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1.
The aim of this study was evaluation of survivability, maturation rate and apoptotic gene expression of preantral follicles after vitrification and slow freezing technique. Normal mouse preantral follicles were randomly divided into three experimental groups. In the control group, follicles were cultured immediately; in the vitrification and slow freezing groups, follicles were cultured after vitrification‐warming and slow freezing‐thawing procedures. Follicular viability was assessed by using 0.4% trypan blue, and molecular evaluation of messenger RNA levels of apoptosis‐related genes was performed by the semi‐quantitative RT‐PCR method after 3 h of culture. Oocyte maturation rates were also evaluated on day 14 of culture. Survival and maturation rate in the slow freezing group were significantly lower than those in control and vitrification groups (P ≤ 0.05). Although there was no difference in Survivin expression among the three experimental groups, Bcl‐2 expression was significantly lower in the slow freezing group compared to the other groups (P ≤ 0.05). The expression of Bax, P53, Fas and Bax/Bcl‐2 ratio in the slow freezing group was significantly higher than control and vitrification groups (P ≤ 0.05). Preantral follicle vitrification seems to be better than slow freezing as seen in the survival, maturation and expression rates of apoptotic gene variants.  相似文献   

2.
The current study examined the protective effects of l ‐glutamine and cytochalasin B during vitrification of immature bovine oocytes. Oocyte vitrification solution (PBS supplemented with 10% FCS, 25% EG, 25% DMSO and 0.5 m trehalose) was the vitrification control. Treatments were the addition of 7 μg/ml cytochalasin B, 80 mm glutamine or both cytochalasin and glutaminine for 30 s. After warming, oocytes were matured in vitro for 24 h, fixed and stained with Hoechst (33342) for nuclear maturation evaluation. l ‐glutamine improved the vitrified/warmed immature bovine oocytes viability (32.8%), increasing the nuclear maturation rates compared to other treatments and the no treatment vitrified control (17.4%). There was, however, no effect of cytochalasin B on in vitro maturation (14.4%).  相似文献   

3.
We tested the effects of resveratrol both as a pre‐treatment and as a recovery treatment after warming during in vitro maturation (IVM) on the viability and developmental competence of porcine oocytes vitrified at the germinal vesicle stage. Pre‐treatment before vitrification of oocytes for 3 hr with 2 μM resveratrol did not affect survival, oocyte maturation and embryo developmental competence to the blastocyst stage after parthenogenetic activation. However, supplementation of the medium with resveratrol during subsequent IVM after vitrification and warming significantly improved the ability of surviving oocytes to develop to the blastocyst stage, and this effect was observed only on vitrified, but not on non‐vitrified oocytes. The intracellular levels of glutathione and hydrogen peroxide in oocytes were not affected by vitrification and resveratrol treatment. Also, there was no significant difference in the occurrence of apoptosis measured by annexin V binding between vitrified and non‐vitrified oocytes, regardless of the resveratrol treatment. In conclusion, resveratrol did not prevent the cellular damages in immature porcine oocytes during vitrification; however, when added to the IVM medium, it specifically improved the developmental competence of vitrified oocytes. Further research will be necessary to clarify the mechanisms of action of resveratrol on the recovery of vitrified oocytes from vitrification‐related damages.  相似文献   

4.
This study was designed to compare the efficiency of porcine oocytes vitrified at the GV and MⅡ stages. The vitrified GV oocytes were matured in vitro and then evaluated their cortical granule distribution with immunofluorescence, CD9 protein level with Western blotting and zona pellucida dissolution time with pronse digestion method, as compared to vitrified MⅡ oocytes.The results showed that the survival percentages of oocytes vitrified at the GV stage were significantly lower than those vitrified at the MⅡ stage after 2 h of warming (P<0.05);However, there was no difference in maturation rate between vitrified and fresh oocytes (P>0.05). The oocytes vitrified at MⅡ stage resulted in significantly decreased normal cortical granule distribution and CD9 protein level (P<0.05), but no such results were found in vitrified GV oocytes after maturation (P>0.05). In addition, vitrification did not affect the dissolution time of zona pellucida in GV and MⅡ oocytes (P>0.05). The data demonstrated that despite of lower survival rate compared with MⅡ oocytes, vitrified porcine GV oocytes after maturation showed normal polar body extrusion, cortical granule distribution and CD9 protein level.  相似文献   

