Endocrine Journal 2008, 55 (6), 1043–1049
Ovarian Histological Findings in an Adult Patient with the Steroidogenic Acute Regulatory Protein (StAR) Deficiency Reveal the Impairment of Steroidogenesis by Lipoid Deposition
UIKO KAKU, KAORI KAMEYAMA*, MASAKO IZAWA, MAKOTO YAMADA, JUNKO MIYAMOTO,
TAKASHI SUZUKI**, HIRONOBU SASANO** AND YUKIHIRO HASEGAWA
Department of Endocrinology and Metabolism, Tokyo Metropolitan Kiyose Children’s Hospital, 1-3-1 Umezono, Kiyose, Tokyo
204-8567, Japan
*Division of Diagnostic Pathology, Keio University School of Medicine 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan
**Department of Pathology, Tohoku University School of Medicine, 1-1 Seiryou, Aoba-ku, Sendai 980-8575, Japan
Abstract. Context: The steroidogenic acute regulatory protein (StAR) is essential for the production of steroid hormones.
The mutations in the StAR gene typically cause congenital lipoid adrenal hyperplasia (lipoid CAH), characterized by
severe adrenal insufficiency in both sexes and complete female external genitalia in genetic males. Affected 46, XX
females feminize at puberty and menstruate but have progressive hypergonadotropic hypogonadism. It has been
hypothesized that the cholesterol accumulation in the steroidogenic cells destroys the residual steroidogenic capacity and
progressive ovarian failure occurs (two-hit model). Additionally, ovulation and luteinization in the patients is supposed to
be impaired. However, those hypotheses have not been confirmed histologically. Objective: We examined whether
pathological findings of the ovary in a patient of lipoid CAH corresponded with two-hit model, and whether ovulation and
luteinization occurred or not in the patient. Subject: The ovary in an adult 46, XX female with a homozygous nonsense
mutation (Q258X) in the StAR gene was examined. When the patient was age 22 yr, the ovary was resected because of
enlargement with polycysts and subsequent torsion. Result: The affected ovary demonstrated remarkable lipoid deposition
and changes of the mitochondrial ultrastructure. Immunohistochemical examination showed decrease of steroidogenic
enzymes such as P450 cholesterol side-chain cleavage (P450scc). Additionally, we detected corpus albicans in the
affected ovary. Conclusion: This is the first detailed report on ovarian histology in an adult 46, XX female with a null type
mutation of the StAR gene (Q258X), which indicates the evidence of the impairment of ovarian StAR-independent
steroidogenesis by lipoid deposition.
Key words: StAR, Ovulation, Corpus albicans, Two-hit model
(Endocrine Journal 55: 1043–1049, 2008)
STEROIDOGENIC acute regulatory protein (StAR) is essential for the production of steroid hormones,
mediating the transport of cholesterol from the outer
to the inner mitochondrial membrane in steroidogenic
tissues. A disorder due to StAR mutations, a clinical
entity traditionally called congenital lipoid adrenal
hyperplasia (lipoid CAH), results in severe impairment
of steroid biosynthesis in the adrenal glands and
gonads. In patients with null type mutations, clinical
manifestations of adrenal insufficiency become appar-
ent in the first few weeks of life. 46, XY individuals
have female external genitalia [1], and 46, XX indi-
Received: April 5, 2008
Accepted: August 5, 2008
Correspondence to: Uiko KAKU, M.D., Department of Endo-
crinology and Metabolism, Tokyo Metropolitan Kiyose Chil-
dren’s Hospital, 1-3-1 Umezono, Kiyose, Tokyo 204-8567, Japan
Abbreviation: StAR, steroidogenic acute regulatory protein;
lipoid CAH, congenital lipoid adrenal hyperplasia; P450scc, P450
cholesterol side-chain cleavage; P450c17, 17α-hydroxylase/
17,20-lyase; 3βHSD, 3β-hydroxysteroid dehydrogenase; Ad4BP/
SF-1, adrenal 4 binding protein/steroidogenic factor-1
KAKU et al.1044
viduals can have breast development and menstruation
at puberty spontaneously [2–4]. Despite null StAR
function, enough estrogen is produced in the ovary to
induce puberty in this disorder.
A two-hit model of lipoid CAH, which partially
explains these clinical phenotypes, was proposed by
Bose; the first hit is the loss of steroid hormone
synthesis which depends on StAR, resulting in hypo-
steroidogenesis. The second hit is the accumulation of
cholesterol, which damages the steroidogenic cells and
eventually disrupts StAR-independent steroidogenesis
[5]. The fetal testis, which synthesizes testosterone in
early gestation, is severely affected, so that virilization
does not occur in affected 46, XY patients with null
type mutations. On the other hand, the fetal ovary
lacks steroidogenic capacity, so cholesterol esters do
not accumulate until puberty. StAR-independent
estrogen synthesis in the ovaries first leads to breast
development and vaginal bleeding [2–4], followed by
a decrease in that residual synthesis with a progressive
accumulation of cholesterol esters.
