scholarly journals Melatonin improves rate of monospermic fertilization and early embryo development in a bovine IVF system

PLoS ONE ◽  
2021 ◽  
Vol 16 (9) ◽  
pp. e0256701
Author(s):  
Juan Carlos Gutiérrez-Añez ◽  
Heiko Henning ◽  
Andrea Lucas-Hahn ◽  
Ulrich Baulain ◽  
Patrick Aldag ◽  
...  
Keyword(s):  
Embryo Development ◽  
Sperm Quality ◽  
Hierarchical Cluster ◽  
Early Embryo ◽  
Developmental Competence ◽  
Control Group ◽  
Computer Assisted ◽  
Sperm Analysis ◽  
Flow Cytometry Data

The developmental competence of male and female gametes is frequently reduced under in vitro conditions, mainly due to oxidative stress during handling. The amino-acid derived hormone melatonin has emerged as a potent non-enzymatic antioxidant in many biological systems. The goal of the present study was to evaluate the effects of melatonin on post-thaw sperm quality, fertilizing ability, and embryo development and competence in vitro after in vitro fertilization. Frozen-thawed bovine spermatozoa were incubated either in the presence of 10−11 M melatonin (MT), or its solvent (ethanol; Sham-Control), or plain Tyrode’s Albumin Lactate Pyruvate medium (TALP, Control). Computer-Assisted Sperm Analysis (CASA) and flow cytometry data after 30 min, 120 min, and 180 min incubation did not reveal any significant effects of melatonin on average motility parameters, sperm subpopulation structure as determined by hierarchical cluster, or on the percentage of viable, acrosome intact sperm, or viable sperm with active mitochondria. Nevertheless, in vitro matured cumulus-oocyte-complexes fertilized with spermatozoa which had been preincubated with 10−11 M melatonin (MT-Sperm) showed higher (P < 0.01) rates of monospermic fertilization, reduced (P < 0.05) polyspermy and enhanced (P < 0.05) embryo development compared to the Control group. Moreover, the relative abundance of MAPK13 in the in vitro-derived blastocysts was greater (P < 0.05) than observed in the Control group. In conclusion, adding melatonin to the sperm-preparation protocol for bovine IVF improved proper fertilization and enhanced embryonic development and competence in vitro.

161 EFFECT OF OVIDUCTAL FLUID ON BOVINE OOCYTE ZONA PELLUCIDA HARDENING AND SPERM BINDING, AND ON EARLY EMBRYO DEVELOPMENT

2016 ◽  
Vol 28 (2) ◽  
pp. 210
Author(s):  
P. Hugon ◽  
J. Lamy ◽  
E. Corbin ◽  
P. Mermillod ◽  
M. Saint-Dizier
Keyword(s):  
Corpus Luteum ◽  
Embryo Development ◽  
Zona Pellucida ◽  
Developmental Stages ◽  
Early Embryo ◽  
Developmental Competence ◽  
Control Group ◽  
Vitro Fertilization ◽  
Oviductal Fluid

