Relationship between equilibration times and the presence of cumulus cells, and effect of Taxol treatment for vitrification of in vitro matured porcine oocytes

Fujihira, Takuma; Nagai, Hiroki; Fukui, Yutaka

doi:10.1016/j.cryobiol.2005.08.002

Cited by 48 publications

(35 citation statements)

References 9 publications

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“…Bogliolo et al (2007) also recently concluded that cumulus cells are not essential to the preservation of meiotic competence during vitrification; however, they did so after finding that immature ovine oocytes survived cryopreservation better if they had previously been denuded. This discrepancy between an apparently protective effect of cumulus cells in our study and a detrimental effect in Bogliolo et al (2007)'s study could relate either to a damaging effect of the large number of dead cumulus cells surrounding Bogliolo et al's vitrified-warmed oocytes or to a non-specific effect of cumulus enclosure in modifying the optimal time of exposure to the cryprotectants, as postulated for MII porcine oocytes by Fujihira et al (2005). That is, in Cumulus protection of horse oocytes during vitrification our study, the cumulus cells may have prevented overexposure to CPA, whereas in Bogliolo et al's study, they may have precluded adequate exposure.…”

Section: Discussioncontrasting

confidence: 39%

“…Nevertheless, the potential protective role of the cumulus during the vitrification of MII oocytes remains controversial, not least because some studies still record better results for cumulus-enclosed than denuded human vitrified oocytes (e.g. Kuwayama et al 2005); there may also be between-species differences since there are reports of reduced oocyte viability and developmental competence in cattle oocytes vitrified with an intact cumulus (Chian et al 2004), no effect of cumulus presence during vitrification of sheep oocytes ) and either a protective effect of the cumulus ( Varga et al 2006) or an effect of cumulus presence on the optimal time for exposing MII oocytes to CPA in pigs ( Fujihira et al 2005). Since there are considerable apparent between-species and study differences, it was appropriate to examine the effect of vitrifying MII equine oocytes with or without their cumulus investment on post-warming meiotic spindle quality.…”

Section: Discussionmentioning

confidence: 99%

“…Although we are unable to elaborate on the precise nature of these damaging effects, it is possible that the apparent need for a protective cumulus barrier during vitrification of equine MII oocytes could relate to the relatively high concentrations of CPAs used (20% ethylene glycol (EG), 20% DMSO, 0.5 M sucrose) compared with current protocols for vitrifying human oocytes (e.g. 15% EG, 15% DMSO, 0.5 M sucrose: Cobo et al 2008) and that it might be negated by reducing the concentrations of, or time of exposure to, the CPAs as reported for porcine oocytes ( Fujihira et al 2005) or by altering the temperature at which CPA exposure takes place. On the other hand, damage to the MII oocytes in the current study appeared to be induced primarily by vitrification and warming per se rather than CPA toxicity since exposing denuded oocytes to CPA without vitrification did not affect meiotic spindle quality.…”

Section: Discussionmentioning

confidence: 99%

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Protective effects of the cumulus-corona radiata complex during vitrification of horse oocytes

Tharasanit

Colleoni²,

Galli³

et al. 2009

Reproduction

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Vitrifying oocytes is a potentially valuable means of preserving the female germ line, but significantly compromises oocyte developmental competence. This study examined the hypothesis that the cumulus complex protects the oocyte during vitrification. Vitrified-warmed immature cumulus oocyte complexes (COCs) were labelled with a plasma membrane impermeant DNA marker (ethidium homodimer-1) to examine the percentage and location of dead cumulus cells, and to investigate the effect of the proportion of dead cells (C1,C2 or C3) on the success of in vitro maturation (IVM). Further, oocytes were labelled for connexin-43 or injected with Lucifer yellow dye to determine whether the integrity of the gap junctions between an oocyte and its cumulus was compromised by vitrification. Finally, the effect of denuding immature and mature oocytes on their ability to withstand vitrification was examined. Cryopreserving immature COCs increased the number of dead cumulus cells (13 vs 2.6% for controls; P!0.05). However, an increased proportion of dead cumulus cells did not affect post-warming maturation rates (w30% MII ) presumably because dead cells were located at the periphery of the cumulus mass and cumulus-oocyte gap junction communication was not disrupted. Moreover, cumulus removal prior to IVM or vitrification indicated that while the cumulus does protect immature oocytes during vitrification it does so by mechanisms other than support during maturation. Cumulus presence was also found to protect mature equine oocytes against vitrification-induced damage since cumulus-enclosed MII oocytes preserved their meiotic spindle quality better during vitrification than denuded oocytes (38.1 vs 3.1% normal spindles; P!0.05).