Thus, the ovary in adult patient with complete StAR
deficiency (complete first hit) is the most appropriate
tissue to verify the second hit which is the disruption
of StAR-independent steroidogenesis. However, his-
tological analysis of the ovary was reported only by
Tanae et al., who demonstrated two female individuals
with lipoid adrenal hyperplasia had bilateral ovarian
cysts and lipid laden cell tumor after puberty [6].
Ovulation and luteinization in 46, XX patients with
null type mutations is supposed to be impaired because
of the following three reasons. First, in the ovary of
StAR knockout mice at 8 weeks of ages, no corpus
luteum was detected [7]. Second, the basal body tem-
perature was non-biphasic and urinaly pregnanediol
was undetectable in the affected human after puberty
begun [2]. Third, successful pregnancy has not been
reported in either StAR KO mice or affected human
females with null type mutations. However, impair-
ment of the corpus luteum formation and anovulation
have not been confirmed in the affected adult human
female.
Hence, we report in detail on the ovarian of an adult
46, XX female with complete StAR deficiency for the
first time. The findings strongly support the two-hit
model of lipoid CAH and demonstrate ovarian lutein-
ization.
Material and Methods
Case report
The patient, a Japanese female, was born after 41
weeks of an uneventful gestation with a birth weight of
3500 g. At the age of 2 days, pigmentation of her lips
was noted. One week later, the baby was admitted be-
cause of vomiting. Initial laboratory studies revealed
hyponatremia (121 mEq/l) and hyperkalemia (7.0
mEq/l). Serum cortisol level was low (5.2 μg/dl) and
plasma ACTH concentration was extremely high
(1746 pg/ml). On the basis of these findings, lipoid
CAH was suspected. A therapy with hydrocortisone
and fluorocortisone was instituted. A karyotype ob-
tained at this time was 46, XX. Later, we detected a
homozygous nonsense mutation (Q258X) in the StAR
gene. Indeed, the expression of the StAR was not
detected in the ovary of this patient by the immuno-
histochemical analysis in this study (data not shown).
This mutation is the most common one in Japanese pa-
tients with lipoid CAH [8]. Menarche was observed at
the age of 11 years and 3 months and her menstrual
cycle was regular for the first seven years. The basal
body temperature was not checked during her puberty.
Right multiple ovarian cysts were detected by ultra-
sonography at the age of 12 yr, when, basal levels of
luteinizing hormone (LH) and follicle-stimulating
hormone (FSH) were 17.6 mIU/ml and 7.5 mIU/ml,
respectively (LH, normal range 1.8–7.6 mIU/ml, FSH,
normal range 5.2–14.8 mIU/ml at pubertal stage) and
serum estradiol concentration was 17 pg/ml. After
that, basal LH levels were elevated to 40 mIU/ml,
whereas serum FSH levels remained within the normal
range. As she menstruated irregularly, replacement
therapy with progesterone was started at the age of
18 yr. By this age, breast development was at Tanner
stage 5, pubic hair was at Tanner stage 2, and axillary
hair was not found. Following the initiation of the
treatment, menstrual cycle became regular again. At
the age of 19 yr, she complained of acute pelvic and
abdominal pain and ultrasonography demonstrated an
enlarged right ovary (8.5 × 6.0 × 8.0 cm) with many
cysts. At laparotomy, we found complete torsion of
the right adnexa, and the left ovary was also enlarged
(about 5.0 × 2.8 × 4.0 cm) with multiple cysts. A right
oophorectomy was performed. One month later, treat-
ment with estrogen and progesterone were initiated to
prevent further enlargement of the left ovarian cysts.
OVARIAN HISTOLOGY IN StAR DEFICIENCY 1045
After the initiation of the treatment, basal LH and FSH
levels determined at 3-month intervals ranged from 9.5
to 22.2 mIU/ml and from 4.5 to 12.8 mIU/ml, respec-
tively and the sizes of the cysts and ovary were not
diminished. Serum estradiol concentration, which had
fluctuated from 17 to 60 pg/ml until the age of 19 yr,
were decreased and remained below 12 pg/ml after the
age of 19 years and 10 months. By the age of 22 yr,
she was admitted to the hospital four times because of
abdominal pain and adrenal crisis due to torsion of the
left enlarged ovary. Thus, left oophorectomy and its
histological analysis were performed after obtaining
informed consent.
