Abstract
Currently, evaluation of sperm quality is primarily based on in vitro measures of sperm function such as motility, viability and/or acrosome reaction. However, results are often poorly correlated with fertility, and alternative diagnostic tools are therefore needed both in veterinary and human medicine. In a recent pilot study, we demonstrated that MS profiles from intact chicken sperm using MALDI-TOF profiles could detect significant differences between fertile/subfertile spermatozoa showing that such profiles could be useful for in vitro male fertility testing. In the present study, we performed larger standardized experimental procedures designed for the development of fertility- predictive mathematical models based on sperm cell MALDI-TOF MS profiles acquired through a fast, automated method. This intact cell MALDI-TOF MS-based method showed high diagnostic accuracy in identifying fertile/subfertile males in a large male population of known fertility from two distinct genetic lineages (meat and egg laying lines). We additionally identified 40% of the m/z peaks observed in sperm MS profiles through a top-down high-resolution protein identification analysis. This revealed that the MALDI-TOF MS spectra obtained from intact sperm cells contained a large proportion of protein degradation products, many implicated in important functional pathways in sperm such as energy metabolism, structure and movement. Proteins identified by our predictive model included diverse and important functional classes providing new insights into sperm function as it relates to fertility differences in this experimental system. Thus, in addition to the chicken model system developed here, with the use of appropriate models these methods should effectively translate to other animal taxa where similar tests for fertility are warranted.
Highlights
From the ‡Institute for Agricultural Research (INRA), UMR85 Physiologie de la Reproduction et des Comportements, F-37380 Nouzilly, France; §CNRS, UMR7247, F-37380 Nouzilly, France; ¶Universite Francois Rabelais de Tours, F-37000 Tours, France; ʈIFCE, F-37380 Nouzilly, France; **INRA, Plateforme d’Analyse Integrative des Biomolecules, Laboratoire de Spectrometrie de Masse, F-37380 Nouzilly, France; ‡‡INRA, UMR1282 Infectiologie et Sante Publique, F-37380 Nouzilly, France; §§Universite Francois Rabelais de Tours, UMR1282 Infectiologie et Sante Publique, F-37000 Tours, France
In an initial attempt to test if this analytical approach could be employed to phenotype sperm cells with different fertility abilities, we observed that subfertile sperm cells ICM-Mass spectrometry (MS) profiles showed characteristic features that could be useful to discriminate individuals regarding their fertility, and that these could be confidently identified by top-down high resolution mass spectrometry (HRMS) [23]
Our previous results indicated that sperm cells Intact Cell MALDI-TOF-Mass Spectrometry (ICM-MS) profiling coupled with top-down sifier-non parametric; HRMS, High Resolution Mass Spectrometry; reversed phase (RP), Reverse Phase; gel filtration (GF), Gel Filtration; HCD, High Collision Energy; PUF, ProSight Upload Format; CASA, Computer-Assisted Sperm Snalysis; HTM, Hamilton-Thorn Motility Analyzer; IVOS, Integrated Visual Optical System; VAP, Average Path Velocity; PPV, Positive Predictive Value; NPV, Negative Predictive Value; LRϩ, Positive Likelihood Ratio; LR, Negative Likelihood Ratio; Receiver operating characteristic (ROC), Receiver Operating Characteristic; AUC, Area Under the Curve; HGNC, Hugo Gene Nomenclature Committee; WB, Western Blotting; ATP, Adenosine Triphosphate
Summary
From the ‡INRA, UMR85 Physiologie de la Reproduction et des Comportements, F-37380 Nouzilly, France; §CNRS, UMR7247, F-37380 Nouzilly, France; ¶Universite Francois Rabelais de Tours, F-37000 Tours, France; ʈIFCE, F-37380 Nouzilly, France; **INRA, Plateforme d’Analyse Integrative des Biomolecules, Laboratoire de Spectrometrie de Masse, F-37380 Nouzilly, France; ‡‡INRA, UMR1282 Infectiologie et Sante Publique, F-37380 Nouzilly, France; §§Universite Francois Rabelais de Tours, UMR1282 Infectiologie et Sante Publique, F-37000 Tours, France. The progress in the general use of MS-based profiling has been somewhat hampered by the difficulties in confidently identifying diagnostic MS signatures, because traditional bottom-up approaches are certainly not adequate In this sense, the development of direct identification of the peptide/protein through top-down high resolution mass spectrometry (HRMS) has opened new possibilities. The objectives of the present study were [1] to acquire sperm cells ICM-MS profiles in a standardized and automated way from a large and representative cohort of roosters of known fertility, [2] to employ sperm cells ICM-MS profiles to construct fertility-predictive mathematical models, [3] to test the diagnostic performance of the ICM-MS -based models versus other traditional sperm cell quality tests, and [4] to analyze the native and endogenous peptides/proteins of spermatozoa by LC-top-down HRMS after two different fractionation modes (gel filtration and reverse phase chromatography), in order to identify represented biomolecules in the ICM-MS spectra
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
