Personne :
Blondin, Patrick.

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Département des sciences animales, Faculté des sciences de l'agriculture et de l'alimentation, Université Laval
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  • Publication
    The use of adenosine to inhibit oocyte meiotic resumption in Bos taurus during pre-IVM and its potential to improve oocyte competence
    (Elsevier Inc., 2019-10-07) Vigneault, Christian; Caballero, Julieta; Richard, François J.; Sirard, Marc-André; Blondin, Patrick.
    One of the major challenges of artificial reproductive technologies is to develop new methods for pro-ducing greater numbers of embryos. An oocyte fosters the ability to develop into an embryo beforeoocyte meiotic resumption. The aim of the present study was to assess the effect of adenosine (ADO), apurine nucleoside found in follicularfluid, on the inhibition of oocyte meiotic resumption and theproduction of blastocysts. The results showed the efficacy of ADO to inhibit oocyte meiotic resumption.The use of ADO (3 mM) during a pre-in vitro maturation (pre-IVM) culture period of 6 h resulted in asignificant increase (p<0.05) of blastocysts compared to control conditions with no pre-IVM cultureperiod. No effect on the percentage of cleavage was observed. The effect of adenosine on blastocyst yieldwas time- and concentration-dependent with an optimum effect at 3 mM for 6 h. Supplementing theADO pre-IVM culture medium with estradiol, follicle-stimulating hormone, progesterone, epidermalgrowth factor, insulin-like growth factor-2 or reelin did not improve the blastocyst yield. Transcriptionalanalyses of ADO-treated cumulus cells revealed that NRP1, RELN, MAN1A1, THRA and GATM were up-regulated. Finally, bioinformatic analysis identified mitochondrial function as the top canonicalpathway affected by ADO. This opens up new opportunities for further investigations.
  • Publication
    Cumulus cell gene expression associated with pre-ovulatory acquisition of developmental competence in bovine oocytes
    (Commonwealth Scientific and Industrial Research Organization, 2013-07-05) Vigneault, Christian; Bunel, Audrey; Nivet, Anne-Laure; Richard, François J.; Sirard, Marc-André; Blondin, Patrick.
    The final days before ovulation impact significantly on follicular function and oocyte quality. This study investigated the cumulus cell (CC) transcriptomic changes during the oocyte developmental competence acquisition period. Six dairy cows were used for 24 oocyte collections and received FSH twice daily over 3 days, followed by FSH withdrawal for 20, 44, 68 and 92 h in four different oestrous cycles for each of the six cows. Half of the cumulus–oocyte complexes were subjected to in vitro maturation, fertilisation and culture to assess blastocyst rate. The other half of the CC underwent microarray analysis (n = 3 cows, 12 oocyte collections) and qRT-PCR (n = 3 other cows, 12 oocyte collections). According to blastocyst rates, 20 h of FSH withdrawal led to under-differentiated follicles (49%), 44 and 68 h to the most competent follicles (71% and 61%) and 92 h to over-differentiated ones (51%). Ten genes, from the gene lists corresponding to the three different follicular states, were subjected to qRT-PCR. Interestingly, CYP11A1 and NSDHL gene expression profiles reflected the blastocyst rate. However most genes were associated with the over-differentiated status: GATM, MAN1A1, VNN1 and NRP1. The early period of FSH withdrawal has a minimal effect on cumulus gene expression, whereas the longest period has a very significant one and indicates the beginning of the atresia process.
  • Publication
    Changes in granulosa cells' gene expression associated with increased oocyte competence in bovine.
    (Journals of Reproduction and Fertility, 2013-05-01) Nivet, Anne-Laure; Vigneault, Christian; Sirard, Marc-André; Blondin, Patrick.
