Biotechnologie de la reproduction chez le bovin Découverte et étude de nouvelles biomolécules d'intérêt

Toute perturbation de la mobilité des spermatozoïdes a un fort impact sur la fécondation etpeut conduire à une sous-fertilité ou infertilité. Des efforts importants ont donc été faits pouridentifier des composés pharmacologiques susceptibles de moduler la mobilité desspermatozoïdes. Les composés activateurs sont particulièrement utiles en azoospermie pouraméliorer l'extraction des spermatozoïdes testiculaires et dans le domaine de lacryoconservation car la mobilité des spermatozoïdes congelés-dégelés est réduite. L’étudeprésentée dans ce manuscrit s’attache plus particulièrement à l’identification d’un composéoriginal, à partir d'une bibliothèque de venins, capable d'améliorer la mobilité desspermatozoïdes de mammifères, y compris chez l'homme. Nous avons d'abord identifié dansle venin d'un scorpion S. m. palmatus un peptide de 73 acides aminés et riche en disulfurecapable d'activer puissamment la mobilité des spermatozoïdes. Nous avons ensuite purifié etcaractérisé ce peptide responsable par chromatographie liquide, spectrométrie de masse etsynthèse peptidique. Enfin, la puissance et la toxicité des versions purifiée et synthétique ducomposé identifié sur la motilité des spermatozoïdes ont été évaluées à l'aide de différentstests in vitro chez différentes espèces de mammifères. Nous avons observé que la puissancedu peptide était plus élevée sur les spermatozoïdes éjaculés frais avec une faible motilité,obtenant une augmentation de 100% de la vitesse curviligne dans les spermatozoïdes peuperformants. Nous avons également démontré que le peptide est efficace sur l'épididyme fraisdes bovins et des souris, l'éjaculation congelée des bovins et les spermatozoïdes frais destesticules de primates non humains.Ce manuscrit évoque également brièvement les travaux réalisés durant ce diplôme sur uncomposé inhibant la mobilité dont l’utilisation permettrait d’élargir la fenêtre de fécondationoptimale en insémination artificielle en retardant le phénomène de capacitation duspermatozoïde

Enzymatic activity of mouse group X-sPLA2 improves in vitro production of preimplantation bovine embryos

International audienceAssisted reproductive technologies (ART) are widely used for both humans and domestic animals. In bovine species, in vitro embryo production is increasingly used and significant efforts are being made to optimize media and culture conditions. Phospholipase A2 (PLA2) are lipolytic enzymes that hydrolyze glycerophospholipids to produce free fatty acids and lysophospholipids that have been found to be critical for many biological processes. Mouse group X secreted PLA 2 (mGX) is abundant in the male reproductive tract and its use during sperm capacitation has been shown to improve in vitro production of viable embryos in a mouse model. Here, we examined its effect in the bovine species, testing the impact of mGX on the three steps involved in vitro production of preimplantation embryos: oocyte maturation, fertilization and preimplantation development. We found that incubating cumulus oocyte complexes (COC) or gametes with mGX resulted in increased blastocyst hatching and blastocyst production , respectively. The increases of embryo production induced by the phospholipase mGX were not observed for the catalytically inactive mutant H48Q-mGX, suggesting that these effects require the enzymatic activity of mGX. We also tested bGIB, a bovine homolog of mGX. bGIB failed to improve blastocyst production, underlining the high specificity of mGX. In conclusion, the results presented show that the effects of mGX are not restricted to the mouse model and that it is potent in the bovine species as well. This result strengthens the potential of mGX as a "pro-fertility drug" for mammalian reproduction

Pantoprazole, a proton-pump inhibitor, impairs human sperm motility and capacitation in vitro

The effects of PPIs on human sperm fertilizing capacity were poorly investigated although these drugs are widely over-used. Two publications retrospectively studied relationships between any PPI intake and sperm parameters from patients consulting at infertility clinics, but the conclusions of these reports were contradictory. Only two reports investigated the effects of lansoprazole and omeprazole on sperm motility and found lansoprazole to be deleterious and omeprazole to be neutral for sperm motility. The inconsistency of the PPI effect in the previous reports emphasizes the need for more basic research on human spermatozoa, taking into account the hypothesis that the different PPI drugs may have different effects on sperm physiology

Is sperm FISH analysis still useful for Robertsonian translocations? Meiotic analysis for 23 patients and review of the literature

Résumé Contexte Le mode de ségrégation chromosomique le plus fréquemment observé chez les patients porteurs de translocation robertsonienne est. un mode équilibré. Les données semblent varier peu selon la translocation analysée. La relative constance des résultats dans le cas de ces translocations robertsoniennes rend elle inutile ces analyses chromosomiques pour ces patients? Patients et méthodes Nous avons analysé de façon rétrospective les données spermatiques et de ségrégation méiotique de 23 patients porteurs de translocation robertsonienne, de 2003 à 2017 et comparé les résultats observés à ceux décrits dans la littérature pour 187 patients. Résultats Le mode de ségrégation alterne est. prépondérant dans notre série de patients avec 73.45% ±8.05 de spermatozoïdes équilibrés (min 50.92%; max 89.99%). Ces résultats sont en accord avec les données de la littérature, toutes translocations confondues et selon le type de translocation (p > 0.05) sauf pour la translocation der(13;15) où ces taux sont significativement plus faibles (p < 0.05 vs der(13;14), der(14;21), (13;21) et der(15;22)). Nous observons également des taux de spermatozoïdes équilibrés significativement plus élevés chez les patients à spermogramme normal (p < 0.01). Conclusions Les différences observées dans les taux d’aneuploïdies entre les translocations der(13;15) et les autres translocations robertsoniennes et entre les porteurs de translocation à spermogramme normal ou altéré, et l’utilité de ces données dans le conseil génétique conduisent à poursuivre l’analyse systématique de la ségrégation méiotique pour les patients porteurs de translocations robertsoniennes et ceci particulièrement pour les translocations rares

