Name: comparable decellularization of fetal and adult cardiac tissue explants as 3d-like platforms for in vitro studies
Uploaded: 2019-03-21T12:30:00
Description: Watch this Scientific Journal Video about Comparable Decellularization of Fetal and Adult Cardiac Tissue Explants as 3D-like Platforms for In Vitro Studies at JoVE.com

The authors are indebted to all members of Pinto-do-&#211; laboratory for relevant critical discussion. This work was supported by Programa MIT-Funda&#231;&#227;o para Ci&#234;ncia e Tecnologia (FCT) under the project &#34;CARDIOSTEM-Engineered cardiac tissues and stem cell-based therapies for cardiovascular applications&#34; (MITP-TB/ECE/0013/2013). A.C.S. is a recipient of a FCT fellowship [SFRH/BD/88780/2012] and M.J.O. is a FCT Fellow (FCT-Investigator 2012).

All the methodologies described were approved by the i3S Animal Ethics Committee and Dire&#231;&#227;o Geral de Veterin&#225;ria (DGAV) and are in accordance with the European Parliament Directive 2010/63/EU.
1. Preparation of the decellularization solutions
NOTE: All decellularization solutions should be filtered through a 0.22 &#956;m membrane filter and stored for a maximum of 3 months, except specified otherwise.
<ol>
	<li>For 1x PBS: Mix 8 g of NaCl, 1.01 g o...