    One of the challenges in mammalian reproduction is to understand the basic physiology of oocyte quality. It is believed that the follicle status is linked to developmental competence of the enclosed oocyte. To explore the link between follicles and competence in cows, previous research at our laboratory has developed an ovarian stimulation protocol that increases and then decreases oocyte quality according to the timing of oocyte recovery post-FSH withdrawal (coasting). Using this protocol, we have obtained the granulosa cells associated with oocytes of different qualities at selected times of coasting. Transcriptome analysis was done with Embryogene microarray slides and validation was performed by real-time PCR. Results show that the major changes in gene expression occurred from 20 to 44 h of coasting, when oocyte quality increases. Secondly, among upregulated genes (20-44 h), 25% were extracellular molecules, highlighting potential granulosa signaling cascades. Principal component analysis identified two patterns: one resembling the competence profile and another associated with follicle growth and atresia. Additionally, three major functional changes were identified: (i) the end of follicle growth (BMPR1B, IGF2, and RELN), involving interactions with the extracellular matrix (TFPI2); angiogenesis (NRP1), including early hypoxia, and potentially oxidative stress (GFPT2, TF, and VNN1) and (ii) apoptosis (KCNJ8) followed by iii) inflammation (ANKRD1). This unique window of analysis indicates a progressive hypoxia during coasting mixed with an increase in apoptosis and inflammation. Potential signaling pathways leading to competence have been identified and will require downstream testing. This preliminary analysis supports the potential role of the follicular differentiation in oocyte quality both during competence increase and decrease phases.
  • Publication
    Accès libre
    Breed specific factors influence embryonic lipid composition : comparison between Jersey and Holstein
    (Commonwealth Scientific and Industrial Research Organization, 2015-01-15) Baldoceda Baldeon, Luis Manuel; Vigneault, Christian; Gilbert, Isabelle; Gagné, Dominic; Robert, Claude; Blondin, Patrick.; Ramires Ferreira, Christina
    Some embryos exhibit better survival potential to cryopreservation than others. The cause of such a phenotype is still unclear and may be due to cell damage during cryopreservation, resulting from overaccumulation and composition of lipids. In cattle embryos, in vitro culture conditions have been shown to impact the number of lipid droplets within blastomeres. Thus far, the impact of breed on embryonic lipid content has not been studied. In the present study were compared the colour, lipid droplet abundance, lipid composition, mitochondrial activity and gene expression of in vivo-collected Jersey breed embryos, which are known to display poor performance post-freezing, with those of in vivo Holstein embryos, which have good cryotolerance. Even when housed and fed under the same conditions, Jersey embryos were found to be darker and contain more lipid droplets than Holstein embryos, and this was correlated with lower mitochondrial activity. Differential expression of genes associated with lipid metabolism and differences in lipid composition were found. These results show genetic background can impact embryonic lipid metabolism and storage.
  • Publication
    Papaverine-sensitive phosphodiesterase activity is measured in bovine spermatozoa
    (Wiley, 2016-11-16) Vigneault, Christian; Poulin, Marie-Pier; Bergeron, Annick; Leclerc, Pierre; Richard, François J.; Sullivan, Robert; Hébert, Audrey; Guillemette, Christine; Aragon, Juan Pablo; Laroche, A.; Blondin, Patrick.
    Cyclic adenosine monophosphate (cAMP) plays a crucial role as a signaling molecule for capacitation, motility, and acrosome reaction in mammalian spermatozoa. It is well‐known that cAMP degradation by phosphodiesterase (PDE) enzyme has a major impact on sperm functions. This study was undertaken to characterize cAMP‐PDE activity in bovine spermatozoa. Total cAMP‐PDE activity in cauda epididymal and ejaculated spermatozoa was 543.2 ± 49.5 and 1252.6 ± 86.5 fmoles/min/106 spermatozoa, respectively. Using different family‐specific PDE inhibitors, we showed that in cauda epididymal and ejaculated spermatozoa, the major cAMP‐PDE activity was papaverine‐sensitive (44.5% and 57.5%, respectively, at 400 nm, papaverine is a specific inhibitor of the PDE10 family). These data are supporting the functional presence of PDE10 in bovine spermatozoa and were further confirmed by western blot to be PDE10A. Using immunocytochemistry, we showed immunoreactive signal for PDE10A present on the post‐acrosomal region of the head and on the flagella of ejaculated spermatozoa. Using papaverine, we showed that it promotes tyrosine phosphorylation of sperm proteins, phosphorylation of Erk1 and Erk2, and Ca²+ release from Ca²+ store. These results suggest that PDE10 is functionally present in bovine spermatozoa and is affecting different molecular events involved in capacitation, most probably by cAMP local regulation.