5.
为比较猪卵母细胞在GV期与MⅡ期的冷冻保存效果,试验在这两个成熟阶段对其进行玻璃化冷冻,GV期卵母细胞解冻后培养至成熟,MⅡ期卵母细胞解冻后恢复2 h,然后采用免疫荧光标记、Western blotting和链霉蛋白酶溶解方法分别检测它们的皮质颗粒分布、CD9蛋白表达水平和透明带消化时间上的差异。结果表明,GV期卵母细胞在解冻后2 h的存活率显著低于MⅡ期卵母细胞（P<0.05）,但极体排出率与对照卵母细胞无明显差异（P>0.05）;在冷冻MⅡ期卵母细胞中,皮质颗粒的皮质区分布比例和CD9的蛋白表达水平显著下降（P<0.05）,但冷冻GV期卵母细胞经体外成熟后则无明显变化（P>0.05）;冷冻GV期与MⅡ期卵母细胞均不会影响透明带的消化时间（P>0.05）。由此可见,猪卵母细胞在GV期的冷冻存活率虽然较MⅡ期低,但其体外成熟后极体排出率、皮质颗粒分布和CD9蛋白表达水平均未受到冷冻的影响。  相似文献   

6.
We report the cryopreservation of oocytes from Ban miniature pigs which are endemic in Vietnam. Immature cumulus‐oocyte complexes were collected from antral follicles of 7–8 mo old female cyclic Ban pigs and vitrified in micro‐drops. Oocyte morphology, lipid content, post‐warming survival, nuclear maturation, and embryo development were compared to those of oocytes from commercially slaughtered Landrace × Large white hybrid pigs. The size of oocytes in the two breeds was similar. However, significantly lower amounts of intracellular lipid were detected in Ban oocytes. There was no difference (p > 0.05) between Ban and Landrace × Large white oocytes in percentages of post‐warming survival (93.1 ± 3.4% vs. 70.7 ± 16.7%, respectively) and nuclear maturation after in vitro maturation (80.4 ± 5.1% vs. 90.0 ± 1.3% respectively). Similarly, cleavage (30.8 ± 7.8% vs. 10.3 ± 6.1%, respectively) and blastocyst development rates (9.4 ± 5.0% vs. 0.79 ± 0.79, respectively) were not different (p > 0.05) between vitrified Ban and Landrace × Large white oocytes after in vitro fertilization and embryo culture. In conclusion, high survival and maturation rates were achieved after vitrification of immature Ban oocytes and their cryo‐tolerance was similar to that of Landrace × Large white oocytes, despite the difference in lipid content. We succeeded to generate reasonable rates of blastocysts from vitrified Ban oocytes by in vitro fertilization.  相似文献   

7.
The present study was conducted to investigate the effect of meiotic stages during in vitro maturation (IVM) on the survival of vitrified-warmed buffalo oocytes, vitrified at different stages of IVM. Cumulus oocyte complexes obtained from slaughterhouse ovaries were randomly divided into 6 groups: control (non-vitrified, matured for 24 h at 38 ± 1°C, 5% CO2 in humidified air), and those matured for 0 h (vitrified before IVM) or 6, 12, 18 and 24 h before vitrification. Cumulus oocyte complexes were vitrified in solution consisting of 40% w/v propylene glycol and 0.25 mol/L trehalose in phosphate-buffered saline supplemented with 4% w/v bovine serum albumin. Vitrified cumulus oocyte complexes were stored at −196∘C (liquid nitrogen) for at least 7 days and then thawed at 37°C; cryoprotectant was removed with 1 mol/L sucrose solution. Cumulus oocyte complexes in the 0, 6, 12, 18 and 24 h groups were then matured for an additional 24, 18, 12, 6 and 0 h, respectively, to complete 24 h of IVM. Among the five vitrification groups, 89–92% of cumulus oocyte complexes were recovered, after warming, of which 84–91% were morphologically normal. Overall survivability of vitrified cumulus oocyte complexes was lower (p < 0.05) than that of non-vitrified cumulus oocyte complexes (94.5%). Survival rates of cumulus oocyte complexes matured 24 h prior to vitrification (61.3%) were higher (p < 0.05) than those matured for 12 h (46.7%), 6 h (40.6%) and 0 h (37.6%). Nuclear status following 24 h IVM was assessed. A higher proportion of non-vitrified (control) oocytes (72.7%) reached metaphase II (M-II) stage in control than oocytes vitrified for 24 h (60.0%), 18 h (54.4), 12 h (42.3%), 6 h (33.3%) and 0 h (31.6%) (p < 0.05). The results suggest that length of time in maturation medium prior to vitrification influences post-thaw survivability of buffalo oocytes; longer intervals resulted in higher survival rates.  相似文献   