<ol>
	<li>Frantz, C., Stewart, K. M., Weaver, V. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+extracellular+matrix+at+a+glance.">The extracellular matrix at a glance.</a> Journal of Cell Science. 123 (24), 4195-4200 (2010).</li><li>Rozario, T., DeSimone, D. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+extracellular+matrix+in+development+and+morphogenesis:+a+dynamic+view.">The extracellular matrix in development and morphogenesis: a dynamic view.</a> Dev Biol. 341 (1), 126-140 (2010).</li><li>Badylak, S. F., Taylor, D., Uygun, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Whole-Organ+Tissue+Engineering:+Decellularization+and+Recellularization+of+Three-Dimensional+Matrix+Scaffolds.">Whole-Organ Tissue Engineering: Decellularization and Recellularization of Three-Dimensional Matrix Scaffolds.</a> Annu Rev Biomed Eng. 13 (1), 27-53 (2011).</li><li>Ott, H. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Perfusion-decellularized+matrix:+using+nature's+platform+to+engineer+a+bioartificial+heart.">Perfusion-decellularized matrix: using nature's platform to engineer a bioartificial heart.</a> Nat Med. 14 (2), 213-221 (2008).</li><li>Silva, A. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Three-dimensional+scaffolds+of+fetal+decellularized+hearts+exhibit+enhanced+potential+to+support+cardiac+cells+in+comparison+to+the+adult.">Three-dimensional scaffolds of fetal decellularized hearts exhibit enhanced potential to support cardiac cells in comparison to the adult.</a> Biomaterials. 104, 52-64 (2016).</li><li>Nakayama, K. H., Batchelder, C. A., Lee, C. I., Tarantal, A. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decellularized+rhesus+monkey+kidney+as+a+three-dimensional+scaffold+for+renal+tissue+engineering.">Decellularized rhesus monkey kidney as a three-dimensional scaffold for renal tissue engineering.</a> Tissue Eng Part A. 16 (7), 2207-2216 (2010).</li><li>Williams, C., Quinn, K. P., Georgakoudi, I., Black, L. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Young+developmental+age+cardiac+extracellular+matrix+promotes+the+expansion+of+neonatal+cardiomyocytes+in+vitro.">Young developmental age cardiac extracellular matrix promotes the expansion of neonatal cardiomyocytes in vitro.</a> Acta Biomater. 10 (1), 194-204 (2014).</li><li>Fong, A. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Three-Dimensional+Adult+Cardiac+Extracellular+Matrix+Promotes+Maturation+of+Human+Induced+Pluripotent+Stem+Cell-Derived+Cardiomyocytes.">Three-Dimensional Adult Cardiac Extracellular Matrix Promotes Maturation of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes.</a> Tissue Eng Part A. 22 (15-16), 1016-1025 (2016).</li><li>Tapias, L. F., Ott, H. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decellularized+scaffolds+as+a+platform+for+bioengineered+organs.">Decellularized scaffolds as a platform for bioengineered organs.</a> Curr Opin Organ Transplant. 19 (2), 145-152 (2014).</li><li>Crapo, P. M., Gilbert, T. W., Badylak, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+overview+of+tissue+and+whole+organ+decellularization+processes.">An overview of tissue and whole organ decellularization processes.</a> Biomaterials. 32 (12), 3233-3243 (2011).</li><li>Gilbert, T. W., Sellaro, T. L., Badylak, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decellularization+of+tissues+and+organs.">Decellularization of tissues and organs.</a> Biomaterials. 27 (19), 3675-3683 (2006).</li><li>Pinto, M. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decellularized+human+colorectal+cancer+matrices+polarize+macrophages+towards+an+anti-inflammatory+phenotype+promoting+cancer+cell+invasion+via+CCL18.">Decellularized human colorectal cancer matrices polarize macrophages towards an anti-inflammatory phenotype promoting cancer cell invasion via CCL18.</a> Biomaterials. 124, 211-224 (2017).</li><li>Garlikova, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+a+close-to-native+in+vitro+system+to+study+lung+cells-ECM+crosstalk.">Generation of a close-to-native in vitro system to study lung cells-ECM crosstalk.</a> Tissue Eng Part C: Methods. , (2017).</li><li>Wong, M. L., Griffiths, L. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immunogenicity+in+xenogeneic+scaffold+generation:+Antigen+removal+vs.+decellularization.">Immunogenicity in xenogeneic scaffold generation: Antigen removal vs. decellularization.</a> Acta Biomaterialia. 10 (5), 1806-1816 (2014).</li><li>Motsavage, V. A., Kostenbauder, H. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+influence+of+the+state+of+aggregation+on+the+specific+acid-catalyzed+hydrolysis+of+sodium+dodecyl+sulfate.">The influence of the state of aggregation on the specific acid-catalyzed hydrolysis of sodium dodecyl sulfate.</a> J Colloid Sci. 18 (7), 603-615 (1963).</li><li>Rahman, A., Brown, C. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+pH+on+the+critical+micelle+concentration+of+sodium+dodecyl+sulphate.">Effect of pH on the critical micelle concentration of sodium dodecyl sulphate.</a> J Appl Polymer Sci. 28 (4), 1331-1334 (1983).</li><li>Brown, B. N., Badylak, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+matrix+as+an+inductive+scaffold+for+functional+tissue+reconstruction.">Extracellular matrix as an inductive scaffold for functional tissue reconstruction.</a> Transl Res. 163 (4), 268-285 (2014).</li><li>Ieda, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiac+fibroblasts+regulate+myocardial+proliferation+through+beta1+integrin+signaling.">Cardiac fibroblasts regulate myocardial proliferation through beta1 integrin signaling.</a> Dev Cell. 16 (2), 233-244 (2009).</li><li>Whitby, D. J., Ferguson, M. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+extracellular+matrix+of+lip+wounds+in+fetal,+neonatal+and+adult+mice.">The extracellular matrix of lip wounds in fetal, neonatal and adult mice.</a> Development. 112 (2), 651-668 (1991).</li><li>Brennan, E. P., Tang, X. H., Stewart-Akers, A. M., Gudas, L. J., Badylak, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemoattractant+activity+of+degradation+products+of+fetal+and+adult+skin+extracellular+matrix+for+keratinocyte+progenitor+cells.">Chemoattractant activity of degradation products of fetal and adult skin extracellular matrix for keratinocyte progenitor cells.</a> J Tissue Eng Regen Med. 2 (8), 491-498 (2008).</li><li>Coolen, N. A., Schouten, K. C., Middelkoop, E., Ulrich, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+between+human+fetal+and+adult+skin.">Comparison between human fetal and adult skin.</a> Arch Dermatol Res. 302 (1), 47-55 (2010).</li></ol>

The extracellular matrix (ECM) is a highly dynamic and complex meshwork of fibrous and adhesive glycoproteins, consisting of a reservoir of numerous bioactive peptides and entrapped growth factors. As the major modulator of cell adhesion, cytoskeleton dynamics, motility/migration, proliferation, differentiation and apoptosis, ECM actively regulates cellular function and behavior. Knowing that cellular behavior differs in 2D and 3D cultures, there have been efforts to develop novel organotypic models that can accurately r...

