Vitrification of in vitro matured ovine oocytes affects in vitro pre-implantation development and mRNA abundance

The impact of vitrification procedures
on in vitro matured (IVM) ovine oocytes mRNA
content and ability to undergo successful fertilization,
cleavage and embronic development was assessed.
Vitrified-warmed (n¼113) and control (n¼140) IVM
oocytes were in vitro fertilized and cultured up to
blastocyst stage under standard conditions. Vitrified
oocytes showed lower cleavage rate (47% vs. 75%,
P<0.001) and development to blastocyst stage
(17% vs. 57%, P<0.001) than controls. In addition,
the timings of the first cleavage and blastocysts
production were significantly delayed in the vitrifiedwarmed
group (P<0.001 in both cases). In parallel,
we analyzed by reverse transcriptase real-time PCR the
relative abundance of b-actin, H2A.Z histone, Poli A
Polimerase (PAP), Heat Shock Protein 90b (HSP90b),
P34cdc2, Cyclin b, Na/K-ATPase and Type I cadherin
(E-Cad) transcripts in single IVM controls (n¼24) and
vitrified-warmed oocytes (n¼40). Results were normalized
against the exogenous rabbit a-globin mRNA
standard and the b-actin housekeeping gene and
similarly described a lower abundance of most mRNAs
in oocytes subjected to vitrification procedures. When
normalized against the exogenous standard mRNA,
all transcripts except for b-actin and H2A.Z showed
a significantly different abundance in the two classes
of oocytes. The same results were obtained after
normalization against the internal standard, except for
HSP90b and E-Cad transcripts, whose lower abundance
in vitrified-warmed oocytes resulted prominent,
but not significant (P¼0.083 and P¼0.068, respectively).
The oocyte lower transcripts abundance following
vitrification might be an early indicator of poor
quality in good correlation with the developmental data
to blastocyst stage