8.
This study was designed to evaluate effects of different combinations of cryoprotectants on the ability of vitrified immature buffalo oocytes to undergo in vitro maturation. Straw and open‐pulled straw (OPS) methods for vitrification of oocytes at the germinal vesicle stage also were compared. The immature oocytes were harvested from ovaries of slaughtered animals and were divided into three groups: (i) untreated (control); (ii) exposed to cryoprotectant agents (CPAs); or (iii) cryopreserved by straw and OPS vitrification methods. The vitrification solution (VS) consisted of 6 m ethylene glycol (EG) as the standard, control vitrification treatment, and this was compared with 3 m EG + 3 m dimethyl sulfoxide (DMSO), 3 m EG + 3 m glycerol, and 3 m DMSO + 3 m glycerol. Cryoprotectants were added in two steps, with the first step concentration half that of the second (and final) step concentration. After warming, oocyte samples were matured by standard methods and then fixed and stained for nuclear evaluation. Rates of MII oocytes exposed to CPAs without vitrification were lower (54.3 ± 1.9% in EG, 47.5 ± 3.4% in EG + DMSO, 36.8 ± 1.2% in EG + glycerol and 29.9 ± 1.0% in DMSO + glycerol; p < 0.05) than for the control group (79.8 ± 1.3%). For all treatments in each vitrification experiment, results were nearly identical for straws and OPS, so all results presented are the average of these two containers. The percentages of oocytes reaching telophase‐I or metaphase‐II stages were lower in oocytes cryopreserved using all treatments when compared with control. However, among the vitrified oocytes, the highest maturation rate was seen in oocytes vitrified in EG + DMSO (41.5 ± 0.6%). Oocytes cryopreserved in all groups with glycerol had an overall low maturation rate 19.0 ± 0.6% for EG + glycerol and 17.0 ± 1.1% for DMSO + glycerol. We conclude that the function of oocytes was severely affected by both vitrification and exposure to cryoprotectants without vitrification; the best combination of cryoprotectants was EG + DMSO for vitrification of immature buffalo oocytes using either straw or OPS methods.  相似文献   

9.
Apoptosis is one of the main drivers of the decline in developmental potential of porcine oocytes after vitrification. However, which apoptotic pathways are engaged after vitrification remains poorly understood. To distinguish among the possible apoptotic pathways induced by vitrification of MII stage porcine oocytes, this study detected activity and expression levels of several key proteins and genes in both the death receptor and mitochondrial pathways using in situ fluorescence staining and real‐time PCR (RT‐PCR) following the addition of specific inhibitors of either the death receptor or the mitochondrial apoptotic pathway (Z‐IETD‐FMK or Z‐LEHD‐FMK, respectively) into the incubation solution. Survival and parthenogenetic developmental ability were also examined. The results showed the following: (i) compared with the vitrified group, the activities of pan‐caspase, caspase 3, caspase 8 and caspase 9 as well as the early apoptotic rate were significantly lower in the Z‐IETD‐FMK and Z‐LEHD‐FMK groups (p < .05); (ii) After vitrification, mitochondrial ΔΨm decreased from 1.33 for fresh oocytes to 0.90 for vitrified oocytes, while the addition of Z‐IETD‐FMK or Z‐LEHD‐FMK following vitrification increased mitochondrial ΔΨm to 1.09 and 1.05, respectively (p < .05); (iii) Relative expression levels of apoptotic‐related genes from both the death receptor pathway (caspase 8 and TNF‐α) and the mitochondrial pathway (caspase 9, Bcl‐2 and CuZnSOD) changed substantially after the addition of Z‐IETD‐FMK or Z‐LEHD‐FMK into the incubation solution; (iv) Survival, cleavage and blastocyst rates were higher in the Z‐IETH‐FMK or Z‐LEHD‐FMK groups than in the vitrified group (p < .05). In summary, both the death receptor and the mitochondrial apoptotic pathways participate in the apoptosis of MII stage porcine oocytes after vitrification.  相似文献   

