Dry storage of mammalian spermatozoa and cells: state-of-the-art and possible future directions

Abstract

This review provides a snapshot of the current state-of-the-art of drying cells and spermatozoa. The major successes and pitfalls of the most relevant literature are described separately for spermatozoa and cells. Overall, the data published so far indicate that we are closer to success in spermatozoa, whereas the situation is far more complex with cells. Critical for success is the presence of xeroprotectants inside the spermatozoa and, even more so, inside cells to protect subcellular compartments, primarily DNA. We highlight workable strategies to endow gametes and cells with the right combination of xeroprotectants, mostly sugars, and late embryogenesis abundant (LEA) or similar ‘intrinsically disordered’ proteins to help them withstand reversible desiccation. We focus on the biological aspects of water stress, and in particular cellular and DNA damage, but also touch on other still unexplored issues, such as the choice of both dehydration and rehydration methods or approaches, because, in our view, they play a primary role in reducing desiccation damage. We conclude by highlighting the need to exhaustively explore desiccation strategies other than lyophilisation, such as air drying, spin drying or spray drying, ideally with new prototypes, other than the food and pharmaceutical drying strategies currently used, tailored for the unique needs of cells and spermatozoa.

Long-term preservation of germ cells and gonadal tissues at ambient temperatures

“…Note that excellent illustrations of the methods described below are available in recent reviews (Loi et al 2021, Weng 2021.…”

“…Lyophilization (freeze-drying) is the most widely used method for germplasm desiccation (Saragusty et al 2020, Loi et al 2021, mainly for sperm cells. The process includes freezing samples followed by primary drying through sublimation of ice at freezing temperature under vacuum and secondary drying through desorption by slowly elevating temperature under vacuum (Weng 2021).…”

“…Unfortunately, desiccation genes or related analogs are not present in vertebrate genomes. It is therefore mandatory to directly supply 'xero-protectants' to the cells from vertebrate species before removing the water content (Loi et al 2021).…”

“…So far, most studies focused on optimizing the drying process to reduce the damage, and little is known about rehydration conditions, which is a critical step (Loi et al 2021). Commonly, rehydration is achieved by simply adding the volume of water lost back to the sample.…”

Damages and stress responses in sperm cells and other germplasms during dehydration and storage at nonfreezing temperatures for fertility preservation

“…The capacity to synthesize and accumulate intracellular disaccharides (mainly trehalose or sucrose) or specialized proteins is the main strategy to survive extreme water loss (Wolkers & Oldenhof, 2021); however, that capacity has been lost in vertebrate species. For cell or tissue samples from more evolved organisms, it is therefore necessary to incorporate the xeroprotectants (agents protecting the organism from damages in very dry conditions) into cells before starting the water removal (Loi et al, 2021). Major advantages of natural nonreducing disaccharides like trehalose also are their low toxicity and high glass transition temperature (possibility to vitrify at nonfreezing temperatures) compared to conventional cryoprotectants such as dimethyl sulfoxide, ethylene glycol, or 1,2‐propanediol (Chen et al, 2000).…”