Chapter 40
Freeze-Drying of Decellularized Heart Valves
for Off-the-Shelf Availability
Willem F. Wolkers and Andres Hilfiker
Abstract
Malfunctioning heart valves can cause severe health problems, which if left untreated can lead to death. One
of the treatment options is to replace a diseased heart valve with a decellularized valve construct prepared
from human or animal material. Decellularized tissue scaffolds closely resemble properties of native tissue,
while lacking immunogenic factors of cellular components. After transplantation, circulating stem and
progenitor cells of the patient adhere to the scaffold resulting in in vivo tissue regeneration of the valve.
Decellularized heart valve scaffold implants need to be stored to be readily available whenever needed,
which can be done by freeze-drying. The advantage of freeze-drying is that it does not require bulky and
energy-consuming freezing equipment for storage and allows easy transport. This chapter outlines the
entire process from decellularization to freeze-drying to obtain dry decellularized heart valves, which after a
simple rehydration step, can be used as implants. The protocol is described for porcine heart valves, but
procedures can easily be adapted for material obtained from other species.
Key words Freeze-drying, Sucrose, Pulmonary heart valve conduits, Matrix implants, Anhydrobiosis
1 Introduction
Malfunctioning heart valves can cause various health problems,
which eventually may result in death. Valve-related diseases result
in a reduction of the forward blood flow limiting tissue oxygenation. Heart valve dysfunction is typically treated by replacing the
diseased valve with mechanical or biological prostheses. Side effects
associated with mechanical valves include hemorrhage and thromboembolism, whereas bioprosthetic valves, which are made by
treating bovine or porcine tissue with glutaraldehyde, have been
implicated in fibrosis, calcification, degeneration, and immunogenic complications [1, 2]. The use of decellularized native heart
valves to replace a malfunctioning heart valve reduces the risk of
immunogenic rejection, and anti-coagulation therapy is generally
not required [3, 4]. Decellularization refers to the removal of cells
and cellular remnants leaving biological scaffold material composed
Willem F. Wolkers and Harrie ¨ tte Oldenhof (eds.), Cryopreservation and Freeze-Drying Protocols, Methods in Molecular Biology,
vol. 2180, https://doi.org/10.1007/978-1-0716-0783-1_40, © Springer Science+Business Media, LLC, part of Springer Nature 2021
731
Freeze-Drying of Decellularized Heart Valves
for Off-the-Shelf Availability
Willem F. Wolkers and Andres Hilfiker
Abstract
Malfunctioning heart valves can cause severe health problems, which if left untreated can lead to death. One
of the treatment options is to replace a diseased heart valve with a decellularized valve construct prepared
from human or animal material. Decellularized tissue scaffolds closely resemble properties of native tissue,
while lacking immunogenic factors of cellular components. After transplantation, circulating stem and
progenitor cells of the patient adhere to the scaffold resulting in in vivo tissue regeneration of the valve.
Decellularized heart valve scaffold implants need to be stored to be readily available whenever needed,
which can be done by freeze-drying. The advantage of freeze-drying is that it does not require bulky and
energy-consuming freezing equipment for storage and allows easy transport. This chapter outlines the
entire process from decellularization to freeze-drying to obtain dry decellularized heart valves, which after a
simple rehydration step, can be used as implants. The protocol is described for porcine heart valves, but
procedures can easily be adapted for material obtained from other species.
Key words Freeze-drying, Sucrose, Pulmonary heart valve conduits, Matrix implants, Anhydrobiosis
1 Introduction
Malfunctioning heart valves can cause various health problems,
which eventually may result in death. Valve-related diseases result
in a reduction of the forward blood flow limiting tissue oxygenation. Heart valve dysfunction is typically treated by replacing the
diseased valve with mechanical or biological prostheses. Side effects
associated with mechanical valves include hemorrhage and thromboembolism, whereas bioprosthetic valves, which are made by
treating bovine or porcine tissue with glutaraldehyde, have been
implicated in fibrosis, calcification, degeneration, and immunogenic complications [1, 2]. The use of decellularized native heart
valves to replace a malfunctioning heart valve reduces the risk of
immunogenic rejection, and anti-coagulation therapy is generally
not required [3, 4]. Decellularization refers to the removal of cells
and cellular remnants leaving biological scaffold material composed
Willem F. Wolkers and Harrie ¨ tte Oldenhof (eds.), Cryopreservation and Freeze-Drying Protocols, Methods in Molecular Biology,
vol. 2180, https://doi.org/10.1007/978-1-0716-0783-1_40, © Springer Science+Business Media, LLC, part of Springer Nature 2021
731