This study was designed to evaluate the effects of oviductal fluid at different periovulatory times on oocyte maturation, modification of the zona pellucida (ZP), fertilization and embryo development. Bovine oviducts were collected at a slaughterhouse and classified as preovulatory (pre-ov: 1 pre-ov follicle and a regressing corpus luteum) or post-ovulatory (post-ov: a corpus haemorrhagicum or recent corpus luteum; n = 10 cows/stage). Both oviducts were flushed with 1 mL of sterile TCM-199, and oviductal flushes (OF) were aliquoted and stored at –80°C. Abattoir-derived bovine ovaries were aspirated and cumulus‐oocyte complexes (COC) with at least 3 cumulus layers and homogeneous oocyte cytoplasm were in vitro matured for 22 h in standard maturation medium (control group, n = 319) or in standard medium with 2× concentrated additives supplemented (50% v/v) with pre-ov OF (n = 255) or post-ov OF (n = 248). After in vitro maturation (IVM), subgroups of COC were denuded, and the time of digestion of the ZP by pronase 0.1% (v/v in TCM-199) was determined to evaluate ZP hardening. After IVM, COC were fertilised in vitro for 18–20 h at a final concentration of 1.106 million spermatozoa (spz)/mL. After in vitro fertilization (IVF), COC were denuded, washed twice and cultured for 8 days more under standard conditions. After IVM, IVF, and embryo culture, oocytes/embryos were fixed with ethanol, stained with Hoescht, and examined under fluorescence microscopy for determination of (1) maturation and developmental stages, (2) numbers of fertilised and polyspermic oocytes, and (3) spz bound to the ZP. Percentages were compared between groups by chi-square. Times of ZP digestion were compared by Kruskal‐Wallis test. Numbers of spz bound to the ZP were compared by ANOVA on normalised data followed by Newman-Keuls tests. Data are presented as mean ± SEM. A P < 0.05 was considered significant. Addition of OF during IVM had no effect on maturation rates compared with the control. However, the digestion time of the ZP by pronase was reduced after IVM with pre-ov OF (313 ± 21 s; n = 26) compared with post-ov OF (459 ± 23 s; n = 23) but not with the control (416 ± 30 s; n = 25). After IVF, the number of spermatozoa bound to the ZP was increased after IVM with pre-ov OF (57 ± 5 spz/oocyte; n = 67) and decreased after IVM with post-ov OF (34 ± 3 spz/oocyte; n = 76) compared with the control (42 ± 5 spz/oocyte; n = 60). Addition of OF during IVM had no effect on rates of IVF and polyspermia. However, the rate of development to the blastocyst stage was less after IVM with post-ov OF (10%, n = 97 cleaved oocytes) compared with control (24%, n = 130) and pre-ov OF (29%, n = 101). In conclusion, the OF collected before ovulation decreased the resistance of the ZP to protease digestion and increased its ability to bind spz, whereas it was the opposite for the post-ov OF. Furthermore, the post-ov OF decreased the developmental competence of fertilised oocytes.

Effect of transfection and co-incubation of bovine sperm with exogenous DNA on sperm quality and functional parameters for its use in sperm-mediated gene transfer

Zygote ◽  
2016 ◽  
Vol 25 (1) ◽  
pp. 85-97 ◽  
Author(s):  
María Elena Arias ◽  
Esther Sánchez-Villalba ◽  
Andrea Delgado ◽  
Ricardo Felmer
Keyword(s):  
Gene Transfer ◽  
Sperm Quality ◽  
Computer Assisted ◽  
Exogenous Dna ◽  
Sperm Analysis ◽  
Functional Parameters ◽  
Transgenic Embryos ◽  
Fertilization Capacity ◽  
Motility Parameters

SummarySperm-mediated gene transfer (SMGT) is based on the capacity of sperm to bind exogenous DNA and transfer it into the oocyte during fertilization. In bovines, the progress of this technology has been slow due to the poor reproducibility and efficiency of the production of transgenic embryos. The aim of the present study was to evaluate the effects of different sperm transfection systems on the quality and functional parameters of sperm. Additionally, the ability of sperm to bind and incorporate exogenous DNA was assessed. These analyses were carried out by flow cytometry and confocal fluorescence microscopy, and motility parameters were also evaluated by computer-assisted sperm analysis (CASA). Transfection was carried out using complexes of plasmid DNA with Lipofectamine, SuperFect and TurboFect for 0.5, 1, 2 or 4 h. The results showed that all of the transfection treatments promoted sperm binding and incorporation of exogenous DNA, similar to sperm incorporation of DNA alone, without affecting the viability. Nevertheless, the treatments and incubation times significantly affected the motility parameters, although no effect on the integrity of DNA or the levels of reactive oxygen species (ROS) was observed. Additionally, we observed that transfection using SuperFect and TurboFect negatively affected the acrosome integrity, and TurboFect affected the mitochondrial membrane potential of sperm. In conclusion, we demonstrated binding and incorporation of exogenous DNA by sperm after transfection and confirmed the capacity of sperm to spontaneously incorporate exogenous DNA. These findings will allow the establishment of the most appropriate method [intracytoplasmic sperm injection (ICSI) orin vitrofertilization (IVF)] of generating transgenic embryos via SMGT based on the fertilization capacity of transfected sperm.