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Section: Discussioncontrasting

confidence: 39%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Protective effects of the cumulus-corona radiata complex during vitrification of horse oocytes

Tharasanit

Colleoni²,

Galli³

et al. 2009

Reproduction

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“…Cryopreservation of oocytes was carried out by vitrification using the minimum volume cooling (MVC) method [24,25]. All solutions used during vitrification and re-warming were prepared with a basal medium composed of TCM199 (Nissui Pharmaceutical) containing 20 mM Hepes, 4.2 mM NaHCO3, 75 μg/ml potassium penicillin G, and 50 μg/ml streptomycin sulfate.…”

Section: Vitrification Of MII Stage Oocytesmentioning

confidence: 99%

Developmental Ability of Porcine In Vitro Matured Oocytes at the Meiosis II Stage After Vitrification

Ogawa

Ueno

Nakayama

et al. 2010

Journal of Reproduction and Development

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Abstract. The aim of the present study was to investigate whether a combination of cytoplasmic lipid removal (delipation) and treatment by a microtubule stabilizer, paclitaxel, would lead to efficient cryopreservation of porcine in vitro matured (IVM) oocytes at the meiosis II (MII) stage. Vitrification and subsequent re-warming and culture of 109 untreated oocytes produced only 9 blastocysts (8.3%). On the other hand, the post-vitrification blastocyst rate was significantly improved (21/113, 18.6%, P<0.05) when oocytes were treated with 1 μM paclitaxel. Oocyte delipation also significantly increased the post-vitrification blastocyst rate compared with the untreated group (15/37, 40.5%, P<0.05). The delipation-and-paclitaxel group exhibited a significantly higher blastocyst rate (34/75, 45.3%, P<0.05) than the paclitaxel group, although it was not significantly higher than that for the delipation group. In transfer experiment, a total of 109 (18.6%) parthenogenetic blastocysts were obtained from 586 oocytes vitrified with the delipation-andpaclitaxel treatment. Transfer of 72 blastocysts to two recipients resulted in 14 (19.4%) somite stage fetuses. In conclusion, we demonstrated for the first time that by removing cytoplasmic lipid droplets from oocytes and performing a microtubule stabilization procedure, vitrified porcine IVM MII-stage oocytes could efficiently develop to the blastocyst stage while retaining the ability to develop into fetuses. Key words: Cryopreservation, Delipation, In vitro matured oocytes, Pig, Vitrification (J. Reprod. Dev. 56: [356][357][358][359][360][361] 2010) ryopreservation techniques for mammalian oocytes and embryos have been used for three main purposes: to preserve the genes of elite livestock animals and increase the efficiency of animal breeding, preserve valuable genetically modified animals and endangered species and use germ cells effectively in assisted reproductive technology. Thus far, embryo cryopreservation as a practical technique has been used in a variety of species, including experimental and livestock animals [1,2]. On the other hand, cryopreservation of oocytes remains impractical compared with embryos, and successful production of progeny from cryopreserved oocytes has been reported in limited species, including rabbits [3] [8] and rats [9]. As with porcine embryos, unfertilized porcine oocytes are highly sensitive to low temperature [10]. Consequently, cryopreserved porcine oocytes have yet to be used to successfully produce viable piglets.One of the reasons for the low-temperature sensitivity of porcine oocytes and embryos is their high intracellular lipid content. Nagashima and colleagues showed that the freezing tolerance of porcine oocytes [11] and early-stage embryos [12,13] can be dramatically increased by removing their cytoplasmic lipid droplets using a process called delipation.It is known that the meiotic spindle of the oocyte is extremely cryosensitive, and the impaired development of oocytes at meiosis II after cryopreservation has been attribut...