The extracellular matrix (ECM) is a dynamic network of molecules that regulate important cellular processes, namely fate-decision, proliferation and differentiation1,2. The investigation of cell-ECM interactions has been performed mainly in two-dimensional (2D) in vitro cultures coated with ECM components, which constitute a simplification of native ECM properties found in vivo. Decellularization generates acellular 3D-like ECM bioscaffolds that largely preserve the extracellular architecture and composition of native tissues and organs3,4. In addition to serving as bioactive scaffolds for tissue engineering, decellularized 3D ECM biomaterials are emerging as novel platforms to assess cell-ECM biology that parallel the in vivo environment.
Assessment of the differential role of the ECM components of distinct tissues, organs and age will benefit with the use of similar protocols of generating native bioscaffolds. In the heart, we have developed a versatile protocol for decellularization of fetal and adult-derived samples, as an alternative approach to perform comparative studies of the organ microenvironment. Using this methodology, we captured the native cardiac microenvironment and showed that fetal ECM promotes higher repopulation yields of cardiac cells5. Decellularization further provided identification of resident structural differences between fetal and adult ECM at the level of basement lamina and pericellular matrix mesh arrangement and fiber composition5. Prior to this work, head-to-head comparison of tissues at different ontogenic stages using the same decellularization approach has only been reported for rhesus monkey kidneys and rodent hearts. In addition, a limited number of studies report fetal tissue/organ decellularization per se5,6,7. This has been achieved using SDS as a unique decellularization agent; however, distinct SDS concentrations were used for the decellularization of fetal and adult cardiac tissue7,8. SDS is one of the most effective ionic detergents for clearance of cytoplasmic and nuclear material, and widely used in the decellularization of different tissues and specimens9,10. Solutions containing high SDS concentrations and extended periods of exposure have been correlated with protein denaturation, glycosaminoglycan (GAGs) loss and disruption of collagen fibrils10,11, and therefore a balance between ECM preservation and cell removal is necessary. To apply the same procedure to fetal and adult heart tissue, the protocol described herein is divided in three sequential steps: cell lysis by osmotic shock (hypotonic buffer); solubilization of lipid-protein, DNA-protein and protein-protein interactions (0.2% SDS); and nuclear material removal (DNase treatment).
Our protocol shows several advantages: i) the possibility of equivalent decellularization of age-specific cardiac tissues by the application of the same decellularization strategy; ii) no requirements for specialized methods or equipment; iii) ready adaptation to other tissues and species as it has been successfully applied with minor alterations in human intestine biopsies12 and mouse lung13; and, importantly, iv) can address ECM biomechanical properties while enabling the assembly of 3D-like organotypic cultures that more closely mimic the molecular features of the native tissue microenvironment.

