356 MODULATION OF PORCINE NUCLEAR MATURATION STATUS USING A SPECIFIC PHOSPHODIESTERASE INHIBITOR

2010 ◽  
Vol 22 (1) ◽  
pp. 334
Author(s):  
M. L. Sutton-McDowall ◽  
R. B. Gilchrist ◽  
J. G. Thompson
Keyword(s):  
Gap Junction ◽  
Time Course ◽  
Phosphodiesterase Inhibitor ◽  
Adenosine Monophosphate ◽  
Developmental Competence ◽  
Nuclear Maturation ◽  
Gap Junction Communication ◽  
Defined Culture ◽  
Type 3

Compared to other livestock species, IVM porcine COCs have poor developmental competence.Thisismost likely due to poor cytoplasmic maturation and asynchronous nuclear maturation. The resumption of nuclear maturation is largely regulated by cyclic adenosine monophosphate(cAMP), with increasing intra-oocyte levels prolonging cumulus-oocyte gap junction communication and delaying meiotic resumption. Modulation of cAMP levels using phosphodiesterase (PDE) inhibitors during bovine IVM significantly improves developmental competence (Thomas et al. 2004 Biol. Reprod. 71, 1142-1149). Hence, the aim of this study was to determine the effect of a type 3 PDE inhibitor (cilostamide, CIL) supplementation during porcine IVM on nuclear maturation, using a defined culture system. COCs derived from follicles of prepubertal gilts were cultured in groups of 10 in 100μL IVM medium (VitroMat, IVF Vet Solutions, Adelaide, Australia) +100 m IU mL-1 rhFSH +4 mg mL-1 BSA and nuclear maturation was assessed using orcein dye. Exp. 1: IVM medium was supplemented with 0, 5, 10, and 20 μM CIL or 0, 0.01, 0.1, 1, and 10 μM CIL and nuclear maturation was determined at 24 h and 44 h. Exp. 2: COCs were cultured in ± 0.1 μM CIL and nuclear maturation was assessed at 24 h, 44 h, 48 h, 52 h, and 56 h. Proportions of COCs at each stage of nuclear maturation were arcsine transformed and differences determined using a general linear model and Bonferroni post hoc test. Four replicates per experiment were performed with 20 COCs used for each treatment and/or time point. Results for Exp. 1 revealed no differences in the rate of nuclear maturation after 24 h of culture. After 44 h, 77% of COCs incubated in the absence of CIL were at metaphase II (MII) compared to 35-45% MII when cultured in the presence of 1, 5, 10, or 20 μM CIL (P < 0.001). Furthermore, CIL supplementation resulted in approximately half the COCs arresting at germinal vesicle stage (GV) after 44 h of culture. While there were no significant differences in the MII rates of COCs cultured in 0, 0.01, and 0.1 μM CIL, significantly more COCs were at metaphase I (MI) at 44 h compared to control COCs (0 μM CIL, P < 0.05). The time course experiment (Exp 2) demonstrated that nuclear maturation was delayed by 12 h with 0.1 μM CIL, compared to the absence of CIL, with comparable MII rates being achieved at 56 h for 0.1 μM CIL (72%) and 44 h for controls (0 μM CIL = 73%). These results demonstrate that porcine oocyte maturation can be induced in vitro by FSH in the presence of a low dose of type 3 PDE inhibitor resulting in a delay in the resumption and completion of nuclear maturation. Further investigations are underway to determine if CIL treatment prolongs gap junction communication and improves the developmental competence of porcine oocytes. This work was supported by the National Institutes of Health (USA) and National Health and Medical Research Council (Australia).