Histological examination
Ovarian tissue was fixed in Bouin’s solution and
embedded in paraffin, according to routine techniques,
and cut at 4–7 μm. Sections were deparaffinized and
stained with hematoxylin and eosin.
Immunohistochemical analysis of P450 cholesterol
side-chain cleavage (P450scc), 17α-hydroxylase/17,20-
lyase (P450c17), 3β-hydroxysteroid dehydrogenase
(3βHSD) and adrenal 4 binding protein/steroidogenic
factor-1 (Ad4BP/SF-1) was performed as described
previously elsewhere [9–11]. For normal control, im-
munohistochemical analysis of these enzymes was
performed in a morphologically normal adult human
ovary.
For electron microscopic analyses, ovary was
trimmed free of fat, fixed in 0.1 M sodium phosphate
(pH 7.4) containing 1% paraformaldehyde and 2.5%
glutaraldehyde, prestained by use of 1% osmium
tetroxide in the above buffer, embedded in epoxy res-
in, and cut in 70-nm sections with an Ultratome 3.
Thin sections stained with uranyl acetate and lead
citrate were examined with a JEOL-1200EX trans-
mission electron microscope (Nihon Denshi, Tokyo).
Results
The left ovary was the size of 5.7 × 5.0 × 2.8 cm. At
least 10 cysts of 2–3 mm in diameter and some hemor-
rhagic regions were observed in the ovary. In spite of
several episodes of torsion, no remarkable necrotic
change was detected.
Histological findings support the two-hit model
The ovary contained abundant lipid deposits, prima-
rily in the theca cells and the stromal cells (Fig. 1).
First, to assess the lipoid accumulation mediated
damage to organelles, we examined the intracellular
ultrastructure of the theca cells. Electron microscopy
demonstrated many lipid droplets of approximately 1–
2 μm diameter accumulated within the cytosol, and the
number of mitochondria was decreased as compared
with normal cells (Fig. 2). Additionally, in some mito-
chondrias their bodies were swollen and their ultra-
structures were perturbed. Remarkable abnormality of
the ultrastructure was not seen in other organelles.
Next, we examined expression of steroidogenic
enzymes and Ad4BP/SF-1 by immunostaining of the
ovarian tissue. P450c17 were detected within the theca
interna cells and the stromal cells with lower expres-
sion level compared with normal ovary. The expres-
sion of 3βHSD and P450scc, which were detected in
the theca cells of normal ovary, were not observed in
the cells of the affected ovary. The immunohisto-
chemical localization and amount of Ad4BP/SF-1
were the same as normal ovary (Fig. 3).
Histological findings demonstrate luteinization
Oocytes with follicles were located in the cortex of
the ovary, and the amount of them was decreased com-
Fig. 1. Hematoxylin and eosin staining of ovarian tissue. The
theca cells had lipoid deposits (white arrow) and the
granulosa cells (arrow head) had no lipid. (magnifica-
tion, ×400)
KAKU et al.1046
pared with those of normal adult female. Although the
development up to secondary follicle with insufficient
development of the theca cells and the granulosa cells
was seen, graafian follicles were not detected in the
ovary. Similarly to StAR KO mice, no corpus luteum
was detected. However, we found corpus albicans,
which is a regressed form of corpus luteum. The size
and the number of corpus albicans were slightly small-
er and less than these in normal ovary (Fig. 4).
Discussion
The histological findings of the ovary in our patient
with complete StAR deficiency support the second hit
of two-hit model theory, that is, lipoid deposition in
the theca cells causes disruption of the StAR-indepen-
dent steroidogenesis. Similarly to past reports about
ovarian histology of human patients [6] and StAR KO
mice [7], abundant lipoid deposition was detected in
the adult female ovary, primarily in the theca cell,
where steroidogenesis begins during preovulatory
phase. In the theca cells mitochondrial ultrastructures
were changed. This change is suspected to result from
not ovarian torsion but lipoid deposition because the
granulosa cells had no deformed mitochondria (date
not shown).
Immunohistochemical analysis in this study demon-
strated that lipoid accumulation damaged not only
mitochondria but also other organelles. Diminished
expression of P450scc represented mitochondrial dys-
function, which was consistent with the findings by
electron microscopy. Furthermore, the expressions of
P450c17 and 3βHSD were decreased. These enzymes
are generally expressed in the microsome of the ste-
roidogenic cells. Because the expression of Ad4BP/
SF-1 was normal, it was speculated that the nucleus
was not severely damaged.
