Abstract
Currently, two conventional freezing techniques are used in sperm cryopreservation: slow freezing (SF) and rapid freezing (RF). Despite the protocolar improvements, cryopreservation still induces significant alterations in spermatozoon that are poorly understood. Here, available proteomic data from human cryopreserved sperm was analyzed through bioinformatic tools to unveil key differentially expressed proteins (DEPs) that can be used as modulation targets or quality markers. From the included proteomic studies, 160 and 555 DEPs were collected for SF and RF groups, respectively. For each group, an integrative network was constructed using gene ontology and protein-protein interaction data to identify key DEPs. Among them, arylsulfatase A (ARSA) was highlighted in both freezing networks, and low ARSA levels have been associated with poor-sperm quality. Thus, ARSA was selected for further experimental investigation and its levels were assessed in cryopreserved samples by western blot. ARSA levels were significantly decreased in RF and SF samples (∼31.97 and ∼39.28%, respectively). The bioinformatic analysis also revealed that the DEPs were strongly associated with proteasomal and translation pathways. The purposed bioinformatic approach allowed the identification of potential key DEPs in freeze-thawed human spermatozoa. ARSA has the potential to be used as a marker to assess sperm quality after cryopreservation.
Highlights
Sperm cryopreservation is a key procedure in reproductive medicine worldwide
From the included proteomic studies (Bogle et al, 2017; Fu et al, 2019; Li et al, 2019), decreased and increased differentially expressed proteins (DEPs) were collected for the two conventional freezing techniques used in sperm cryopreservation: slow freezing (SF) and rapid freezing (RF)
Decreased protein levels are explained by the cryopreservation process, which leads to protein degradation, plasmatic membrane damage, and osmotic stress induced by the cryoprotectant
Summary
Sperm cryopreservation is a key procedure in reproductive medicine worldwide. This technique has been used extensively to preserve male fertility in situations that can lead to testicular dysfunction and ejaculatory failures such as some pathological scenarios (malignant cancers and severe spinal cord injuries), corrective surgeries, or chemo-/radio-therapies (Di Santo et al, 2012; Tournaye et al., Abbreviations: ARSA, Arylsulfatase A; DEPs, Differentially Expressed Proteins; GO, Gene Ontology; PPI, Protein-Protein Interaction; RF, Rapid Freezing; SF, Slow Freezing; ZP, Zona-Pellucida.Key DEPs in Cryopreserved Human Sperm2014; Tomlinson et al, 2015). Sperm cryopreservation is a key procedure in reproductive medicine worldwide. This technique has been used extensively to preserve male fertility in situations that can lead to testicular dysfunction and ejaculatory failures such as some pathological scenarios (malignant cancers and severe spinal cord injuries), corrective surgeries, or chemo-/radio-therapies Two conventional freezing techniques are used in sperm cryopreservation: slow freezing (SF) and rapid freezing (RF) (World Health Organization, 2010). Sperm samples frozen through a RF protocol are exposed to nitrogen vapors (-80°C) during 10–15 min before being placed in liquid nitrogen (-196°C) (World Health Organization, 2010; Di Santo et al, 2012). The efforts made in the last decades, the current cryopreservation protocols are still far from ideal inducing significant deleterious structural and molecular alterations in spermatozoa (Hezavehei et al, 2018; Rienzi et al, 2018; Kumar et al, 2019)
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