176. INHIBITION OF PORCINE OOCYTE NUCLEAR MATURATION IN VITRO USING A PHOSPHODIESTERASE INHIBITOR AND AN ADENYLATE CYCLASE ACTIVATOR

2010 ◽  
Vol 22 (9) ◽  
pp. 94
Author(s):  
M. Bertoldo ◽  
T. Sellens ◽  
C. G. Grupen
Keyword(s):  
Adenylate Cyclase ◽  
Phosphodiesterase Inhibitor ◽  
Phosphodiesterase Inhibitors ◽  
Extended Period ◽  
Nuclear Maturation ◽  
Porcine Oocyte ◽  
Defined Culture ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

Asynchronous nuclear and cytoplasmic maturation is thought to contribute to poor embryo production in vitro. Nuclear arrest is mediated by cAMP and can be maintained within the oocyte using non-specific phosphodiesterase inhibitors (3-isobutyl-1-methylxanthine ; IBMX) and the adenylate cyclase activator forskolin (FSK) (1). The aim of this study was to investigate the effect of IBMX and FSK supplementation on porcine oocyte nuclear maturation during COC recovery and IVM using a defined culture system. In all experiments, cAMP modulators were added to Hepes-buffered media held in collection tubes. COCs recovered from 3–5 mm diameter follicles of prepubertal ovaries were cultured in basic maturation media in the absence of FSH. Nuclear maturation was assessed using orcein dye. In Experiment 1, IVM media was supplemented with 0, 50 or 500 µM IBMX. In Experiment 2, IVM media was supplemented with 0, 5, 10, 50 and 100µM FSK. In Experiment 3, IVM medium was supplemented with combinations of IBMX and FSK to give the treatments; control, 50IBMX/50FSK, 50IBMX/100FSK, 500IBMX/50FSK and 500IBMX/100FSK. Nuclear maturation was assessed at 0, 2, 4 and 18 h after the onset of IVM. At 18 h of culture, there were no differences in the proportion of oocytes supplemented with 0, 50 or 500 µM IBMX reaching MII. Incubation with 10, 50 or 100 µM FSK resulted in 8-16% of oocytes at MII at 18 h compared to the other groups (25–29%; P < 0.001). The combinations of IBMX and FSK resulted in greater proportions (86–98%) of oocytes remaining at the GV stage at 18 h compared to the control (16%; P < 0.001). There were no differences in the proportion of oocytes remaining at the GV stage at the earlier time points (P > 0.05). The results demonstrate that these cAMP modulators, in combination, are highly effective in maintaining porcine oocyte meiotic arrest in vitro for an extended period. (1) Albuz FK et al., Proceedings of the 25th Annual Meeting of ESHRE, Amsterdam, The Netherlands, 2009.

Insulin influences developmental competence of bovine oocytes cultured in α-MEM plus follicle-simulating hormone

Zygote ◽  
2014 ◽  
Vol 23 (4) ◽  
pp. 563-572 ◽  
Author(s):  
Gustavo Bruno Mota ◽  
Ingrid Oliveira e Silva ◽  
Danielle Kaiser de Souza ◽  
Flavia Tuany ◽  
Michele Munk Pereira ◽  
...  
Keyword(s):  
Basal Medium ◽  
Developmental Competence ◽  
Cleavage Rate ◽  
Serum Free ◽  
Oocyte Competence ◽  
Nuclear Maturation ◽  
Blastocyst Rate ◽  
Defined Culture ◽  
Bovine Oocytes

