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1.
本研究观察了不同超排处理方法对羔山羊卵母细胞体外成熟及体外受精的影响，并将体外受精胚胎进行了移植，以便探讨利用羔山羊卵母细胞生产体外受精胚胎的技术途径。对２～４月龄羔山羊用ＦＳＨ、ＦＳＨ＋ＬＨ（静脉注射）和ＦＳＨ＋ＬＨ（肌肉注射）三种方法进行超排处理，平均分别回收１６．０、２０．３和１５．０枚卵母细胞。将这些卵母细胞进行２０～２４ｈ的成熟培养后，其成熟率分别为４５．５％，４４．４％和７６．５％。成熟卵母细胞经体外受精，体外发育培养后其２～４细胞胚胎的发生率分别为５．６％ａ、６．７％和３１．７％ｂ（ｂ，ｖｓ，ａ，Ｐ＜０．０５）。将９枚２～４细胞期胚胎移植给６只受体母羊，结果有２只妊娠并产羔２只。  相似文献   

2.
This study aims to assess the efficiency of in vivo oocyte and embryo recovery after a recombinant human FSH (rhFSH) treatment in rabbit does. Females were distributed in two experimental groups: donor does were treated with rhFSH (superovulation group) for 3 days prior to artificial insemination (embryo recovery) or ovulation induction (oocyte recovery) and does without treatment remained as the control group. Mature oocytes or embryos were collected with the laparoscopy technique 16 h after ovulation induction (oocytes) or 72 h after artificial insemination (embryos). Up to four recoveries were performed with each doe. Recovery efficiencies differed significantly between embryos (84%) and oocytes (58%). Yet, the recovery rates for the superovulation and control groups did not differ. The rhFSH group was associated with a significant increase (p < 0.05) in the number of oocytes and embryos recovered in comparison with the control group (10.2 ± 1.0 and 14.3 ± 1.2 vs 6.0 ± 2.7 and 8.4 ± 2.3 for oocytes and embryos, respectively). Results from this study indicate that repeated in vivo oocyte and embryo recovery from rhFSH superovulated does maximizes the number of oocytes or embryos collected from the same female.  相似文献   

3.
卵母细胞和胚胎冷冻技术可与超数排卵和胚胎移植等繁殖技术相结合,打破生理、地区和时间对母畜繁殖潜力的限制,对国际种源流通、优良家畜扩繁和濒危动物种质资源保存具有重要意义。然而,卵母细胞和胚胎中丰富的脂质在降温过程中容易引起膜结构损伤、内质网和线粒体损伤、脂质过氧化,降低其解冻后发育能力,这极大地限制了冷冻卵母细胞和胚胎的应用。众多研究表明,在冷冻前降低卵母细胞和胚胎的脂质含量有利于提高其解冻后的存活率、囊胚率和妊娠率。目前,常用的降脂方法包括3种,即离心极化脂滴后通过显微操作去除脂滴、添加化学物质促进脂质代谢以及通过调控脂质代谢相关基因的表达降低脂质含量。作者简要阐述了过高的脂质含量引起卵母细胞和胚胎冷冻损伤的机制,总结了离心去脂法与化学去脂法的原理、应用效果以及局限性,并探讨了通过调控脂质代谢相关基因的表达从而去脂的可能性,以期为开发更稳定的卵母细胞和胚胎冷冻技术提供一定的参考。  相似文献   

4.
Embryo transfer: its uses and recent developments   总被引：3，自引：0，他引：3  
The technique of embryo transfer is now routinely employed in several species and with several objectives. In laboratory animals the technique is used mainly in investigations of reproductive biology, whereas in human beings it is used to overcome specific forms of impaired fertility. Because embryo transfer combined with superovulation of the donor can significantly increase the female reproductive rate its greatest application to date has been in farm animals, in which it is widely used in both research and commercial production. Within the past few years there have been many advances in the techniques used in farm animals, particularly in the area of embryo manipulation. The supply of embryos can now be increased by repeated superovulation and by embryo bisection and there has been significant progress in in vitro fertilisation technology. Deep freeze (-196 degrees C) storage of embryos is now routine and may be combined with direct one-step thawing and removal of cryoprotectant. This technique allows the routine non-surgical transfer of embryos in the field. There is also evidence that a routine non-traumatic procedure for sexing embryos may soon be developed. Identical multiplets or clones have been produced by the microdissection of embryos and more recently by the transfer of nuclei from embryos into unfertilized oocytes. Transgenic animals have been produced by the microinjection of recombinant DNA into one of the pronuclei of single-cell unfertilized ova.  相似文献   

