MINI REVIEW

Transcripts Encoding the Enzymes that Convert
Acetyl-CoA to Cholesterol are Inducedin Cumulus Cells and are Essential for Progesterone
Biosynthesis and Meiotic Resumption of Porcine Oocytes
Masayuki SHIMADA, Yasuhisa YAMASHITA and Masahide NISHIBORIIntroduction

otic resumption of oocytes and improved the rate of early
embryonic development to the blastocyst stage after in vi-

The surge of luteinizing hormone (LH) induces marked

tro fertilization. We also reported that progesterone pro-functional (endocrine, biochemical and molecular) changes

duced by porcine COCs bound PR, another FSH- and LH-

in the preovulatory follicle. Estrogen concentrations decline

induced factor in cumulus cells. LH also reducedin follicular fluid as a consequence of reduced transcription

proliferative activity of cumulus cells and closed the gapα) genes injunctional communication within cumulus cells [11, 13].

granulosa and theca cells, respectively, whereas progester-

Conversely, when COCs were cultured with FSH, LH andone concentrations rise in association with induction of

PR antagonist RU486, prolilferation and gap junctional com-A ) in granulosa cells [1,2]. The LH surge

munication was restored [13,14]. These results suggest that

also initiates ovulation, a process that is inhibited in rats byprogesterone produced by cumulus cells acts within cumu-

treatment with either anti-progesterone antiserum [3] orlus cells and/or at the oocyte cell surface to facilitate in vitro

epostane [4], a compound that blocks the synthesis of pro-meiotic resumption of porcine oocytes. Thus, in order togesterone. Female mice null for progesterone receptor

obtain oocytes with a high developmental competence to(PR) (PRKO) fail to ovulate, even in response to exoge-

blastocyst stage in vitro, it is essential to analyze the factors

nous hormones [5], suggesting that the LH-induced in-controlling progesterone biosynthesis in cumulus cells.

of aromatase (CYP

P450scc (CYP

) and 17αHSD (CYP

crease of progesterone is essential for ovulation.
In association with in vitro meiotic maturation of cumulus
enclosed oocytes in human [6], rat [7], cattle [8] and pig[9-11], progesterone production by cumulus cells is in-Serum supplementation is not required forprogesterone production by cumulus cells during
meiotic resumption of porcine oocytes

creased in response to FSH and LH stimulation. In our previous studies [12], we showed that the high concentrations

In our in vitro oocyte maturation system, COCs culturedof progesterone secreted by porcine COCs accelerated mei-

with both 0.02 µg/ml FSH and 1.0 µg/ml LH produced moreprogesterone than was detected in cultures using either go-

Correspondence：Masayuki SHIMADA, Department of Applied Animal Science, Graduate School of Biosphere Science,
Hiroshima University, 1-4-4, kagamiyama Higashi-Hiroshima,739-8528, Japan
TEL: +81-82-424-7899
FAX: +81-82-424-7899
E-mail: mashimad@hiroshima-u.ac.jp
This work was supported in part, by Grant-in-Aid for Scientific
Research (M.S., No.14760179, 15380222, 16780195) and a Research Fellowship for Young Scientist (Y.Y., No.08254) from theJapan Society for the Promotion of Science (JSPS).

nadotropin alone [15]. Since progesterone is synthesizedfrom free cholesterol by cytochrme P450 cholesterol sidechain cleavage (CYPA ) [16, 17] and 3β-hydrosteroid

dehydorogenase /isomerase ( βHSD) enzymes [18], we
analyzed levels of mRNA encoding these proteins in FSHand LH-stimulated COCs. The data showed that mRNAlevels were up-regulated in cumulus cells cultured with
FSH and LH for 10 or 20-hr cultivation (Fig. 1A). In follicular cells, it has been shown that lipoprotein-bound choles-

Japanese Journal of Reproductive Endocrinology（2005）10：15-20

15Masayuki SHIMADA et al.