SummaryThe aim of this study was to evaluate the dose–response effect of insulin, plus follicle-simulating hormone (FSH) at a fixed concentration, in a serum-free defined culture medium (DCM) on the in vitro maturation of bovine cumulus–oocyte complexes (COCs). For oocyte nuclear maturation, the expression levels of GDF9, GLUT1, PRDX1 and HSP70.1 transcripts related to oocyte and embryo developmental competence were analysed. For in vitro maturation (IVM), cumulus–oocyte complexes from slaughterhouse ovaries were distributed into four groups based on insulin concentration added to serum-free DCM, which was composed of alpha minimum essential medium (α-MEM), as basal medium: (1) DCM control: 0 ng/ml; (2) DCM1: 1 ng/ml; (3) DCM10: 10 ng/ml; and (4) DCM100: 100 ng/ml. After IVM, the nuclear status of a sample of oocytes was analysed and the other oocytes were submitted for in vitro fertilization (IVF) and in vitro culture (IVC). Different concentrations of insulin did not affect significantly the nuclear maturation and cleavage rate (72 h post-insemination) across all groups. Blastocyst rate (192 h post-insemination) did not differ in DCM control (24.3%), DCM1 (27.0%) and DCM10 (26.3%) groups, but the DCM100 (36.1%) group showed a greater blastocyst rate (P < 0.05) than the DCM control. Insulin concentrations of 1, 10, or 100 ng/ml decreased the relative levels of GDF9 and HSP70-1 transcripts in oocytes at the end of IVM (P < 0.05). The transcripts levels of PRDX1 decreased (P < 0.05) only when 10 or 100 ng/ml insulin was added to the DCM medium. No difference in levels of GLUT1 transcripts (P > 0.05) was observed at the different insulin concentrations. The results indicated that insulin added to DCM influenced levels of transcripts related to cellular stress (HSP70-1 and PRDX1) and oocyte competence (GDF9) in bovine oocytes and at higher concentrations enhanced blastocyst production.

199 PREMATURATION OF BOVINE OOCYTES WITH BUTYROLACTONE I AND/OR CILOSTAMIDE: EFFECTS ON MEIOSIS PROGRESSION, CYTOPLASMIC MATURATION AND GENE EXPRESSION

2012 ◽  
Vol 24 (1) ◽  
pp. 212
Author(s):  
G. Z. Mingoti ◽  
F. Filion ◽  
P. Vincent ◽  
L. C. Smith
Keyword(s):  
Cell Communication ◽  
Phosphodiesterase Inhibitor ◽  
Developmental Competence ◽  
Mitochondrial Activity ◽  
Additional Time ◽  
Chi Square ◽  
Developmental Potential ◽  
Nuclear Maturation ◽  
Cytoplasmic Maturation

Meiotic block during a prematuration culture (pre-IVM) before in vitro maturation (IVM) is suggested as a way to provide additional time to synchronize oocyte-somatic cell communication, leading to improved cytoplasmic maturation and nuclear meiotic competence and the acquisition of critical cellular functions necessary for developmental competence. This study was designed to evaluate the effects of the inhibitors butyrolactone I (Bl-I) and cilostamide on oocyte nuclear and cytoplasmic maturation. For that, 2 well-established methods of pre-IVM and IVM for cilostamide (Albuz et al. 2010 Hum. Reprod. 25, 2999–3011) or Bl-I (Hashimoto et al. 2002 Biol Reprod. 66, 1696–1701) were used. Abattoir-collected oocytes were IVM in maturation medium (MM: mSOF with 0.8% BSA and hormones) for 24 h, without a previous pre-IVM culture (control). Cilostamide-group oocytes were treated for the first 2 h in vitro (pre-IVM) with 100 μM of the adenylate cyclase activator forskolin and 500 μM IBMX (nonspecific phosphodiesterase inhibitor) and then oocytes underwent extended IVM for 30 h in the presence of 20 μM cilostamide (type 3 phosphodiesterase inhibitor; pre-IVM 2 h + IVM 30 h). The Bl-I-group oocytes were pre-IVM for 24 h with 100 μM Bl-I diluted in TCM-199 supplemented with 0.2 mM pyruvate and then were IVM in MM for 20 h (pre-IVM 24 h + IVM 20 h). The Bl-I + Cilost group was a combination of both procedures: oocytes were first pre-IVM with Bl-I for 24 h and then cultured as described for the cilostamide group (pre-IVM 24 h + IVM 30 h). Cultures were carried out at 38.5°C in 5% CO2 in humidified air. After IVM, oocytes were stained with 500 nM mitotracker red to assess the mitochondrial membrane potential (Δψm) and with 10 μg mL–1 of Hoescht 33342 to evaluate the nuclear maturation (n = 207). Images were captured by an Olympus confocal microscope and analyzed with Fluoview software. The TUNEL assay was used to detect oocyte DNA fragmentation (n = 74). Relative amounts of mRNA for apoptotic-related genes were quantified after IVM in individual oocytes using real-time PCR (Kameyama et al. 2007 Reproduction 133, 423–32). Means were compared by ANOVA and Tukey's test or by chi-square (P ≤ 0.05). The percentage of oocytes reaching metaphase II after IVM did not differ among groups (73.8 to 90.4%; P ≥ 0.05), indicating that in vitro meiotic resumption was normal. The Δψm, expressed in arbitrary units of fluorescence, was 1.0 ± 0.1a (control), 2.7 ± 0.4b (Bl-I), 3.2 ± 0.5b (Cilost) and 2.1 ± 0.3ab (Bl-I + Cilost). The percentage of TUNEL-positive oocytes (18.8–41.3%) did not differ among groups (P ≥ 0.05). The relative abundance of BAX (1.0 ± 0.4 to 2.3 ± 0.4) and BCL-XL (1.0 ± 0.3 to 0.3 ± 0.1) transcripts was unaffected by pre-IVM and IVM (P ≥ 0.05). In conclusion, except for an increase in mitochondrial activity, pre-IVM with cilostamide and/or Bl-I did not affect cytoplasmic and nuclear oocyte maturation. However, oocyte developmental potential needs to be better evaluated in a future study through assessment of embryonic development. We acknowledge FAPESP and NSERC.