The effects of brain-derived neurotrophic factor and metformin on in vitro developmental competence of bovine oocytes

Zygote ◽  
2009 ◽  
Vol 17 (3) ◽  
pp. 187-193 ◽  
Author(s):  
So Gun Hong ◽  
Goo Jang ◽  
Hyun Ju Oh ◽  
Ok Jae Koo ◽  
Jung Eun Park ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Embryo Development ◽  
Neurotrophic Factor ◽  
Brain Derived Neurotrophic Factor ◽  
Early Embryo ◽  
Developmental Competence ◽  
Blastocyst Development ◽  
Cell Stage ◽  
Significant Difference

SummaryBrain-derived neurotrophic factor (BDNF) signalling via tyrosine kinase B receptors may play an important role in ovarian development and function. It has been reported that metformin elevates the activity of Tyrosine kinase receptors and may amplify BDNF signalling. The objective of this study was to investigate the effect of BDNF during in vitro maturation (IVM) and/or in vitro culture (IVC) (Experiment 1), and to evaluate the collaborative effect of BDNF and metformin treatment on the developmental competence of bovine in vitro fertilized (IVF) embryos (Experiment 2). In Experiment 1, BDNF, which was added to our previously established IVM systems, significantly increased the proportions of MII oocytes at both 10 ng/ml (86.7%) and 100 ng/ml (85.4%) compared with the control (64.0%). However, there was no statistically significant difference in blastocyst development between the control or BDNF-supplemented groups. In Experiment 2, in order to investigate the effect of BDNF (10 ng/ml) and/or metformin (10−5 M) per se, TCM-199 without serum and hormones was used as the control IVM medium. The BDNF (48.3%) and BDNF plus metformin (56.5%) significantly enhanced the proportions of MII oocytes compared with the control (34.4%). Although, BDNF or metformin alone had no effect in embryo development, BDNF plus metformin significantly improved early embryo development to the 8–16-cell stage compared with the control (16.5 vs. 5.5%). In conclusion, the combination of BDNF and metformin may have a collaborative effect during the IVM period. These results could further contribute to the establishment of a more efficient bovine in vitro embryo production system.

168 INHIBITION OF BOVINE OOCYTE CUMULUS CELL PROGESTERONE SYNTHESIS DURING IN VITRO MATURATION AFFECTS CHROMOSOME ALIGNMENT AND SPINDLE INTEGRITY IN FERTILIZED EGGS AND EARLY EMBRYOS

2014 ◽  
Vol 26 (1) ◽  
pp. 198
Author(s):  
E. Daly ◽  
A. G. Fahey ◽  
M. M. Herlihy ◽  
T. Fair
Keyword(s):  
Embryo Development ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Early Embryo ◽  
Developmental Competence ◽  
Bovine Oocyte ◽  
Spindle Formation ◽  
Vitro Fertilization ◽  
Time Treatment

