title: Identification of protein changes in human spermatozoa throughout the cryopreservation process.
abstract: Cryoinjury is a consequence of cryopreservation and may have a negative impact on sperm quality regarding motility, morphology, and viability. This study was designed to identify potential proteomic changes in human sperm cells throughout the cryopreservation process. Comparisons made within this study included the detection of the sperm proteomic changes induced by incubation of the sperm cells with a protein-free cryoprotectant (with and without CryoSperm), and the proteomic changes induced by freezing, thawing, and subsequent after-thawing incubation at two different temperatures (0 °C vs. 23 °C). Tandem Mass Tag (TMT) peptide labeling coupled with LC-MS/MS was used for protein quantification. LC-MS/MS resulted in the identification of 769 quantifiable proteins. The abundance of 105 proteins was altered upon CryoSperm incubation. Freezing and thawing also induced substantial protein changes. However, fewer changes were observed when semen was thawed and then maintained after-thawing at approximately 0 °C than when it was maintained after-thawing at 23 °C, with 60 and 99 differential proteins detected, respectively, as compared to unfrozen semen incubated in CryoSperm. Collectively, these differences indicate that substantial changes occur in the sperm proteome at every stage of the cryopreservation process which may ultimately impair the sperm fertilizing capability. This is the first study to compare protein levels in fresh and cryopreserved semen using the TMT technology coupled to LC-MS/MS.
authors.affiliation: Molecular Biology of Reproduction and Development Research Group, Department of Biomedicine, Faculty of Medicine, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Biochemistry and Molecular Genetics Service, Hospital Clínic, University of Barcelona, Barcelona, Spain.
authors.forename: O A
authors.initials: OA
authors.lastname: Bogle
authors.affiliation: Centre for Public Health, Reproductive Medicine, Institute of Clinical Science, Queen's University Belfast, Northern Ireland, UK.
authors.forename: K
authors.initials: K
authors.lastname: Kumar
authors.affiliation: Molecular Biology of Reproduction and Development Research Group, Department of Biomedicine, Faculty of Medicine, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Biochemistry and Molecular Genetics Service, Hospital Clínic, University of Barcelona, Barcelona, Spain.
authors.forename: C
authors.initials: C
authors.lastname: Attardo-Parrinello
authors.affiliation: Centre for Public Health, Reproductive Medicine, Institute of Clinical Science, Queen's University Belfast, Northern Ireland, UK.
authors.forename: S E M
authors.initials: SE
authors.lastname: Lewis
authors.affiliation: Proteomics Unit, Scientific Technical Services, University of Barcelona, Barcelona, Spain.
authors.forename: J M
authors.initials: JM
authors.lastname: Estanyol
authors.affiliation: Clinic Institute of Gynecology, Obstetrics and Neonatology, Clinic Hospital, Barcelona, Spain.
authors.forename: J L
authors.initials: JL
authors.lastname: Ballescà
authors.affiliation: Molecular Biology of Reproduction and Development Research Group, Department of Biomedicine, Faculty of Medicine, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Biochemistry and Molecular Genetics Service, Hospital Clínic, University of Barcelona, Barcelona, Spain.
authors.forename: R
authors.initials: R
authors.lastname: Oliva
mesh_terms: D000328:Adult; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents; D005306:Fertilization / Q000502:physiology; D006801:Humans; D008297:Male; D040901:Proteomics; D012662:Semen Preservation / Q000379:methods*; D013081:Sperm Motility / Q000502:physiology; D013094:Spermatozoa / Q000166:cytology / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D003451:Cryoprotective Agents
keywords: LC-MS/MS; cryoinjury; cryopreservation; proteomics; spermatozoa