One study exhibited high levels of the P450scc in
the ovaries of StAR KO mice compared with the wild-
type ovaries at 8 weeks of age [7]. The reason the ex-
pression of P450scc, which was not observed in our
patient, was detected in StAR KO mice at 8 weeks is
currently unknown. One possible explanation is that
mitochondria damage by the accumulation of choles-
terol was more severe in the ovary of our patient than
StAR KO mice at 8 weeks. Indeed, serum estradiol
had been below the measurable level for half a year
before the ovary was resected in our patient, whereas
estradiol levels did not differ significantly from wild-
type values in StAR KO mice at 8 weeks [7]. Addi-
tionally, we speculated that the reason of the increased
expression of P450scc in KO mice was due to stimula-
tion by LH [12, 13].
The presence of the corpus luteum or corpus albi-
cans in patients with congenital lipoid CAH has not
been described in the past. Hasegawa et al. reported
the ovaries of StAR KO mice exhibited impaired folli-
cular maturation and absent corpus luteum at 8 weeks
of age, when sexual maturation normally has occurred.
In this case, we revealed the StAR deficient ovary had
corpus albicans. Corpus albicans is a regressed form
of corpus luteum, which normally develops from an
ovarian follicle following ovulation. Thus, the pres-
Fig. 2. Mitochondrial ultrastructure of the theca cell. Panel A indicates lower magnification view of the theca cell, whereas panel B
shows a high-power view of mitochondrial ultrastructure. L, lipid droplet; M, mitochondria. Multiple small lipid droplets of
approximately 1 to 2 μm diameter were detected in the theca cell. Mitochondria was swollen and the ultrastructure was
distorted.
OVARIAN HISTOLOGY IN StAR DEFICIENCY 1047
ence of the corpus albicans gave an indication of the
possibility of ovulation in the patient with StAR defi-
ciency. However, the indication is not infallible. A
rare disease, luteinized unruptured follicle syndrome,
is known to undergo luteinization without ovulation
[14]. In addition, progesterone receptor-null mice are
able to form corpus luteum containing trapped oocyte
[15]. Thus, because we had not monitored ovarian
follicle directly or ultrasonographically, we could not
confirm whether or not ovulation had actually oc-
curred in our patient.
Low level of progesterone does not necessarily
demonstrate the failure of luteinization. Despite the
formation of corpus albicans and presumably corpus
luteum, in our patient all of serum progesterone levels
had always been undetectable (date not shown) from
early in puberty as in past reports [2, 7]. Similarly, no-
biphasic basal body temperature does not necessarily
Fig. 3. Immunohistochemistry for human p450c17, 3βHSD, p450scc, and Ad4BP/SF-1 of the ovaries. A–D, normal ovary; E–H,
ovary of the patient with StAR deficiency. A and E, immunohistochemistry for p450c17; B and F, for 3βHSD; C and G, for
p450scc; D and H, for Ad4BP/SF-1. In the normal ovary, p450c17 remarkably expressed in the theca cells (A), but in the
ovary of the patient, the expression was decreased (E). Expression of 3βHSD and p450scc were detected mainly in the theca
cells and slightly in granulosa cells in the normal ovary (B and C), but not detected in the theca cells of affected ovary (F and
G). Expression of Ad4BP/SF-1 in the theca cells was comparable in normal ovary (D) and affected ovary (H). (magnification,
×200)
KAKU et al.1048
deny luteinization.
Increased secretion of LH is speculated to be re-
sponsible for the development of ovarian cyst in the
patient of lipoid CAH. The transgenic mice over-
expressing LH showed cysts formation and marked
enlargement of ovaries [16]. Additionally, hormonal
replacement initiated at the age of 14 yr diminished or
suppressed ovarian cysts [2, 4]. However, in our pa-
tient the progesterone and estrogen treatments initiated
at the age of 18 yr were unable to suppress her LH se-
cretion fully and to prevent the progressive ovarian
cyst formation. If we had started the hormonal treat-
ment earlier, the cyst enlargement and torsion of the
ovary would have been prevented.
In summary, our report about the ovarian histology
of an adult female with a complete StAR deficiency
(Q258X), which demonstrated markedly lipoid depos-
its, decreased expression of both mitochondrial and
microsomal steroidogenic enzymes, and deformation
of mitochondrial ultrastructure, strongly supports two-
hit model of lipoid CAH.
Acknowledgements
We thank Dr. Tomonobu Hasegawa (Department of
Endocrinology and Metabolism, Keio University
School of Medicine), and Dr. Masahiko Morikawa
(Department of Pathology, Tokyo Metropolitan
Kiyose Children’s Hospital) for supporting the histo-
logical analysis.
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