Introduction

INTRODUCTION
The Polycystic Ovary Syndrome (PCOS) is one of the most
common endocrine/metabolic disorders of women. This
syndrome was first described by Stein and Leventhal in 1935,
(1)
although the presence of sclerocystic ovaries had been
recognized for at least 90 years before the publication of that
seminal work. (1, 2)

PCOS is still a syndrome, namely a collection of signs

and features that characterize a disorder, where no single test
is diagnostic. It is a heterogeneous disorder, (2)
whose
principal features include androgen excess, ovulatory
dysfunction, and/or polycystic ovaries. (2, 3)

PCOS is a functional disorder of unclear etiology, and, it is

a diagnosis of exclusion, other androgen excess and ovulatory
disorders of clearly defined etiologies excluded. (4)

PCOS should be considered as a lifespan disorder. The

clinical features may change throughout the lifespan, starting
from adolescence to postmenopausal age. (4, 5)

Pathophysiology:

No single etiologic factor fully explains all the features of

PCOS. Excess androgen production is primarily from the ovary.
(6)
Increased testosterone production by theca cells of
polycystic ovaries and Increased LH secretion by the pituitary
gland are consistently demonstrated. (7)

1
 Introduction

Increased LH secretion: It is not clear if this is a

primary defect in the GnRH pulse generator or if this is a
secondary phenomenon. Increased LH stimulation to the
theca cells drives increased androgen production. (7, 8)

The mechanisms of LH increase secretion:

A particular characteristic of LH secretion in PCOS is

increased pulse frequency, the periodicity of which is
approximately 1 hour. This implies that corresponding
pulsatile release of hypothalamic GnRH is increased, (9)
or
increased LH sensitivity to subsequent GnRH stimulation, or
an abnormality of gonadotropin secretion in PCOS may be a
primary consequence of increased hypothalamic GnRH
activity. (10)

Chronic estrogen secretion associated with PCOS may

bring about an increase in LH, either by a direct effect as it
increases the fraction of individual gonadotropes responding
to GnRH, or indirectly by facilitating GnRH pulse frequency by
its effect on the hypothalamus. (11)

Hyperandrogenemia has also been implicated as a

potential cause of increased LH secretion in PCOS as high
circulating levels of androgen prevent the negative feedback
effects of estrogen and progesterone on LH pulse release. (12)

Insulin has a facilitative role for on GnRH-stimulated LH

release. Reduction of hyperinsulinemia by the administration
of insulin-lowering drugs to patients with PCOS resulted in

2
 Introduction

decreased mean serum levels of androgens and LH in some

cases. (13)

Excessive androgen production:

Both the ovary and the adrenal glands contribute to the

pool of increased circulating androgens, however, in PCOS, (14)

the major serum androgens, androstenedione and

testosterone, are produced by the ovary, whereas elevated
concentrations of DHEAS are derived from the adrenal glands.
(15)

Theca cell function

Numerous antral follicles are surrounded by hyperplastic

theca cells that are the predominant site of androgen
overproduction. PCOS women, to a degree, have abnormal
androgen production by the theca cell is an inherent defect of
steroid genesis. (16)

That excess ovarian androgen production is driven by:

abnormally increased secretion of pituitary LH or Increased
theca cell sensitivity to LH or Insulin may increase androgen
production from theca cells of women with PCOS though an
alternative pathway involving inositol glycan, a downstream
mediator of insulin signaling, (17)
or presence of gonadotropic
growth factors as insulin and insulin like growth factor (IGF).
Receptors for insulin, IGF-I, and IGF-II have been localized to
the theca compartment of ovaries of both normal women and
patients with PCOS. (18)
these growth factors are capable of

3
 Introduction

enhancing androgen responses to LH as IGF-I has been

attributed to increased LH receptor induction, greater
expression of steroidogenic enzymes, (19)
and enhanced LH-
induced cyclic AMP production.

Fig. 1. Ovarian steroid genesis in the theca and granulosa cells

Granulosa cell function

Ovarian follicle development is arrested at the midantral

stage of growth, and the granulosa cells that line these
follicles appear to be in various stages of degeneration. A
deficiency of aromatase activity may explain the lack of
follicle development as E2 concentrations were low in
follicular fluid, (20)
also an inability to sustain peak levels, in
contrast to the pattern of normal cells, which implied
suboptimal granulosa cell function. (21)
These findings link

4
 Introduction

granulosa cell sensitivity to increased FSH receptor binding in

granulosa cells from polycystic ovaries and provide insight
into a possible mechanism for increased E2 responsiveness to
FSH stimulation. (22)

PCOS adrenal function

Approximately 50% of women with PCOS exhibit

increased levels of DHEAS and 11-hydroxyandrostenedione,
indicating excess androgen production by the zona reticularis
of the adrenal gland. (23)

Circadian rhythms of serum dehydroepiandrosterone

(DHEA) and cortisol in women with PCOS were not different
from those exhibited by normal women. The mechanism for
adrenal hyperandrogenemia may arise from either altered
adrenal responsiveness to ACTH or abnormal adrenal
stimulation by factors other than ACTH. (12, 24)

In women with PCOS, serum 17-hydroxyprogesterone and

androstenedione responses to ACTH were significantly greater
in those with hyperinsulinemia compared with those with
normal insulin levels, (25)

Genetic considerations:

Multiple studies have shown evidence of familial

clustering of PCOS and the strong link between
hyperinsulinemia and hyperandrogenism also suggests that
the stimulatory effect of insulin on ovarian androgen

5
 Introduction

production is influenced by genetic predisposition; However,

the mode of inheritance of PCOS is autosomal dominant. (26)

Fig. 2. Multiple factors included in PCOS pathogenesis.

6
 Introduction

Specific criteria for PCOS:

Three major diagnostic criteria for PCO have been

proposed by the National Institute of Health (NIH 1990), the
Rotterdam European Society for Human Reproductive and
Embryology. PCOS could be diagnosed, after the exclusion of
related disorders, by two of three features: (a) oligo- or
anovulation, (b) clinical and/or biochemical signs of
hyperandrogenism, or (c) polycystic ovaries, (4, 27)
so:

Menstrual irregularity and chronic anovulation:

In PCOS, menstrual dysfunction is primarily characterized

by irregular, infrequent, or absent menstrual bleeding.
Commonly, bleeding is not preceded by premenstrual
symptoms, which is typical for anovulatory bleeding and
therefore is unpredictable, and monthly menstrual cyclisty is
never established. (28)

In PCOS, approximately 20% of women have complete

absence of menses, whereas 5% to 10% of patients show
regular ovulatory function. Recognition of normal ovulation in
PCOS is significant in that a history of regular menstrual
cycles does not exclude the diagnosis. (29)

Typically, this irregular pattern of bleeding is an extension

of postmenarchal irregularity, (30)
but in some PCOS women,
the onset of chronic anovulation emerges beyond
adolescence, but this is unusual so they apparently have