5.
Crossbred heifers (n = 103) were synchronized to estrus with prostaglandin (PGF2α) and superovulated with follicle stimulating hormone (FSH-P). Animals were ovariectomized every 12 hr after the PGF2α injection (n = 7 to 9/time) up to 108 hr to monitor the follicular, hormonal, and oocyte changes associated with follicular development and ovulation. Twenty-eight animals were implanted with Norgestomet implants 12 hr before PGF2α and ovariectomized at 72, 84, 96, and 108 hr post PGF2α injection to monitor effects of progesterone and suppression of the luteinizing hormone (LH) surge on oocyte maturation and quality. Follicular fluid was collected and analyzed for progesterone, estradiol, prolactin, and glycosaminoglycan content in conjunction with cumulus maturation and nuclear stage of oocyte maturation. Analysis of in vivo matured oocytes by in vitro fertilization was carried out at 60, 72, 84, and 96 hr post PGF2α and in vitro matured oocytes at 12 to 108 hr post PGF2α. No developmental changes in cumulus cells surrounding the oocyte of small follicles was noted (≤ 4 mm dia) indicating a static population. Medium (> 4 ≤ 8 mm) and large size (> 8 mm) follicles developed to the corona radiata and loose cumulus stages in animals in which an LH surge was detected but cumulus status remained primarily in the tight cumulus stage for animals without an LH surge. The estradiol-to-progesterone ratio for tight cumulus (TC), corona radiata (CR), and loose cumulus (LC) stages was 1.8 ± .1, 1.0 ± .1, and .4 ± .2, respectively (P < .01). Nuclear maturation of oocytes in small follicles from animals without a detectable LH surge seem to indicate early maturation (48 to 72 hr post PGF2α) in conjunction with a high percent of degenerate oocytes not seen in animals exhibiting an LH surge. Oocytes from medium size follicles matured to germinal vesicle breakdown (GVBD) and early meiosis (metaphase I; MI) stages of development in all treatments. Most oocytes were degenerate in Norgestomet-implanted animals. Oocytes from large follicles (> 8 mm dia) from animals exhibiting an LH surge were in MI and metaphase II (MII) stages (48 to 84 hr post PGF2α) in preparation of ovulation whereas oocytes from animals not exhibiting an LH surge had oocytes that early matured to MII (48 to 72 hr post PGF2α), later regressing to degenerate oocytes (84 to 108 hr). Follicular progesterone, estradiol, and prolactin increased with oocyte maturation, particularly in medium and large follicles. In vivo matured oocytes for fertilization (60, 72, 84, and 96 hr post PGF2α) were nude (from the oviduct) and primarily CR from follicles. Tubal oocytes (37%) were fertilized more frequently by a single sperm than follicular oocytes (14.3%; P < .01) and single sperm penetration peaked at 72 hr post PGF2α. Follicular hormone concentrations were not related to sperm penetration. Oocytes (n = 101) matured in vivo had lower fertilization potential from ovaries producing < 14 or > 50 follicles (39.3%) as compared to 21 to 45 aspirated follicles (68.2%; P < .05), with a peak penetration at 32 follicles (86.7% penetration). No treatment differences (LH surge or no detectable LH surge) were noted in relation to in vivo matured oocytes. Oocytes with single sperm penetration had the lowest estradiol/progesterone ratio of 2.2 vs polyspermic penetration of 13.7.  相似文献   

