Freeze-Dried Heart Valve Scaffolds

Abstract

In this study, structure and biomechanical properties of freeze-dried decellularized porcine pulmonary heart valves were investigated. Heart valves were dissected from porcine hearts. The tissues were decellularized and separated in three groups: (1) without lyoprotectant, (2) with 5% sucrose, and (3) with a mixture of 2.5% sucrose and 2.5% hydroxyl ethylene starch (HES), and then underwent freeze-drying. Freeze-drying in the absence of lyoprotectants caused an overall more disintegrated appearance of the histological architecture of the porcine valves, especially between the fibrosa and the ventricularis layers. Freeze-dried tissues with lyoprotectants have a looser network of collagen and elastic fibers with bigger pore sizes. Tissue freeze-dried in the absence of lyoprotecants had the largest pore sizes, whereas the tissue freeze-dried in the presence of protectants showed pores of intermediate sizes between the decellularized tissue and the unprotected freeze-dried samples. Tissue freeze-dried with sucrose alone displayed less porosity than tissue freeze-dried with the sucrose/HES mixture, whereas no significant differences in biomechanical properties were observed. Decellularization decreased the elastic modulus of artery tissue. The elastic modulus of freeze-dried tissue without protectants resembled that of decellularized tissue. The elastic modulus values of freeze-dried tissue stabilized by lyoprotectants were greater than those of decellularized tissue, but similar to those of native tissue.

Optimum Parameters for Freeze-Drying Decellularized Arterial Scaffolds

“…Lyophilization (freeze drying) offers a potential solution to these issues, as it produces a completely dry, stable scaffold that can be easily sterilized with ethylene oxide or gamma radiation. 10,12,13 Controlling the crystal nucleation rate and crystal size is essential when freeze drying biological tissue, as excessive crystal growth and size can disrupt extracellular matrix (ECM) components and create tissue heterogeneity. 14,15 The importance of determining the ideal freeze-drying parameters to preserve the ECM has been demonstrated for other biological tissues such as decellularized heart valves 12 and bovine pericardium.…”

“…10,12,13 Controlling the crystal nucleation rate and crystal size is essential when freeze drying biological tissue, as excessive crystal growth and size can disrupt extracellular matrix (ECM) components and create tissue heterogeneity. 14,15 The importance of determining the ideal freeze-drying parameters to preserve the ECM has been demonstrated for other biological tissues such as decellularized heart valves 12 and bovine pericardium. 16 Some studies have investigated the use of additional agents to preserve structure during both freezing and freeze drying.…”

“…16 Some studies have investigated the use of additional agents to preserve structure during both freezing and freeze drying. 12,17,18 However, cryoprotective agents have largely been used to preserve cell function, which is not relevant with the use of decellularized tissue. Lyoprotectants have been shown to stabilize proteins and liposomes during freeze drying, 18 but their use with whole tissue structure has not been widely studied.…”

Heart Valve Replacements with Regenerative Capacity

“…Similarly, lyophilization can improve the shelf life of decellularized tissues or engineered matrix. To retain the structural in- tegrity and the final geometry of the product, the use of sucrose solution and/or the optimization of the freezing cycle temperatures may be necessary [77]. Finally, biodegradable polymer-based TEHVs for in situ applications do not require special processing, gaining further interest in the community.…”

Combining endocytic and freezing‐induced trehalose uptake for cryopreservation of mammalian cells

“…It has been known for decades that disaccharides have good cryoprotective properties and also protect during drying. Nevertheless, they are not frequently used for cryopreservation of cells and have been primarily used for drying or freeze‐drying of proteins and other biological compounds . Disaccharides such as sucrose and trehalose have been used as secondary cryoprotectants in conjunction with other cryoprotectants .…”