10.
This study was undertaken to compare cryotolerance, in terms of viability and resumption of meiosis after warming and culture (24 and 48 h), of ex situ (isolated) and in situ (enclosed in the ovarian tissue) feline cumulus–oocyte complexes (COCs) vitrified with DAP 213 (2 m DMSO, 1 m acetamide, 3 m propylene glycol) in cryotubes or Cryotop method. Ovaries were harvested from 49 pubertal queens. Of each pair of ovaries, one was dissected to release COCs randomly divided into three groups: fresh COCs (control), ex situ COCs vitrified with DAP 213 and Cryotop. The cortex of the other ovary was sectioned into small fragments (approximately 1.5 mm3) and randomly assigned to be vitrified by DAP 213 or Cryotop. After warming, ex situ and in situ (retrieved form vitrified ovarian tissue) COCs were matured in vitro. Viability of oocytes was highly preserved after warming and culture in all treatments. Proportions of oocytes surrounded by complete layers of viable cumulus cells were remarkably decreased (p < 0.00001) in both vitrification procedures compared to fresh oocytes. Resumption of meiosis occurred in all treatments. After 24 h of culture, results were similar in ex situ and in situ vitrified oocytes regardless of the vitrification protocol used (range 29–40%), albeit lower (p < 0.05) than those of fresh oocytes (65.8%). After 48 h of culture, ex situ oocytes vitrified with Cryotop achieved the rates of meiosis resumption similar to fresh oocytes (53.8% vs 67.5%; p > 0.05) and ex situ and in situ oocytes vitrified with DAP 213 showed similar rates of resumption of meiosis. These findings demonstrated that DAP 213 and Cryotop preserve the viability of ex situ and in situ oocytes, but cumulus cells are highly susceptible to vitrification. However, the capability to resume meiosis evidences that feline immature oocytes vitrified as isolated or enclosed in the ovarian cortex have comparable cryotolerance.  相似文献   

11.
The aim of the present study was to compare the efficiency of the solid surface (SSV), cryotop (CT) vitrification methods and cytochalasin B (CB) pretreatment for cryopreservation of immature buffalo oocytes. Cumulus‐oocyte complexes (COCs) were placed for 1 min in TCM199 containing 10% dimethylsulfoxide (DMSO), 10% ethylene glycol (EG), and 20% fetal bovine serum, and then transferred for 30 s to base medium containing 20% DMSO, 20% EG and 0.5 mol/L sucrose. CB pretreated ((+)CB) or non‐pretreated ((?)CB) COCs were vitrified either by SSV or CT. Surviving vitrified COCs were selected for in vitro maturation (IVM) and in vitro fertilization (IVF). The rate of viable oocytes after vitrification in CT groups (82%) was significantly lower (P < 0.05) than that in a fresh control group (100%), but significantly higher (P < 0.05) than those in SSV groups (71–72%). Among vitrified groups, the highest maturation rate was obtained in the CT (?)CB group (32%). After IVF, the cleavage and blastocyst formation rates were similar among vitrified groups but significantly lower than those of the control group. In conclusion, a higher survival rate of oocytes after vitrification and IVM was obtained in the CT group compared with that in the SSV group, indicating the superiority of the CT method. Pretreatment with CB did not increase the viability, maturation or embryo development of vitrified oocytes.  相似文献   

12.
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   

13.
  相似文献   

14.
The objectives of this study were to assess the efficiency of polarized light microscopy (PLM) in detecting microtubule‐polymerized protein in in vitro‐matured bovine oocytes; to examine its effects on oocyte developmental competence; and to assess the meiotic spindle of in vitro‐matured oocytes after vitrification/warming and further assessment of oocyte developmental competence. In the first experiment, the presence of microtubule‐polymerized protein (MPP) was confirmed as a positive PLM signal detected in 99.1% of analysed oocytes (n = 115), which strongly correlated (r = 1; p < 0.0001) with the presence of MPP as confirmed by immunostaining. In the second experiment, oocytes (n = 651) were exposed or not (controls) to PLM for 10 min and then fertilized and cultured in vitro. Oocytes exposed to PLM did not significantly differ from controls with regard to cleavage, total blastocyst and expanded blastocyst rates and cell numbers. In the third experiment, meiotic spindles were detected in 145 of 182 oocytes (79.6%) following vitrification and warming. Interestingly, after parthenogenetic activation and in vitro culture, oocytes that displayed a positive PLM signal PLM(+) differed significantly from PLM(?) in cleavage and Day 8 blastocyst rates. These results suggest that polarized light microscopy is an efficient system to detect microtubule‐polymerized protein in in vitro‐matured bovine oocytes and does not exert detrimental effects on bovine oocyte developmental competence. Moreover, PLM could be used as a tool to assess post‐warming viability in vitrified bovine oocytes.  相似文献   

