IMPACT OF AMPK ACTIVATOR METFORMIN ON SPERM QUALITY

Semen cryopreservation allows crucial management of animal genetic diversity. However, the freeze-thaw process causes biochemical and physical alterations, impairing sperm energy-dependent functions. Currently, many chemicals are added to the media to enhance frozen-thawed sperm quality during artificial insemination. The aims of this study was to determine the effects of Metformin (Metf) on fresh chicken sperm motility and ability to perform acrosome reaction, and to evaluate Metf’s effects on the functions of cryopreserved sperm. Chicken semen was diluted and incubated at 35°C in media supplemented with or without different doses of 5’-AMP-Activated Protein Kinase (AMPK) activator, Metf (0,5 to 5 mM). We then looked for the concentration improving the most sperm quality to use it in the cryopreservation media used for chicken sperm. Our results show that 1 mM Metf is the concentration giving the best results regarding sperm quality. AMPKα phosphorylation, viability, acrosome reaction ability (AR), and various motility parameters, were negatively affected by the freeze-thaw process, and that Metf partially restored them. Sperm quality improved (mean increased by 23% for motility, by 10% for viability) as well as AMPKα phosphorylation (mean increased by 30%). Moreover, fluorescent intensity levels of phospho-AMPK were also stronger with Metf than in the control. These results show that the presence of Metf in fresh semen has a positive impact on the quality of sperm and helps reducing the gradual decline in sperm motility caused by cryopreservation by partially restoring several essential sperm functions, and thus leads to a better overall quality of cryopreserved sperm

Relationships between sterol/phospholipid composition and xenobiotic transport in nematodes

International audienceTherapeutic failure limits prophylaxis of nematode diseases and has been mainly attributed to mutations in cellular targets of anthelmintics. Besides these specific mechanisms, alterations of drug transport also occur in parasites resistant to anthelmintics and depend on both the presence of membrane pumps such as P-glycoprotein (Pgp) and on the lipid composition of membranes. We recently showed in the nematode Haemonchus contortus, using eggs as a model, that the total cholesterol (TC) concentration alters the transport of lipophilic molecules due to membrane pumps such as P-glycoprotein and the resistance to anthelmintics. The effect of TC may depend on the presence of other lipids interacting with TC. Therefore, we analysed the lipid composition and its relationship with Pgp and resistance to anthelmintics. Better correlations were found between Pgp and free cholesterol (FC) than with TC. We also showed that the relationships between lipid composition and resistance to anthelmintics or Pgp depended on the equilibrium between FC and phospholipids (PLs), mainly PLs known to be present primarily in either the external leaflets of cell membranes or the internal leaflets. The PLs phosphatidylcholine and phosphatidylethanolamine played the most significant role, but phosphatidic acid also influenced drug resistance

Search for the genes involved in oocyte maturation and early embryo development in the hen

<p>Abstract</p> <p>Background</p> <p>The initial stages of development depend on mRNA and proteins accumulated in the oocyte, and during these stages, certain genes are essential for fertilization, first cleavage and embryonic genome activation. The aim of this study was first to search for avian oocyte-specific genes using an <it>in silico </it>and a microarray approaches, then to investigate the temporal and spatial dynamics of the expression of some of these genes during follicular maturation and early embryogenesis.</p> <p>Results</p> <p>The <it>in silico </it>approach allowed us to identify 18 chicken homologs of mouse potential oocyte genes found by digital differential display. Using the chicken Affymetrix microarray, we identified 461 genes overexpressed in granulosa cells (GCs) and 250 genes overexpressed in the germinal disc (GD) of the hen oocyte. Six genes were identified using both <it>in silico </it>and microarray approaches. Based on GO annotations, GC and GD genes were differentially involved in biological processes, reflecting different physiological destinations of these two cell layers. Finally we studied the spatial and temporal dynamics of the expression of 21 chicken genes. According to their expression patterns all these genes are involved in different stages of final follicular maturation and/or early embryogenesis in the chicken. Among them, 8 genes (<it>btg4</it>, <it>chkmos</it>, <it>wee</it>, <it>zpA</it>, <it>dazL</it>, <it>cvh</it>, <it>zar1 </it>and <it>ktfn) </it>were preferentially expressed in the maturing occyte and <it>cvh</it>, <it>zar1 </it>and <it>ktfn </it>were also highly expressed in the early embryo.</p> <p>Conclusion</p> <p>We showed that <it>in silico </it>and Affymetrix microarray approaches were relevant and complementary in order to find new avian genes potentially involved in oocyte maturation and/or early embryo development, and allowed the discovery of new potential chicken mature oocyte and chicken granulosa cell markers for future studies. Moreover, detailed study of the expression of some of these genes revealed promising candidates for maternal effect genes in the chicken. Finally, the finding concerning the different state of rRNA compared to that of mRNA during the postovulatory period shed light on some mechanisms through which oocyte to embryo transition occurs in the hen.</p

