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1.
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2.
Melatonin enhances in vitro embryo development in several species by improving the oocyte developmental competence during in vitro maturation (IVM). Melatonin has a wide range of actions, from scavenging reactive oxygen species (ROS) to regulating gene expression, and it can also act by way of melatonin receptors. The aim of this study was to determine the mechanism of action of melatonin during the IVM of juvenile goat oocytes and the role of the membrane receptors. Melatonin receptor 1 was immunolocalized in cumulus cells and oocytes before and after 24 hr of IVM. The effect of melatonin on oocyte developmental competence was tested in three experimental IVM groups: (a) control, (b) 10?7 M melatonin, and (c) 10?7 M melatonin +10?7 M luzindole (an inhibitor of both melatonin receptors). After IVM oocytes were assessed for ROS levels, mitochondrial activity, adenosine 5′‐triphosphate (ATP) concentration and relative gene expression (ACTB, SLC1A1, SOD1, GPx1, BAX, DNMT1, GCLC and GDF9). IVM‐oocytes were in vitro fertilized and cultured under conventional conditions. Blastocyst rate and quality (differential cell count) were assessed at 8 days post‐fertilization. Melatonin decreased ROS levels, increased mitochondrial activity and ATP content and increased blastocyst quality compared to control group (55.8 vs. 30.4 inner cell mass ICM, p < 0.05). There was no effect on the relative gene expression due to treatment with melatonin. In conclusion, we have showed that melatonin improves oocyte developmental competence in juvenile goats by reducing ROS levels and improving mitochondrial activity.  相似文献   

3.
Melatonin has been reported to improve the in vitro development of embryos in some species. This study was conducted to investigate the effect of melatonin supplementation during in vitro maturation (IVM) and development culture on the development and quality of porcine embryos. In the first experiment, when the in vitro fertilized embryos were cultured with different concentrations of melatonin (0, 10, 25 and 50 ng/ml) for 8 days, the blastocyst formation rate of embryos cultured with 25 ng/ml melatonin (10.7%) was significantly increased (p < 0.05) compared to the control embryos cultured without melatonin (4.2%). The proportion of DNA‐fragmented nuclei in blastocysts derived from embryos cultured with 50 ng/ml melatonin was significantly lower (p < 0.05) than that of embryos cultured without melatonin (2.1% vs 7.2%). In the second experiment, when oocytes were cultured in the maturation medium supplemented with different concentrations of melatonin (0, 10, 25 and 50 ng/ml), fertilized and then cultured with 25 ng/ml melatonin for 8 days, there were no significant differences in the rates of cleavage and blastocyst formation among the groups. However, the proportions (2.7–5.4%) of DNA‐fragmented nuclei in blastocysts derived from oocytes matured with melatonin were significantly decreased (p < 0.05) compared to those (8.9%) from oocytes matured without melatonin, irrespective of the concentration of melatonin. Our results suggest that supplementation of the culture media with melatonin (25 ng/ml) during IVM and development has beneficial effects on the developmental competence and quality of porcine embryos.  相似文献   

4.
Effects of adding different concentrations of melatonin (10?7, 10?9 and 10?11 M) to maturation (Experiment 1; Control, IVM  + 10?7, IVM  + 10?9, IVM  + 10?11) and culture media (Experiment 2; Control, IVC  + 10?7, IVC  + 10?9, IVC  + 10?11) were evaluated on in vitro bovine embryonic development. The optimal concentration of melatonin (10?9 M) from Experiments 1–2 was tested in both maturation and/or culture media of Experiment 3 (Control, IVM  + 10?9, IVC  + 10?9, IVM /IVC  + 10?9). In Experiment 1, maturated oocytes from Control and IVM  + 10?9 treatments showed increased glutathione content, mitochondrial membrane potential and percentage of Grade I blastocysts (40.6% and 43%, respectively). In Experiment 2, an increase in the percentage of Grade I blastocysts was detected in IVC  + 10?7 (43.5%; 56.7%) and IVC  + 10?9 (47.4%; 57.4%). Moreover, a lower number and percentage of apoptotic cells in blastocysts were observed in the IVC  + 10?9 group compared to Control (3.8 ± 0.6; 3.6% versus 6.1 ± 0.6; 5.3%). In Experiment 3, the IVC  + 10?9 treatment increased percentage of Grade I blastocysts with a lower number of apoptotic cells compared to IVM /IVC  + 10?9 group (52.6%; 3.0 ± 0.5 versus 46.0%; 5.4 ± 1.0). The IVC  + 10?9 treatment also had a higher mRNA expression of antioxidant gene (SOD 2) compared to the Control, as well as the heat shock protein (HSPB 1) compared to the IVM  + 10?9. Reactive oxygen species production was greater in the IVM /IVC  + 10?9 treatment group. In conclusion, the 10?9 M concentration of melatonin and the in vitro production phase in which it is used directly affected embryonic development and quality.  相似文献   

