Drying and temperature induced conformational changes of nucleic acids and stallion sperm chromatin in trehalose preservation formulations

Abstract

Even though dried sperm is not viable, it can be used for fertilization as long as its chromatin remains intact. In this study, we investigated drying- and temperature-induced conformational changes of nucleic acids and stallion sperm chromatin. Sperm was diluted in preservation formulations with and without sugar/albumin and subjected to convective drying at elevated temperatures on glass substrates. Accumulation of reactive oxygen species was studied during storage at different temperatures, and the sperm chromatin structure assay was used to assess DNA damage. Fourier transform infrared spectroscopy was used to identify dehydration and storage induced conformational changes in isolated DNA and sperm chromatin. Furthermore, hydrogen bonding in the preservation solutions associated with storage stability were investigated. Reactive oxygen species and DNA damage in dried sperm samples were found to accumulate with increasing storage temperature and storage duration. Non-reducing disaccharides (i.e., trehalose, sucrose) and albumin counteracted oxidative stress and preserved sperm chromatin during dried storage, whereas glucose increased DNA damage during storage. When sperm was dried in the presence of trehalose and albumin, no spectral changes were detected during storage at refrigeration temperatures, whereas under accelerated aging conditions, i.e., storage at 37 °C, spectral changes were detected indicating alterations in sperm chromatin structure.

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

“…Lee et al, 2019; Silva et al, 2020). Importantly, in addition to the immediate effect of dehydration, the incidence of DNA damage increases with higher storage temperatures and longer storage duration in air‐dried equine sperm (measured by Fourier transform infrared spectroscopy) (Brogna et al, 2021), and in lyophilized murine sperm (comet assay) (Kamada et al, 2018; Kawase et al, 2005).…”

“…We have already mentioned trehalose as one of the main xeroprotectants that mitigates most of the damages, by stabilizing cellular membranes, proteins, and DNA during dehydration (Brogna et al, 2021; Crowe, 2012; Wolkers & Oldenhof, 2021; M. Zhang et al, 2016). Trehalose substitutes water molecules during dehydration to form hydrogen bonds with a lipid membrane, proteins, or nucleic acids, which maintains three‐dimensional (3D)/ordered conformations and structures (Golovina et al, 2010; Jain & Roy, 2009).…”

“…As a result, drying in the presence of trehalose has been shown to protect DNA, membranes, and structural integrity. It also reduces reactive oxygen species formation and improves the developmental potential of spermatozoa in humans (Shahmoradi et al, 2022), macaque (Sánchez‐Partida et al, 2008), mouse (Ito et al, 2019; McGinnis et al, 2005), cat (Patrick et al, 2017), stallion (Brogna et al, 2021), and bull (Martins et al, 2007; Sitaula et al, 2009). Moreover, mouse sperm nuclei lyophilized in the presence of trehalose can withstand short exposure to a wide range of temperatures from −196°C to 150°C and still retain the capacity to produce offspring (S. Wakayama et al, 2019).…”

“…Cells are found to exert natural protective measures to overcome the dehydration stress. For instance, Fourier transform infrared spectroscopic analysis revealed that stallion sperm DNA transitions to a more compact conformation upon air dehydration (Brogna et al, 2021). Such conformational change has been suggested to play a role in resistance to dry conditions in eukaryotic cells (Whelan et al, 2011).…”

Desiccated cat spermatozoa retain DNA integrity and developmental potential after prolonged storage and shipping at non-cryogenic temperatures

“…Trehalose is the most commonly used disaccharides due to its high glass transition temperature (T g ; the temperature at which molecular property changes from liquid to amorphous glass). The beneficial effect of trehalose in protecting spermatozoa against desiccation stress has been demonstrated in spermatozoa from the mouse [11][12][13], cat [14], stallion [15], macaque [16], and human [17]. For example, McGinnis et al reported a significant higher developmental potential of mouse sperm when desiccated in medium with trehalose than in medium without, resulting in a four-to tenfold increase in blastocyst formation [12].…”