Tissue-engineered human embryonic stem cell-containing cardiac patches: evaluating recellularization of decellularized matrix

Abstract

Decellularized cardiac extracellular matrix scaffolds with preserved composition and architecture can be used in tissue engineering to reproduce the complex cardiac extracellular matrix. However, evaluating the extent of cardiomyocyte repopulation of decellularized cardiac extracellular matrix scaffolds after recellularization attempts is challenging. Here, we describe a unique combination of biochemical, biomechanical, histological, and physiological parameters for quantifying recellularization efficiency of tissue-engineered cardiac patches compared with native cardiac tissue. Human embryonic stem cell-derived cardiomyocytes were seeded into rat heart atrial and ventricular decellularized cardiac extracellular matrix patches. Confocal and atomic force microscopy showed cell integration within the extracellular matrix basement membrane that was accompanied by restoration of native cardiac tissue passive mechanical properties. Multi-electrode array and immunostaining (connexin 43) were used to determine synchronous field potentials with electrical coupling. Myoglobin content (~60%) and sarcomere length measurement (>45% vs 2D culture) were used to evaluate cardiomyocyte maturation of integrated cells. The combination of these techniques allowed us to demonstrate that as cellularization efficiency improves, cardiomyocytes mature and synchronize electrical activity, and tissue mechanical/biochemical properties improve toward those of native tissue.

Layer-By-Layer Fabrication of Large and Thick Human Cardiac Muscle Patch Constructs With Superior Electrophysiological Properties

“…These costameres and their subsequent integrins contribute to signal transduction, transmitting force signals between the contractile apparatus and ECM via interactions with primarily titin and other Z-line associated structures (Peter et al, 2011). This process not only supports the mechanical integrity of the sarcolemma, contributes to mechanical signaling but also provides spatial prompts for muscle fiber organization (Hochman-Mendez et al, 2020b). The upregulation in the levels of laminin observed in the tri-lineage tissues could, thus, likely be associated with the structural and functional maturation of the iCMs, as well as the enhanced tissue compaction and alignment noted in Figures 2C,D.…”

Cues from human atrial extracellular matrix enrich the atrial differentiation of human induced pluripotent stem cell-derived cardiomyocytes

“…Proteomics were performed as previously described. [26][27][28] Protein samples from five different dECM-hA and dECM-hV powders were macerated in the presence of liquid nitrogen, precipitated with cold 10% (1 : 4 v/v) trichloroacetic acid (Sigma-Aldrich) in acetone (Sigma-Aldrich), and centrifuged for 15 minutes at 4°C and 15 000 rpm. The samples were washed three times with cold acetone and air-dried.…”

Characterization of perfusion decellularized whole animal body, isolated organs, and multi‐organ systems for tissue engineering applications

“…To decellularize different types of organs used in this study we adapted our previously reported perfusion decellularization protocols (Hochman-Mendez et al, 2020;Ott et al, 2008). The renal artery of kidney, the portal vein of liver, the descending aorta of heart and lung, and the iliac artery of the thigh (gracilis cranialis and the gluteus superficialis) were cannulated and perfused with 1% SDS.…”