Preimplantation development and expression of Hsp-70 and Bax genes in bovine blastocysts derived from oocytes matured in alpha-MEM supplemented with growth factors and synthetic macromolecules.
In vitro culture conditions affect both the maternal and embryonic expression of genes and is likely to alter both oocyte and embryo developmental competence. The search for better and less variable culture conditions simulating those in vivo has led to the development of defined culture media, with lower impact on the molecular reprogramming of oocytes and embryos. We evaluated embryo development and relative abundance (RA) of Hsp-70 and Bax transcripts in bovine blastocysts produced from oocytes matured in a chemically defined IVM system with synthetic polymers. Immature cumulus oocyte complexes (COCs) were matured for 22-24h in alpha-MEM supplemented with IGF-1, insulin, 0.1% polyvinyl alcohol (PVA), or 0.1% polyvinylpyrrolidone (PVP), but without FSH or LH. The control group consisted of COCs matured in TCM plus FSH and 10% estrous cow serum. After fertilization, presumptive zygotes were co-cultured with cumulus cells until 224 h post-insemination. Total RNA was isolated from embryo pools, reverse transcribed into cDNA, and subjected to transcript analysis by real-time PCR. Cleavage rate was higher (P<0.05) for the control group (68.3%) than for the PVA (54.4%) and PVP-40 (58.3%) groups. Nevertheless, there was no difference among the PVA, PVP-40 and control groups in blastocyst or hatching rates. Similarly, no difference in relative abundance of Hsp-70 and Bax transcripts was detected in comparison to the control group. We inferred that bovine oocytes can be matured in serum- and gonadotrophin-free medium supplemented with PVA or PVP, enriched with IGF-I and insulin, without altering post-cleavage development and relative abundance of some genes associated with stress and apoptosis.
Vireque
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AA
Department of Obstetrics and Gynecology, School of Medicine of Ribeirão Preto, University of São Paulo, Bandeirantes anevue 3900, 14049-900 Ribeirão Preto, SP, Brazil. alessandra_vireque@yahoo.com.br
Camargo
L S A
LS
Serapião
R V
RV
Rosa E Silva
A A M
AA
Watanabe
Y F
YF
Ferreira
E M
EM
Navarro
P A A S
PA
Martins
W P
WP
Ferriani
R A
RA
D000818:Animals; D001755:Blastocyst / Q000378:metabolism*; D002417:Cattle; D003470:Culture Media / Q000737:chemistry; D046149:Embryo Culture Techniques; D004624:Embryo Transfer / Q000662:veterinary; D005307:Fertilization in Vitro / Q000662:veterinary; D018507:Gene Expression Regulation, Developmental / Q000502:physiology; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D036341:Intercellular Signaling Peptides and Proteins / Q000737:chemistry / Q000494:pharmacology*; D046911:Macromolecular Substances / Q000737:chemistry / Q000494:pharmacology*; D009865:Oocytes / Q000378:metabolism; D009930:Organic Chemicals / Q000737:chemistry; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism*
D003470:Culture Media; D018840:HSP70 Heat-Shock Proteins; D036341:Intercellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D009930:Organic Chemicals; C420642:alpha minimal essential medium; D051028:bcl-2-Associated X Protein