Mammalian gametes and embryos are particularly vulnerable to oxidative stress-induced damage, which is mainly caused by reactive oxygen species（ROS） originating from normal metabolism and/or the external environment...Mammalian gametes and embryos are particularly vulnerable to oxidative stress-induced damage, which is mainly caused by reactive oxygen species（ROS） originating from normal metabolism and/or the external environment. Several researchers have implicated the role of oxidative stress in the activation of apoptosis, causing peroxidative damage to sperms/oocytes and inducing embryo fragmentation, arrest, or demise. Melatonin is a tryptophan derivative that is known for its powerful free radical-scavenging activity and broad-spectrum antioxidant property. Numerous studies have shown that melatonin and its metabolic derivatives can sequentially detoxify ROS in an antioxidant cascade, and modulate various antioxidant enzymes via its receptors to prevent radical-mediated damage. The identification of melatonin receptors in cumulus/granulosa cells, oocytes, and epididymal tissues implies that melatonin has protective actions on gametes and embryos. Enriching the semen extender or culture medium with melatonin significantly benefits sperm characteristics, improves oocyte maturation potential and quality, and enhances the developmental competence of preimplantation embryos. Certainly, further comparative studies are needed to show the unique antioxidant role and the advantage of melatonin in this field. This review summarizes the harmful effects of ROS and the beneficial role of melatonin against oxidative damage of gametes and embryos.展开更多
To our knowledge,no single study has systemically compared cryopreservation efficiencies of bovine blastocysts derived from in vitro fertilization （IVF）,intracytoplasmic sperm injection （ICSI） and somatic cell nuc...To our knowledge,no single study has systemically compared cryopreservation efficiencies of bovine blastocysts derived from in vitro fertilization （IVF）,intracytoplasmic sperm injection （ICSI） and somatic cell nuclear transfer （SCNT） by controlled freezing and vitrification.This experiment,therefore,was designed to compare the cryopreservation of these blastocysts with controlled freezing and OPS vitrification.Adenosine-5’-triphosphate （ATP） content and reactive oxygen species （ROS） level in blastocysts were also analyzed.Firstly,for each type of blastocyst （IVF,ICSI or SCNT）,significant differences were observed between the survival rates of the controlled freezing （（81.56±2.33）,（68.18±4.72） or （47.89±5.83）%） and OPS vitrification groups （（92.24±4.54）,（82.40±3.76） or （78.71±5.91）%;P〈0.05）.Secondly,for each type of blastocyst （IVF,ICSI or SCNT）,ATP content was significantly decreased after controlled freezing or vitrification,and the ATP content in the controlled freezing group （0.43±0.06）,（0.35±0.05） or （0.21±0.02） pmol） was significantly lower than that found in the OPS vitrification group （0.62±0.04）,（0.46±0.03） or （0.30±0.01） pmol;P〈0.05）.Thirdly,ROS level in fresh IVF （（47.33±3.56） c.p.s （counted photons per second）,ICSI （（36.51±2.58） c.p.s） or SCNT blastocysts （（26.44±1.49） c.p.s） was significantly lower than that found in the OPS vitrification group （（72.14±4.31）,（58.89±3.89） or （40.11±5.73） c.p.s;P〈0.05）,but higher than that of the controlled freezing group （34.41±3.32）,（23.13±1.26） or （15.46±2.45） c.p.s;P〈0.05）.The present study indicated that vitrification is more efficient in the cryopreservation of bovine blastocysts derived from IVF,ICSI or SCNT than controlled freezing.Furthermore,both vitrification and controlled freezing significantly altered the ATP content and ROS level in those blastocysts.展开更多
The antioxidant of reduced glutathione（GSH） is the most abundant thiol in cells for the maintenance of the intracellular redox balance. The study aimed to assay the effect of sperm treatment with GSH before incubati...The antioxidant of reduced glutathione（GSH） is the most abundant thiol in cells for the maintenance of the intracellular redox balance. The study aimed to assay the effect of sperm treatment with GSH before incubation with oocytes on the development potential of embryos obtained by in vitro fertilization（IVF）. Also the mitochondrial membrane potential（ΔΨm）, plasma membrane integrity（viability）, DNA fragmentation, reactive oxygen species（ROS） content, superoxide dismutase（SOD）, catalase（CAT） and glutathione peroxidase（GPx） activities, methane dicarboxylic aldehyde（MDA） level as indices of lipid peroxidation in sex-sorted and unsorted sperm from three bulls were investigated using flow cytometry and enzyme-labeled instrument individually. The results showed that 2 mm ol L^–1 GSH increased significantly the cleavage rate（86.68% vs. 82.78%）, 4- to 8-cell rate（82.30% vs. 73.43%） and blastocyst rate（43.15% vs. 35.24%） of IVF embryos compared with untreated group. Furthermore, addition of GSH increased significantly the ΔΨm and viability, decreased the ratio of DNA fragmentation in sex-sorted or unsorted semen（P〈0.05）, except the sex-sorted semen from bull 019. Similarly, activities of SOD, CAT and GPx were increased significantly. However, the contents of MDA were decreased significantly both in sex-sorted and unsorted semen treated with GSH（P〈0.05）. These results suggest that sperm pretreatment with GSH during IVF can maintain better the viability and fertility of sperm through reducing apoptosis and increasing the antioxidant capacity, which improves the IVF embryos development.展开更多