Early germinal vesicle breakdown is a predictor of high preimplantation developmental competent oocytes in mice

Zygote ◽  
2016 ◽  
Vol 25 (1) ◽  
pp. 41-48 ◽  
Author(s):  
Shogo Higaki ◽  
Masao Kishi ◽  
Keisuke Koyama ◽  
Masashi Nagano ◽  
Seiji Katagiri ◽  
...  
Keyword(s):  
Time Course ◽  
Evaluation Method ◽  
Germinal Vesicle ◽  
Oocyte Quality ◽  
Developmental Competence ◽  
Germinal Vesicle Breakdown ◽  
Nuclear Maturation ◽  
Non Invasive ◽  
Germinal Vesicle Break Down

SummaryThe preselection of highly developmentally competent oocytes for in vitro maturation (IVM) is crucial for improving assisted reproductive technology. Although several intrinsic markers of oocyte quality are known to be closely related to the onset of nuclear maturation (germinal vesicle break down, GVBD), a direct comparison between GVBD timing and oocyte quality has never been reported. In this study, we established a non-invasive oocyte evaluation method based on GVBD timing for preselecting more developmental competent oocytes in mice. Because the O2 concentration during IVM may affect the nuclear kinetics, all experiments were performed under two distinct O2 concentrations: 20% and 5% O2. First, we determined the time course of changes in nuclear maturation and preimplantation developmental competence of in vitro-matured oocytes to estimate GVBD timing in high developmental competent oocytes. Two-thirds of oocytes that underwent GVBD in early IVM seemed to mainly contribute to the blastocyst yield. To confirm this result, we compared the preimplantation developmental competence of the early and late GVBD oocytes. Cleavage and blastocyst formation rates of early GVBD oocytes (80.2% and 52.7% under 20% O2, respectively, and 67.6% and 47.3% under 5% O2, respectively) were almost double those of late GVBD oocytes (44.8% and 26.0% under 20% O2, respectively, and 40.4% and 17.9% under 5% O2, respectively). With no observable alterations by checking the timing of GVBD in preimplantation developmental competence, oocyte evaluation based on GVBD timing can be used as an efficient and non-invasive preselection method for high developmental competent oocytes.

scholarly journals Oocyte Competence Biomarkers Associated With Oocyte Maturation: A Review

2021 ◽  
Vol 9 ◽  
Author(s):  
Batara Sirait ◽  
Budi Wiweko ◽  
Ahmad Aulia Jusuf ◽  
Dein Iftitah ◽  
R. Muharam
Keyword(s):  
Oocyte Maturation ◽  
Blastocyst Stage ◽  
Current Approach ◽  
Developmental Competence ◽  
Matrix Remodeling ◽  
Oocyte Competence ◽  
Nuclear Maturation ◽  
Oocyte Developmental Competence ◽  
Cumulus Oocyte Complex