7
 Introduction

regular cycles at first and subsequently develop menstrual

irregularity in association with weight gain. (31)

Menses irregularities include mild or severe

oligomenorrhea (cycle length more than 3540 days or 8 or
less menstrual cycles per year) or amenorrhea (no cycles for
6 or more consecutive months). (32)
Chronic anovulation is one
of the most important criteria in the diagnosis of PCOS,
leading to persistent estrogen production. The thickened
endometrium is prone to superficial sloughing or tissue
breakdown in response to persistent secretion or spontaneous
decreases in circulating estrogen. (33)

The volume of blood loss associated with menstrual

irregularity is generally mild, but in women with significant
endometrial proliferation, (34)
the bleeding can be substantial
and may result in anemia with transient orthostatic
hypotension. Prolonged heavy bleeding should raise
consideration of abnormal endometrial hyperplasia and even
endometrial adenocarcinoma. (35, 36)

Amenorrhea or severe oligomenorrhea should prompt

evaluation for hypothalamic or pituitary causes for
anovulation by measuring serum FSH and estradiol levels,
must be considered if FSH levels are low or in the normal
range and estradiol level is _30 pg/mL. (37)

Hyperandrogenism or hyperandrogenemia:

8
 Introduction

In women with polycystic ovary syndrome (PCOS),

hyperandrogenism is clinically manifested by hirsutism acne
and androgenic alopecia, and it contributes to chronic
anovulation and menstrual dysfunction. Biochemically,
hyperandrogenism is established by elevated circulating
levels of serum total or unbound testosterone or
androstenedione. (38, 39)

The ovary is considered the main source of excess

androgens in women with PCOS, although excess adrenal
androgen production may also occur. Ovarian theca cells
produce androgen under the influence of LH, and it can be
modulated by a number of local growth factors, hormones
and cytokines. Theca cells display alterations in steroidogenic
activities, including increased expression and differential
regulation of genes required for steroidogenesis. (40)

A certain amount of intra ovarian androgen is essential

for normal follicular growth and for the synthesis of estradiol.
Nonetheless, when the synthesis of androgens is not co-
ordinated with the needs of a developing follicle, and is in
excess, poor follicle maturation and increased follicular
atresia results. In the normal ovary, LH acts on the theca
interstitialstromal cells, whereas FSH acts on granulosa cells.
According to the two-gonadotrophin, two-cell theory of
estrogen biosynthesis, the thecal compartment secretes
androgens in response to LH,(41) and the produced
androstenedione is converted in the granulosa cell to

9
 Introduction

oestrogens by the action of aromatase, which in turn is under

the influence of FSH. In PCOS, the ovarian theca cells are
increased in number, and they have increased steroidogenic
capacity caused by increased transcription and mRNA
synthesis. (42)

Nearly a half of the circulating testosterone in normal

adult women is derived from the peripheral conversion of
androstenedione; the remainder is derived from the ovary and
adrenal cortex. Testosterone in women varies not only with
the menstrual cycle but also with age, race, and body mass
index. Because Sex Hormone Binding Globulin (SHBG) is
present in excess in women, (43)
Free Testosterone
concentrations are driven mainly by SHBG abundance. The
Free Androgen Index depends on accurate measurements for
testosterone and SHBG. Having measured both testosterone
and SHBG, Free Testosterone should be calculated, which is
easily done using a fixed formula. Calculated Free
Testosterone is the most useful and a sensitive marker of
hyperandrogenemia in women. (44)

The access of androgens to target tissues is regulated by

SHBG. Serum SHBG levels are usually decreased in women
with hyperandrogenism, especially in association with PCOS.
Both high androgens and high insulin have been known to
lower SHBG. Obesity is also known to be associated with
decreased serum SHBG levels, but SHBG values may also be
genetically determined. (45)

10
 Introduction

The most distinctive and visible clinical feature of PCOS is

mild to severe hirsutism. The rate of hair growth is important
clinically because gradual and progressive growth indicates a
functional etiology, (39)
whereas the rapid, and or aggressive
appearance of thick, pigmented hair often suggests a
neoplastic source of androgen production. (43)

In PCOS, increased hair growth is commonly found on the

side of the face, upper lip, and chin, extending down to the
neck region, lower back, and inner thighs. This pattern of
hirsutism invariably is accompanied by extension of pubic hair
growth toward the umbilicus and may resemble a male
escutcheon. (46)

More severe cases include the appearance of hair on the

chest. Progressive hyperandrogenism may be associated with
temporal balding and male pattern baldness. Excessive hair
also may be found on the extremities and abdominal flank,
although these areas are not considered specific sites of
sexual hair growth. The degree of hair growth is commonly
determined subjectively, although standardized measurement
may be achieved using the Ferriman-Gallwey method, which
quantifies terminal hair over nine body areas. (46, 47)

A Ferriman-Gallwey score of greater than 7 is usually

regarded as hirsutism. In PCOS, the amount of hirsutism has
been correlated to serum androgen concentrations. However,
The rate and distribution of hair growth may vary among
individuals due to altered responses to androgens based on

11
 Introduction

ethnic differences, which may account for subtle changes in

the prevalence of PCOS in different parts of the world. (47)

Coexisting conditions that alter the bioactivity of

androgens, such as hypothyroidism and obesity, may also
give rise to excessive hair growth. These conditions are
associated with lowered SHBG, which provides increased
availability of free testosterone. (47, 48)

Patients with dermatologic signs suggestive of androgen

excess, principally hirsutism, should also be investigated for
the presence of PCOS. Between 85 and 95% of patients with
frank hirsutism (i.e., a modified Ferriman-Gallwey [mF-G]
score of 68) will have PCOS when evaluated. (49)

We should also note that the sole complaint of

unwanted hair growth in the absence of frank hirsutism is a
strong predictor of PCOS. Minimal unwanted hair growth and
an mF-G score of 5 or less, 50% demonstrated PCOS; among
patients with menstrual irregularities, 65% had an underlying
androgen excess disorder, while 22% of those women who
reported being eumenorrheic were so affected. (48, 49)

Alternatively, while complaints or signs of acne and scalp

hair thinning are suggestive of PCOS, their predictive value
for the disorder is somewhat less than is the presence of
hirsutism or a complaint of unwanted hair, between 20 and
40% of patients with persistent acne-only , and only 10% of
those women with alopecia-only will have PCOS . (50)

12
 Introduction

Exclusion of other androgen excess or ovulatory

disorders:

As PCOS is a diagnosis of exclusion, the diagnosis can

only be arrived at after other disorders have been excluded.
These include 21-hydroxylase deficient NCAH (by a basal
and/or stimulated 17-hydroxyprogesterone [17-HP] level),
androgen-secreting neoplasms (by history and clinical exam
and appropriate studies in selected patients), (22)

adrenocortical hyperactivity (by clinical exam and appropriate

testing), and drug-induced hyperandrogenism (by history).
Overall, these disorders account for 510% of all women with
androgen excess. (51)

In addition, we should exclude thyroid dysfunction and

hyperprolactinemia by measuring a TSH, and prolactin level,
although the prevalence of these endocrine abnormalities in
patients with apparent PCOS is relatively low, on the order of
13%. Likewise, if polycystic ovaries are detected, particularly
in the absence of overt signs of androgen excess, disorders
that can result in this ovarian morphology need to be also
excluded, such as hypothalamic amenorrhea. (52)

Ovarian morphology

Ultrasound assessment of the ovaries:

Assessment of the size and number of follicles, and

volume are key to making the sonographic diagnosis of PCO.
The consensus definition is that a polycystic ovary should

13
 Introduction

have 12 or more follicles of 29 mm in diameter (figure 5&6).

