Biomedical Engineering Reference
In-Depth Information
7.5.2 StorageofTissues
To cryopreserve successfully a large size tissue, one would rather avoid the del-
eterious effects of a cooling rate, similar to single cells. The power of the cold to
destroy undesirable cells like cancer cells is used to surgically operate on cells in
cryosurgery. In addition to the two types of cellular damage discussed earlier (solu-
tion injury and intracellular ice injury), exposure to the cold can eliminate blood
circulation (called ischemic injury) in the undesirable cells, thereby preventing the
delivery of nutrients and oxygen as well as the removal of wastes and CO
2
; as
blood contains a large amount of water, ice will readily form and grow throughout
the interconnected tubes. One of the practices for the short-term storage of vascu-
larized tissues is to purfuse the tissue with preservative solutions such as noncol-
loidal histidine-tryptophan-ketoglutarate (HTK) solution, hyperkalaemic N-Tris
(hydroxymethyl) methyl-2-aminoethane sulphonate balanced (anionic buffer), or
salt solution (CPTES). The key areas that need advancement are additives to pres-
ervation solutions, alternatives/adjuncts to preservation solutions, and optimum
perfusion technology.
Cryopreserving tissues for the long term poses more complex scenarios, al-
though equations developed for single cells are used to predict changes in tissue
volume. Tissues normally contain more than one cell type, which could demand
different freezing rates. Furthermore, the presence of an extracellular matrix and
a large amount of water make the penetration of CPA into the cell more difficult.
First, the microenvironment surrounding each cell needs to be equilibrated with the
CPA and then the CPA can enter the cell. However, the prolonged exposure of cells
to hypertonic conditions (caused by a high CPA content) and a high concentration
of CPA at temperatures are also deleterious. An emerging technology is vitrification
(glass formation), which allows cells to be preserved in their existing state without
going through the dehydration that is achieved in slow cooling. In vitrification,
high concentrations of CPAs (three to four times higher) are used with a cooling
rate nearly 10,000 times faster by plunging them directly into liquid nitrogen. The
rapid cooling is necessary for preventing the toxicity of the high levels of CPA at
room temperature and achieving vitrification. The glass transition occurs even at a
low cooling rate, if the concentration of CPA is high enough. However, introduc-
ing such a high concentration of CPA into cells, as well as into a tissue, is difficult
unless new CPAs (low viscosity, low toxicity) are discovered. Alternatively, adding
a pressure during a freezing process is made to reduce the concentration of CPA,
which can vitrify.
The cryopreservation procedures that are currently in use for tissues can vary
greatly from tissue bank to tissue bank. Furthermore, some of the current practices
are protected by trade secrets. Although the American Association of Tissue Banks
(AATB), a nonprofit organization, publishes standards for tissue banking, it does
not recommend any specific cryopreservation procedures. Rather, the AATB im-
poses guidelines for establishing procedures and implementing standard operations
in retrieval, processing, storage, and/or distribution of tissues that meets the needed
quality control for safe clinical usage. Thus, individual tissue banks need to develop
an optimal cryopreservation protocol for a particular tissue. Then the AATB can
help the tissue bank in implementing the protocol and accrediting the tissue bank.