The objective of this study was to establish an efficient system of producing early monozygotic twin bovine embryos in vitro using the blastomere separation and coculture technique. In this study, early eight-cell emb...The objective of this study was to establish an efficient system of producing early monozygotic twin bovine embryos in vitro using the blastomere separation and coculture technique. In this study, early eight-cell embryos were chosen to optimize the separation method, and multi-coculture tactics were applied to improve the efficiency of this production system. Bovine embryo blastomeres（groups of at least 30 at the eight-cell stage） were separated into eight segments（to regard an eight-cell embryo as a tangerine, a blastomere as one segment） and one, two and four segments（blastomeres） were cultured respectively in microwells on the bottom of the four-well dish（Nunc, Denmark） with 400 μL of culture medium under paraffin oil. Four different types of coculture tactics（cocultured with nothing, intact embryos, bovine cumulus cells（b CCs）, intact embryos ＆ b CCs） were applied to the group of four segments（blastomeres）. Finally, diameter and inner cell mass（ICM）：trophectoderm（TE） cell ratio was measured as a criterion to assess the quality of the twin embryos which were derived from bovine separated blastomeres. Our results showed that rate of blastocyst formation of the four segments group was significantly greater than one or two group（P〈0.05）. In addition, rate of blastocyst formation was significantly increased when the four segments were cocultured with intact embryo ＆ b CCs（P〈0.05）. Although the ICM, TE and total cells of blastocysts derived from separated blastomeres was less than the control group from intact embryo（P〈0.05）, more important quality indicator of the blastocyst diameter and ICM：TE cell ratio was similar between our experimental group and the control group（P〉0.05）. Thus, these results suggest that combined with intact embryos ＆ b CCs coculture system, culturing four isolated segments（blastomeres） per microwell is an efficient system of producing early monozygotic twin bovine embryos. Furthermore, our results also indicate that the quality of blastocysts derived from separated blastomere may be similar to those derived from intact eight-cell embryos.展开更多
基金supported by the grants from the Agricultural Science and Technology Innovation Program, China (ASTIPIAS06)the earmarked fund for China Agriculture Research System (CARS-36)
文摘Mammalian gametes and embryos are particularly vulnerable to oxidative stress-induced damage, which is mainly caused by reactive oxygen species（ROS） originating from normal metabolism and/or the external environment. Several researchers have implicated the role of oxidative stress in the activation of apoptosis, causing peroxidative damage to sperms/oocytes and inducing embryo fragmentation, arrest, or demise. Melatonin is a tryptophan derivative that is known for its powerful free radical-scavenging activity and broad-spectrum antioxidant property. Numerous studies have shown that melatonin and its metabolic derivatives can sequentially detoxify ROS in an antioxidant cascade, and modulate various antioxidant enzymes via its receptors to prevent radical-mediated damage. The identification of melatonin receptors in cumulus/granulosa cells, oocytes, and epididymal tissues implies that melatonin has protective actions on gametes and embryos. Enriching the semen extender or culture medium with melatonin significantly benefits sperm characteristics, improves oocyte maturation potential and quality, and enhances the developmental competence of preimplantation embryos. Certainly, further comparative studies are needed to show the unique antioxidant role and the advantage of melatonin in this field. This review summarizes the harmful effects of ROS and the beneficial role of melatonin against oxidative damage of gametes and embryos.
基金supported by the Fund of China Agriculture Research System(CARS-37)the Genetically Modified Organisms Breeding Major Projects of China(2009ZX08011-031B)+1 种基金the Basic Research Fund of IAS,CAAS(2010jc-3-1)the National Natural Science Foundation of China(31001011)
文摘To our knowledge,no single study has systemically compared cryopreservation efficiencies of bovine blastocysts derived from in vitro fertilization （IVF）,intracytoplasmic sperm injection （ICSI） and somatic cell nuclear transfer （SCNT） by controlled freezing and vitrification.This experiment,therefore,was designed to compare the cryopreservation of these blastocysts with controlled freezing and OPS vitrification.Adenosine-5’-triphosphate （ATP） content and reactive oxygen species （ROS） level in blastocysts were also analyzed.Firstly,for each type of blastocyst （IVF,ICSI or SCNT）,significant differences were observed between the survival rates of the controlled freezing （（81.56±2.33）,（68.18±4.72） or （47.89±5.83）%） and OPS vitrification groups （（92.24±4.54）,（82.40±3.76） or （78.71±5.91）%;P〈0.05）.Secondly,for each type of blastocyst （IVF,ICSI or SCNT）,ATP content was significantly decreased after controlled freezing or vitrification,and the ATP content in the controlled freezing group （0.43±0.06）,（0.35±0.05） or （0.21±0.02） pmol） was significantly lower than that found in the OPS vitrification group （0.62±0.04）,（0.46±0.03） or （0.30±0.01） pmol;P〈0.05）.Thirdly,ROS level in fresh IVF （（47.33±3.56） c.p.s （counted photons per second）,ICSI （（36.51±2.58） c.p.s） or SCNT blastocysts （（26.44±1.49） c.p.s） was significantly lower than that found in the OPS vitrification group （（72.14±4.31）,（58.89±3.89） or （40.11±5.73） c.p.s;P〈0.05）,but higher than that of the controlled freezing group （34.41±3.32）,（23.13±1.26） or （15.46±2.45） c.p.s;P〈0.05）.The present study indicated that vitrification is more efficient in the cryopreservation of bovine blastocysts derived from IVF,ICSI or SCNT than controlled freezing.Furthermore,both vitrification and controlled freezing significantly altered the ATP content and ROS level in those blastocysts.