<table>
	<tbody>
		<tr>
			<td colspan="4">Equipment</td>
		</tr>
		<tr>
			<td>Incubated Benchtop Shaker</td>
			<td>Orbital Shakers</td>
			<td>IKA:3510001</td>
			<td>Recommended</td>
		</tr>
		<tr>
			<td>Fluorimeter</td>
			<td>-</td>
			<td>-</td>
			<td>Equipment available</td>
		</tr>
		<tr>
			<td>Digital weight scale</td>
			<td>-</td>
			<td>-</td>
			<td>Equipment available</td>
		</tr>
		<tr>
			<td>Inverted Microscope</td>
			<td>-</td>
			<td>-</td>
			<td>Equipment available</td>
		</tr>
		<tr>
			<td>Cell culture incubator</td>
			<td>-</td>
			<td>-</td>
			<td>Equipment available</td>
		</tr>
		<tr>
			<td>Fridge (4&#186;C)</td>
			<td>-</td>
			<td>-</td>
			<td>Equipment available</td>
		</tr>
		<tr>
			<td>Deep freezer (-80&#186;C)</td>
			<td>-</td>
			<td>-</td>
			<td>Equipment available</td>
		</tr>
		<tr>
			<td>Microtome</td>
			<td>-</td>
			<td>-</td>
			<td>Equipment available</td>
		</tr>
		<tr>
			<td colspan="4">Cirurgical Instruments</td>
		</tr>
		<tr>
			<td>Vannas Spring Scissors - 2.5mm Cutting Edge</td>
			<td>Fine Science Tools</td>
			<td>5000-08</td>
			<td>Recommended</td>
		</tr>
		<tr>
			<td>Dumont 5 Fine Forceps - Biologie/Inox</td>
			<td>Fine Science Tools</td>
			<td>11254-20</td>
			<td>Recommended</td>
		</tr>
		<tr>
			<td>Dumont 7 forceps</td>
			<td>Fine Science Tools</td>
			<td>11272-30</td>
			<td>Recommended</td>
		</tr>
		<tr>
			<td>Dissecting Scissors, straight</td>
			<td>-</td>
			<td>-</td>
			<td>Tool available</td>
		</tr>
		<tr>
			<td>Forceps, serrated, curved</td>
			<td>-</td>
			<td>-</td>
			<td>Tool available</td>
		</tr>
		<tr>
			<td colspan="4">Materials</td>
		</tr>
		<tr>
			<td>24 well plates, individually wrapped</td>
			<td>VWR</td>
			<td>29442-044</td>
			<td>-</td>
		</tr>
		<tr>
			<td>96 well plates, individually wrapped</td>
			<td>VWR</td>
			<td>71000-078</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Steriflip-GV, 0.22&#181;m, PVDF, Radio-Sterilized</td>
			<td>Millipore</td>
			<td>SE1M179M6</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Eppendorff</td>
			<td>-</td>
			<td>-</td>
			<td>Material available</td>
		</tr>
		<tr>
			<td>15 mL Falcon tubes</td>
			<td>Fisher Scientific</td>
			<td>430791</td>
			<td>-</td>
		</tr>
		<tr>
			<td>50 mL Falcon tubes</td>
			<td>Fisher Scientific</td>
			<td>430829</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Four-Compartment Biopsy Processing/Embedding Cassettes with Lid</td>
			<td>Electron Microscopy Science</td>
			<td>70075-B</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Fisherbrand Superfrost Plus Microscope Slides</td>
			<td>Thermo Fisher Scientific</td>
			<td>22-037-246</td>
			<td>-</td>
		</tr>
		<tr>
			<td colspan="4">Tissue cryopreservation</td>
		</tr>
		<tr>
			<td>Shandon Cryomatrix embedding resin</td>
			<td>Thermo Scientific</td>
			<td>6769006</td>
			<td>-</td>
		</tr>
		<tr>
			<td>2-METHYLBUTANE ANHYDROUS 99+% (isopentane)</td>
			<td>Sigma-Aldrich</td>
			<td>277258-1L</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Dry ice</td>
			<td>-</td>
			<td>-</td>
			<td>-</td>
		</tr>
		<tr>
			<td colspan="4">Decellularization</td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>BDH Prolabo</td>
			<td>27810.364</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Na2HPO4</td>
			<td>Sigma-Aldrich</td>
			<td>S-31264</td>
			<td>-</td>
		</tr>
		<tr>
			<td>KH2PO4</td>
			<td>Sigma-Aldrich</td>
			<td>P5379-100g</td>
			<td>-</td>
		</tr>
		<tr>
			<td>KCl</td>
			<td>Sigma-Aldrich</td>
			<td>P8041-1KG</td>
			<td>-</td>
		</tr>
		<tr>
			<td>TrisBASE</td>
			<td>Sigma-Aldrich</td>
			<td>T6066-500G</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Sodium dodecyl sulfate</td>
			<td>Sigma-Aldrich</td>
			<td>L-4390</td>
			<td>-</td>
		</tr>
		<tr>
			<td>MgCl2</td>
			<td>MERCK</td>
			<td>1.05833.1000</td>
			<td>-</td>
		</tr>
		<tr>
			<td>DNAse I</td>
			<td>AplliChem</td>
			<td>A3778,0050</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Gentamicin</td>
			<td>Gibco</td>
			<td>15710-049</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Fungizone</td>
			<td>Gibco BRL</td>
			<td>15290-026</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Deionized water (DI water)</td>
			<td>-</td>
			<td>-</td>
			<td>-</td>
		</tr>
		<tr>
			<td colspan="4">Histology</td>
		</tr>
		<tr>
			<td>10 % formalin neutral buffer</td>
			<td>Prolabo</td>
			<td>361387P</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Eosin Y AQUEOUS</td>
			<td>Surgipath</td>
			<td>01592E</td>
			<td>Can be replaced by alcoholic eosin</td>
		</tr>
		<tr>
			<td>Richard-Allan Scientific HistoGel Specimen Processing Gel</td>
			<td>Thermo Fisher Scientific</td>
			<td>HG-4000-012</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Ethanol ethilic alcohol 99,5% anydrous</td>
			<td>Aga</td>
			<td>4,006,02,02,00</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Deionized water (DI water)</td>
			<td>-</td>
			<td>-</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Clear Rite 3</td>
			<td>Richard-Allan Scientific</td>
			<td>6915</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Shandon Histoplast</td>
			<td>Thermo Fisher Scientific</td>
			<td>RAS.6774006</td>
			<td>-</td>
		</tr>
		<tr>
			<td colspan="4">Kits</td>
		</tr>
		<tr>
			<td>PureLink Genomic DNA Mini Kit</td>
			<td>Thermo Fisher Scientific</td>
			<td>K182001</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Quant-iT PicoGreen&#160;dsDNA kit</td>
			<td>Invitrogen</td>
			<td>P11496</td>
			<td>-</td>
		</tr>
		<tr>
			<td colspan="4">Cell culture</td>
		</tr>
		<tr>
			<td>DPBS</td>
			<td>VWR</td>
			<td>45000-434</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Penicillin-Streptomycin Solution 100X</td>
			<td>Labclinics</td>
			<td>L0022-100</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Fungizone</td>
			<td>Gibco BRL</td>
			<td>15290-026</td>
			<td>-</td>
		</tr>
		<tr>
			<td>Cell culture media of the cell of interest</td>
			<td>-</td>
			<td>-</td>
			<td>-</td>
		</tr>
	</tbody>
</table>

comparable decellularization of fetal and adult cardiac tissue explants as 3d-like platforms for in vitro studies