Oocyte developmental competence is one of the determining factors that influence the outcomes of an IVF cycle regarding the ability of a female gamete to reach maturation, be fertilized, and uphold an embryonic development up until the blastocyst stage. The current approach of assessing the competency of an oocyte is confined to an ambiguous and subjective oocyte morphological evaluation. Over the years, a myriad of biomarkers in the cumulus-oocyte-complex has been identified that could potentially function as molecular predictors for IVF program prognosis. This review aims to describe the predictive significance of several cumulus-oocyte complex (COC) biomarkers in evaluating oocyte developmental competence. A total of eight acclaimed cumulus biomarkers are examined in the study. RT-PCR and microarray analysis were extensively used to assess the significance of these biomarkers in foreseeing oocyte developmental competence. Notably, these biomarkers regulate vital processes associated with oocyte maturation and were found to be differentially expressed in COC encapsulating oocytes of different maturity. The biomarkers were reviewed according to the respective oocyte maturation events namely: nuclear maturation, apoptosis, and extracellular matrix remodeling, and steroid metabolism. Although substantial in vitro evidence was presented to justify the potential use of cumulus biomarkers in predicting oocyte competency and IVF outcomes, the feasibility of assessing these biomarkers as an add-on prognostic procedure in IVF is still restricted due to study challenges.

Reversible effects of phenothiazines on frog gastric stimulation

1984 ◽  
Vol 247 (4) ◽  
pp. G366-G376
Author(s):  
N. Raphael ◽  
E. B. Ekblad ◽  
T. E. Machen
Keyword(s):  
Adenylate Cyclase ◽  
Time Course ◽  
Phosphodiesterase Inhibitor ◽  
Secretory Activity ◽  
Gastric Stimulation ◽  
Camp Content ◽  
Oxyntic Cell ◽  
Camp Generation ◽  
Stimulus Secretion Coupling

The calmodulin inhibitors trifluoperazine (TFP), chlorpromazine (CPZ), and promethazine (PZ) were tested for effects on stimulus-secretion coupling in in vitro bullfrog gastric mucosa. When added to histamine-stimulated tissues, the drugs caused H+ secretion to decrease and transepithelial resistance to increase over a 2-h time course. The potency sequence was TFP (IC50 = 40 microM) greater than CPZ (IC50 = 72 microM) congruent to PZ (IC50 = 72 microM). Anesthetics and other phenothiazines with weak anticalmodulin activity had no effect on secretory parameters. In the presence of histamine, further addition of isobutylmethylxanthine (IBMX, a phosphodiesterase inhibitor) plus dibutyryl cAMP (DBcAMP), IBMX alone, or forskolin (a specific activator of adenylate cyclase) to phenothiazine-inhibited tissues caused full resumption of secretory activity. If TFP (50 microM) was added before stimulation with histamine, the normal increases in tissue cAMP content (which occurs primarily in oxyntic cells), oxyntic cell apical membrane elaboration (morphometric analysis of electron micrographs), and H+ secretion were all blocked. Subsequent addition of IBMX or IBMX plus DBcAMP completely reversed the TFP effect. These results indicate that the histamine-sensitive adenylate cyclase may be the site of TFP inhibition and Ca2+-calmodulin regulation; since these drugs inhibited stimulation by DBcAMP plus IBMX, they may also be exerting additional effects distal to cAMP generation.

scholarly journals Factors affecting developmental competence of equine oocytes after intracytoplasmic sperm injection

Reproduction ◽  
2004 ◽  
Vol 127 (2) ◽  
pp. 187-194 ◽  
Author(s):  
Y H Choi ◽  
L B Love ◽  
D D Varner ◽  
K Hinrichs
Keyword(s):  
Embryo Culture ◽  
Intracytoplasmic Sperm Injection ◽  
Room Temperature ◽  
Culture Media ◽  
Developmental Competence ◽  
Glucose Medium ◽  
Factors Affecting ◽  
Nuclear Maturation ◽  
High Glucose Medium