5.
The purpose of oocyte in vitro maturation is generation of mature oocytes that could support future development. Efforts have been made to enhance oocyte developmental competence by developing optimal culture conditions. The present study is conducted to determine melatonin effects on quality of polycystic ovarian syndrome (PCOS) oocytes when it has been added during in vitro maturation, and immature oocytes were cultured in defined conditioned medium with and without different melatonin concentrations. Melatonin could significantly improve nuclear maturation of PCOS oocytes (81.1% vs. 56.3%, P < 0.05 were achieved with 10?6 mol/L concentration). Cleavage rate was significantly higher in 10?5 mol/L concentration compared to untreated oocytes in PCOS (54% vs. 35%, respectively) and it was significantly higher with 10?6 mol/L concentration in the control group, 55% versus 38%, compared to untreated oocytes. This study showed that melatonin has the potential to induce oocyte nuclear maturation and guarantee fertilization potential. © 2016 Japanese Society of Animal Science  相似文献   

6.
Growing porcine oocytes from early antral follicles can acquire meiotic and developmental competence under suitable culture conditions, but at lower rates compared to full‐grown oocytes. We postulated that estradiol‐17β (E2) supported the acquisition of meiotic and developmental competence as well as cumulus‐expansion ability during growth culture. Growing oocytes from early antral follicles (1.2 to 1.5 mm in diameter) were grown in vitro for 5 days in a medium containing 0, 10?7, 10?6, 10?5 or 10?4 mol/L E2; after in vitro maturation, 35, 58, 47, 74 and 49% of oocytes matured to metaphase II, 25, 79, 77, 90 and 97% acquired cumulus‐expansion ability, and 23, 54, 63, 89 and 64% were fully surrounded by cumulus cells, respectively. Following maturation, electro‐stimulation was applied to the oocytes grown with 10?5 mol/L E2. After 6 days of culture, in vitro‐grown oocytes developed to the blastocyst stage at a rate similar to that for full‐grown oocytes (31% and 40%, respectively). Therefore, we suggest that the use of E2 during growth culture improves the meiotic and developmental competence of oocytes, cumulus‐expansion ability, and cumulus cell attachment to the oocytes.  相似文献   

7.
This study was conducted to determine the adequate medium for a serum‐free culture system of domestic cat embryos produced by in vitro maturation (IVM) and fertilization (IVF). Cumulusoocyte complexes recovered from cat ovaries were matured in vitro for 24 h, and then inseminated in vitro for 12 h. After insemination, the oocytes were cultured in five media [Ham's F10, Waymouth 752/1 (Waymouth), TCM199, modified Earle's balanced salt solution (MK‐1) and CR1aa], each of which contained 0.4% bovine serum albumin. There were no significant differences among the rates of fertilization of oocytes cultured in five media following IVF. The rate of oocytes/embryos developed to at least the morula stage was significantly lower (p < 0.05) in Waymouth than in MK‐1, TCM199 and CR1aa. Moreover, none of the embryos cultured in Ham's F10 and Waymouth developed to the blastocyst stage. There were no differences among the rates of development to the blastocyst stage of oocytes/embryos cultured in MK‐1, TCM199 and CR1aa. These results indicate that the type of serum‐free medium has a major impact on in vitro development of domestic cat embryos derived from IVM/IVF, and MK‐1, TCM199 and CR1aa media are suitable for in vitro culture of cat embryos in a serum‐free culture system.  相似文献   