(53)

Fig. 4. Typical polycystic ovary. Fig.5.Typical polycystic ovary as

described by Adams, with
more than 12 follicles of 29
mm in diameter in one plane,
arranged peripherally around
an echo dense stroma.

In the past, it was thought that the follicles in polycystic

ovaries were arranged peripherally just beneath the surface
of the ovary, but it is now accepted that the distribution of the
follicles is unimportant. (53, 54)

Increased stromal echogenicity of the ovary is a

subjective assessment depending on the operator, the
settings of the ultrasound machine and the size of the patient.
However, it has been shown that ovarian volume correlates
well to the increase in stroma and therefore measurement of
the ovarian volume remains crucial. (54)

The Rotterdam ESHRE/ASRM-sponsored PCOS consensus

workshop group defines the increased volume as greater than
10 cm3. They specify that the ovarian volume should be

14
 Introduction

calculated using the simplified formula for a prorate ellipsoid

(0.53 x length x width x thickness). In practice, modern
ultrasound machines have this formula built in to the software
program for volume calculation, the normal ovaries never
have a volume of greater than 8.0 cm3. The consensus
definition states that only one ovary fulfilling the criteria is
sufficient to define PCO. (55)

The ultrasound findings fulfilling the criteria above have

to be taken in the context of the clinical presentation,
together with appropriate endocrine, biochemical and
metabolic tests, For example, abnormalities of basal serum
prolactin or FSH levels may indicate a coexistent
hypothalamic-pituitary disorder or incipient ovarian failure. (56)

In women taking the combined oral contraceptive pill, the

ovarian volume may be within the normal range, but the
appearance may still be polycystic. (54, 55)
Polycystic ovaries
may also be found incidentally in post-menopausal women
and whilst, not surprisingly, they are smaller than in pre-
menopausal women. (56)

Laboratory evaluation:

Patients suspected of having PCOS can be subdivided into

four groups:

a) Women with overt long-term oligomenorrhea and

hirsutism: these women basically have PCOS, pending

15
 Introduction

exclusion of related disorders. At a minimum, these

women should undergo measurement of:

1- Circulating TSH, prolactin.

2- 17-HP levels.

If these values are normal, then the patient is

presumed to have PCOS. (22, 57)

b) Women with overt long-term oligomenorrhea, but no

obvious sign of androgen excess:

These women should undergo measurement of

circulating androgen levels (generally total and free
testosterone, and DHEAS) and, if elevated, assessment
of TSH, prolactin, and 17-HP levels. If these latter
values are normal: then the patient is presumed to
have PCOS.(22, 57)

c) Women with hirsutism but apparent eumenorrhea:

These women should undergo confirmation of

ovulation (most simply by measuring a P4 level in the
luteal phase of the menstrual cycle, i.e., day 2024 of
the cycle) on one or two cycles. They should also
undergo ovarian ultrasonography. If the patient is
found to have anovulation or polycystic ovaries on
ultrasonography, they should undergo measurement of
TSH, prolactin, and 17-HP levels. If these values are
normal, then the patient is presumed to have PCOS
(either classic PCOS if anovulatory, or ovulatory

16
 Introduction

PCOS if she has polycystic ovaries but normal

ovulation). (58)

The use of 17-HP levels to screen for NCAH: 21-

hydroxylase deficient NCAH affects 110% (depending on
ethnicity) of patients with androgen excess. Approximately
90% of NCAH patients can be detected by a basal level of 17-
HP > 2 ng/mL. The blood sample should be obtained in the
morning, and, most importantly, in the follicular
(preovulatory) phase of the menstrual cycle. Obviously, the
use of any corticosteroids will artificially suppress 17-HP
levels and should not be used prior to screening. (58)

The Value less than 2 ng/mL virtually exclude NCAH. If

the screening 17-HP level is more than 2 ng /mL, and the
investigator is certain that the sample was not drawn in the
luteal (postovulatory) phase of the cycle. ACTH stimulation
test is performed. Values of 17-HP > 1012 ng/mL at 3060
min. after the IV administration of ACTH are diagnostic for
NCAH. (22, 59)

The reversed ratio of serum luteinizing hormone (LH) to

follicle-stimulating hormone (FSH) is no longer accepted as a
criterion for diagnosis of PCOS. (60)

Related disorder to PCOS

Insulin resistance in PCOS:

Insulin resistance results in hyperinsulinemia which in

turn stimulates androgen secretion by theca cells, and

17
 Introduction

suppresses the hepatic production of SHBG. Thus, both

obesity and insulin resistance lead to lower SHBG levels and
higher bioavailable levels of androgens. In fact, SHBG may
become a measure in the future that reflects both
abnormalities in ovarian production and insulin resistance. (61)

The prevalence of insulin resistance in PCOS has been

reported to range from 20% to 40%. The common occurrence
of insulin resistance in obesity may account, in part, for the
rather wide prevalence that insulin resistance may worsen the
clinical manifestations of PCOS. Administration of insulin-
lowering drugs has been shown to improve insulin sensitivity,
reduce androgen levels, and restore ovulation in some, but
not all, patients. (61, 62)

Obesity:

Obesity was present in slightly more than 50% of PCOS

cases. However, of recent note, the rate of obesity associated
with PCOS has not been corroborated and there is a growing
impression that the incidence may be greater. (62)

Commonly an increase in the upper body or central

distribution of fat gives rise to an increased waist-to-hip ratio
compared with obese women without PCOS. This fat
distribution pattern has been termed android obesity and can
be found in other hyperandrogenic states, diabetes, and
hyperlipidemia. Notably, there is a preponderance of visceral
fat compared with peripheral fat. (63)

Infertility& pregnancy complication in PCOS:

18
 Introduction

A significant number of patients have infertility as a

presenting feature of PCOS, due to chronic anovulation , also
women with PCOS have a higher incidence of spontaneous
pregnancy loss, recurrent miscarriage was ranging from 25 to
37% in PCOS patients compared with 1825% in normal
women.(64, 65)
The mechanism of which remains unclear but
insulin resistance may be the cause. Hyperinsulinaemia in
PCOS, particularly in the obese , is strongly associated with
elevated concentrations of plasminogen activator inhibitor-1
(PAI-1) which is a potent inhibitor of fibrinolysis, and high
serum concentrations may be a factor in the etiology of early
pregnancy loss. (65)

Treatment of PCOS:

Generally there are four issues in the management of

PCOS patients: regulation of menses, control of hirsutism,
fertility issues, and the management of the insulin resistance
IR syndrome and its associated risks (type II diabetes mellitus,
dyslipidemia, and cardiovascular disease). (66)

19
 Introduction

Table (1): Treatment of women with polycystic ovary

syndrome.

complaint Treatment options

Metformin; clomiphene or tamoxifen;
Infertility
letrozole; gonadotropins; ovarian cautery
Contraceptive + antiandrogen
Skin manifestations ( spironolactone, flutamide' finastride);
GnRh agonists
Dysfunctional bleeding Cyclic progestogen; oral contraceptives
Weigh / metabolic
Diet/lifestyle management; metformin
concerns

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 Introduction

Tamoxifen

Tamoxifen was initially developed as a fertility agent, and

has been used for this indication in some countries. In the
early 1970 tamoxifen was found to be an effective treatment
of breast cancer and has since become a primary therapy for
metastatic breast cancer (67)

Tamoxifen the first triphenylethylene antiestrogen was

found to be antiestrogenic effects in the treatment of
infertility and breast cancer. Although these agents are widely
described as antiestrogens, they are best characterized
pharmacologically as partial estrogen agonists. (67, 68)

Chemistry:

Tamoxifen Citrate is the trans-isomer of a

triphenylethylene derivative. The structural and empirical
formulas are:

Fig. 6. Tamoxifen chemical structure

Tamoxifen Citrate has a molecular weight of 563.62, the

equilibrium solubility in water at 37 oC is 0.5 mg/mL and in
0.02 N HCl at 37oC (figure 7).

21
 Introduction

Pharmacodynamics:

Tamoxifen, is a nonsteroidal agent, binds to estrogen

receptor (ER) which in turn interacts with DNA. The
ER/tamoxifen complex recruits other proteins known as co-
repressors to stop genes being switched on by estrogen.
Tamoxifen function can be regulated by a number of different
variables including growth factors. (69)

Tamoxifen itself is a prod rug, having relatively little

affinity for its target protein, the estrogen receptor.It is
metabolized in the liver by the cytochrome P450 isoform
CYP2D6 and CYP3A4 into active metabolites such as 4-
hydroxytamoxifen and N-desmethyl-4-hydroxytamoxifen
(endoxifen) which have 30-100 times more affinity with the
estrogen receptor than tamoxifen itself. These active
metabolites compete with estrogen in the body for binding to
the estrogen receptor. (70)

The estrogenic or antiestrogenic effects of these agents

depend on level of endogenous estrogen and the target organ
affected, these agents display antiestrogen effects such as
augmentation of ovarian folliculogenesis, inhibition of
endometrial proliferation, vasodilatation, and increased bone
resorption. (70, 71)
Tamoxifen has potent antiestrogenic
properties which compete with estrogen for binding sites in
breast and other tissues. Tamoxifen causes cells to remain in
the G0 and G1 phases of the cell cycle. Because it prevents

22
 Introduction

(pre)cancerous cells from dividing but does not cause cell

death, tamoxifen is cytostatic rather than cytocidal. (72)

Tamoxifen and ovulation induction:

Tamoxifen is generally considered to increase fertility

rates in a similar way to clomiphene. In contrast to
clomiphene, however, tamoxifen does not increase follicular
phase FSH and LH levels, although there is an increase in
estradiol levels and luteal phase progesterone. It has been
postulated that tamoxifen improves follicular development by
direct action on the ovary rather than through the
hypothalamic-pituitary axis. (72, 73)

Tamoxifen acts as an agonist on the estrogen receptors of

the vaginal mucosa and endometrium. The effects of
tamoxifen on cervical mucus remains contradictory. (74)

Patients with variant forms of the gene CYP2D6 (also

called simply 2D6) may not receive full benefit from
tamoxifen because of too slow metabolism of the tamoxifen
pro drug into its active metabolite 4-hydroxytamoxifen, For
example; Selective serotonin reuptake inhibitor (SSRI)
antidepressants such as Paxil, Prozac, etc., can decrease the
effectiveness of tamoxifen, because these drugs compete for
the CYP2D6 enzyme which is needed to metabolize tamoxifen
into the active form endoxifen. (75)

Pharmacokinetics:

23
 Introduction

Tamoxifen is metabolized by hydroxylation reactions. The

metabolite 4-OH-tamoxifen is a potent ER antagonist that
circulates at nanomolar concentrations during chronic
tamoxifen therapy. Tamoxifen has a long elimination half-time
of approximately 4 to 7 days, whereas its major circulating
metabolite, N-desmethyl-tamoxifen, has an elimination half-
life of 7 to 14 days. Tamoxifen binds to plasma albumin, and is
extensively bound in tissues such as the liver, uterus, and
breast. (75, 76)

Although several hypotheses of tamoxifen's mechanism

of action have been proposed, the primary effect of tamoxifen
is to inhibit estrogen binding to the estrogen receptor. (76)

24
 Introduction

Volume and distribution:

Following the oral dose of 20 mg tamoxifen, an average

peak plasma concentration of 40 ng /mL (range 35 to 45
ng /mL) occurred approximately 5 hours after dosing. The
decline in plasma concentrations of tamoxifen is biphasic with
a terminal elimination of the half-life is about 5 to 7 days. The
average peak plasma concentration for N-desmethyl
tamoxifen is 15 ng /mL (range 10 to 20 ng /mL). (77)

Excretion:

Fecal excretion is the primary route of elimination. The

drug is excreted mainly as polar conjugates, with unchanged
drug and unconjugated metabolites accounting for less than
30% of the total fecal radioactivity. (78)

Adverse Effects of Tamoxifen Therapy:

Tamoxifen has adverse effects that are consistent with its

pharmacologic classification as a partial estrogen agonist.
Chronic tamoxifen treatment leads to endometrial thickening
in the majority of postmenopausal breast cancer patients.
And has been associated with an increased risk of
endometrial cancer. (78)

Tamoxifen also shows partial estrogen agonist effects on

lipoprotein concentrations, bone mineral density, and vaginal
epithelial cytology in postmenopausal women, Tamoxifen
causes increased venous thromboembolic events and shows

25
 Introduction

alterations in plasma hemostatic parameters that are similar

to estrogen hormone replacement therapy. Tamoxifen use has
been associated with an increased incidence of ovarian cysts
in both premenopausal and postmenopausal women. (79)

In premenopausal women, tamoxifen produces

moderate decreases in bone mineral density and causes hot
flushes. (78, 79)

Toxicity:

Acute neurotoxicity manifested by tremor, hyperreflexia,

unsteady gait and dizziness were noted. These symptoms
occurred within 3-5 days of beginning tamoxifen and cleared
within 2-5 days after stopping therapy. No permanent
neurologic toxicity was noted. (80)

Contraindications:

Tamoxifen is contraindicated in patients with a history of

stroke, deep venous thrombosis or pulmonary embolism, and
in patients who are at an increased risk of developing
endometrial cancer. (81)

26
 Introduction

Anti-Mllerian hormone (AMH)

Anti-Mllerian hormone (AMH) is a member of the

transforming growth factor family of growth and
differentiation factors.