(A)

(B)

3βHS D
DHCR14

CYP11A1
DHCR14

DHCR7

DHCR7

βactinForβactin

H89

10h
0h

Free FL

Free FL

10h

20h

For

H89

20h

Fig. 1. Levels of mRNA encoding β,,and
in cumulus
cells of porcine COCs cultured with FSH and LH.
(A)Porcine COCs were cultured without (control) or with FSH (F) and LH (L)(FL) for 10 or 20 hr. (B) Porcine COCs were cultured with 10µM forskolin (For)and/or 10 µM of PKA inhibitor H89 for 10 or 20 hr.z 0h: COCs were collectedfrom small antral follicles.

terolester and stored cholesterolester comprise the majoramounts and/or activities of enzymes that involve in choles-

source for the cholesterolester used for steroid hormone

terol biosynthesis, the major source of progesterone is de-biosynthesis [19-21]. In contrast, luteinized follicular cells

livered from de-novo synthesized cholesterol during meiotic

contain numerous lipid droplets, and the stored cholester-

resumption of porcine oocytes in vitro.

olesters are hydrolyzed by cholesterol esterase to provide
the substrate for progesterone biosynthesis [22]. Furthermore, Armstrong [23] and Armstrong et al. [24] reported
that the increased production of progesterone in rabbit ova-

Gonadotropins induce the expression of enzymes
that convert acetyl-CoA to cholesterol as well as
CYP11A1 and 3 βHSD

ry and bovine corpus luteum was exerted at a point subsequent to the biosynthesis of cholesterol. However, in cumu-

The cholesterol biosynthetic pathway is regulated bylus cells just after collection from the antral follicles, theover 30 cytochrome P450 superfamily enzymes that requirelevel of cholesterolesters was much lower than that in lu-

NADPH as functional coactivator (Fig. 2B) [25]. The mostteinized cells (our unpublished data). Therefore, we hy-

important reaction is catalyzed by HMG-CoA reductase

pothesized that exogenous cholesterol might be used as the(HMGR), the rate limiting enzyme of the cholesterol bio-

substrate for progesterone biosynthesis in cumulus cells.

synthetic pathway [26]. The enzyme activity is regulated by

When COCs were cultured in medium without serum for 20

a negative feedback mechanism mediated by sterols andhr, the level of progesterone was lower than that in serum-

non-sterol metabolites derived from mevalonate, the prod-

containing medium (Fig. 2A). Unexpectedly, the addition of

uct of the reaction catalyzed by HMGR [27-30]. When lev-cholesterolester to the serum free medium did not increaseels of the pathway endproducts such a cholesterol are low,

the progesterone production, whereas the level was in-activity of HMGR is high. Conversely when the levels ofcreased by exogenous essential amino acids (Fig. 2A).

endproducts are low, enzyme activity increases. This feed-

These results indicated that exogenous cholesterol did notback mechanism involves sterol regulation of the HMGR

serve as an immediate substrate. Rather, the addition ofgene (Hmgcr) promoter [31]. Specifically, when sterol binds

specific inhibitors to enzymes in the cholesterol biosynthet-a sterol regulatory enhancer element (SRE), transcription

ic pathway (shown Fig. 2B) significantly suppressed pro-of this gene is repressed whereas in the absence of sterols

gesterone production (Fig. 2C), suggesting that cholesteroltranscription of the promoter is activated [31]. Thus, sterols

was derived by de novo synthesis from acetate. These di-including cholesterol regulate both gene expression andvergent results suggest that gonadotropins increase theserve as substrate for enzyme activity of HMGR.

16

Japanese Journal of Reproductive Endocrinology Vol.10 2005

The Role of Cholesterol Biosynthesis Pathway in Cumulus Cells

Thaddeus and Strauss [31] have shown that exposure ofProgesterone concentration(ng/ml)(A)
30

cultured granulosa cells to 8-bromoadenosine cyclic 3 , 525-phosphate (8-bromo-cAMP) results in a rapid increase in20

the content of the message for Hmgcr. 14α-sterol demeth-15
10

ylase (CYP

5alpha-methyl group (C32) of lanosterol (Fig. 2B) was also

0

serum

free

cholesterolAA

AA+
cholesterol

without serum

HMG-CoA reductase(Hmgrc )ing stimulates transcription of the Hmgcr and CYP
Lanosterol

AY9944-A -7transcription of two additional genes in the biosynthesis of

FF-MAScholesterol is induced in cumulus cells: delta 14-reductase

Delta 14-reductase(DHCR14)(DHCR) and delta 7-reductase (DHCR ) (Fig. 1A).

