Cryopreservation and vitrification: recent advances in fertility preservation technologies

Abstract

Over the last half the 20th Century, reproductive medicine has become a critically important branch of modern medical science. Fertility preservation is a vital branch of reproductive medicine and involves the preservation of gametes (sperm and oocytes), embryos, and reproductive tissues (ovarian and testicular tissues) for use in artificial reproduction. This technology gives millions of people suffering from reproductive ailments, cancer patients who have their reproductive functions destroyed by therapy (chemotherapy and radiation) and people undergoing sterilization, a chance to conceive. The most common fertility preservation technique is cryopreservation, which involves freezing cells and tissues at cryogenic temperatures. Cryopreserved cells and tissues can endure storage for centuries with almost no change in functionality or genetic information, making this storage method highly attractive. However, developing efficient cryopreservation techniques is challenging, as both freezing and thawing exposes cells to severe stresses, potentially causing cell death. There are two major techniques for cryopreservation: freeze-thaw processes and vitrification. The major difference between them is the total avoidance of ice formation in vitrification. The use of both theoretical models that describe cell response to freezing and thawing, and experimental investigations of freezing behavior, has led to the development of successful freeze-thaw and vitrification procedures for a number of cell types. Among reproductive cells, there exist efficient cryopreservation techniques for spermatozoa and embryos. Oocytes, however, present significant hurdles in achieving successful cryopreservation, primarily due to their sensitive microtubule structure. Recently, cryopreservation of ovarian and testicular tissues has been investigated with success reported. Ovarian cryopreservation can help circumvent many of the problems associated with oocyte cryopreservation, while testicular tissue preservation may be helpful when insufficient sperm counts are available for routine semen preservation.

Live birth after SrCl2 oocyte activation in previous repeated failed or low fertilization rates after ICSI of frozen-thawed testicular spermatozoa: case report

“…Especially, cryopreservation of testicular spermatozoa prior to ICSI is routinely performed in patients with OA and NOA [21], because if pregnancy is not achieved in the first TESE-ICSI cycle, a repeat TESE may be necessary. The results of TESE-ICSI are determined by the availability of motile spermatozoa and the type of azoospermia, either OA or NOA.…”

Cryopreservation of Testicular Tissue

“…Therefore, optimal thawing and CPA removal procedures are also critical factors for cell/tissue survival after freezing (Bagchi et al, 2008). Earlier studies pointed out that consistent cooling and thawing rates (slow-freezing followed by slow-thawing, or fast-freezing followed by fast-thawing) can improve cell/tissue survival after cryopreservation (Whittingham et al, 1972).…”

“…Unlike cell suspensions, the cellular organization and structural composition of the tissue may be seriously affected by cryogenic damage through widespread extracellular ice formation (Hunt et al, 1982;Taylor & Pegg, 1983). Ice formation within a tissue, initiated in the extracellular space, leads to an osmotic gradient across the cell membranes, causing intracellular water to move toward the concentrated extracellular space surrounding the cells (Bagchi et al, 2008;Fuller, 2004). Due to the differential destructive effects of extracellular ice formation between cell suspensions and complex tissues, conventional approaches to cryopreservation of cells, even testis cells for instance, may not necessarily be suitable for multicellular tissues such as the testis tissue.…”

“…These two cryoprotectants have mainly been used since then as classic cryoprotective additives, although many other CPAs have been introduced. Permeating CPAs, such as DMSO, glycerol, methanol, propanediol, ethylene glycol, and dimethyl acetaldehyde, as well as non-permeating CPAs, including sucrose, dextran, albumin, polyvinyl pyrollidone, and hydroxyethyl starch, have also been shown to afford effective cryoprotection (Bagchi et al, 2008;Fuller, 2004).…”

“…Some CPAs, such as DMSO or glycerol, inhibit nucleation by increasing the high viscosity of intracellular water (Fuller, 2004). Nonpermeating CPAs, on the other hand, increase and promote cellular dehydration by increasing the extracellular solute concentration thereby reducing intracellular crystallization (Bagchi et al, 2008).…”