15.
This work analyses the effects of a high hydrostatic pressure (HHP) treatment on in vitro survival of in vitro produced (IVP) bovine embryos vitrified with the Cryologic Vitrification Method (CVM). Consequences on embryo quality in terms of cell proliferation and differentiation, and levels of embryonic Heat Shock Protein 70 (Hsp‐70) were also examined. Day 7 and 8 bovine in vitro‐produced blastocysts were submitted to an HHP treatment (60 MPa, at 32°C for 1 h) and allowed to recover for 1 or 2 h in culture medium. The HHP treatment did not improve blastocyst survival rates after vitrification/warming. Survival (24 h post‐warming) and hatching (48 h post‐warming) rates were 79.3 ± 4.9 and 51.8 ± 4.2 vs 73.9 ± 4.2 and 44.7 ± 4.1 for untreated controls and HHP‐treated embryos, respectively. Total cell numbers measured in fresh embryos were reduced after 1 h at 32°C, with or without HHP treatment, indicating that cell proliferation was stopped as a result of stress. Vitrified HHP‐treated embryos that hatched at 48 h after warming showed increased cell numbers in their ICM compared with untreated controls (50.2 ± 3.1 vs 38.8 ± 2.7), indicating higher embryo quality. Treatment of blastocysts with HHP did not alter the level of the Hsp‐70 protein. In our conditions, HHP treatment did not affect the cryoresistance of these embryos. However, combination of HHP treatment and vitrification in fibreplugs resulted in an increase in the ICM cell number of hatched embryos 48 h post‐warming.  相似文献   

16.
The aim of the work was to evaluate the in vitro developmental competence of in vitro‐matured buffalo oocytes after Cryotop vitrification (CTV) and in vitro fertilization (IVF). To optimize parameters, two cryoprotectant (CP) concentrations and two warming–dilution procedures were applied. Oocytes were vitrified in 16.5% ethylene glycol (EG), 16.5% dimethylsulphoxide (DMSO) and 0.5 m sucrose in Groups A and C, and in higher CP concentrations (20% EG, 20% DMSO and 0.5 m sucrose) in Groups B and D. Warming was performed in 1.25 m sucrose for 1 min, then in 0.62, 0.42 and 0.31 m sucrose, 30 s each (Groups A and B), or in 0.25 m sucrose for 1 min and in 0.15 m sucrose for 5 min (Groups C and D). After warming, the oocytes were fertilized and cultured in vitro. Survival rate post‐warming was lower in Group D (83.6%) than in Groups A and B (92.4 and 92.8%, respectively), while intermediate values were found in Group C (85.7%). Survival rates at 24 h decreased in Groups C and D (52.0% and 50%, respectively) and remained high in Groups A and B (84.0% and 85.6%, respectively), thus indicating that the dilution of CP after warming is critical for buffalo oocyte cryopreservation. Similar differences were also observed in cleavage rates (42.7%, 55.3%, 28.4% and 36.3% for Groups A, B, C and D, respectively) whereas no differences in blastocyst rates were found among groups (6.4%, 7.8%, 5.9% and 6.9% for Groups A, B, C and D, respectively). Blastocyst production after IVF of vitrified oocytes proves the feasibility of CTV in buffalo species.  相似文献   

17.
Vitrification has been the method of choice for the cryopreservation of bovine oocytes, as rapid cooling decreases chilling sensitivity. The aim of this study was to determine the in vitro and in vivo survival and the viability of immature oocytes vitrified using super‐cooled liquid nitrogen. Immature oocytes were randomly allocated to three groups: (i) non‐vitrified control group, (ii) vitrified in normal (?196°C) liquid nitrogen (LN2) and (iii) vitrified in super‐cooled LN2 (≤?200°C). Open‐pulled glass micropipettes were used as vitrification containers. Immature oocytes were in vitro‐matured, fertilized and cultured to the blastocyst stage. In vitro viability was assessed by cleavage and blastocyst rates on days 2 and 7 of culture respectively. Vitrified blastocysts derived from the immature vitrified oocytes were directly transferred to synchronous recipients. The in vitro embryo development of vitrified immature oocytes was not influenced by the LN2 state. After direct transfer (one embryo per recipient) of 16 embryos obtained from immature vitrified oocytes (eight from each vitrified group), two healthy calves were born in each group. These results indicated that vitrification of immature bovine oocytes using glass micropipettes under normal or super‐cooled LN2, resulted in viable blastocysts and live calves following in vitro embryo production.  相似文献   