8.
This study was carried out to investigate the effect of supplementing culture medium with different concentrations of taurine and melatonin, on buffalo oocyte in vitro meiotic maturation and embryo development. In experiment 1, oocytes were matured in vitro and the cleaved embryos were cultured in the same following seven culture medium; (i) control (TCM 199 + 10% SS); (ii) control + 0.5 m m taurine; (iii) control + 1 m m taurine; (iv) control + 3 m m taurine; (v) control + 5 μ m melatonin; (vi) control + 10 μ m melatonin and (vii) control + 50 μ m melatonin. In experiment 2, based on the results of experiment 1, to examine the synergistic effect of antioxidants, the oocytes were matured in culture medium (TCM199 + 10% SS), supplemented with both taurine at 1 m m and melatonin at 10 μ m concentration and the cleaved embryos were cultured in the same medium. Supplementation of taurine at 1 m m concentration in the culture medium resulted in a higher (p < 0.05) transferable embryo (TE) yield when compared with control (20.6% vs 14.1%). Supplementation of melatonin at 10 and 50 μ m concentration in the culture medium resulted in a higher (p < 0.05) meiotic maturation rate (90.3% and 88.8% respectively) and TE yield (28.4% and 27.2% respectively), than the other treatments. In experiment 2, the TE yield did not improve by supplementing the culture medium with both taurine and melatonin, when compared with melatonin alone. In conclusion, the results of this study demonstrated that, enriching the culture medium with taurine and melatonin, improves in vitro embryo production efficiency in buffaloes. In particular, a high TE yield was obtained by enriching the culture medium with 10 μ m melatonin.  相似文献   

9.
Chlorogenic acid (CGA) is known to protect oocytes from oxidative stress. Here we investigated the effects of CGA on porcine oocyte maturation under heat stress and subsequent embryonic development after parthenogenetic activation. For in vitro maturation (IVM) at 41.0°C (hyperthermic condition), supplementation of the maturation medium with 50 μM CGA significantly improved the percentage of matured oocytes and reduced the rate of apoptosis relative to oocytes matured without CGA (p < .05). CGA treatment of oocytes during IVM under hyperthermia tended to increase (p < .1) percentage of blastocyst formation after parthenogenesis and significantly increased (p < .05) the total cell number per blastocyst relative to oocytes matured without CGA. For IVM at 38.5°C (isothermic condition), CGA significantly improved the rate of blastocyst development compared with oocytes matured without CGA (p < .05), but did not affect oocyte maturation, apoptosis rate or the number of cells per embryo. Omission of all antioxidants from the IVM medium significantly reduced the rate of oocyte maturation, but the rate was restored upon addition of CGA. These results demonstrate that CGA is a potent antioxidant that protects porcine oocytes from the negative effects of heat stress, thus reducing the frequency of apoptosis and improving the quality of embryos.  相似文献   

10.
We evaluated the effects of polyethylene glycol (PEG) and Supercool X‐1000 (SC) as supplements during the vitrification of immature cumulus‐enclosed porcine oocytes in a solution based on 17.5% ethylene glycol + 17.5% propylene glycol. After warming, the oocytes were subjected to in vitro maturation, fertilization and embryo culture. In Experiment 1, equilibration and vitrification solutions were supplemented with or without 2% (w/v) PEG (PEG+ and PEG‐, respectively). The survival rate, cleavage and blastocyst development were similar between PEG+ and PEG‐ groups; however, all values were lower than those in the non‐vitrified control. In Experiment 2, vitrification solution was supplemented with or without 1% (v/v) SC (SC+ and SC‐, respectively). The percentages of survival and blastocyst development were similar between SC+ and SC‐ groups but lower than those in the non‐vitrified control. The percentage of cleavage in the SC‐ group was significantly lower than the control and the SC+ groups, which were in turn similar to one another. In both experiments, the cell numbers in blastocysts were not significantly different among the non‐vitrified and vitrified groups. In conclusion, PEG did not improve oocyte survival and embryo development, whereas SC improved the ability of surviving oocytes to cleave but not to develop into blastocysts.  相似文献   