Furthermore, agonist induced expression of these two

Delta 7-reductase(DHCR7)

genes suppressed by the PKA inhibitor H89 (Fig. 1B). Se-

Cholesterol

(C)

. Our

study showed that in response to FSH and LH or forskolin,

T-MAS
AY9944-A -7
BM15.766

gonadotropin (eCG) priming [32]. The promoter regions ofwell as SRE, suggesting that in follicular cells cAMP signal-

Acethyl-CoA

14α-demethylase(CYP51)expressed in rat ovary within 48 h after equine chorionicboth genes contain a cAMP-responsive element (CRE) as(B)

Ketoconazole) which catalyzes the oxidative removal of 14

Progesterone concentration(ng/ml)rum used as a source of cholesterol and other sterols did
25

not suppress expression of these genes [33], suggesting

20

that DHCR

15

and DHCR

promoters are regulated bycAMP-dependent pathway, but not negative feedback via a

10

SRE mechanism. Thus, the gonadotropin stimulation induc-5

es transcription of several genes regulating sterol and ste-

0
C

FL

Keto

AY

BM

roid biosynthesis in cumulus cells: Hmgcr, CYP
DHCR

FL

Fig. 2. (A) Progesterone production by COCs cultured with FSH and
LH (FL) for 20 hr. SerμM: COCs were cultured with 10% fetal
calf, contorl: COCs were cultured without serμM, Cholesterol:
COCs were cultured with 1% cholesterol ester solution (Gibco), AA: COCs were cultured with non-essential and essential
amino acid (Gibco), AA+cholesterol: COCs were cultured withcholesterol ester and amino acid. Values are mean +/- SEM of
3 replicates.
(B) Simplified outline of the cholesterol biosynthetic pathway
Arrows denote enzymatic conversions, either a single step (onearrow) or multienzymatic steps (2 arrows). Ketoconazole
blocks the activity of 14α-demethylase, AY9944-A-7 blocks
the activity of both the delta 14-reductase and delta 7-reductase, and BM15.766 only blocks the activity of delta 7-reductase.
(C) Effects of Ketoconazole, AY9944-A-7 or BM15.766 on theprogesterone production by COCs cultured for 20 hr. Values aremean +/- SEM of 3 replicates. Control: COCs were cultured
without FSH and LH for 20 h. FL: COCs were cultured with
FSH and LH for 20 h. Keto: COCs were cultured with FSH, LH,
and Ketoconazole (1mM) for 20 h. AY: COCs were cultured
with FSH, LH, and AY9944-A-7 (1μM) for 20 h. BM: COCs
were cultured with FSH, LH, and BM15.766 (20μM) for 20 h.

, DHCR , CYP,

A and βHSD. Importantly,

the induced expression of these genes leads to increased
local concentrations of progesterone in cumulus cells that
impact meiotic resumption of porcine oocytes (Fig. 3).

The expression of DHCR14 and DHCR7 plays animportant role in meiotic resumption of porcine
cumulus-enclosed oocytes
Meiosis-activating sterol, FF-MAS, first reported to be
purified from human follicular fluid (4,4-dimethyl-5αcholesta-8,14,24-trien-3β-ol), was synthesized by COCs
in response to FSH stimulation [34]. This type of sterol is
an intermediate in the cholesterol biosynthetic pathway
produced by demethylation of lanosterol by 14α-demethylase(Fig. 2B). Byskov et al. [34] showed that FF-MAS canstimulate in a dose-dependent manner in vitro meiotic maturation of mouse denuded oocytes. Addition of an inhibitorof FF-MAS synthesis, ketoconazole, resulted in the inhibition of GVBD of mouse oocytes in a dose-dependent fashion [35]. Our results also showed that GVBD in porcine oo-

MINI REVIEW

17

Masayuki SHIMADA et al.

100

Acethyl-CoA

HMG-CoA reductase(Hmgcr)FSH

14alpha-demethylase(CYP51)

ATP

Delta 14-reductase(DHCR14)cAMP

Delta 7-reductase(DHCR7)

ATP

Lanosterol

FF-MAS

T-MAS

Rate of GVBD oocytes (%)

Cumulus cells

80

Keto

BM

60
40
20
0

Cholesterol

CFL

P450scc(CYP11A1)

LH

AY

-

+

-

+

-

+

Progesterone (20ng/ml)
Pregnenolone

3βHS D
Progesterone

Progesterone receptor

Secretion

(a)

50

(a�)

Progesterone

Meiotic maturation

Fig. 3. Simplified outline of the progesterone biosynthesis pathway
and its role in oocyte meiotic maturation.
Three sources of cholesterol can be used as substrate: 1) exogenous cholesterol (HDL or LDL); 2) hydrolysis of storedcholesterol ester by cholesterol esterase; 3)
synthesized cholesterol from acetate.