6.
超数排卵在动物胚胎工程技术的体内胚生产中起关键作用,不同品种甚至个体对超数排卵的反应不同。人绝经期促性腺激素(HMG)是从绝经期妇女尿液中提纯得到的具有促卵泡素(FSH)活性的蛋白质激素,在医学上广泛应用于诱导排卵和治疗不育不孕症。本试验以安格斯青年母牛为超排供体,采用分次肌肉注射激素方法进行超排处理,比较了HMG与FSH用于安格斯母牛超数排卵的效果。B超检查发现,HMG和FSH超排处理后牛只两侧卵巢上直径>1cm的卵泡数分别为10.33±0.52和9.25±0.46个,二者差异不显著(P>0.05),表明HMG与FSH同样安全有效;总剂量1 400IU的HMG或FSH用于安格斯母牛超排后B超检查卵巢没有发现异常变化,说明该剂量是用于安格斯母牛超数排卵的安全有效剂量。  相似文献   

7.
以昆明小鼠为实验动物,经过超数排卵后得到卵丘卵母细胞复合体(COCs),以卵丘细胞为核供体细胞,卵母细胞为核受体细胞进行核移植,分析比较了不同蔗糖浓度对小鼠卵母细胞去核时间、去核率及重构胚体外发育的影响,并探讨了供体细胞的保存温度和融合温度对重构胚体外发育的影响.结果表明,各蔗糖浓度组的去核率均达到100%,但3%、4...  相似文献   

8.
Plasma progesterone- and LH concentrations were studied in lactating dairy cows around the superovulatory heat. The pattern of both hormones accurately reflected the donor animals' endocrine balance and could be used to evaluate and predict the subsequent potentials or quality of a given animal as embryo- and oocyte donor. It was the general impression that both PMSG- and FSH treatment caused endocrine changes in progesterone- and LH patterns that would account for improper oocyte- and embryo development.  相似文献   

9.
The aims of this study were to evaluate the chronology of periovulatory events (oestrus behaviour, LH surge and ovulation) in 16 superovulated Manchega sheep and to determine whether follicular status at start of the FSH supply might affect their occurrence. Mean timing for onset of oestrus behaviour was detected at 28.1 +/- 0.7 h after sponge withdrawal; the preovulatory LH surge and ovulation started at 37.2 +/- 0.7 h and 65.4 +/- 0.7 h after progestagen withdrawal, respectively. The intervals between oestrus, LH surge and ovulation were affected by a high individual variability, which might be the cause for reported decreased efficiency in embryo production. Current results also addressed the role of follicular status at start of the superovulatory treatment on the preovulatory LH surge and the ovulation. The interval LH surge-ovulation was increased in ewes with a growing dominant follicle at starting the FSH treatment (32.3 +/- 0.9 vs 28.6 +/- 0.5 h, p < 0.05). The developmental stage of the largest follicle at starting the superovulatory treatment also affected occurrence of LH surge and ovulation; follicles in growing phase advanced the occurrence of the LH surge and ovulation when compared to decreasing follicles (33.0 +/- 1.0 vs 43.5 +/- 1.1 h, p < 0.05, for LH peak and 60.7 +/- 1.1 vs 72.8 +/- 1.2 h, p < 0.05, for ovulation). Thus, only ewes with growing follicles ovulated prior to 55 h after sponge withdrawal; conversely, no sheep with decreasing follicles ovulated earlier than 67 h, when an 85.7% of the ewes bearing growing follicles has ovulated at 63 h.  相似文献   

10.
The aim of this study was to investigate whether plasma anti-Muellerian hormone (AMH) levels of Holstein-Friesian heifers could be used to predict ovum pick-up (OPU) and embryo production outcomes. Plasma samples and data were collected from 64 heifers, which underwent repeated OPU with subsequent in vitro embryo production followed by embryo flushing after superovulation. AMH levels were significantly positively correlated with the number of follicles aspirated per OPU session (r = 0.45), recovered oocytes per OPU (r =0.43) and in vitro produced embryos per OPU (r = 0.28). No significant correlations between AMH and in vivo produced embryos were ascertained. Our results suggest that correlations between AMH and outcomes of an OPU-IVF program are too low to use AMH as a precise predictive parameter for the success of a particular OPU procedure in Holstein-Friesian heifers. However, AMH can help to identify groups of very good or very poor oocyte donors.  相似文献   