11.
The availability of cow ovaries from the slaughterhouse has been very limited in Taiwan. To maximize the use of cow ovaries for research purposes, whole ovary dissection was performed and the developmental competence of the oocytes derived from different sizes of follicles was assessed by the rates of in vitro maturation (IVM) and parthenogenetic activation of the oocytes in Experiment 1 (Exp 1). Cumulus-oocyte complexes (COCs) derived from small (1-2 mm) and large (3-8 mm) follicles were subjected to standard IVM culture for 24 h. Mature oocytes were selected and then parthenogenetically activated using A23187 (5 microm, 5 min) or thimerosal (200 microm, 10 min) alone or combined with 6-dimethylaminopurine (2.5 mm and 3.5 h, respectively). Activation rates of the oocytes, neither from the large nor small follicles, were affected by different activation treatments (single or combined stimuli). Whereas maturation rates for the oocytes from large follicles were superior to those from small follicles in both the single (59% vs 45%) and combined treatments (76% vs 40%; p < 0.05). To understand how prolonged heat shock (HS) influences cytoskeletal configurations of mature bovine oocytes, in Experiment 2 (Exp 2), matured oocytes derived from large follicles were randomly allocated to different durations of HS treatments at 41.5 degrees C for 0 (C0h, control, n = 12), 1 (HS1h, n = 28), 2 (HS2h, n = 31), and 4 h (HS4h, n = 30). An additional control group was cultured for 4 h without HS (38.5 degrees C, 4 h, n = 35). Alterations in nuclear structures, microtubules (MTs), and microfilaments (MFs) of the oocytes were examined. Abnormalities in the chromosomes, spindle MTs and the percentages of oocytes with cytoplasmic MTs increased with time of HS treatment. The intensity of the MF distribution in the HS oocytes was also altered. Significant changes in the cytoskeleton after HS may be associated with the reduced development under hyperthermia and, perhaps, with the low pregnancy rates of the animals during hot seasons.  相似文献   

12.
The developmental potential of nuclei from a bovine mammary epithelial cell line (BMEC) in nuclear transfer was investigated. For nuclear transfer donors, BMEC cells (passage 15) were cultured for 4–5 days after seeding at cell densities of 1.0 × 105 cells/cm2 (high‐density group) or 0.8 × 104 cells/cm2 (low‐density group). First, the effective electric stimulation for fusion of enucleated oocytes with BMEC cells was examined. Fusion rates reached maximum with two DC pulses of 30 V/150 µm for 20 µs in the high‐density group and with two DC pulses of 25 V/150 µm for 10 µs in the low‐density group. The fusion rate (37.5%) in the high‐density group was significantly (P < 0.005) lower than in the low‐density group (71.4%). Second, the in vitro developmental potential of nuclear transfer embryos derived from BMEC cells was examined. In the high‐density and low‐density groups, 18.8% and 24.1% of fused oocytes, respectively, developed to blastocyst stage. The results of this study indicate that nuclei from BMEC cells support the development of nuclear transfer embryos to the blastocyst stage and that the efficiency of oocyte–cell fusion is affected by the culture conditions of the donor BEMC cells before nuclear transfer.  相似文献   