This study was conducted to evaluate the effect of initial cumulus morphology (expanded or compact) and duration of in vitro maturation (24, 30 or 42 h) on the developmental competence of equine oocytes after intracytoplasmic sperm injection (ICSI). The effect of manipulation temperature (room temperature vs 37 °C) at the time of ICSI and concentration of glucose (0.55 vs 5.5 mM) during embryo culture was also investigated. The nuclear maturation rates of expanded (Ex) oocytes were significantly (P < 0.001) higher than those of compact (Cp) oocytes at all maturation times (61–72 vs 23–25% respectively). Forty-eight hours after ICSI of mature Ex oocytes, the rate of cleavage with normal nuclei was significantly (P < 0.05) higher for oocytes matured for 24 h than for those matured for 30 or 42 h (73 vs 57–59% respectively). For Cp oocytes, the morphologic cleavage rates for oocytes matured for 30 h were significantly higher (P < 0.05) than for those matured for 24 or 42 h (86 vs 55–61% respectively). The overall proportion of embryos having more than four normal nuclei at 48 h culture was significantly higher (P < 0.05) for Cp than for Ex oocytes. Manipulation temperature did not affect development of embryos from Ex or Cp oocytes at 96 h after ICSI. Culture in high-glucose medium significantly increased morphologic cleavage of Cp, but not Ex, oocytes (P < 0.05). Embryos from Cp oocytes had a significantly higher average nucleus number after 96-h culture than did embryos from Ex oocytes. These data indicate that developmental competence differs between Ex and Cp equine oocytes, and is differentially affected by the duration of maturation and by composition of embryo culture media.

scholarly journals Effect of Hyaluronan on Developmental Competence and Quality of Oocytes and Obtained Blastocysts fromIn VitroMaturation of Bovine Oocytes

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Jolanta Opiela ◽  
Joanna Romanek ◽  
Daniel Lipiński ◽  
Zdzisław Smorąg
Keyword(s):  
Dna Fragmentation ◽  
Oocyte Maturation ◽  
Developmental Competence ◽  
Meiotic Maturation ◽  
Nuclear Maturation ◽  
Significant Difference ◽  
The Mean ◽  
Bovine Oocytes

The objective of the present study was to evaluate the effect of hyaluronan (HA) during IVM on meiotic maturation, embryonic development, and the quality of oocytes, granulosa cells (GC), and obtained blastocysts. COCs were maturedin vitroin control medium and medium with additional 0.035% or 0.07% of exogenous HA. The meiotic maturity did not differ between the analysed groups. The best rate and the highest quality of obtained blastocysts were observed when 0.07% HA was used. A highly significant difference (P<0.001) was noted in the mean number of apoptotic nuclei per blastocyst and in the DCI between the 0.07% HA and the control blastocysts (P<0.01). Our results suggest that addition of 0.035% HA and 0.07% HA to oocyte maturation media does not affect oocyte nuclear maturation and DNA fragmentation. However, the addition of 0.07% HA during IVM decreases the level of blastocysts DNA fragmentation. Finally, our results suggest that it may be risky to increase the HA concentration during IVM above 0.07% as we found significantly higherBaxmRNA expression levels in GC cultured with 0.07% HA. The final concentration of HA being supplemented to oocyte maturation media is critical for the success of the IVP procedure.