Current knowledge of extracellular matrix (ECM)-cell communication translates to large two-dimensional (2D) in vitro culture studies where ECM components are presented as a surface coating. These culture systems constitute a simplification of the complex nature of the tissue ECM that encompasses biochemical composition, structure, and mechanical properties. To better emulate the ECM-cell communication shaping the cardiac microenvironment, we developed a protocol that allows for the decellularization of the whole fetal heart and adult left ventricle tissue explants simultaneously for comparative studies. The protocol combines the use of a hypotonic buffer, a detergent of anionic surfactant properties, and DNase treatment without any requirement for specialized skills or equipment. The application of the same decellularization strategy across tissue samples from subjects of various age is an alternative approach to perform comparative studies. The present protocol allows the identification of unique structural differences across fetal and adult cardiac ECM mesh and biological cellular responses. Furthermore, the herein methodology demonstrates a broader application being successfully applied in other tissues and species with minor adjustments, such as in human intestine biopsies and mouse lung.

The decellularization efficiency should be assessed through three main techniques: macroscopic observation, histology and DNA quantification. The macroscopic appearance of samples post-SDS treatment indirectly affects the efficacy of cell removal. After SDS incubation, samples should appear as translucent to whitish (Figure 1C). Fetal (E18) decellularized tissues are characterized by a highly translucent structure while adult explants have a translucent to wh...

Watch this Scientific Journal Video about Comparable Decellularization of Fetal and Adult Cardiac Tissue Explants as 3D-like Platforms for In Vitro Studies at JoVE.com

Comparable Decellularization of Fetal and Adult Cardiac Tissue Explants as 3D-like Platforms for In Vitro Studies

The cardiac extracellular matrix (ECM) is a complex network of molecules that orchestrate key processes in tissues and organs while enduring physiological remodeling throughout life. Standardized decellularization of fetal and adult hearts permits comparative experimental studies of both tissues in a 3D context by capturing native architecture and biomechanical properties.

Current knowledge of extracellular matrix (ECM)-cell communication translates to large two-dimensional (2D) in vitro culture studies where ECM components ...

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Culturing cells in a three dimensional hydrogel environment is an important technique for developing constructs for tissue engineering as well as studying ...

Capillary Force Lithography for Cardiac Tissue Engineering

Cardiovascular disease remains the leading cause of death worldwide1. Cardiac tissue engineering holds much promise to deliver groundbreaking medical ...

Tissue Engineering of a Human 3D in vitro Tumor Test System

Tissue Engineering of a Human 3D in vitro Tumor Test System

Cancer is one of the leading causes of death worldwide.  Current therapeutic strategies are predominantly developed in 2D culture  systems, which ...

Eye Irritation Test (EIT) for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial (RhCE) Tissue Model

Eye Irritation Test EIT for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial RhCE Tissue Model

To comply with the Seventh Amendment to the EU Cosmetics Directive and EU REACH legislation, validated non-animal alternative methods for reliable and ...

Neonatal Cardiac Scaffolds: Novel Matrices for Regenerative Studies

The only definitive therapy for end stage heart failure is orthotopic heart transplantation.  Each year, it is estimated that more than 100,000 donor ...

Fabrication of Extracellular Matrix-derived Foams and Microcarriers as Tissue-specific Cell Culture and Delivery Platforms

Cell function is mediated by interactions with the extracellular matrix (ECM), which has complex tissue-specific composition and architecture. The focus ...

Protocol for MicroRNA Transfer into Adult Bone Marrow-derived Hematopoietic Stem Cells to Enable Cell Engineering Combined with Magnetic Targeting

While CD133+ hematopoietic stem cells (SCs) have been proven to provide high potential in the field of regenerative medicine, their low retention rates ...

Two Methods for Decellularization of Plant Tissues for Tissue Engineering Applications

The autologous, synthetic, and animal-derived grafts currently used as scaffolds for tissue replacement have limitations due to low availability, poor ...

Advanced 3D Liver Models for In vitro Genotoxicity Testing Following Long-Term Nanomaterial Exposure

Due to the rapid development and implementation of a diverse array of engineered nanomaterials (ENM), exposure to ENM is inevitable and the development of ...