13.
The development of cleavage stage preimplantation embryos is disrupted by exposure to heat shock, such as high temperatures in the summer season. In this study, we investigated whether addition of anthocyanins, which are strong scavengers of reactive oxygen species (ROS), improves development and intracellular redox status of heat-exposed bovine preimplantation embryos by reduction of heat shock-derived oxidative stress. After in vitro fertilization (IVF), embryos were cultured at 38.5 C through Day 8 (Day 0=day of IVF) with 0, 0.1, 1 and 10 microg/ml anthocyanins (non-heat-shocked group). On Day 2, embryos were cultured at 41.5 C for 6 h with 0, 0.1, 1 and 10 microg/ml anthocyanins followed by culture at 38.5 C until Day 8 (HS group). After exposure to heat shock, the intracellular ROS and glutathione (GSH) contents of individual embryos were measured in the non-heat-shocked and HS groups using fluorescent probes. On Day 8, the blastocysts formation rates of the embryos and total cell numbers of blastocysts were evaluated. Embryos exposed to heat shock without anthocyanins showed a significant decrease in blastocyst formation rate and GSH content (P<0.05) and an increase in intracellular ROS (P<0.05) compared with non-heat-shocked embryos. In contrast, addition of 0.1 microg/ml anthocyanins significantly (P<0.05) improved the blastocyst formation rate of the heat-shocked embryos. Addition of any dose of anthocyanins produced a significant decrease in the ROS levels (P<0.05) and tended to increase the GSH levels under heat-shock conditions. However, addition of higher concentrations (1 and 10 microg/ml) of anthocyanins to the culture media under heat shock did not improve the development of embryos. These results indicate that anthocyanins maintain the intracellular redox balance of heat-shocked bovine embryos by reducing intracellular oxidative stress and increasing the GSH levels. Thus, alterations of the redox state using natural antioxidative polyphenols is a useful approach for reducing heat shock-derived oxidative stress.  相似文献   

14.
This study examined the presence of immunoreactivity and mRNA for different nitric oxide synthase (NOS) isoforms in immature and in vitro matured oocytes and in embryos at two‐, four‐ and eight‐cell, and morula and blastocyst stages in buffalo. Oocytes obtained from slaughterhouse buffalo ovaries were subjected to in vitro maturation in TCM‐199 + 10% FBS + 5 μg/ml pFSH + 1 μg/ml estradiol‐17β + 0.81 mm sodium pyruvate + 10% buffalo follicular fluid + 50 μg/ml gentamycin sulphate for 24 h in a CO2 incubator (5% CO2 in air) at 38.5°C. Following in vitro fertilization carried out by incubating them with 2–4 million spermatozoa/ml for 18 h, the presumed zygotes were cultured in mCR2aa medium containing 0.6% BSA and 10% FBS for up to 8 days post insemination. Immunofluorescence staining of NOS using antibodies that cross‐reacted either with all the NOS isoforms i.e., universal (uNOS) or specifically with inducible (iNOS) or endothelial (eNOS) isoforms revealed that NOS was present in oocytes and embryos at all the stages examined. Examination of the semi‐quantitative expression of NOS genes by RT‐PCR revealed that the iNOS, eNOS and nNOS mRNA was present in the immature and mature oocytes and in all the embryonic stages examined. In conclusion, it was demonstrated in the present study that immunoreactivity and mRNA for different NOS isoforms was present in buffalo oocytes and pre‐implantation stage embryos.  相似文献   

15.
Fibroblast growth factor (FGF10) acts at the cumulus oocyte complex, increasing the expression of cumulus cell expansion‐related genes and oocyte competency genes. We tested the hypothesis that addition of FGF10 to the maturation medium improves oocyte maturation, decreases the percentage of apoptotic oocytes and increases development to the blastocyst stage while increasing the relative abundance of developmentally important genes (COX2, CDX2 and PLAC8). In all experiments, oocytes were matured for 22 h in TCM‐199 supplemented with 0, 2.5, 10 or 50 ng/ml FGF10. In Experiment 1, after maturation, oocytes were stained with Hoechst to evaluate meiosis progression (metaphase I, intermediary phases and extrusion of the first polar body) and submitted to the TUNEL assay to evaluate apoptosis. In Experiment 2, oocytes were fertilized and cultured to the blastocyst stage. Blastocysts were frozen for analysis of COX2, CDX2 and PLAC8 relative abundance. In Experiment 1, 2.5 ng/ml FGF10 increased (p < 0.05) the percentage of oocytes with extrusion of the first polar body (35%) compared to 0, 10 and 50 ng/ml FGF10 (21, 14 and 12%, respectively) and FGF10 decreased the percentage of oocytes that were TUNEL positive in all doses studied. In Experiment 2, there was no difference in the percentage of oocytes becoming blastocysts between treatments and control. Real‐time RT‐PCR showed a tendency of 50 ng/ml FGF10 to increase the relative abundance of COX2 and PLAC8 and of 10 ng/ml FGF10 to increase CDX2. In conclusion, the addition of FGF10 to the oocyte maturation medium improves oocyte maturation in vitro, decreases the percentage of apoptotic oocytes and tends to increase the relative abundance of developmentally important genes.  相似文献   