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“…We pioneered use of the Cryotop for vitrification of pronuclear rat embryos at a high success rate [8]. To date, Cryotop methods have been used for vitrification of oocytes and/or embryos in various other species including rabbits [11], humans [20], cattle [21], minke whales [22], pigs [23], buffalo [24], cats [25], horses [26] and sheep [27,28]. These reports strongly suggest that the Cryotop is one of the most powerful devices for vitrification of mammalian oocytes and embryos.…”

mentioning

confidence: 99%

Ethylene Glycol-supplemented Calcium-free Media Improve Zona Penetration of Vitrified Rat Oocytes by Sperm Cells

et al. 2010

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Abstract. Cryopreservation of matured oocytes is a useful technique because the oocytes can be used for some assisted reproductive technologies after warming. Even though rats, like mice, have been used in various research fields including reproductive technology, information about cryopreservation of rat oocytes is limited. The objective of the present study was to improve the vitrification protocol for matured rat oocytes. To determine the optimal equilibration time, oocytes were equilibrated in 7.5% ethylene glycol (EG) + 7.5% dimethyl sulfoxide (DMSO) + 20% fetal calf serum (FCS) for 1, 4, 7 or 10 min at 24 C and then 15% EG + 15% DMSO + 0.5 M sucrose + 20% FCS for 1 min at 24 C before being plunged into liquid nitrogen on Cryotops. Oocytes exposed to equilibration medium for 4 min showed higher survival and cleavage rates after artificial activation than those of oocytes exposed for 1, 7 or 10 min. The survival and cleavage rates of vitrified oocytes after activation were 98.3 and 78.4%, respectively. However, the perivitelline spaces of most of the vitrified/warmed oocytes (6/168, 3.6%) could not be penetrated by sperm after in vitro fertilization, and cortical granule exocytosis (CGE) was observed in the oocytes. Therefore, the inhibitory effect of calcium and cryoprotectants in vitrification medium on CGE was examined. In most of the oocytes vitrified in calcium-free media, CGE was strongly suppressed independent of cryoprotectants. Oocytes vitrified in EG-supplemented calcium-free media showed high survival rates after warming (79.4%). After artificial activation, the cleavage and blastocyst formation rates of the oocytes were also high (72.8 and 23.1%, respectively). The zona penetration rate of vitrified/ warmed oocytes was dramatically improved by using EG-supplemented calcium-free media after in vitro fertilization (111/155, 63.9%). Thus, our data suggest that EG-supplemented calcium-free media improve zona penetration of vitrified rat oocytes by sperm cells. Key words: Cryopreservation, Oocytes, Rat, Vitrification (J. Reprod. Dev. 56: [169][170][171][172][173][174][175] 2010) ryopreservation of germ cells and reproductive organs is a useful and important technology for efficient production and maintenance of transgenic, mutant, knock-out and other animals. In particular, embryo and sperm cryopreservation has been routinely used for not only efficient production of experimental animals but also clinical medicine. On the other hand, the most desired germ cells for freezing or vitrification are matured oocytes because successfully cryopreserved oocytes can be used for in vitro fertilization (IVF), intracytoplasmic sperm injection (ICSI) or somatic-cell nuclear transfer (SCNT) [1,2]. Although successful freezing or vitrification of oocytes has been reported in several mammalian species [3][4][5][6], the survival is generally low. Especially in rats, a high rate of successful vitrification of early-stage embryos has not been obtained until recent years, possibly because rat germ cells seem to be sens...

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Relationship between equilibration times and the presence of cumulus cells, and effect of Taxol treatment for vitrification of in vitro matured porcine oocytes

Cited by 48 publications

References 9 publications

Protective effects of the cumulus-corona radiata complex during vitrification of horse oocytes

Protective effects of the cumulus-corona radiata complex during vitrification of horse oocytes

Developmental Ability of Porcine In Vitro Matured Oocytes at the Meiosis II Stage After Vitrification

Ethylene Glycol-supplemented Calcium-free Media Improve Zona Penetration of Vitrified Rat Oocytes by Sperm Cells

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