Spermaurin, an La1-like peptide from the venom of the scorpion Scorpio maurus palmatus , improves sperm motility and fertilization in different mammalian species

International audienceSTUDY QUESTIONIs it possible to identify original compounds that are able to enhance sperm motility from the venom of the scorpion Scorpio maurus palmatus?SUMMARY ANSWERWe identified a potent disulfide-rich peptide (DRP) of 73 amino acids that significantly improved the motility of fresh and frozen-thawed sperm in different mammalian species, including human, and improved fertilization outcome in mouse IVF experiments.WHAT IS KNOWN ALREADYAny disturbance of sperm motility has a strong impact on fertilization and can lead to subfertility or infertility. Significant efforts have, therefore,  been made to identify pharmacological drugs that might improve sperm motility. Such compounds are particularly useful in azoospermia to improve testicular sperm extraction and in the domain of cryopreservation because the motility of frozen-thawed sperm is reduced.STUDY DESIGN, SIZE, DURATIONThis was a basic science/medical research study aimed at identifying original compounds from a library of venoms able to enhance mammalian sperm motility, including human. We first identified in the venom of a scorpion S. m. palmatus a fraction able to potently activate sperm motility. We next purified and characterized the compound by liquid chromatography, mass spectrometry and peptide synthesis. Finally, the potency and toxicity of both purified and synthetic versions of the identified compound on sperm motility were assessed using different in vitro tests in different mammalian species.PARTICIPANTS/MATERIALS, SETTING, METHODS For human sperm, biological samples were collected from normozoospermic donors and subfertile patients attending a reproduction department for diagnostic semen analysis. Testicular sperm was collected from cynomolgus monkeys (Macaca fascicularis) euthanized for the needs of specific authorized research projects. The peptide was also tested on bovine and mouse epidydimal sperm. We measured different sperm motility parameters with a computer-assisted sperm analysis system in the presence or absence of the peptide.MAIN RESULTS AND THE ROLE OF CHANCESize exclusion chromatography enabled us to isolate a fraction of the venom of S. m. palmatus able to increase sperm motility. By liquid chromatography and mass spectrometry, a peptide comprising 73 amino acids with 4 disulfide bridges was identified as responsible for the biological activity and called ‘spermaurin’. The identity of spermaurin was confirmed by chemical synthesis. We showed that the peptide increased the motility of fresh and frozen-thawed human sperm. We observed that the potency of the peptide was higher on fresh ejaculated spermatozoa with a low motility, achieving a 100% increase of curvilinear velocity in poorly performing sperm. We also demonstrated that peptide is effective on bovine and mouse fresh epididymal, bovine frozen-thawed ejaculated and fresh non-human primate testicular sperm. Finally, in mouse IVF, the production of 2-cell embryos was increased by 24% when sperm were treated with the peptide.LIMITATIONS, REASONS FOR CAUTIONThis work is an in vitro evaluation of the ability of spermaurin to improve sperm motility parameters. Another limitation of this study is the small number of human sperm samples tested with the natural (n = 36) and synthetic (n = 12) peptides. Moreover, the effect of the peptide on IVF outcome was only tested in mouse and further tests with human and bovine gametes are required to confirm and extend this result in other mammalian species.WIDER IMPLICATIONS OF THE FINDINGSThis work confirms our initial study showing that venoms represent an interesting source of molecules that are able to modify sperm physiology. Moreover, this work presents the first demonstrated biological action of a venom peptide from the scorpion S. m. palmatus with sequence similarities to La1 peptide from Liocheles australasiae (Wood scorpion), a widespread family of DRPs.STUDY FUNDING/COMPETING INTEREST(S)This work is part of the project ‘LAB COM-14 LAB7 0004 01-LIPAV’, funded by the program LabCom 2014 from t e French Research Agency (ANR). Dr Arnoult reports grants from IMV Technologies during the conduct of the study. In addition, Drs Arnoult, Martinez, Ray and Schmitt have a patent EP16305642.7 pending containing some of the information presented in this manuscript

Identification, Characterization and Synthesis of Walterospermin, a Sperm Motility Activator from the Egyptian Black Snake Walterinnesia aegyptia Venom

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Oligogenic heterozygous inheritance of sperm abnormalities in mouse

Male infertility is an important health concern that is expected to have a major genetic etiology. Although high-throughput sequencing has linked gene defects to more than 50% of rare and severe sperm anomalies, less than 20% of common and moderate forms are explained. We hypothesized that this low success rate could at least be partly due to oligogenic defects - the accumulation of several rare heterozygous variants in distinct, but functionally connected, genes. Here, we compared fertility and sperm parameters in male mice harboring one to four heterozygous truncating mutations of genes linked to multiple morphological anomalies of the flagellum (MMAF) syndrome. Results indicated progressively deteriorating sperm morphology and motility with increasing numbers of heterozygous mutations. This first evidence of oligogenic inheritance in failed spermatogenesis strongly suggests that oligogenic heterozygosity could explain a significant proportion of asthenoteratozoospermia cases. The findings presented pave the way to further studies in mice and man

Whole-exome sequencing identifies mutations in FSIP2 as a recurrent cause of multiple morphological abnormalities of the sperm flagella

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