11.
The objective of this study was to determine the relationship between the number of transferable embryos (TE) and various blood chemistry parameters as a reflection of the metabolic state of cows after superovulatory treatment. Forty-nine Holstein cows were subjected to superovulatory treatment for commercial embryo production. At the time of embryo harvest, individual blood samples were taken from cows for biochemical analysis. All embryos including dead ones as well as non-fertilized oocytes were counted in uterine lavage. Feed samples collected daily for a period of two weeks before embryo harvest, were analyzed for mycotoxins: vomitoxin, zearalenone and T-2 toxin. On average, cows produced 9.45 ± 5.60 embryos and oocytes of which 5.27 ± 4.20 were TE, 0.37 ± 0.80 were dead embryos and 3.82 ± 3.78 were non-fertilized oocytes. Higher concentrations of Mg and K were associated with a higher production of TE (p = 0.005 and p = 0.043, respectively) and higher activity of creatinine kinase was associated with a lower production of TE (p = 0.011).  相似文献   

12.
In vitro oocyte maturation can be influenced by oocyte source and maturation media composition. The aim of the present study was to compare the efficiency of a defined in vitro maturation medium (TCM199 supplemented with cysteamine and epidermal growth factor; Cys + EGF) with an undefined medium (TCM199 supplemented with follicle-stimulating hormone and follicular fluid; FSH + FF) for in vitro production (IVP) of ovine embryos, using oocytes obtained by laparoscopic ovum pick-up from FSH-stimulated [n=11; 158 cumulus-oocyte complexes (COCs)] and non-stimulated (n=16; 120 COCs) live ewes, as well as abattoir-derived oocytes (170 COCs). The produced blastocysts were vitrified and some of them were transferred to synchronized recipients. The best and the worst final yields of embryo IVP observed in this study were obtained using oocytes from FSH-stimulated ewes matured in FSH + FF (41.3%; 33/80) and in Cys + EGF (19.2%; 15/78) medium, respectively (p<0.01). No significant differences between both media were attained in the blastocyst development rate or in the final yield of embryo IVP using oocytes from non-stimulated ewes or abattoir-derived oocytes. The overall in vivo survival rate of the transferred vitrified blastocysts was 13.1% (8/61), without significant differences between oocyte sources or maturation media. In conclusion, under the experimental conditions of the present study, TCM199 supplemented with cysteamine and EGF is a convenient defined maturation medium for IVP of embryos from oocytes of live non-stimulated ewes or from oocytes of abattoir-derived ovaries. However, the best final yield of embryo IVP observed in this study was attained when oocytes came from FSH-stimulated donors and TCM199 was supplemented with FSH and follicular fluid.  相似文献   

13.
To investigate the impact of porcine oocytes in vivo and in vitro maturation (IVM) on the development of porcine somatic cell cloned embryos,the somatic cell cloned embryos cultured in vitro and the sows were treated with hormones to collect mature oocytes in vivo,and the cleavage rate, blastocyst rate and embryo implantation were compared. The results showed that the average number of ovulation in PGC+PMSG+HCG group was significantly higher than that of PGC+HCG,PMSG+HCG and the natural estrus groups (P<0.05). The oocytes collected in vivo could be used for the construction, and the available oocytes rate reached more than 90%,and there was no significant difference among the four groups (P>0.05),which indicated that groups treated by hormone could obtain more available oocytes and the quality of oocytes was not significant different. In vivo and in vitro matured oocytes were used as nuclear transfer embryos of recombinant receptor,the fusion efficiency (80.31% and 79.29%) and cleavage rate (90.40% and 86.51%) were not significant different (P>0.05), but the proportion of in vivo matured oocytes cloned embryos developed into the blastocyst stage was significantly higher (P<0.05). The reconstructed embryos made from in vivo and in vitro matured oocytes were transplanted into surrogate sows (transferred 30 or 60 embryos),10 piglets were born in in vivo maturation of cloned embryo transfer group,while there was no implantation in in vitro maturation of cloned embryo transfer group. The results showed that high quality oocytes obtained by superovulation could significantly increase the blastocyst rate of embryos,reduce the number of embryos transferred and improve the pregnancy rate of surrogate sows.  相似文献   