We have previously demonstrated the importance of progesterone (P4) synthesis by cumulus cells during oocyte maturation in vitro (IVM) for bovine oocyte acquisition of developmental competence and subsequent embryo development (Aparicio et al. 2011 Biol. Reprod. 84). The aim of this study was to identify key processes that may be deregulated by the inhibition of P4 signalling in the cumulus–oocyte complex (COC) during IVM. To this end, good quality immature COC were placed in IVM medium [TCM-199 supplemented with 10% (vol/vol) FCS and 10 ng mL–1 epidermal growth factor] and cultured at 39°C for 22 h in a humidified atmosphere containing 5% CO2, in the presence or absence of 10 μM trilostane (which blocks P4 synthesis by inhibiting 3 β-hydroxysteroid dehydrogenase; Stegram Pharmaceuticals Ltd., Surrey, UK). Matured COC were washed and placed in 250 μL of fertilization medium (25 mM bicarbonate, 22 mM Na-lactate, 1 mM Na-pyruvate, 6 mg mL–1 fatty acid-free BSA, and 10 mg mL–1 heparin). In vitro fertilization (IVF) was performed with 250 μL of frozen–thawed semen at a final concentration of 1 × 106 spermatozoa mL–1 at 39°C under 5% CO2 during 20 h. Presumptive zygotes were denuded, washed, and transferred to 25-μL culture droplets (SOF + 5% FCS) at 39°C under 5% CO2, 90% of N2, and 5% O2 atmosphere with maximum humidity. Subsets of presumptive fertilized eggs and developing embryos were recovered at 6, 72, 120, and 192 h postinsemination (hpi) and processed for confocal whole-mount immunocytochemistry. The meiotic and mitotic spindles and chromosomes were visualised by immunofluorescent labelling of α-tubulin and 4′,6-diamindino-2-phenylindole (DAPI), respectively, and classified as normal if the chromosomes were correctly aligned or appropriately segregated, or abnormal if lagging chromosomes or abnormal chromosome segregation were observed. Samples were collected from 5 replicates (n = 50 zygotes/embryos per treatment, per timepoint) and a total of 157 spindles were observed. Logistic regression analysis was conducted to determine the probability of abnormal spindle formation. The incidence of spindle abnormality was regressed on time, treatment, and treatment by time. For all time points, there was significant reduction in the odds of abnormal spindle formation in control samples versus trilostane-treated samples (P < 0.001). In conclusion, our data imply a role for P4 signalling in maintaining spindle integrity during oocyte meiotic maturation and progression through the initial mitotic divisions of early embryo development in cattle.

Morphology, developmental stages and quality parameters of in vitro-produced equine embryos

2019 ◽  
Vol 31 (12) ◽  
pp. 1758 ◽  
Author(s):  
Elaine M. Carnevale ◽  
Elizabeth S. Metcalf
Keyword(s):  
Embryo Development ◽  
Developmental Stages ◽  
Inner Cell Mass ◽  
Early Stage ◽  
Cell Mass ◽  
Quality Parameters ◽  
Early Embryo ◽  
Developmental Competence ◽  
Limited Information

Intracytoplasmic sperm injection (ICSI) is used to produce equine embryos invitro. The speed of embryo development invitro is roughly equivalent to what has been described for embryos produced invivo. Morphological evaluations of ICSI-produced embryos are complicated by the presence of debris and the dark nature of equine embryo cytoplasm. Morulas and early blastocysts produced invitro appear similar to those produced invivo. However, with expansion of the blastocyst, distinct differences are observed compared with uterine embryos. In culture, embryos do not undergo full expansion and thinning of the zona pellucida (ZP) or capsule formation. Cells of the inner cell mass (ICM) are dispersed, in contrast with the differentiated trophoblast and ICM observed in embryos collected from uteri. As blastocysts expand invitro, embryo cells often escape the ZP as organised or disorganised extrusions of cells, probably through the hole incurred during ICSI. Quality assessment of invitro-produced early stage equine embryos is in its infancy, because limited information is available regarding the relationship between morphology and developmental competence. Early embryo development invivo is reviewed in this paper, with comparisons made to embryo development invitro and clinical assessments from a laboratory performing commercial ICSI for &gt;15 years.

scholarly journals mRNA Expression and Role of PPARγ and PPARδ in Bovine Preimplantation Embryos Depending on the Quality and Developmental Stage

Animals ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 2358
Author(s):  
Katarzyna Suwik ◽  
Emilia Sinderewicz ◽  
Dorota Boruszewska ◽  
Ilona Kowalczyk-Zięba ◽  
Joanna Staszkiewicz-Chodor ◽  
...  
Keyword(s):  
Early Development ◽  
Embryo Development ◽  
Embryo Quality ◽  
Early Embryo ◽  
Developmental Competence ◽  
Preimplantation Embryos ◽  
Peroxisome Proliferator ◽  
Early Embryo Development ◽  
Quality Markers