Physiology of anti-mullerian hormone:

In the ovary, AMH has an inhibitory effect on primordial

follicle recruitment as well as on the responsiveness of
growing follicles to follicle-stimulating hormone (FSH). The
ovary-specific expression pattern in granulosa cells of growing
non selected follicles makes AMH an ideal marker for the size
of the ovarian follicle pool. (82)

AMH has regulatory effect in male sex differentiation.

AMH, produced by the Sertoli cells of the fetal testis induces
the regression of the Mullerian ducts. (82)
In the ovary AMH
expression starts in the columnar granulosa cells of primary
follicles immediately after differentiation from the flattened
pregranulosa cells of primordial follicles. Expression is highest

27
 Introduction

in granulosa cells of preantral and small antral follicles. AMH

is no longer expressed during the FSH dependent final stages
of follicle growth. AMH expression disappears when follicles
become atretic. (83, 84)

In the absence of AMH, primordial follicles are recruited at

a faster rate. Consequently, the primordial follicle pool is
prematurely exhausted. In the absence of AMH, follicles are
more sensitive to FSH. (84)

AMH attenuates the FSH-dependent increase in

aromatase activity and LH receptor expression, this inhibitory
effect of AMH on FSH sensitivity of follicles could play a role in
the process of selection. In women, AMH expression can first
be observed in granulose cells of primary follicles, and
expression is strongest in preantral and small antral follicles
(#4 mm).(85) AMH expression disappears in follicles of
increasing size and is almost lost in follicles larger than 8 mm,
where only very weak staining remains, restricted to the
granulosa cells of the cumulus . This expression pattern
suggests that, also in human AMH may play a role in initial
recruitment and in the selection of the dominant follicle. (84, 85)

AMH receptors:

From the chemical point of view AMH is a peptide

homodimer of molecular weight 140 kDa, consisting of two
identical glycoprotein subunits, connected by disulfide
bridges. The gene for human AMH is localized on the short

28
 Introduction

arm of chromosome 19. Target organs for AMH in males are

Mllerian ducts, and in both sexes gonads. (86)

The receptors for AMH are Trans membrane heteromeric

proteins, composed from two subunits, denoted Type I and II.
As all the receptors for growth factors of the TGF beta family,
they do not use G-proteins and possess intrinsic kinase
activity. Type II (better subunit) binds specifically the ligand
leading thus to activation of the type I, the intracellular part
of which acts as threonine kinase. Activation of the latter
starts a signal cascade resulting in a respective biological
response. The gene encoding for AMH receptor is localized on
chromosome 12. (86, 87)

The changes of AMH expression follow the development

of the male reproductive system, first of all changes in the
activity of hypothalamo-pituitary-gonadal axis. They may be
divided into four main stages: fetal and early postnatal
period, childhood, puberty and adulthood. The variations in
AMH formation are well reflected by its blood levels. (88)

Fetal and early postnatal period:

As mentioned above, AMH is synthesized in sertoli cells of

fetal testes already in early stage of embryonic development.
In this period hypothalamus already produces gonadotrophic
releasing hormone (Gn-RH), which stimulates secretion of

29
 Introduction

pituitary gonadotropins follicle stimulating hormone (FSH)

and luteinizing hormone (LH). (89)

By action of LH on Leydig cells of fetal testes via present

receptors relatively high amounts of testosterone are formed
(as detected for instance in newborns circulation).
Testosterone is responsible for differentiation of Wolfian ducts.
At the same time testosterone would inhibit AMH formation in
Sertoli cells through androgen receptors (AR). On the other
hand, FSH through its receptors on the membrane of Sertoli
cells, stimulates AMH expression. (90)

FSH utilizes here the classical mechanism involving

binding to the receptor and activation of adenylatecyclase
effector through G-protein.(90,91) Cyclic
adenosinemonophoshate (cAMP), formed by action of
adenylate cyclase, activates a number of kinases, among the
first protein kinase A, thus starting signaling cascade, leading
to activation and following translocation of nuclear
transcription factors. They bind then to the respective
responsive elements in the promoter region of the AMH gene,
resulting in its expression. (91) Inhibin B is also formed and
Sertoli cells, constituting in this stage about one half of the
whole testicular tissue, which undergo proliferation. Serum
levels of AMH are relatively high in this stage, comparable
with the levels in childhood (roughly up to the eighth year of
boys, as shown below. (92)

30
 Introduction

Childhood and early pre-pubertal period:

Childhood, until pre-pubertal- stage is a period of a

relative rest of the HPG axis and it is sometimes called a
stage of hypogonadotropic hypogonadism. Leydig cells
produce only very low amounts of testosterone, almost a half
of which in addition originates from adrenals. Sertoli cells are
still immature and spermatogenesis is arrested in a pre-
meiotic stage. The latter cells, however represent the main
portion of testicular tissue and thanks to FSH stimulation
produce AMH in amounts, comparable with prenatal period.
(91, 93)

Puberty:

With the onset of puberty the secretion of hypothalamic

Gn-RH and both gonadotropins increase again, but the effect
of LH is much more pronounced, due to inhibitory effect of
inhibin B on FSH secretion. Leydig cells undergo further
differentiation and dramatically increases testosterone
formation, which invokes also maturation of Sertoli cells. (93)

The inhibitory effect of testosterone prevails over FSH

stimulation, resulting in down-regulation of AMH expression,
the levels of which rapidly sink. Germinal cells undergo
meiosis and spermatogenesis begins. In the late puberty
stage (Tanner V), the germinal cells, in contrast to Sertoli
cells, represent already the major portion of testicular tissue.