16.
Ban is an endangered miniature pig breed in Vietnam. This study aimed to set up an in vitro embryo production (IVP) system for this breed. Ban's epididymal sperm concentration (1240 ± 35 × 106/mL) was lower (P < 0.01) compared with Landrace (4160 ± 42 × 106/mL). However, sperm characteristics before and after freezing in Ban and Landrace were similar. The numbers of follicles with diameter larger than 2 mm per ovary in Ban females treated with equine chorionic gonadotropin and human chorionic gonadotropin (27.1 ± 1.3) were higher (P < 0.05) than those in Landrace (12.9 ± 2.0) and in non‐hormone stimulated Ban (no > 2 mm follicles). After in vitro maturation, the percentages of oocytes with expanded cumulus cells and the first polar body (matured oocytes) were not different among Ban, hormone‐stimulated Ban and Landrace. The percentages of two‐cell embryos and morulae derived from oocytes collected from three sources did not differ. However, the rate of blastocysts derived from oocytes in non‐stimulated Ban (4.0 ± 3.8%) was lower (P < 0.05) than that in Landrace (15.3 ± 1.8%). In conclusion, an effective IVP system for good quality embryos in Ban, that is essential for genetic conservation of this breed, was established.  相似文献   

17.
为探讨表皮生长因子(epidermal growth factor,EGF)的添加浓度及脱卵丘细胞时间对猪卵母细胞体外成熟及孤雌胚胎体外发育的影响.试验通过在体外成熟液中添加不同浓度(0、10、15、20、30、40 ng/mL)的EGF来研究其对培养44 h的卵母细胞成熟率以及孤雌胚胎发育的影响;在培养开始后的不同时间(18、24、38、44 h)进行脱卵丘细胞处理来研究不同时间脱卵丘处理对培养44 h的卵母细胞成熟率以及孤雌胚胎发育的影响.结果表明,成熟培养基中添加10 ng/mL EGF能显著提高卵母细胞的卵裂率和囊胚率(P <0.05).共培组和独培组卵母细胞培养18 h后脱卵丘细胞成熟率均低于44 h,但差异不显著(P >0.05);共培组卵母细胞培养18 h后脱卵丘细胞的卵裂率和囊胚率显著高于培养44 h(P <0.05);独培组卵母细胞培养18 h后脱卵丘细胞的卵裂率与44 h无显著差异(P >0.05),但囊胚率显著高于培养44 h后脱卵丘细胞(P <0.05).添加10 ng/mL EGF对猪卵母细胞体外成熟及孤雌胚胎体外发育较好;卵母细胞培养18 h后脱卵丘细胞可提高孤雌胚胎早期发育能力.  相似文献   

