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Developmental competence of human in vitro aged oocytes as host cells for nuclear transfer

Hall, Vanessa LU ; Compton, D.; Stojkovic, P.; Nesbitt, M.; Herbert, M.; Murdoch, A. and Stojkovic, M. (2007) In Human Reproduction 22(1). p.52-62
Abstract
BACKGROUND: Improving human nuclear transfer (NT) efficiencies is paramount for the development of patient-specific stem cell lines, although the opportunities remain limited owing to difficulties in obtaining fresh mature oocytes. METHODS: Therefore, the developmental competence of aged, failed-to-fertilize human oocytes as an alternate cytoplasmic source for NT was assessed and compared with use of fresh, ovulation-induced oocytes. To further characterize the developmental potential of aged oocytes, parthenogenetic activation, immunocytochemical analysis of essential microtubule proteins involved in meiotic and mitotic division, and RT-PCR in single oocytes (n = 6) was performed to determine expression of oocyte-specific genes... (More)
BACKGROUND: Improving human nuclear transfer (NT) efficiencies is paramount for the development of patient-specific stem cell lines, although the opportunities remain limited owing to difficulties in obtaining fresh mature oocytes. METHODS: Therefore, the developmental competence of aged, failed-to-fertilize human oocytes as an alternate cytoplasmic source for NT was assessed and compared with use of fresh, ovulation-induced oocytes. To further characterize the developmental potential of aged oocytes, parthenogenetic activation, immunocytochemical analysis of essential microtubule proteins involved in meiotic and mitotic division, and RT-PCR in single oocytes (n = 6) was performed to determine expression of oocyte-specific genes [oocyte-specific histone 1 (H1FOO), growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15), zygote arrest 1 (ZAR1)] and microtubule markers [nuclear mitotic arrest (NuMA), minus-end directed motor protein HSET and the microtubule kinesin motor protein EG5]. RESULTS: For NT, enucleation and fusion rates of aged oocytes were significantly lower compared with fresh oocytes (P < 0.05). Cleavage rates and subsequent development were poor. In addition, parthenote cleavage was low. Immunocytochemical analysis revealed that many oocytes displayed aberrant expression of NuMA and EG5, had disrupted meiotic spindles and tetrapolar spindles. One of the six oocytes misexpressed GDF9, BMP15 and ZAR1. Two oocytes expressed EG5 messenger RNA (mRNA), and HSET and NuMA were not detectable. RT-PCR of mRNA for oocyte specific genes and microtubule markers in single aged oocytes. CONCLUSIONS: Thus, aneuploidy and spindle defects may contribute to poor parthenogenetic development and developmental outcomes following NT. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
somatic cell nuclear, human, gene expression, oocyte, microtubule, transfer
in
Human Reproduction
volume
22
issue
1
pages
52 - 62
publisher
Oxford University Press
external identifiers
  • wos:000243065300007
  • scopus:33845666701
ISSN
0268-1161
DOI
10.1093/humrep/del345
language
English
LU publication?
yes
id
0ebd06ce-28fe-490e-9b2d-05d48af34a43 (old id 681693)
date added to LUP
2007-12-11 11:33:26
date last changed
2017-10-29 03:35:48
@article{0ebd06ce-28fe-490e-9b2d-05d48af34a43,
  abstract     = {BACKGROUND: Improving human nuclear transfer (NT) efficiencies is paramount for the development of patient-specific stem cell lines, although the opportunities remain limited owing to difficulties in obtaining fresh mature oocytes. METHODS: Therefore, the developmental competence of aged, failed-to-fertilize human oocytes as an alternate cytoplasmic source for NT was assessed and compared with use of fresh, ovulation-induced oocytes. To further characterize the developmental potential of aged oocytes, parthenogenetic activation, immunocytochemical analysis of essential microtubule proteins involved in meiotic and mitotic division, and RT-PCR in single oocytes (n = 6) was performed to determine expression of oocyte-specific genes [oocyte-specific histone 1 (H1FOO), growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15), zygote arrest 1 (ZAR1)] and microtubule markers [nuclear mitotic arrest (NuMA), minus-end directed motor protein HSET and the microtubule kinesin motor protein EG5]. RESULTS: For NT, enucleation and fusion rates of aged oocytes were significantly lower compared with fresh oocytes (P &lt; 0.05). Cleavage rates and subsequent development were poor. In addition, parthenote cleavage was low. Immunocytochemical analysis revealed that many oocytes displayed aberrant expression of NuMA and EG5, had disrupted meiotic spindles and tetrapolar spindles. One of the six oocytes misexpressed GDF9, BMP15 and ZAR1. Two oocytes expressed EG5 messenger RNA (mRNA), and HSET and NuMA were not detectable. RT-PCR of mRNA for oocyte specific genes and microtubule markers in single aged oocytes. CONCLUSIONS: Thus, aneuploidy and spindle defects may contribute to poor parthenogenetic development and developmental outcomes following NT.},
  author       = {Hall, Vanessa and Compton, D. and Stojkovic, P. and Nesbitt, M. and Herbert, M. and Murdoch, A. and Stojkovic, M.},
  issn         = {0268-1161},
  keyword      = {somatic cell nuclear,human,gene expression,oocyte,microtubule,transfer},
  language     = {eng},
  number       = {1},
  pages        = {52--62},
  publisher    = {Oxford University Press},
  series       = {Human Reproduction},
  title        = {Developmental competence of human in vitro aged oocytes as host cells for nuclear transfer},
  url          = {http://dx.doi.org/10.1093/humrep/del345},
  volume       = {22},
  year         = {2007},
}