31
 Introduction

The secretion of AMH reaches adult values and it is

maintained almost constant until the rest of life. (94)

Clinical utility of AMH measurement:

1. AMH as a marker for ovarian aging.

Direct measurement of the primordial follicle pool is

impossible. However, the number of primordial follicles is
indirectly reflected by the number of growing follicles. (95)

Hence, a factor primarily secreted by growing follicles will

reflect the size of the primordial follicle pool. Since AMH is
expressed by growing follicles up to selection, and can be
detected in serum it is a good candidate. (95, 96)

In young normal ovulatory women, early follicular phase

hormone measurements at 3-year intervals revealed that
serum AMH levels decline significantly whereas serum levels
of FSH and inhibin b and the number of antral follicles do not
change during this interval. Stratification of age revealed that
both serum AMH levels and numbers of antral follicles decline
with age. Importantly, a strong correlation of serum AMH
levels with AFC was observed. (97)

Substantially elevated serum levels of FSH are not found

until cycles have already become irregular. Therefore, a
marker that already shows a considerable change when
cyclisty is still normal would better identify women with
declining fertility. (96, 98)
The usefulness of serum AMH levels as
a measure of the ovarian reserve was recently shown in

32
 Introduction

young women after treatment for childhood cancer.

Chemotherapy and radiotherapy treatment have adverse
effects on the ovary in particular, resulting in loss of
primordial follicles. Indeed, in cancer survivors, the partial
loss of the ovarian reserve is reflected by increased FSH
levels and decreased ovarian volume. Unexpectedly, the
number of small antral follicles is unchanged, (96)
a finding that
may reflect the low accuracy and observer dependency of
AFC measurements. Nevertheless, serum AMH levels were
decreased in these patients, supporting the use of serum AMH
levels as an early predictor of the ovarian reserve. (96)

2. AMH as a marker of ovarian responsiveness

In women undergoing treatment for infertility, ovarian

aging is characterized by decreased ovarian responsiveness
to exogenous gonadotropin administration and poor
pregnancy outcome. On the one hand, correct identification of
poor responders by assessment of their ovarian reserve
before entering an in vitro fertilization (IVF) program is
important. On the other hand, assessment of the ovarian
reserve may also benefit patients that would generally be
excluded from IVF programs because of advanced age. (99)

Several studies have shown that AMH is an excellent marker

to determine ovarian responsiveness also in an IVF program
AMH serum levels were shown to be highly correlated with the
number of antral follicles before treatment and number of

33
 Introduction

oocytes retrieved upon ovarian stimulation. Measurement of

serum AMH levels has several advantages over other serum
markers such as FSH, inhibin B and E2. To achieve a reliable
predictive outcome, one single hormone measurement for
AMH seems sufficient. Furthermore, in contrast to FSH, inhibin
B and E2, AMH levels
remain relatively constant during the follicular phase and
entire menstrual cycle (99, 100)

3. AMH as a marker for ovarian pathophysiology

Serum AMH level can also serve as a marker in ovarian

pathophysiology, such as polycystic ovary syndrome (PCOS),
in which the antral follicle pool is enlarged. (101)The defective
selection mechanism results in an accumulation of small
antral follicles, which contribute significantly to the production
of AMH. (102)
AMH lowers the sensitivity of follicles to FSH,
possibly contributing to deranged follicle selection. It has
been suggested that aromatase activity in PCOS patients
might be decreased because follicles from PCOS women do
not produce large amounts of E2. AMH also inhibits aromatase
activity; AMH contributes to the severity of PCOS. It shows
that follicular fluid and serum of PCOS women contained
increased AMH levels. (102, 103)

In agreement with previous results obtained in normal

cycling women, also in PCOS women serum AMH levels were
correlated with antral follicle number. The two- to threefold

34
 Introduction

increase in the number of growing follicles is reflected by a

two- to threefold increase in serum AMH level. (104)

In PCOS, the follicular excess is mainly caused by an

increase of small antral follicles up to 25mm in size.
Interestingly, in follicles beyond this stage, AMH expression
diminishes. Therefore, it is not surprising that serum AMH
levels positively correlate with the number of 25 mm, but not
69 mm, follicles in PCOS women. (105)
The finding that AMH
levels are also increased in the follicular fluid of PCOS women
suggests that the increase in serum AMH levels is not only
due to an increase in the number of growing follicles, but may
also result from increased AMH production per follicle. As in
cycling women, AMH levels decline with increasing age in
PCOS. However, the decrease in serum levels is significantly
different from that in controls. (106)

4. AMH as a marker of subfertility due to

postponement of childbearing:

Measurement of AMH levels to assess the ovarian reserve

may also be of interest in women in general. Assessment of
the ovarian reserve, at least of the size of the ovarian follicle
pool, may provide insight into the number of fertile years a
woman has left. However, in order to determine whether
serum AMH level has prognostic value, additional prospective
studies in a normal population are necessary to provide
definite proof for this concept. (106, 107)
In conclusion, recent
studies have validated the use of serum AMH levels as a

35
 Introduction

marker for the quantitative aspect of ovarian reserve.

Because AMH levels are strongly correlated with the size of
the follicle pool, and because of the lack of cycle variations,
serum levels of AMH are a good candidate for inclusion in
standard diagnostic procedures to assess other ovarian
dysfunctions, such as premature ovarian failure. Knowledge of
the serum AMH levels in such conditions might provide more
insight into the possible cause or effect of altered AMH levels.
(108)

36
 Aim of the Work

AIM OF THE WORK

The aim of this work was to determine if there is a level of
both antimullrian hormone and total testosterone hormone at
which women with polycystic ovaries ovulate in response to
the standard dose of tamoxifen.

37
 Patients

PATIENTS
This study was carried out upon 80 women diagnosed to
have polycystic ovary syndrome for ovulation induction, all
patient were given tamoxifen 20 mg per day from the day two
of the cycle for five days. Blood samples were collected from
the patients (day 3-5) to estimate the level of the
antimullerian homone and the total testosterone hormone.
They were recruited from the infertility clinic of El Shatby
University Maternity Hospital from March 2014 to June 2015.

The sample size was calculated using Epi-Info 2002

software. Using a power of 80% to detect a significant ovulation
rate after ovarian induction by tamoxifen.

Inclusion criteria:

1 Age group (20 34 years).

2 Primary infertility.
3 Polycystic ovaries (12 or more follicles measuring 2-9
mm in diameter or increased ovarian volume (>10 cm3)
in each ovary diagnosed by ultrasound examination.

4 Oligomenorrhea due to chronic anovulation.

Exclusion criteria:

1. History of ovarian surgery (cyst or ovarian drilling or

chocolate cyst).
2. Patient with galactorrhea or documented
hyperprolactinemia.

38
 Patients

3. Patient who received ovarian stimulation in the last

three monthes.

39
 Methods

METHODS
All cases were subjected to the following after informed
consent was signed:

I. History taking:

II. Physical Examination:

1. General Examination:
2. Abdominal and pelvic examination

III. Transvaginal Ultrasound (MINDRAY D.P. 2200): which

done in the day 2 of the cycle, basal scanning to assess
the uterus (as regards the endometrial thickness), the
ovaries (as regards the ovarian volume, the antral
follicular count and the size of the follicles) and to exclude
the presence of any ovarian or adnexal pathology.