Effect of lanosterol on the in vitro maturation in semi-defined culture system of prepubertal ewe oocytes

Zygote ◽  
2011 ◽  
Vol 22 (1) ◽  
pp. 50-57 ◽  
Author(s):  
F. Marco-Jiménez ◽  
J.S. Vicente ◽  
M.P. Viudes-de-Castro
Keyword(s):  
Oxygen Concentration ◽  
Cell Function ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Low Oxygen ◽  
Oocyte Growth ◽  
Important Indicator ◽  
Nuclear Maturation ◽  
Defined Culture

SummaryThe choice of medium and supplements can affect meiotic regulation and may have an impact on the regulation of mammalian oocyte growth and embryonic cell function. The aim of the present study was to assess the effects of oxygen concentration and endogenous lanosterol on the in vitro maturation (IVM) media without serum and based on recombinant human chorionic gonadotrophin in prepubertal ewe oocytes. Firstly, the effect of varying oxygen concentrations (5% and 20%) during IVM in TCM-199 supplemented (4 mg/ml bovine serum albumin (BSA), 100 μM cysteamine, 0.3 mM sodium pyruvate, 0.1 UI/ml recombinant follicle-stimulating hormone (r-FSH; Gonal-F® 75 UI, Serono, Italy), 0.1 UI/ml recombinant leuteinizing hormone (r-LH; Lhadi® 75 UI, Serono, Italy) and 1 μg/ml estradiol-17β) on subsequent nuclear maturation of oocytes examined under ultraviolet light following staining with bisbenzimide (Hoechst 33342) was investigated. Secondly, two concentrations of lanosterol (0, 10 and 50 μM) were added to the IVM medium. Nuclear maturation of oocytes was examined as previously. Lipid content in oocytes, an important indicator of cytoplasmic maturity, was also measured using Nile red fluorescent stain. The results showed that low oxygen concentration affected the nuclear maturation. Similarly, a significantly higher rate of meiosis resumption was observed with 10 μM (72.3%) of lanosterol compared with the control (51.8%) or 50 μM of lanosterol (59.4%). A significantly higher content of lipids was also observed with 10 and 50 μM of lanosterol (7.3 ± 0.2 × 106 and 7.4 ± 0.2 × 106 arbitrary units of fluorescence) compared with the control (6.7 ± 0.2 × 106 arbitrary units of fluorescence). The results indicate that 10 μM lanosterol during IVM in medium without serum and based on recombinant human chorionic gonadotrophin has a positive effect on maturation of prepubertal ewe oocytes.

Improvement of the developmental competence of canine oocyte using caffeine supplementation during IVM at different maturation time

Zygote ◽  
2018 ◽  
Vol 26 (2) ◽  
pp. 162-167 ◽  
Author(s):  
Mohamed Fathi ◽  
A. Salama ◽  
Magdy R. Badr
Keyword(s):  
Developmental Competence ◽  
Control Group ◽  
Free Medium ◽  
Maturation Time ◽  
Fluid Medium ◽  
Developmental Potential ◽  
Nuclear Maturation ◽  
Maturation Medium ◽  
Oviductal Fluid

SummaryThe aim of the current study was to investigate the effect of caffeine supplementation during in vitro maturation (IVM) for different maturation times on the developmental potential of canine oocytes recovered from ovariohysterectomized bitches. The recovered cumulus–oocytes complexes were in vitro matured for 72 h. Here, 10 mM caffeine was added to the maturation medium for different incubation times (caffeine from 0–72 h maturation, caffeine for the first 24 h of maturation only, caffeine addition from 24 to 48 h maturation time, caffeine addition from 48 to 72 h maturation or in caffeine-free medium, control group). The matured oocytes were in vitro fertilized using frozen–thawed spermatozoa. The presumptive zygotes were in vitro cultured in synthetic oviductal fluid medium for 5 days. The results showed that both maturation and fertilization rates were significantly higher (P ˂ 0.05) using caffeine-treated medium for the first 24 h of maturation compared with the control and other two groups of caffeine treatment (from 24 to 48 h and from 48 to 72 h), whereas use of caffeine-treated medium for a 0–72 h incubation time did not affect these rates (P > 0.05). Interestingly, the matured oocytes in caffeine-supplemented medium for the first 24 h or from 0–72 h showed a significant (P ˂ 0.05) increase in the total number of cleaved embryos compared with the control group. In conclusion, supplementation of the maturation medium with 10 mM caffeine for the first 24 h of maturation or during the whole maturation time (0–72 h) improved nuclear maturation and subsequent embryo development preimplantation following in vitro fertilization.

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