  • Publication
    Cyclic nucleotide phosphodiesterases in human spermatozoa and seminal fluid : presence of an active PDE10A in human spermatozoa
    (Elsevier, 2016-11-09) Leclerc, Pierre; Richard, François J.; Guillemette, Christine; Goupil, Serge; Maréchal, Loïze; Blondin, Patrick.
    Background: Cyclic adenosine monophosphate (cAMP) plays a crucial role as a signaling molecule for sperm functions such as capacitation, motility and acrosome reaction. It is well known that cAMP degradation by phosphodiesterase (PDE) enzyme has a major impact on sperm functions. The present study was undertaken to characterize cAMP-PDE activity in human semen. Methods: cAMP-PDE activity was measured in human sperm and seminal plasma using family specific PDE inhibitors. Three sperm fractionation methods were applied to assess cAMP-PDE activity in spermatozoa. Western blots were used to validate the presence of specific family in sperm and seminal plasma. Results: Using three sperm fractionation methods, we demonstrated that in human sperm, the major cAMP-PDE activity is papaverine-sensitive and thus ascribed to PDE10. In seminal plasma, total cAMP-PDE activity was 1.14 ± 0.39 fmol of cAMP hydrolyzed per minute per μg of protein. Using specific inhibitors, we showed that the major cAMP-PDE activity found in human seminal plasma is ascribed to PDE4 and PDE11. Western blot analysis, immunoprecipitation with a specific monoclonal antibody, and mass spectrometry confirmed the presence of PDE10 in human spermatozoa. Conclusion: This study provides the first demonstration of the presence of functional PDE10 in human spermatozoa and functional PDE4 and PDE11 in human seminal plasma. General significance: Since the contribution of cyclic nucleotides in several sperm functions is well known, the finding that PDE10 is an active enzyme in human spermatozoa is novel and may lead to new insight into fertility.
  • Publication
    Characterization of cAMP-phosphodiesterase activity in bovine seminal plasma
    (American Society of Andrology, 2016-08-27) Bergeron, Annick; Richard, François J.; Sullivan, Robert; Hébert, Audrey; Guillemette, Christine; Aragon, Juan Pablo; Blondin, Patrick.
    The second messenger cyclic adenosine monophosphate (cAMP) has a central role in sperm physiology. Extracellular cAMP can besequentially degraded into 50AMP and adenosine by ecto-phosphodiesterases (ecto-PDE) and ecto-nucleotidases, a phenomenoncalled extracellular cAMP-adenosine pathway. As cAMP-adenosine pathway is involved in sperm capacitation, we hypothesize thatextracellular PDEs are functionally present in seminal plasma. Exclusively measuring cAMP-PDE activity, total activity in bovineseminal plasma was 10.1 1.5 fmoles/min/lg. Using different family-specific PDE inhibitors, we showed that in seminal plasma,the major cAMP-PDE activity was papaverine sensitive (47.5%). These data support the presence of PDE10 in bovine seminal plasmaand was further confirmed by western blot. In epididymal fluid, total cAMP-PDE activity was 48.2 14.8 fmoles/min/lg and weshowed that the major cAMP-PDE activity was 3-isobutyl-methylxanthine insensitive and thus ascribed to PDE8 family. PDE10AmRNAs were found in the testis, epididymis, and seminal vesicles. cAMP-PDE activity is present in bovine seminal plasma and epi-didymal fluid. The results suggest a role for ecto-PDEs present in those fluids in the signaling pathways involved in sperm functions.