IV. Estimation of the Serum level of antimullrian

hormone (109)
done by ELIZA and total testosterone
hormone (110) done by ELIZA technique, it will be
estimated day 3 -5 of the cycle.

V. A standard dose of tamoxifen (NOLVADEX 10 milligram

tablet, Astra Zeneca) twenty milligram per day (two
tablets), starting on day 2 of the cycle for 5 days is to be
given.

VI. Follow up by transvaginal ultrasound from the day

8 of the cycle every 3 days for the following two weeks

40
 Methods

will be done to evaluate the ovarian response by defining

the number of growing follicles in both ovaries and
detection of the ovulation by either disappearance of the
mature follicle, Corpus luteum formation, trilaminar
endometrium and minimal fluid collection in the doglus
pouch.

VII. Serum progesterone was done one week after the

predicted time of ovulation by ultrasound.

41
 Results

RESULTS
This study included 80 women with polycystic ovaries, determining the
serum level of antimullerian hormone (AMH) and total testosterone level at
which ovulation occurs in response to standard dose of tamoxifen (20 mg per
day) for 5 days , all attending EL-Shatby Maternity Hospital.

Distribution according to demographic data (age &BMI):

Table (2) and figure (1) shows Distribution of the studied cases
according to the age and the B.M.I. The mean age between the studied cases
was 26.46 4.93 and the median was 27.0 years.

The mean B.M.I between the studied cases was 30.76 4.08 and the
median was 30.60.

Table (2): Distribution of the studied cases according to demographic

data (n= 80)

No. %
Age (years)
25 years 29 cases 36.3
>25 years 51 cases 63.8
Min. Max. 17.0 36.0
Mean SD. 26.46 4.93
Median 27.0
BMI (kg/m2)
Min. Max. 22.26 39.06
Mean SD. 30.76 4.08
Median 30.60

42
 Results

Years >25

Years
Years
25
Cases

Years
Cases

Fig. 7: Distribution of the studied cases according to age

43
 Results

Distribution of the studied cases according to outcome:

Table (3) and figure (2) show Distribution of the studied cases according
to outcome of response to ovarian stimulation by tamoxifen drug.

The number of ovulated cases was 23 (28.8%) and the number of the
cases did not ovulated was 57 (71.3%).

Table (3): Distribution of the studied cases according to outcome (n=

80)

No. %
Outcome
Ovulated 23 cases 28.8
Did not ovulate 57 cases 71.3

44
 Results

Not
ovulated
Ovulated

Cases

Cases

Fig. 8: Distribution of the studied cases according to outcome

45
 Results

Comparison between the two studied groups according to different

parameters

Table (4) and figure (3, 4) show comparison between the two studied
groups according to different parameters, it demonstrated that A.M.H level
ranged between 1.80 7.80 and 1.50 10.0 with the mean of 3.97 1.51 and
5.15 2.07 with the median 4.0 and 4.9 for (ovulated) and (didn't ovulate)
groups respectively. And total testosterone level ranged between 0.10 2.40
and 0.10 2.70 with the mean of 0.72 0.43 and 0.70 0.41 with the median
0.60 and 0.59 for (ovulated) and (didn't ovulate) groups respectively.

There was statistically significant differences between the two studied

groups regarding AMH, (P 0.021), but there was no statistically significant
differences between the two studied groups regarding total testosterone levels.
(P 0.559).

Table (4): Comparison between the two studied groups according to

different parameters.

Did not
Ovulated
ovulate Z p
(n= 23)
(n= 57)
A.M.H (ng /ml).
Min. Max. 1.80 7.80 1.50 10.0
Mean SD. 3.97 1.51 5.15 2.07 2.302* 0.021*
Median 4.0 4.90
Total testosterone
(ng /ml)
Min. Max. 0.10 2.40 0.10 2.70
Mean SD. 0.72 0.43 0.70 0.41 0.585 0.559
Median 0.60 0.59

46
 Results

Fig. 9: Comparison between the two studied groups according to AMH

There is significant difference in AMH level between ovulated cases and

the cases didn't ovulate in PCO patients.

Fig.10: Comparison between the two studied groups according to Total testosterone.

No significance difference between Ovulated and the cases didn't-

ovulate in relation to Total testosterone hormone level.

47
 Results

48
 Discussion

DISCUSSION
This study included 80 patients diagnosed as PCOS, all
attending EL-Shatby Maternity Hospital. All patients received
standard dose of tamoxifen 20 mg /day from day 2 of the
menstrual cycle for 5 days, with estimation of the serum level
of AMH and total testosterone day 3-5 of the menstrual cycle.

There were statistically significant differences between

the two studied groups, (+ve ovulation group) and (-ve
ovulation group), regarding the serum level of antimullerian
hormone (AMH), but the study shows that there no
statistically significance between the two studied groups
regarding the total testosterone level. The studied cases
mean age was 26.46 4.93 years, mean BMI was 30.76
4.08, and mean ovarian volume was 16.26 5.40 cm 3 and
mean AFC was17.94 4.39.

In agreement with the present study, Elhalawaty S, et al,

(111)
found that AMH level in women with PCOS responded to
ovulation induction by clomiphene was significantly higher
than those who didn't respond. A total of 68 women were
recruited, 17 women of them ovulated and 51 not responded,
the mean level of serum AMH was (4.38 3.30 ng/ml versus
1.89 1.33 ng/ml P= 0.0081) in ovulated and non-ovulated
groups respectively. They found that there were statistically
significant differences between the two studied groups
regarding AMH level.

49
 Discussion

In Elhalawaty S study, the sample size of PCOS patients is

near to that of this study, moreover the clomiphene received
as a standard dose (150mg divided on 3 doses) and for fixed
period of time (5 days only) similar to this study, a standard
dose of tamoxifen was given (20 mg daily divided on 2 days)
and for 5 days.

Moreover, Tnl Vuong, et al, (112)

in their published study,
the PCOS patients were undergoing ART by administration of
rFSH injection. The study included 790 women, the mean
level of serum AMH was (2.7 1.7 ng/ml versus 0.7 0.8
ng/ml P= 0.01) in normal responders of and poor responders
groups respectively, so the study reported that the AMH has
significant difference between the both groups.

Although both studies found significant difference in

serum level of AMH between responders and non- responders,
the patients in Tnl Vuong study received rFSH injection which
is more potent than any oral medication and the sample size
is much greater in Tnl Vuong study (790 PCOS patients) than
that of this study (80 PCOS patients).

In disagreement with the present study, Anna Dbkowska-

Hu, et al, (113) in their published study, reported no significant
difference in AMH level between women with PCOS who were
responsive to ovulation induction by recombinant follicle-
stimulating hormone and those who were non-responsive. A
total studied PCOS cases was 26. There were no significant
difference between the ovulated (20 cases) and non-ovulated

50
 Discussion

(6 cases) groups in serum AMH level with mean (7.72 ng/ml

versus 6.62 ng/ml P= 0.78974) respective.