18.
This study investigated the influence of heat shock during in vitro maturation on embryo development following in vitro fertilization (IVF) or parthenogenesis (Part). Immature bovine cumulus–oocyte complexes were exposed to heat shock (41.0°C) during the first 12 hr of in vitro maturation (IVM), followed by 12 hr at 38.5°C. Control group consisted of in vitro maturation for 24 hr at 38.5°C. Oocytes were in vitro‐fertilized or activated with ionomycin and cultured in vitro for 192 hr post‐in vitro insemination or parthenogenetic activation (hpia). There was an interaction (p < .01) between temperature of IVM and method of oocyte activation (IVF or Part) for cleavage at 48 hpia. Heat shock had a negative impact (p < .01) on cleavage of IVF embryos, whereas no (p > .05) effect was found in the Part embryos. Embryo development towards blastocyst stage at 168 and 192 hpia decreased in both IVF and Part embryos derived from heat‐shocked oocytes. Heat shock increased (p < .05) the apoptotic index in Part blastocysts, but no effect (p > .05) was found in IVF counterparts. Heat shock also down‐regulated the expression of AQP3 (p < .01) and up‐regulated the expression of HSP70.1 (p < .01) in Part blastocysts, whereas it down‐regulated the expression of ATP1A1 (p < .05) in IVF blastocysts. In conclusion, the effects of heat shock during IVM on early embryo cleavage and blastocyst apoptosis are influenced by the method of oocyte activation and expression of some genes can be disturbed in embryos derived from heat‐shocked oocytes.  相似文献   

19.
This study investigated the effect of porcine follicular fluid (PFF) and dibutyryl cyclic adenosine monophosphate (dbcAMP) during in vitro maturation (IVM) of porcine oocytes on meiotic maturation, fertilization and embryo development, and compared the effect of supplementing the embryo culture media with PFF or foetal bovine serum (FBS) on embryo development. Oocytes from pre‐pubertal gilts were IVM for 44 h, and parthenogenetically activated or in vitro‐fertilized. Embryos were cultured in porcine zygote medium (PZM3) for 7 days. Cleavage and blastocyst rates were evaluated at 48 h and 7 days of culture. The supplementation of the IVM medium with 25% PFF and 1 mm dbcAMP for the first 22 h resulted in more (p < 0.05) embryos developing to the blastocyst stage as compared with the inclusion of dbcAMP alone. The dbcAMP + PFF combination increased (p < 0.05) the average number of nuclei per blastocyst as compared with either of these components alone or in its absence. A synergistic effect of dbcAMP + PFF during IVM was also reflected in the capacity of oocytes to regulate sperm penetration and prevent polyspermy, as twice as many oocytes from the control group were penetrated by more than one sperm as compared with those matured in the presence of both dbcAMP and PFF. The supplementation of PZM3 with 10% FBS from days 5 to 7 of culture significantly improved the total cell quantity in embryos derived either from control or dbcAMP + PFF matured oocytes. There was no effect on the total cell quantity when FBS was replaced by the same concentration of PFF. These studies showed that dbcAMP, PFF and FBS can improve both the quantity (57.3% vs 41.5%) and quality (74.8 vs 33.3 nuclei) of porcine blastocysts derived from oocytes recovered of pre‐pubertal gilts.  相似文献   

20.
Studies were conducted to examine the possibility of preserving slaughterhouse‐derived buffalo ovaries at 4°C for 0 (control), 12 and 24 h to maintain the developmental competence of the oocytes (experiment 1), to assess the effect of incubation temperature during oocyte maturation on rates of in vitro maturation (IVM) and in vitro fertilization (IVF) of buffalo oocytes and embryo development (experiment 2), and to examine the effect of storage at 25°C for 0 (control), 4 and 8 h of frozen–thawed buffalo sperm and BO and H‐TALP as sperm processing and fertilization media on cleavage and embryo development in vitro of buffalo oocytes (experiment 3) in order to optimize the IVF technology in buffalo. Results suggested that storage of ovaries at 4°C for 12 or 24 h significantly (p < 0.05) reduced the developmental potential of oocytes. Incubation temperatures during the IVM influenced the fertilization rate but had no significant effect on maturation and subsequent embryo development. The incubation temperature of 38.5°C during IVM was found to be optimum for embryo production in vitro. Storage of frozen–thawed sperm at 25°C for 8 h significantly (p < 0.05) decreased its ability to cleave the oocytes. Sperm processed in BO medium had significantly (p < 0.05) higher ability to cleave the oocytes than the H‐TALP medium.  相似文献   

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