14.
Ceftiofur sodium is a third-generation cephalosporin antibiotic with broad spectrum bactericidal activity against Gram-positive and Gram-negative bacteria including the beta-lactamase producing strains. In this study, we use in vitro techniques to examine the effects of low and high levels of ceftiofur sodium on the development of bovine oocytes/embryos during oocyte maturation, oocyte fertilization and embryo culture. A total of 8590 oocytes was used in six independent experiments, each in a randomized complete block design. Each replication within each experiment consisted of oocytes from the same abattoir collection of ovaries. There was no difference in embryo development when oocytes were exposed to ceftiofur sodium during oocyte maturation or fertilization at low (10 and 50 μg/mL) or high (100 and 200 μg/mL) concentrations. However, when fertilized oocytes were exposed to concentrations 50 μg/mL during culture, ceftiofur sodium significantly retarded embryo development (e.g. the numbers of ova developing to the morula and blastocyst stages were reduced, and a large proportion of embryos were blocked at the 8-cell stage). We conclude that ceftiofur sodium does not appear to have detrimental effects on oocyte maturation and fertilization. However, long term exposure to high dosages of ceftiofur sodium during post-fertilization culture adversely affects embryo development in vitro.  相似文献   

15.
Melatonin (MLT) is an endogenous hormone with roles in animal germ cell development. However, the effect of MLT on porcine oocyte maturation and its underlying mechanisms remain largely unknown. Here, we investigated the effects of exogenous MLT on oocyte maturation, histone acetylation, autophagy and subsequent embryonic development. We found that 1 nmol/L MLT supplemented in maturation medium was the optimal concentration to promote porcine oocyte maturation and subsequent developmental competence and quality of parthenogenetic embryos. Interestingly, the beneficial effects of 1 nmol/L MLT treatment on porcine oocyte maturation and embryo development were mainly attributed to the first half period of in vitro maturation. Simultaneously, MLT treatment could also improve maturation of small follicle‐derived oocytes, morphologically poor (cumulus cell layer ≤1) and even artificially denuded oocytes and their subsequent embryo development. Furthermore, MLT treatment not only could decrease the levels of H3K27ac and H4K16ac in metaphase II (MII) oocytes, but also could increase the expression abundances of genes associated with cumulus cell expansion, meiotic maturation, histone acetylation and autophagy in cumulus cells or MII oocytes. These results indicate that MLT treatment can facilitate porcine oocyte maturation and subsequent embryonic development probably, through improvements in histone acetylation and autophagy in oocytes.  相似文献   

16.
为讨论羔羊自身因素与其超排后卵母细胞数量的关系,选取6～8周龄的健康杜晋绵羊（杜泊羊（o）×晋中绵羊♀）母羔31只,进行超数排卵,手术法收集卵母细胞,统计其卵母细胞数量,分析其与全同胞羔羊数、初生重、超排时体重的相关性.结果表明：全同胞羔羊数与卵母细胞数量呈极显著负相关（P＜0.01）,初生重与卵母细胞数量呈极显著正相关（P＜0.01）;超排时体重与卵母细胞数量相关不显著（P＞0.05）.全同胞羔羊数和初生重是羔羊超排卵母细胞数量的影响因素,但仍需进一步增加羔羊数量来验证.  相似文献   

17.
This communication describes the in vitro maturation and in vitro fertilization of bovine follicular oocytes. Further development of the embryos was achieved by using a granulosa cell culture system. The in vitro development of oocytes to morula/blastocyst stages obtained from individual cows was compared to the results of pooled simultaneous cultured oocytes and to our over-all results of this method. While there were no statistical differences in the developmental rates between these three groups (individual cows: 28.1%, simultaneous pool: 34.0%, over-all results: 32.7%) marked differences were found between the 22 animals investigated separately. These results indicate that there were great individual variabilities due to the oocyte population comparable to the variations in ovarian response to superovulation.  相似文献   