Peroxisome proliferator-activated receptors (PPARs), a nuclear receptors for prostacyclin (PGI2) have been recognized as being essential for early embryo development. The objectives of the present study were to determine if the bovine early- and late-cleaved embryos in different stages of early development express PPARγ and PPARδ. Since embryo developmental competence depends on numerous biological factors, we evaluated if the expression of PPARγ and PPARδ correlate with selected embryo quality markers (SOX2, OCT4, PLAC8, IGF1R) in the in vitro produced embryos at different stages of their development. Developmental rates and embryo quality for early- and late-cleaved embryos were provided according to International Embryo Transfer Society (IETS; developmental stages: 2-, 4-, 16-cell embryo, morula, blastocyst (1—early, 2—developing, 3—expanded, 4—hatched); quality stages: A—high quality, B—moderate quality, C—low quality). We found that bovine embryos expressed mRNA of PPARδ and PPARγ at all stages of early development, independently of their quality. In addition, the expression of PPARδ and PPARγ correlated with the expression of quality markers in bovine blastocysts. Positive correlations were stronger and more frequent in the group of early-cleaved embryos, whereas the negative correlations were typical for the group of late-cleaved embryos. Obtained results and available literature reports may indicate the participation of PGI2, via PPARδ and PPARγ, in the processes related to the early embryo development, through the participation of this factor in the modulation of blastocyst hatching, implantation, and post-implantation development.

329 SUPPRESSION OF BOVINE OOCYTE IVM BY SERUM AND FSH DEPRIVATION OR BY α-AMANITIN: EFFECT ON FERTILIZATION AND EMBRYO DEVELOPMENT

2006 ◽  
Vol 18 (2) ◽  
pp. 272
Author(s):  
K. Kananen-Anttila ◽  
M. Eronen ◽  
J. Matilainen ◽  
M. Kallio ◽  
J. Peippo ◽  
...  
Keyword(s):  
Embryo Development ◽  
Cumulus Cell ◽  
Developmental Competence ◽  
Control Group ◽  
Blastocyst Development ◽  
Nuclear Maturation ◽  
Embryo Cleavage ◽  
Bovine Oocytes ◽  
Sperm Aster

We have studied the effect of suppressed IVM on the developmental competence of bovine oocytes, aiming at elucidating the importance of cytoplasmic maturation in fertilization and embryo development. Six replicates of abattoir-derived oocytes were randomly divided into three IVM groups. Control (n = 950): TCM-199 with glutamax-I (Gibco, Grand Island, NY, USA), 0.25 mM Na-pyruvate, 100 IU mL−1 penicillin and 100 μg mL−1 streptomycin, 50 ng mL−1 FSH, and 10% fetal bovine serum (FBS) (Gibco); Serum+FSH-free (n = 944): same as control but without FSH and FBS; α-amanitin (n = 977): same as control but with 10 μg mL−1 α-amanitin. Nuclear maturation of oocytes was studied 24 h after the onset of IVM, the formation of sperm aster structure 10 hours post-insemination (hpi) and the formation of pronuclei 20 hpi. Sperm aster was visualized with β-tubulin antibody (modified from Navara et al. 1999 Dev. Biol. 162, 29–40). Presumptive zygotes were cultured until Day 7 in modified SOFaaci + 4 mg mL−1 fatty acid-free BSA in 5% O2. Cumulus cell expansion was seen only in the control group. The results of nuclear maturation, fertilization, and embryo development are summarized in Table 1. Serum and FSH deprivation did not have a statistically significant effect on the parameters studied (vs. control). α-amanitin exposure during IVM reduced nuclear maturation, fertilization, and Day 3 embryo cleavage vs. control, and resulted in total blockage of Day 7 blastocyst development. The treatment groups had significantly smaller mean diameters of male pronuclei (control: 14 ± 0.6 μ­m; serum+FSH-free: 12 ± 0.5 μ­m, P < 0.05; α-amanitin: 10 ± 0.6 μ­m, P < 0.001) and sperm asters (control: 86 ± 4 μ­m; serum+FSH-free: 82 ± 4 μ­m, P < 0.01; α-amanitin: 49 ± 7 μm, P < 0.001) (nonparametric Kruskall Wallis and Mann-Whitney U tests) vs. control group. Despite reduction in pronucleus and sperm aster diameter, serum and FSH deprivation during IVM did not affect in vitro developmental competence of bovine oocytes, suggesting a need for re-evaluation of the components of IVM. α-Amanitin exposure in IVM disturbed nuclear maturation, fertilization, and embryo development, indicating the essence of early transcription. Table 1. Average percentages ± (n) for nuclear maturation, fertilization (min two pronuclei), embryo cleavage, and blastocyst development

