Molecular and Cellular Endocrinology 315 (2010) 299–307
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Molecular and Cellular Endocrinology
journal homepage: www.elsevier.com/locate/mce
Female infertility due to anovulation and defective steroidogenesis
in NPC2 deficient mice
D. Busso
a,b,∗
, M.J. O
˜
nate-Alvarado
a
, E. Balboa
b
, S. Zanlungo
b
, R.D. Moreno
a
a
Departamento de Fisiología, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, Avenida Bernarndo O‘Higgins 340, 8331010 Santiago, Chile
b
Departamento de Gastroenterología, Facultad de Medicina, Pontificia Universidad Católica de Chile, Avenida Bernarndo O‘Higgins 340, 8331010 Santiago, Chile
article info
Article history:
Received 14 August 2009
Received in revised form 16 October 2009
Accepted 20 October 2009
Keywords:
Female fertility
Ovary
Ovulation
Steroids
NPC2
Cholesterol
abstract
Niemann Pick C2 (NPC2) and NPC1 proteins function cooperatively to catalyze cholesterol efflux from
lysosomes. NPC1 is expressed in ovarian cells and female NPC1 mice are infertile. This work addressed for
the first time the localization and function of murine NPC2 protein in the ovary. Ovarian NPC2 was local-
ized to theca and luteal cells, which use cholesterol as a substrate to produce estradiol and progesterone,
respectively. NPC2 deficient (NPC2−/−) females had abnormal estrous cycles and were infertile, with
normal folliculogenesis until the antral stage, but a complete absence of corpora lutea and many zonae
pellucidae remnants, indicative of anovulation. Serum estradiol was reduced and ovarian cholesterol
was accumulated in NPC2−/− mice, suggesting a defect in cholesterol export from intracellular stores.
After superovulation, NPC2−/− mice ovulated less eggs than their wild type littermates, showed ovaries
with less corpora lutea and numerous unruptured follicles, and lower serum progesterone concentra-
tion. Together, these results suggest that NPC2 participates in the traffic of ovarian cholesterol required
to provide the substrate for steroid synthesis and support follicle maturation, ovulation and luteinization.
© 2009 Elsevier Ireland Ltd. All rights reserved.
1. Introduction
The production of mature oocytes capable of undergoing fertil-
ization starts when primordial follicles give rise to primary follicles,
which in turn origin secondary follicles, then tertiary antral folli-
cles and finally preovulatory follicles. After ovulation, the rupture
of Graafian follicles releases the cumulus oophorus containing the
oocyte, leaving behind follicular cells which luteinise and give rise
to the corpus luteum. These complex events of differentiation are
coordinated by sex steroids that are cyclically produced under the
regulation of the pituitary gland and that interact with ovarian
locally produced growth factors (Drummond, 2006). Steroids are
synthesized de novo using cholesterol as a substrate by the con-
certed action of the two different follicular cell types that surround
the oocyte. The most external thecal cells use cholesterol to pro-
duce androgens which are in turn transformed into estrogens in
granulosa cells under follicular stimulating hormone (FSH) control.
As the folliculogenesis process progresses, granulosa cells mature
and express luteinizing hormone (LH) receptors. At the LH surge,
ovulation takes place and follicular cells differentiate into lutein
cells that now metabolize cholesterol into progesterone.
∗
Corresponding author. Tel.: +56 2 686 2708; fax: +56 2 222 5515.
E-mail addresses: dbussoar@yahoo.com, dbusso@bio.puc.cl (D. Busso).
Cells obtain cholesterol by de novo synthesis and/or by receptor-
mediated uptake from blood (Liscum and Munn, 1999). Cholesterol
from plasma is obtained by selective uptake from high-density
lipoprotein (HDL), in a process regulated by the scavenger receptor
type BI (SR-BI), and by endocytosis of cholesterol-rich low density
lipoproteins (LDL). LDL-borne cholesterol is stored in lysosomes,
where two proteins named Niemann Pick C1 (NPC1) and NPC2 have
been shown to be key players in the mobilization of this lipid to
other cell compartments (Sleat et al., 2004). These two proteins
share similar names as mutations in either of these produce a fatal
disease known as Niemann Pick Type C, characterized by a general
lipidosis in the organism which among other consequences gives
rise to neurodegeneration (Liscum and Klansek, 1998; Sturley et
al., 2004). Despite their similar names, the two NPC proteins are
completely different in size and aminoacidic sequence. Whereas
NPC1 is a large transmembrane protein (Carstea et al., 1997), NPC2
is a small, soluble, cholesterol-binding protein found not only in
the lumen of lysosomes but also abundantly in secretions such as
milk and epididymal fluid (Naureckiene et al., 2000). Although still
hypothetical, the current model to explain the function of NPC pro-
teins proposes that NPC2 catalyzes the delivery of cholesterol to
NPC1 for the efflux from lysosomes (Infante et al., 2008). Despite
the involvement of NPC1 and NPC2 as a “tag team duo” in the export
of lysosomal cholesterol, these two proteins have been recently
shown to exert separate, sequential functions given by their differ-
ent biochemical properties (Subramanian and Balch, 2008).
0303-7207/$ – see front matter © 2009 Elsevier Ireland Ltd. All rights reserved.
doi:10.1016/j.mce.2009.10.011