Protein expression reveals a molecular sexual identity of avian primordial germ cells at pre-gonadal stages

International audienceIn poultry, in vitro propagated primordial germ cells (PGCs) represent an important tool for the cryopreservation of avian genetic resources. However, several studies have highlighted sexual differences exhibited by PGCs during in vitro propagation, which may compromise their reproductive capacities. To understand this phenomenon, we compared the proteome of pregonadal migratory male (ZZ) and female (ZW) chicken PGCs propagated in vitro by quantitative proteomic analysis using a GeLC-MS/MS strategy. Many proteins were found to be differentially abundant in chicken male and female PGCs indicating their early sexual identity. Many of the proteins more highly expressed in male PGCs were encoded by genes localised to the Z sex chromosome. This suggests that the known lack of dosage compensation of the transcription of Z-linked genes between sexes persists at the protein level in PGCs, and that this may be a key factor of their autonomous sex differentiation. We also found that globally, protein differences do not closely correlate with transcript differences indicating a selective translational mechanism in PGCs. Male and female PGC expressed protein sets were associated with differential biological processes and contained proteins known to be biologically relevant for male and female germ cell development, respectively. We also discovered that female PGCs have a higher capacity to uptake proteins from the cell culture medium than male PGCs. This study presents the first evidence of an early predetermined sex specific cell fate of chicken PGCs and their sexual molecular specificities which will enable the development of more precise sex-specific in vitro culture conditions for the preservation of avian genetic resources

Extension des possibilités de collections de la cryobanque aviaire à des espèces autres que Gallus gallus

National audienc

Current status in avian semen cryopreservation

International audienc

Reproduction and reproductive biotechnologies for the preservation of endangered avian species and breeds

International audienc

Freezing avian semen

 Semen cryopreservation is an important tool for the storage of reproductive cells used for the ex situ management of genetic diversity in birds. Recent advances in poultry semen cryopreservation technology have resulted in the emergence of cryobanking, which is now being developed in an increasing number of countries. In addition, semen freezing methods are now effective for various domestic and wild bird species, although species such as guinea fowls are still highly affected by the process. The methods of freezing avian semen now use a small number of cryoprotectants, (mainly glycerol, dimethyl or N-methylacetamide) and cell packaging (such as straws or pellets). Temperature curves of freezing and thawing remain the most variable points as very slow or very rapid curves are sometimes used in the same species. Specific features of bird reproductive physiology are very important for sperm cryopreservation and application. The characteristics of initial semen quality, including cell membrane properties, mobility capacity and ability to undergo the acrosome reaction are critical points. The specific features of the oviparity system of reproduction and the internal fertilization process affect the conditions of semen use. The in vivo storage of spermatozoa in the sperm storage tubules of the female genital tract, and the conditions of the drastic selection of sperm in the highly specialized female oviduct constitute major factors that are highly species-specific. These constraints involve adaptations of the semen media used for freezing and of the zootechnical parameters of semen use. This review highlights the main factors that are critical for the success of semen cryopreservation in bird species

Froid de canard sur les semences

Communiqué de presse Les artifices de la reproductionabsen

Semence et cryopréservation de cellules de reproduction chez les oiseaux

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