Veterinary Record

1997

DOI: 10.1136/vr.141.20.522

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Influence of cumulus cells on in vitro maturation of denuded buffalo oocytes

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M. S. Chauhan²,

et al.

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Cited by 18 publications

(12 citation statements)

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“…However, the molecular changes, visible as differential expression profiles of genes and proteins responsible for COC maturation, are still not fully recognized. It was shown in several studies that the achievement of a fully mature stage of oocytes is regulated via an appropriate communication between the oocyte and the surrounding cumulus-granulosa cells, and it is assumed that attainment of the MII stage by the oocyte is accompanied by CC expansion [8, 18, 19, 24, 25]. The mechanisms responsible for this process are not entirely known.…”

Section: Discussionmentioning

confidence: 99%

Short-term Cultivation of Porcine Cumulus Cells Influences the Cyclin-dependent Kinase 4 (Cdk4) and Connexin 43 (Cx43) Protein Expression—A Real-time Cell Proliferation Approach

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²

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³

et al. 2013

J. Reprod. Dev.

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The CC (cumulus cell) proliferation index in relation to the expression and distribution of Cdk4 and Cx43 proteins, which are crucial factors for oocyte maturation, was investigated. Cumulus-oocyte complexes (COCs) were recovered from pubertal crossbred Landrace gilts and treated with collagenase, and separated CCs were cultured in standard TCM199 medium for 44 h. At each step of in vitro cultivation (IVC) of CCs (0, 12, 24 and 44 h), a normalized proliferation index was assessed. Cdk4 and Cx43 protein expression and the CC-specific cellular distribution were analyzed by confocal microscopic observation. The normalized proliferation index (number of cells attached, measured by impedance) was increased in the first 12 h of IVC (P<0.01) and differed between 12 h and 24 h of cultivation (P<0.001). Later, between 24 h–44 h of IVC, the CC proliferation rate was stable, and no significant differences were observed. Based on the confocal microscopic observation, increased expression of both Cdk4 and Cx43 was found after 44 h of IVC compared with the expression of these proteins before IVC. Moreover, after IVC, a substantial translocation of Cdk4 and Cx43 was noted from the nucleus to the cytoplasm of CCs. In conclusion, it was demonstrated for the first time that CCs can be cultured in vitro separately without oocytes and that the proliferation index was significantly increased in the first 12 h of IVC, which may reflect the process of ordinary cumulus cell expansion. Furthermore, the expression of both Cdk4 and Cx43 in CCs suggested that these proteins may be regarded as markers not only of proper oocyte maturation but also of CC differentiation. Translocation of these proteins into the cytoplasm of CCs after 44 h of IVC may be related to the expansion process.

“…However, the molecular changes, visible as differential expression profiles of genes and proteins responsible for COC maturation, are still not fully recognized. It was shown in several studies that the achievement of a fully mature stage of oocytes is regulated via an appropriate communication between the oocyte and the surrounding cumulus-granulosa cells, and it is assumed that attainment of the MII stage by the oocyte is accompanied by CC expansion [8, 18, 19, 24, 25]. The mechanisms responsible for this process are not entirely known.…”

Section: Discussionmentioning

confidence: 99%

Short-term Cultivation of Porcine Cumulus Cells Influences the Cyclin-dependent Kinase 4 (Cdk4) and Connexin 43 (Cx43) Protein Expression—A Real-time Cell Proliferation Approach

¹

,

²

,

³

et al. 2013

J. Reprod. Dev.

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The CC (cumulus cell) proliferation index in relation to the expression and distribution of Cdk4 and Cx43 proteins, which are crucial factors for oocyte maturation, was investigated. Cumulus-oocyte complexes (COCs) were recovered from pubertal crossbred Landrace gilts and treated with collagenase, and separated CCs were cultured in standard TCM199 medium for 44 h. At each step of in vitro cultivation (IVC) of CCs (0, 12, 24 and 44 h), a normalized proliferation index was assessed. Cdk4 and Cx43 protein expression and the CC-specific cellular distribution were analyzed by confocal microscopic observation. The normalized proliferation index (number of cells attached, measured by impedance) was increased in the first 12 h of IVC (P<0.01) and differed between 12 h and 24 h of cultivation (P<0.001). Later, between 24 h–44 h of IVC, the CC proliferation rate was stable, and no significant differences were observed. Based on the confocal microscopic observation, increased expression of both Cdk4 and Cx43 was found after 44 h of IVC compared with the expression of these proteins before IVC. Moreover, after IVC, a substantial translocation of Cdk4 and Cx43 was noted from the nucleus to the cytoplasm of CCs. In conclusion, it was demonstrated for the first time that CCs can be cultured in vitro separately without oocytes and that the proliferation index was significantly increased in the first 12 h of IVC, which may reflect the process of ordinary cumulus cell expansion. Furthermore, the expression of both Cdk4 and Cx43 in CCs suggested that these proteins may be regarded as markers not only of proper oocyte maturation but also of CC differentiation. Translocation of these proteins into the cytoplasm of CCs after 44 h of IVC may be related to the expansion process.