18.
Our aim was to optimize a cryoprotectant treatment for vitrification of immature porcine cumulus-oocyte complexes (COCs). Immature COCs were vitrified either in 35% ethylene glycol (EG), 35% propylene glycol (PG) or a combination of 17.5% EG and 17.5% PG. After warming, the COCs were in vitro matured (IVM), and surviving oocytes were in vitro fertilized (IVF) and cultured. The mean survival rate of vitrified oocytes in 35% PG (73.9%) was higher (P<0.05) than that in 35% EG (27.8%). Oocyte maturation rates did not differ among vitrified and non-vitrified control groups. Blastocyst formation in the vitrified EG group (10.8%) was higher (P<0.05) than that in the vitrified PG group (2.0%) but was lower than that in the control group (25.0%). Treatment of oocytes with 35% of each cryoprotectant without vitrification revealed a higher toxicity of PG on subsequent blastocyst development compared with EG. The combination of EG and PG resulted in 42.6% survival after vitrification. The maturation and fertilization rates of the surviving oocytes were similar in the vitrified, control and toxicity control (TC; treated with EG+PG combination without cooling) groups. Blastocyst development in the vitrified group was lower (P<0.05) than that in the control and TC groups, which in turn had similar development rates (10.7%, 18.1% and 23.3%, respectively). In conclusion, 35% PG enabled a higher oocyte survival rate after vitrification compared with 35% EG. However, PG was greatly toxic to oocytes. The combination of 17.5% EG and 17.5% PG yielded reasonable survival rates without toxic effects on embryo development.  相似文献   

19.
Although cryopreservation of mammalian oocytes is an important technology, it is well known that unfertilized oocytes, especially in pigs, are highly sensitive to low temperature and that cryopreserved oocytes show low fertility and developmental ability. The aim of the present study was to clarify why porcine in vitro matured (IVM) oocytes at the metaphase II (MII) stage showed low fertility and developmental ability after vitrification. In vitro matured cumulus oocyte complexes (COCs) were vitrified with Cryotop and then evaluated for fertility through in vitro fertilization (IVF). Although sperm‐penetrated oocytes were observed to some extent (30–40%), the rate of pronuclear formation was low (9%) and none of them progressed to the two‐cell stage. The results suggest that activation ability of cryopreserved oocytes was decreased by vitrification. We examined the localization and expression level of the type 1 inositol 1,4,5 trisphosphate receptor (IP3R1), the channel responsible for Ca2+ release during IVF in porcine oocytes. Localization of IP3R1 close to the plasma membrane and total expression level of IP3R1 protein were both decreased by vitrification. In conclusion, our present study indicates that vitrified‐warmed porcine COCs showed a high survival rate but low fertility after IVF. This low fertility seems to be due to the decrease in IP3R1 by the vitrification procedure.  相似文献   

20.
In this study, the effects of the addition of L‐carnitine in in vitro maturation (IVM) medium for bovine oocytes on their nuclear maturation and cryopreservation were investigated; they were matured in IVM medium supplemented with 0.0, 0.3, 0.6 and 1.2 mg/mL of L‐carnitine (control, 0.3, 0.6 and 1.2 groups, respectively) and some of them were vitrified by Cryotop. Moreover, the effects of L‐carnitine during in vitro fertilization (IVF) and in vitro culture (IVC) on the developmental potential and quality of IVF embryos were also examined. A significantly higher maturation rate of oocytes was obtained for 0.3 and 0.6 mg/mL groups compared with the control (P < 0.05). The blastocyst formation rate in the 0.6 group was significantly improved, whereas the rate in the 1.2 group was significantly decreased when compared with the control group (P < 0.05). No significant difference was found in embryo development between the control and the L‐carnitine group after oocyte vitrification. Supplementation of IVF and IVC media with L‐carnitine had no effect on development to the blastocyst stage of IVM oocytes treated with 0.6 mg/mL L‐carnitine. In conclusion, the supplementation of L‐carnitine during IVM of bovine oocytes improved their nuclear maturation and subsequent embryo development after IVF, but when they were vitrified the improving effects were neutralized.  相似文献   

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