(b)

(c)

(b�)

(c�)

apoptosis cells (%)

40

oocyte

30
20
10
0
C

FL

AY

BM

FL

cytes cultured with ketoconazole for 20 hr was significantly
lower than that of oocytes cultured without this drug (Fig.
4A). Although these data suggest that sterol biosynthesis
from lanosterol in cumulus cells is a factor involved in theacceleration of GVBD in mammalian oocytes, metabolism of
FF-MAS was also required for meiotic progression of cumulus-enclosed oocytes. Fig. 4A showed that the additionof 1 µM AY9944-A-7 or 20 µM BM15.766 to FSH+LHmedium significantly decreased GVBD as well as progesterone production. Furthermore, we showed that progesterone could reverse the negative effects of ketoconazole,
AY9944-A-7 or BM15.766 on GVBD (Fig. 4A). Judging

Fig. 4. (A) The effects of Ketoconazole, AY9944-A-7 or BM15.766,
and progesterone on GVBD in oocytes cultured for 20 hr. Values are mean +/- SEM of 3 replicates. (B) The effects of
AY9944-A-7 or BM15.766 on the cumulus cells expansion.
(a) COCs were cultured with FSH and LH for 28 h (a ) COCs
were cultured with FSH, LH, and progesterone (20ng/ml) for
28 h (b) COCs were cultured with FSH, LH, and AY9944-A-7(1μM) for 28 h (b ) COCs were cultured with FSH, LH,
AY9944-A-7 (1μM), and progesterone (20ng/ml) for 28 h (c)COCs were cultured with FSH, LH, and BM15.766 (20μM) for
28 h (c ) COCs were cultured with FSH, LH, BM15.766(20μM), and progesterone (29ng/ml) for 28 h.
(C) The effects of AY9944-A-7 or BM15.766 on the apoptosisof cumulus cells cultured for 28 h. Control: COCs were cultured
without FSH and LH for 28 hr. FL: COCs were cultured with
FSH and LH for 28 h. AY: COCs were cultured with FSH, LH,
and AY9944-A-7 (1uM) for 28 h. BM: COCs were cultured with
FSH, LH, and BM15.766 (20μM) for 28 h. Values are mean +/SEM of 3 replicates.

from these results, we propose that FF-MAS is synthesizedin cumulus cells but does not accumulate. Rather it is metabolized to cholesterol, which is then converted into progesterone. The secreted progesterone accelerates ongoing

icity (Fig. 4B), since the rate of apoptosis was as same as

GVBD of porcine oocytes in vitro (Fig. 3). Since the se-in control. Furthermore, cumulus expansion was restored

creted progesterone also impacts the process of cumulusby addition of 20 ng/ml progesterone. As such, we propose

expansion of porcine COCs [36], we analyzed cumulus ex-that the expression and functions of DHCR

pansion of porcine COCs which were cultured with choles-

in cumulus cells are required for gonadotropin-induced cu-

terol biosynthesis inhibitor. The cumulus expansion ofmulus cells differentiation and oocyte maturation via a pro-

COCs was also significantly suppressed when COCs were

gesterone- and PR-dependent pathway.

cultured with FSH, LH and either 1μ M AY9944-A-7 or
20μ M BM15.766 (Fig. 4B). The negative effects by both
inhibitors were not mediated by cumulus cell death by tox-

18

Japanese Journal of Reproductive Endocrinology Vol.10 2005

and DHCRThe Role of Cholesterol Biosynthesis Pathway in Cumulus Cells

Conclusion
Gonadotropins induce transcription of several genes of

10.

the Cyp superfamily that encode the enzymes essential forthe conversion acetyl-CoA to cholesterol and cholesterol toprogesterone. Progesterone, in turn, acts via receptor (nu-11.

clear and membrane) mediated mechanisms to support nuclear and cytoplasmic maturation of cumulus-enclosed oocytes, as well as COC expansion, not only in pig but also in

12.

rodent oocytes [6, 10, 11, 37, 38]. Thus, the up-regulationof the gene expression in COCs is required for proper meiotic maturation of oocytes with high developmental compe-

13.

tence in vitro.
Acknowledgment

14.

Thanks to Dr. JoAnne S. Richards, Professor of Baylor Collegeof Medicine for insightful comments and suggestions concerning15.

the manuscript. Mr. Tetsuji Okazaki and Mrs. Kaori Matsuo gave
us numerous helps. We thank the staff of the Meat Inspection
Office in Hiroshima City for supplying the porcine ovaries.
16.