“…Follicular CC play an important role in supporting maturation of mammalian oocytes as well as promoting their subsequent developmental competence. Removal of CC before buffalo oocyte maturation in vitro resulted in a significant reduction of nuclear maturation (Das et al. 1997).…”

Section: Discussionmentioning

confidence: 99%

“…Follicular CC play an important role in supporting maturation of mammalian oocytes as well as promoting their subsequent developmental competence. Removal of CC before buffalo oocyte maturation in vitro resulted in a significant reduction of nuclear maturation (Das et al 1997). The gap junctions between CC and the oocyte enable transport of small molecules (e.g., nutrients and messengers), possibly regulating meiotic resumption of oocytes enclosed with CC (Luciano et al 2005).…”

Section: Discussionmentioning

confidence: 99%

Effects of Cumulus Cells on In Vitro Maturation of Oocytes and Development of Cloned Embryos in the Pig

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²

2011

Reprod Domestic Animals

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The purpose of this study was to investigate the role of porcine cumulus cells (CC) in oocyte maturation and somatic cell nuclear transfer (SCNT) embryo development in vitro. Denuded pig oocytes were co-cultured with CC or routinely cultured in maturation medium without a feeder layer. Porcine CC inactivated with mitomycin C or non-inactivated were used for the feeder layer in co-culture with porcine SCNT embryos to investigate comparatively the developmental competence of cloned embryos. The DNA damage aspects of apoptosis and expression pattern of genes implicated in apoptosis (Fas/FasL) as well as the mRNA expression of DNA methyltransferase (Dnmt1, Dnmt3a) of porcine SCNT embryos were also evaluated by comet assay or real-time RT-PCR, respectively. The results showed that co-culture with CC improved the extrusion rate of pbI (49.3% vs 31.5%, p<0.05) and survival rate (75.7% vs 53.3%, p<0.05) of denuded oocytes, but had no effects on blastocyst developmental rate or 2-cell-stage survival rate of in vitro fertilization embryos. Co-culture with CC inactivated by mitomycin C improved the blastocyst developmental rate (26.6% vs 13.0%, p<0.05) and decreased the apoptotic incidence (27.6% vs 46.2%, p<0.05) of porcine cloned embryos. Co-culture with inactivated CC reduced Fas and FasL mRNA expression of cloned embryos at the blastocyst stage compared with NT controls (p<0.05), but there were no differences in Dnmt1 and Dnmt3a mRNA expression among groups. Co-culture with inactivated cumulus cell monolayer significantly increased blastocyst formation and decreased the apoptotic incidence in porcine cloned embryos during in vitro development.

“…Studies in various species (mice, rats, pigs and cattle) have shown that some cumulus‐denuded oocytes can complete meiotic resumption (Binor and Wolf 1979; Magnusson 1980; Yamauchi and Nagai 1999; Geshi et al. 2000), but the removal of cumulus cells before oocyte maturation results in a significant reduction of the nuclear maturation rate (Das et al. 1997).…”

Section: Discussionmentioning

confidence: 99%

“…When cumulus cells were removed from COCs at 24 and 42 h after the onset of maturation culture, the maturation rates were significantly improved (52.5 and 78.1%, respectively). Studies in various species (mice, rats, pigs and cattle) have shown that some cumulus-denuded oocytes can complete meiotic resumption (Binor and Wolf 1979;Magnusson 1980;Yamauchi and Nagai 1999;Geshi et al 2000), but the removal of cumulus cells before oocyte maturation results in a significant reduction of the nuclear maturation rate (Das et al 1997). The interaction of cumulus cells with the oocyte provides the local production of glycosaminoglycans, steroid hormones, nutrients and other factors to support the oocyte maturation (Brower and Schultz 1982;Danforth 1995;Yamauchi and Nagai 1999;Dode and Graves 2002).…”

Section: Discussionmentioning

confidence: 99%

Effect of the Removal of Cumulus Cells on the Nuclear Maturation, Fertilization and Development of Porcine Oocytes

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²

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³

et al. 2005

Reprod Domestic Animals

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The present study was conducted to investigate the effects of attachment of cumulus cells to porcine oocytes during the process of maturation and fertilization on the nuclear maturation, fertilization and subsequent development after in vitro fertilization (IVF). In the first experiment, the cumulus cells were removed from cumulus-oocyte complexes (COCs) at 0, 24 and 42 h after the onset of maturation culture and were then cultured until reaching 42 h of cultivation. In the second experiment, COCs were denuded as described in the first experiment, then fertilized and cultured for 7 days. As a control, cumulus cells were allowed to maintain attachment to the oocytes until the end of IVF. The proportion of oocytes reaching metaphase II significantly increased with the delay in the removal treatment of cumulus cells. The proportion of normal fertilization gradually increased with delay in the removal treatment of cumulus cells from COCs until the end of IVF. However, no significant difference in the proportion of normal fertilization was found between the 42-h and control groups. The removal treatment of cumulus cells in the 0- and 24-h group significantly (p < 0.05) decreased the proportion of cleaved embryos when compared with the control, and none of them developed to the blastocyst stage. The proportion of development to the blastocyst stage was significantly higher (p < 0.05) in the control group than in the 42-h group (18.1% vs 12.4%; p < 0.05). The present study indicates that the attachment of cumulus cells to the oocyte during maturation and fertilization is important to support oocyte nuclear maturation, fertilization and subsequent embryo development. Particularly, the attachment of cumulus cells to the oocyte during IVF promotes embryonic development.

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