The extraction of proteins from a tissue into an aqueous solution during antigen removal is dependent upon its solubility in the antigen removal buffer as a protein can only be extracted into a solution in which it is soluble [98, 99]. We tested the ability of reducing agent and salt, used to promote hydrophilic protein solubility for extraction of proteins from homogenized bovine pericardium [45, 100], to enhance the removal of antigens. Intracellular thioredoxin and glutathione systems maintain a reducing environment within the cell, facilitating solubilization of hydrophilic, cytoplasmic proteins [101].…”

Section: Antigen Removalmentioning

confidence: 99%

Immunogenicity in xenogeneic scaffold generation: Antigen removal vs. decellularization

2014

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Decades of research have been undertaken towards the goal of tissue engineering using xenogeneic scaffolds. The primary advantages associated with use of xenogeneic tissue-derived scaffolds for in vitro development of replacement tissues and organs stem from the inherent extracellular matrix (ECM) composition and architecture. Native ECM possesses appropriate mechanical properties for physiological function of the biomaterial and signals for cell binding, growth, and differentiation. Additionally, xenogeneic tissue is readily available. However, translation of xenogeneic scaffold-derived engineered tissues or organs into clinical therapies requires xenoantigenicity of the material to be adequately addressed prior to implantation. Failure to achieve this goal will result in a graft-specific, host immune rejection response, jeopardizing in vivo survival of the resultant scaffold, tissue, or organ. This review explores (1) the appropriateness of scaffold acellularity as an outcome measure for assessing reduction of the immunological barriers to the use of xenogeneic scaffolds for tissue engineering applications and (2) the need for tissue engineers to strive for antigen removal during xenogeneic scaffold generation.

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Protein extraction and 2‐DE of water‐ and lipid‐soluble proteins from bovine pericardium, a low‐cellularity tissue

Cited by 11 publications

References 72 publications

Head-to-head comparison of two engineered cardiac grafts for myocardial repair: From scaffold characterization to pre-clinical testing

Head-to-head comparison of two engineered cardiac grafts for myocardial repair: From scaffold characterization to pre-clinical testing

Cardiac Extracellular Matrix Scaffold Generated Using Sarcomeric Disassembly and Antigen Removal

Immunogenicity in xenogeneic scaffold generation: Antigen removal vs. decellularization

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