基金supported by the grants from the National Science and Technology Support Program of China (2012BAD12B01)the Basic Research Fund of Institute of Animal Science, Chinese Academy of Agricultural Sciences (2013ywf-zd-2)+1 种基金the Agricultural Science and Technology Innovation Program, China (ASTIP-IAS06)the China Agriculture Research System (CARS-37)
文摘The antioxidant of reduced glutathione（GSH） is the most abundant thiol in cells for the maintenance of the intracellular redox balance. The study aimed to assay the effect of sperm treatment with GSH before incubation with oocytes on the development potential of embryos obtained by in vitro fertilization（IVF）. Also the mitochondrial membrane potential（ΔΨm）, plasma membrane integrity（viability）, DNA fragmentation, reactive oxygen species（ROS） content, superoxide dismutase（SOD）, catalase（CAT） and glutathione peroxidase（GPx） activities, methane dicarboxylic aldehyde（MDA） level as indices of lipid peroxidation in sex-sorted and unsorted sperm from three bulls were investigated using flow cytometry and enzyme-labeled instrument individually. The results showed that 2 mm ol L^–1 GSH increased significantly the cleavage rate（86.68% vs. 82.78%）, 4- to 8-cell rate（82.30% vs. 73.43%） and blastocyst rate（43.15% vs. 35.24%） of IVF embryos compared with untreated group. Furthermore, addition of GSH increased significantly the ΔΨm and viability, decreased the ratio of DNA fragmentation in sex-sorted or unsorted semen（P〈0.05）, except the sex-sorted semen from bull 019. Similarly, activities of SOD, CAT and GPx were increased significantly. However, the contents of MDA were decreased significantly both in sex-sorted and unsorted semen treated with GSH（P〈0.05）. These results suggest that sperm pretreatment with GSH during IVF can maintain better the viability and fertility of sperm through reducing apoptosis and increasing the antioxidant capacity, which improves the IVF embryos development.
基金supported by the China Agriculture Research System (CARS-37)the Agricultural Science and Technology Innovation Program, China (ASTIP-IAS06-2015)
文摘The objective of this study was to establish an efficient system of producing early monozygotic twin bovine embryos in vitro using the blastomere separation and coculture technique. In this study, early eight-cell embryos were chosen to optimize the separation method, and multi-coculture tactics were applied to improve the efficiency of this production system. Bovine embryo blastomeres（groups of at least 30 at the eight-cell stage） were separated into eight segments（to regard an eight-cell embryo as a tangerine, a blastomere as one segment） and one, two and four segments（blastomeres） were cultured respectively in microwells on the bottom of the four-well dish（Nunc, Denmark） with 400 μL of culture medium under paraffin oil. Four different types of coculture tactics（cocultured with nothing, intact embryos, bovine cumulus cells（b CCs）, intact embryos ＆ b CCs） were applied to the group of four segments（blastomeres）. Finally, diameter and inner cell mass（ICM）：trophectoderm（TE） cell ratio was measured as a criterion to assess the quality of the twin embryos which were derived from bovine separated blastomeres. Our results showed that rate of blastocyst formation of the four segments group was significantly greater than one or two group（P〈0.05）. In addition, rate of blastocyst formation was significantly increased when the four segments were cocultured with intact embryo ＆ b CCs（P〈0.05）. Although the ICM, TE and total cells of blastocysts derived from separated blastomeres was less than the control group from intact embryo（P〈0.05）, more important quality indicator of the blastocyst diameter and ICM：TE cell ratio was similar between our experimental group and the control group（P〉0.05）. Thus, these results suggest that combined with intact embryos ＆ b CCs coculture system, culturing four isolated segments（blastomeres） per microwell is an efficient system of producing early monozygotic twin bovine embryos. Furthermore, our results also indicate that the quality of blastocysts derived from separated blastomere may be similar to those derived from intact eight-cell embryos.