133 EFFECTS OF ACTIVINA ON mRNA EXPRESSION IN IN VITRO FERTILIZED BOVINE EMBRYOS CULTURED FROM CHEMICALLY DEFINED TWO-STEP CULTURE MEDIUM

2008 ◽  
Vol 20 (1) ◽  
pp. 147
Author(s):  
J. E. Park ◽  
G. Jang ◽  
H. J. Oh ◽  
S. G. Hong ◽  
I. S. Yang ◽  
...  
Keyword(s):  
Embryo Development ◽  
Culture Medium ◽  
Early Stage ◽  
Activin A ◽  
Developmental Competence ◽  
Control Group ◽  
Hatching Rate ◽  
Bovine Embryos ◽  
Defined Culture

During the preimplantation stage, embryo development occurs in a maternal environment within the oviducts and uterine horns. It has been speculated that both the embryo itself and the maternal reproductive tract provide paracrine factors that influence embryo development (Jones et al. 2006 Reproduction 132(5), 799–810). Activins are known for FSH releasers, and several previous studies have reported that activin subunits and activin receptors mRNA were expressed in oocytes, zygotes, and oviduct (Yoshioka et al. 1998 Reprod. Fertil. Dev. 10(3), 293–298; Gandolfi et al. 1995 Mol. Reprod. Dev. 40(3), 286–291). The purposes of the present study were Experiment 1) to evaluate the effects of activin A on developmental competence of bovine embryos derived from two-step defined culture medium (Lim et al. 2007 Theriogenology 67(2), 293–302) and Experiment 2) to analyze the effects of activin A on transcriptional level of the genes in IVF embryos. Cumulus–oocyte complexs were harvested from ovaries obtained from a local slaughter house, matured, and fertilized in vitro. In vitro fertilized zygotes cultured in media supplemented with activin A in the early stage at the concentrations of 0, 10, or 100 ng mL–1 or in the later stage medium at the concentrations of 0, 10, or 100 ng mL–1. Data were analyzed using the Statistical Analysis System (SAS) program. In Exp. 1, although the development competence of embryos that cultured with activin A in the early stage medium was not significantly different, development to blastocysts on day 8 in the later stage medium with 100 ng mL–1 activin A was significantly higher than the control group [22.4% (54/264) v. 34.7% (76/233); P < 0.05]. Hatching rate of blastocyst on day 8 was significantly higher in the presence of 100 ng mL–1 activin A in the later stage culture medium compared with the control group [9.3% (5/54) v. 22.4% (17/76); P < 0.05]. In Exp. 2, the relative expression of 3 genes (Na/KATPase, E-cad, Glut-1) related to blastocyst hatching and implantation was analyzed. The relative abundance (ratio to GAPDH mRNA) of gene transcripts in blastocysts was measured by conventional semi-quantitative reverse transcription-polymerase chain reaction. The expression level of the Na/K ATPase, E-cad, and Glut-1 gene were higher in the presence of activin A in the culture medium compared with the control group. In conclusion, this study suggests that activin A during the later stage of in vitro bovine embryo development can enhance the developmental competence of preimplantation embryos, increase the hatching rate, and affect expression level of genes related to hatching and implantation in defined culture medium. This study was financially supported by KOSEF (grant ? M10625030005-07N250300510) and the Korean MOE, through the BK21 program for Veterinary Science.