18.
To expand the breeding flock of Poll Dorset sheep in China, multiple ovulation and embryo transfer breeding program was applied to the limited number of imported Australian Poll Dorset sheep. This study investigated the effects of FSH from three different manufacturers, parity (nulliparous vs multiparous), repeated superovulation, oestrus induction, corpus luteum regression and oestrus delay on Poll Dorset superovulation. The results showed that gonadotropin FSH from Canada Folltropin‐V (Ca‐FSH) was successfully used for superovulatory treatment with 160 mg–200 mg dosage per ewe and recovered 12.91 ± 7.80 embryos. Multiparous ewes for superovulation treatment were significantly better nulliparous ewes (p < 0.05). The successive superovalution treatment reduced significantly embryo collection but did not affect transferable embryo number. Ewes with natural oestrus resulted in significantly higher number of embryos (13.83 ± 4.64) and of transferable embryos (12.00 ± 5.76) than ewes with induced oestrus (7.00 ± 4.92; 4.22 ± 3.42) and unknown oestrus cycle (5.94 ± 3.38; 3.19 ± 2.56, p < 0.05). The delayed oestrus ewes at 24 h after superovulatory treatment produced significantly fewer embryos and transferable embryos (0.92 ± 1.51 vs 0.42 ± 0.90) than those with normal oestrus (p < 0.01). Furthermore, the more transferable embryos were recovered from ewes with normal corpus luteum than those with corpus luteum regression (5.88 ± 5.09 vs 3.59 ± 4.30 and 8.83 ± 5.75 vs. 6.66 ± 5.41, p < 0.01). These results suggest that in our farm practice, a comprehensive treatment method by using the Canadian FSH (Folltropin‐V), plus choosing multiparous and natural oestrus ewes with normal corpus luteum might obtain an optimum embryo collection and embryos transfer in sheep.  相似文献   

19.
Eight cows were used to study the feasibility of transvaginal ultrasound-guided puncture of follicles as a method for the collection of immature oocytes for embryo production in vitro. In six trials at intervals of seven days, 104 oocytes were collected. After in vitro maturation and fertilisation the 104 oocytes were transferred to the oviducts of sheep. Six days later, 75 oocytes were recovered by flushing the oviducts. Twenty-four per cent of the recovered oocytes/embryos had developed into transferable and viable morulae and, or, blastocysts. The data show that this non-surgical and repeated collection of immature oocytes can be used successfully for the in vitro production of bovine embryos. The procedure may produce yields of embryos comparable to those obtainable by conventional superovulation procedures.  相似文献   

20.
Assisted reproduction technologies (ART) are well developed in humans and cattle and are gaining momentum also in the equine industry because of the fact that the mare does not respond to superovulation but can donate large numbers of oocytes through ovum pick up (OPU). After collection, the oocytes can be fertilized by intracytoplasmic sperm injection (ICSI) using a variety of stallion semen samples, even of poor quality, and the resulting embryos can establish high pregnancy rates after cryopreservation and transfer. The discoveries that equine oocytes can be held at room temperature without loss of viability and that an increase in vitro maturation time can double the number of embryos produced are fueling the uptake of the OPU technique by several clinics that are shipping oocytes of their client’s mares to specialized ICSI laboratories for embryo production and freezing. In this article, we present a retrospective analysis of 10 years of work at Avantea with a special focus on the last 3 years. Based on our data, an average production of 1.7 to 2 embryos per OPU-ICSI procedure can be obtained from warmblood donor mares with a pregnancy rate of 70% and a foaling rate in excess of 50%. OPU-ICSI offers the added value of freezing embryos that allows the development of embryo commercialization worldwide to the benefit of top horse breeders who are endorsing this technology as never before.  相似文献   

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