References
１. Ainsworth L, Tsang BK, Downey BR, Marcus GJ, Armstrong DT(1980) Interrelationships between follicular fluid steroid levels,
gonadotropic stimuli, and oocyte maturation during preovulatory
development of porcine follicles. Biol Reprod 23, 621-627.
２. Richards JS (1994) Hormonal control of gene expression in theovary. Endocr Rev 15, 725-751.
３. Mori T, Suzuki A, Nishimura T, Kambegawa A (1977) Inhibitionof ovulation in immature rats by anti-progesterone antiserum. J
Endocrinol 73, 185-186.
４. Snyder BW, Beecham GD, Schane HP (1984) Inhibition of ovulation in rats with epostane, an inhibitor of 3 beta-hydroxysteroid
dehydrogenase. Proc Soc Exp Biol Med 176, 238-242.
５. Lydon JP, DeMayo FJ, Funk CR, Mani SK, Hughes AR, Montgomery CA Jr, Shyamala G, Conneely OM, O Malley BW (1995)
Mice lacking progesterone receptor exhibit pleiotropic reproductive abnormalities. Genes Dev 9, 2266-2278.
６. Chian RC, Ao A, Clarke HJ, Tulandi T, Tan SL (1999) Productionof steroids from human cumulus cells treated with different concentrations of gonadotropins during culture in vitro. Fertil Steril
71, 61-66.
７. Zhang X, Armstrong DT (1989) Effects of follicle-stimulating
hormone and ovarian steroids during in vitro meiotic maturation
on fertilization of rat oocytes. Gamete Res 23, 267-277.
８. Armstrong DT, Xia P, de Gannes G, Tekpetey FR, Khamsi F(1996) Differential effects of insulin-like growth factor-I and follicle-stimulating hormone on proliferation and differentiation ofbovine cumulus cells and granulosa cells. Biol Reprod 54,
331-338.
９. Xia P, Tekpetey FR, Armstrong DT (1994) Effect of IGF-I on pig

17.

18.

19.

20.

21.

22.
23.
24.

25.

26.
27.

oocyte maturation, fertilization, and early embryonic development
in vitro, and on granulosa and cumulus cell biosynthetic activity.
Mol Reprod Dev 38, 373-379.
Coskun S, Uzumcu M, Lin YC, Friedman CI, Alak BM (1995)
Regulation of cumulus cell steroidogenesis by the porcine oocyte
and preliminary characterization of oocyte-produced factor(s).
Biol Reprod 53, 670-675.
Shimada M, Terada T (2002) FSH and LH induce progesterone
production and progesterone receptor synthesis in cumulus cells:
a requirement for meiotic resumption in porcine oocytes. Mol
Hum Reprod 8, 612-618.
Shimada M, Kawano N, Terada T (2002) Delay of nuclear maturation and reduction in developmental competence of pig oocytes
after mineral oil overlay of in vitro maturation media. Reproduction 124, 557-564.
Okazaki T, Nishibori M, Yamashita Y, Shimada M (2003) LH reduces proliferative activity of cumulus cells and accelerates
GVBD of porcine oocytes. Mol Cell Endocrinol 209, 43-50.
Shimada M, Yamashita Y, Ito J, Okazaki T, Kawahata K, Nishibori
M (2004) Expression of two progesterone receptor isoforms incumulus cells and their roles during meiotic resumption of porcine oocytes. J Mol Endocrinol 33, 209-225.
Shimada M, Samizo N, Yamashita Y, Matsuo K, Terada T (2002)
Both Ca2+-PKC pathway and cAMP-PKA pathway require forprogesterone production in FSH- and LH-stimulated cumulus
cells during in vitro maturation of porcine oocytes. J Mamm Ova
Res 19, 81-88.
Simpson ER, Boyd GS (1967) The cholesterol side-chain cleavage system of bovine adrenal cortex. Eur J Biochem 2, 275-285.
Goldschmit D, Kraicer P, Orly J (1989) Periovulatory expressionof cholesterol side-chain cleavage cytochrome P-450 in cumulus
cells. Endocrinology 124, 369-378.
Strauss JF 3rd, Miller WL (1991) Molecular basis of ovarian steroid synthesis. In Hillier SG (ed), Ovatian Endocrinology Blackwell Scientific Publications, Oxford UK 25-72.
Gwynne JT, Strauss JF 3rd (1982) The role of lipoproteins in steroidogenesis and cholesterol metabolism in steroidogenic glands.
Endocr Rev 3, 299-329.
Reaven E, Chen YD, Spicher M, Azhar S (1984) Morphologicalevidence that high density lipoproteins are not internalized bysteroid-producing cells during in situ organ perfusion. J Clin Invest 74, 1384-1397.
Azhar S, Stewart D, Reaven E (1989) Utilization of cholesterolrich lipoproteins by perfused rat adrenals. J Lipid Res 30,
1799-1810.
Niswender GD (2002) Molecular control of luteal secretion ofprogesterone. Reproduction 123, 333-339.
Armstrong DT (1967) On the site of action of luteinizing hormone. Nature 213, 633-634.
Armstrong DT, Lee TP, Miller LS (1970) Stimulation of progesterone biosynthesis in bovine corpora lutea by luteinizing hormone in the presence of an inhibitor of cholesterol synthesis. Biol
Reprod 2, 29-36.
Rozman D, Waterman MR (1998) Lanosterol 14alpha-demethylase (CYP51) and spermatogenesis. Drug Metab Dispos 26,1199-1201.
Brown MS, Goldstein JL (1990) Atherosclerosis. Scavenging forreceptors Nature 343, 508-509.
Reynolds GA, Basu SK, Osborne TF, Chin DJ, Gil G, Brown MS,