Calcium-sensing receptor (CASR) is involved in porcine in vitro fertilisation and early embryo development

2018 ◽  
Vol 30 (2) ◽  
pp. 391 ◽  
Author(s):  
C. Liu ◽  
Y. Liu ◽  
K. Larsen ◽  
Y. P. Hou ◽  
H. Callesen
Keyword(s):  
Embryo Development ◽  
Early Embryo ◽  
Extracellular Calcium ◽  
In Vitro Fertilisation ◽  
Control Group ◽  
Blastocyst Formation ◽  
Calcium Sensing Receptor ◽  
Early Embryo Development ◽  
Calcium Sensing

It has been demonstrated that extracellular calcium is necessary in fertilisation and embryo development but the mechanism is still not well understood. The present study mainly focussed on the extracellular calcium effector called the calcium-sensing receptor (CASR) and examined its expression in porcine gametes and embryos and its function during fertilisation and early embryo development. By using reverse transcription polymerase chain reaction, CASR was found to be expressed in porcine oocytes, spermatozoa and embryos at different developmental stages. Functionally, medium supplementation with a CASR agonist or an antagonist during in vitro fertilisation (IVF) and in vitro culture (IVC) was tested. During fertilisation, the presence of a CASR agonist increased sperm penetration rate and decreased polyspermy rate leading to an increased normal fertilisation rate. During embryo development, for the IVF embryos, agonist treatment during IVC significantly increased cleavage rate and blastocyst formation rate compared with the control group. Furthermore, parthenogenetically activated embryos showed similar results with lower cleavage and blastocyst formation rates in the antagonist group than in the other groups. It was concluded that CASR, as the effector of extracellular calcium, modulates porcine fertilisation and early embryo development.

Sonic hedgehog promotes porcine oocyte maturation and early embryo development

2009 ◽  
Vol 21 (6) ◽  
pp. 805 ◽  
Author(s):  
Ngoc Tan Nguyen ◽  
David Pei-Cheng Lin ◽  
Shih-Ying Yen ◽  
Jung-Kai Tseng ◽  
Jui-Fen Chuang ◽  
...  
Keyword(s):  
Embryo Development ◽  
Sonic Hedgehog ◽  
Oocyte Maturation ◽  
Calcium Release ◽  
Cyclin B1 ◽  
Early Embryo ◽  
Control Group ◽  
Early Embryo Development ◽  
Porcine Oocyte

In the present study, we investigated the effects of the Sonic hedgehog (Shh) protein on porcine oocyte maturation and early embryo development. Immunohistochemistry showed activation of Shh signalling in cumulus–oocyte complexes (COCs), as reflected by Patched (Ptc), Smoothened (Smo) and Gli1 expression in oocytes, cumulus cells and granulosa cells, particularly those of small follicles (<2 mm in diameter). Western blot analysis showed Smo expression in COCs and in denuded oocytes derived from small and medium (3–7 mm)-sized follicles. Small follicles contained the highest concentration of Shh in follicular fluid compared with medium-sized and large (>7 mm in diameter) follicles. Supplementation with Shh (0.5 or 1 μg mL–1) enhanced oocyte maturation compared with the control group (92.4% and 90.4% v. 81.9%, respectively; P < 0.05). This effect was reversed by the simultaneous addition of cyclopamine (1–2 μm), an Shh inhibitor. Similar to intact COCs, denuded COCs showed enhanced maturation following Shh supplementation. Furthermore, cyclin B1 content, extracellular signal-regulated kinase 1/2 phosphorylation, intracellular calcium release, blastocyst rate and total cell numbers were greater (P < 0.05) in oocytes matured in the presence of 0.5 and 1 μg mL–1 Shh compared with control oocytes. The findings of the present study provide the first evidence that the Shh signalling pathway is active, or at least partially activated, in the porcine ovary and is likely to promote oocyte cytoplasmic and nuclear maturation, as well as subsequent in vitro development, although the underlying mechanisms remain to be elucidated.

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