FTIR spectral signatures of mouse antral oocytes: Molecular markers of oocyte
maturation and developmental competence
Diletta Ami
a
, Paolo Mereghetti
b,c
, Antonino Natalello
d
, Silvia Maria Doglia
d,
⁎
, Mario Zanoni
e
,
Carlo Alberto Redi
a,
⁎⁎
, Manuela Monti
a
a
Fondazione IRCCS Policlinico San Matteo, V.le C. Golgi 19, 27100 Pavia, Italy
b
BIOMS (Center for Modeling and Simulation in the Biosciences), University of Heidelberg, Im Neuenheimer Feld 36, 69120 Heidelberg, Germany
c
Molecular and Cellular Modeling Group, HITS gGmbH, Schloss Wolfsbrunnenweg 35, 69118, Heidelberg, Germany
d
Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Piazza della Scienza 2, 20126 Milano, Italy
e
Laboratorio di Biologia dello Sviluppo, Dipartimento di Biologia Animale, Università degli Studi di Pavia, Via A. Ferrata 1, 27100 Pavia, Italy
abstractarticle info
Article history:
Received 12 October 2010
Received in revised form 15 February 2011
Accepted 15 March 2011
Available online 22 March 2011
Keywords:
FTIR microspectroscopy
SN oocytes
NSN oocytes
Oocyte maturation
Developmental competence
Polyadenylation
Mammalian antral oocytes with a Hoescht-positive DNA ring around the nucleolus (SN) are able to resume
meiosis and to fully support the embryonic development, while oocytes with a non-surrounded nucleolus
(NSN) cannot. Here, we applied FTIR microspectroscopy to characterize single SN and NSN mouse oocytes in
order to try to elucidate some aspects of the mechanisms behind the different chromatin organization that
impairs the full development of NSN oocyte-derived embryos. To this aim, oocytes were measured at three
different stages of their maturation: just after isolation and classification as SN and NSN oocytes (time 0); after
10 h of in vitro maturation, i.e. at the completion of the metaphase I (time 1); and after 20 h of in vitro
maturation, i.e. at the completion of the metaphase II (time 2). Significant spectral differences in the lipid
(3050–2800 cm
−1
) and protein (1700–1600 cm
−1
) absorption regions were found between the two types of
oocytes and among the different stages of maturation within the same oocyte type. Moreover, dramatic
changes in nucleic acid content, concerning mainly the extent of transcription and polyadenylation, were
detected in particular between 1000 and 800 cm
− 1
. The use of the multivariate principal component–linear
discriminant analysis (PCA–LDA) enabled us to identify the maturation stage in which the separation between
the two types of oocytes took place, finding as the most discriminating wavenumbers those associated to
transcriptional activity and polyadenylation, in agreement with the visual analysis of the spectral data.
© 2011 Elsevier B.V. All rights reserved.
1. Introduction
Murine oocytes isolated from the ovarian antral compartment are
characterized by two different types of chromatin organization [1,2] as
in most mammalian species like the rat [3], the pig [4], the monkey [5]
and the human [6]. In the Surrounded Nucleolus (SN) type, chromatin
is highly condensed and forms a Hoechst positive ring around the
nucleolus, while in the Not Surrounded Nucleolus (NSN) oocytes the
chromatin is more dispersed and less condensed around the nucleolus
[7]. The important issue of chromatin organization has been studied
by several complementary techniques, such as confocal fluorescence
microscopy [8] and transmission electron microscopy [6,9].For
oocytes, chromatin organization and regulation of transcription are
strictly related to each other, as it is well known that heterochromatic
chromatin is associated with low level of transcription. For this reason,
SN oocytes are considered transcriptionally inactive while the NSN
types are transcriptionally active [1,2,6]. Another important difference
between SN and NSN antral oocytes concerns their ability to resume
meiosis and complete, after fertilization, the embryonic development:
only the SN type is able to develop till the blastocyst stage while the
NSN type arrests its development at the two cell stage.
It is still unknown how the well orchestrated functional ovary
originates oocytes with different destiny and which are the molecular
events accounting for the two different chromatin organizations and
thus two different prospective zygotic developments.
It has been suggested that the NSN-derived zygotic epigenome
shows reduced levels of expression of some important genes involved
Biochimica et Biophysica Acta 1813 (2011) 1220–1229
Abbreviations: A, Adenine; CPE, cytoplasmicpolyadenylation element; CpG, Cytosine-
phosphate-Guanine; FTIR, Fourier Transform InfraRed; GV, Germinal vesicle; GVBD,
Germinal vesicle break down; MCT, Mercury cadmium telluride; MI, Metaphase I; MII,
Metaphase II; mRNA, messenger RNA; NSN, not surrounded nucleolus; PCA–LDA, Principal
component analysis–linear discriminant analysis; polyA, polyAdenyne; SN, Surrounded
nucleolus; U, Uracil
⁎ Correspondence to: S.M. Doglia, Dipartimento di Biotecnologie e Bioscienze,
Università di Milano-Bicocca, Piazza della Scienza 2, 20126 Milano, Italia. Tel.: +39
02 64483459; fax: +39 02 64483565.
⁎⁎ Correspondence to: Prof. Carlo Alberto Redi, Dipartimento di Biologia Animale,
Università degli Studi di Pavia, Via Ferrata 1, 27100 Pavia, Italy. Tel.: +39 0382 986306;
fax: +39 0382 986270.
E-mail addresses: silviamaria.doglia@unimib.it (S.M. Doglia),
carloalberto.redi@unipv.it (C.A. Redi).
0167-4889/$ – see front matter © 2011 Elsevier B.V. All rights reserved.
doi:10.1016/j.bbamcr.2011.03.009
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journal homepage: www.elsevier.com/locate/bbamcr