MINI REVIEW

19

Masayuki SHIMADA et al.

33.

Goldstein JL, Luskey KL (1984) HMG CoA reductase: a negatively regulated gene with unusual promoter and 5 untranslated
regions. Cell 38, 275-285.
Osborne TF, Gil G, Brown MS, Kowal RC, Goldstein JL (1987)
Identification of promoter elements required for in vitro transcription of hamster 3-hydroxy-3-methylglutaryl coenzyme A reductase gene. Proc Natl Acad Sci USA 84, 3614-3618.
Osborne TF (1991) Single nucleotide resolution of sterol regulatory region in promoter for 3-hydroxy-3-methylglutaryl coenzyme A reductase. J Biol Chem 266, 13947-13951.
Metherall JE, Goldstein JL, Luskey KL, Brown MS (1989) Lossof transcriptional repression of three sterol-regulated genes inmutant hamster cells. J Biol Chem 264, 15634-15641.
Thaddeu GG, Strauss JF 3rd (1988) 8-bromoadenosine cyclic 3
5 -phosphate rapidly increases 3-hydoxy-2-methylglutaryl coenzyme A reductase mRNA in human granulusa cells: role of cellular sterol balance in controlling the response to tropic stimulation. Biochemistry 27, 3503-3506.
Yoshida Y, Yamashita C, Noshiro M, Fukuda M, Aoyama Y (1996)
Sterol 14-demethylase P450 activity expressed in rat gonads:
contribution to the formation of mammalian meiosis-activating
sterol. Biochem Biophys Res Commun 223, 534-538.
Yamashita Y, Nishibori M, Terada T, Isobe N and Shimada M

20

Japanese Journal of Reproductive Endocrinology Vol.10 2005

28.

29.

30.

31.

32.

34.

35.

36.

37.

38.

(2005) Gonadotropin-induced delta 14-reductase and delta 7-reductase gene expression in cumulus cells during meiotic resumption of porcine oocytes. Endocrinology 146, 186-194.
Byskov AG, Andersen CY, Nordholm L, Thogersen H, Xia G,
Wassmann O, Andersen JV, Guddal E, Roed T (1995) Chemical
structure of sterols that activate oocyte meiosis. Nature 374,559-562.
Downs SM, Ruan B, Schroepfer GJ Jr (2001) Meiosis-activating
sterol and the maturation of isolated mouse oocytes. Biol Reprod
64, 80-89.
Shimada M, Nishibori M, Yamashita Y, Ito J, Mori T, Richards JS(2004) Down-regulated expression of a disintegrin and metalloproteinase with thrombospondin-like repeats-1 by progesterone
receptor antagonist is associated with impaired expansion of porcine cumulus-oocyte complexes. Endocrinology 145, 4603-4614.
Mori T, Morimoto N, Kohda H, Nishimura T, Kambegawa A(1983) Meiosis-inhibiting effects in vivo of antiserum to progesterone on follicular ova in immature rats treated with gonadotropins. Endocrinol Jpn 30, 593-599.
Osborn JC, Moor RM, Crosby IM (1986) Effect of alterations infollicular steroidogenesis on the nuclear and cytoplasmic maturation of ovine oocytes. J Embryol Exp Morphol 98, 187-208.