In Anna Dbkowska-Hu study, the number of the PCOS

patients was 26 which are much less than in this study (80
patients) which makes its result less accurate. Also the Anna
Dbkowska-Hu study chose clomiphene resistant PCOS
patients in contrary to this study which select just PCOS
patients.

In this study, the period of ovarian stimulation is fixed

(only 5 days) but in Anna Dbkowska-Hu study the ovarian
stimulation wasn't fixed and much longer (up to 14 days in
some cases). All this makes the result of AMH in this study
more accurate than those of Anna Dbkowska-Hu study.

In this study, it showed that there was no statistically

significance between the two studied groups regarding the
total testosterone level.

In agreement with the present study, Huber-Buchholz, et

al, (114)
there was no significant difference in total testosterone
concentration between obese women with PCOS who were
responsive to ovulation by lifestyle modification and those
who were non-responsive. The total women were recruited
and complete the study was 15 PCOS patient, 9 women of
them ovulated and 6 didn't ovulated, the mean level of serum
total testosterone was (2.2 0.2 ng/ml versus 2.3 0.4
ng/ml) in ovulated and non-ovulated groups respectively.

51
 Discussion

The level of the total testosterone hormone in the Huber-

Buchholz study in both responders and non-responders are
higher than that of this study, the sample size in this study
larger(80 patients) than that Huber-Buchholz (15 patients)
which makes the result of this study more accurate but both
of them still found no significant difference in the level of total
testosterone between responders and non-responders.

Also in agreement with the present study, Mervat Sheikh-

El-Arab, et al,(115) there was no significant difference in total
testosterone concentration between women with PCOS who
were responsive to ovulation induction by clomiphen. A total
studied PCOS cases subjected to clomiphene cases was 57
cases , 35 women of them ovulated and 22 not responded,
the mean level of serum total testosterone was (0.870.26
ng/ml versus 0.930.32 ng/ml P= 0.36) in ovulated and non-
ovulated groups respectively.

Also in the same study, Mervat Sheikh-El-Arab, et al, (115)

there was no significant difference in total testosterone

concentration between women with PCOS who were
responsive to ovulation induction by letrizole. A total studied
PCOS cases subjected to clomiphene cases was 59 cases , 41
women of them ovulated and 18 not responded, the mean
level of serum total testosterone was (0.910.41ng/ml versus
0.890.35 ng/ml P= 0.27) in ovulated and non-ovulated
groups respectively.

52
 Discussion

Although the both studies used close numbers of PCOS

patients and both of them found no significant difference in
the level of total testosterone between PCOS responders and
non-responders and the level of the total testosterone in both
of them are almost close to each other, Mervat Sheikh-El-Arab
study; was double blind study as neither the patient nor the
ultrasound operator knows the drug the patient took which
gives more accuracy for its result.

In disagreement with the present study, Babek Imani, et

al, (116)
in their published study, found that total testosterone
level concentrations between women with PCOS who were
responsive to ovulation induction by clomiphene were
significant higher than those were non-responsive. A total
studied PCOS cases was 201. There were significant
difference between the ovulated (156 cases) and non-
ovulated (45 cases) groups in serum total testosterone level
with mean (2.1 0.9 ng/ml versus 2.7 1.0 ng/ml P=0.001)
respective. They found that there were statistically significant
differences between the two studied groups regarding total
testosterone level.

In the Babek Imani study, the number of the patients

were 201 patients which much more than those of this study
(80 patients), Also the study follow up the non-responder
PCOS patients for 3 consecutive cycles with increasing the
dose of clomiphene (50mg, 100 mg to 150mg) while in our

53
 Discussion

study, patients received standard dose of tamoxifen (20 mg)

for 5 days, for only one month.

More studies including larger samples of PCOS patients

are recommended for more accurate and determined results
specially for the significance of the total testosterone level
among tamoxifen responders and non- responders of PCOS
patients.

In summary, the large degree of variability between

studies reflects, on one hand, the complexity of this hormonal
system and, on the other hand, the involvement of additional
(hormonal or other) factors that can either positively or
negatively impinges upon response of ovulation induction of
polycystic ovaries women.

The current study reported, AMH has significant difference

between PCOS patients ovulated and those who did not
ovulate PCOS patients in response to the standard dose of the
tamoxifen (3.97 1.51, 5.15 2.07) but there is no cut off
level has been detected to discriminate the predictive
response of the tamoxifen as ovulation induction in PCOS
patients, but the total testosterone hormone has no
significance between two groups.

54
 Summary

SUMMARY
The Polycystic Ovary Syndrome (PCOS) is one of the most common
endocrine/metabolic disorders of women. This syndrome was first described
by Stein and Leventhal in 1935, (1) although the presence of sclerocystic
ovaries had been recognized for at least 90 years before the publication of
that seminal work. (1, 2). PCOS is a functional disorder of unclear etiology,
and, it is a diagnosis of exclusion, other androgen excess and ovulatory
disorders of clearly defined etiologies excluded. (4) Three major diagnostic
criteria for PCO have been proposed by the National Institute of Health
(NIH 1990), the Rotterdam European Society for Human Reproductive
and Embryology/American. PCOS could be diagnosed, after the exclusion
of related disorders, by two of three features: (a) oligo- or anovulation, (b)
clinical and/or biochemical signs of hyperandrogenism, or (c) polycystic
ovaries, (4, 27) A significant number of patients have infertility as a
presenting feature of PCOS,chronic anovulation , also women with PCOS
have a higher incidence of spontaneous pregnancy loss, recurrent
miscarriage was ranging from 25 to 37%(64, 65).

Tamoxifen the first triphenylethylene antiestrogen was found to be

antiestrogenic effects in the treatment of infertility and breast cancer.
Although these agents are widely described as antiestrogens, they are best
characterized pharmacologically as partial estrogen agonists. (67, 68) The
estrogenic or antiestrogenic effects of these agents depend on level of
endogenous estrogen and the target organ affected, Tamoxifen is generally
considered to increase fertility rates in a similar way to clomiphene. It has
been postulated that tamoxifen improves follicular development by

55
 Summary

direct action on the ovary rather than through the hypothalamic-pituitary

axis. (72, 73)

This study was conducted aiming to determine the level of both AMH and
the total Testosterone at which women with PCO ovulation in response to
a standard dose of Tamoxifen.

The study was conducted upon 80 PCO women indicated for ovulation
induction, each patient was given Nolvadex 20 mg twice daily. They were
recruited from the infertility clinic of El Shatby University Maternity
Hospital from January 2013 to December 2014.

All patients were subjected to:

Informed consent then complete history taking and Complete
examination.

Transvaginal Ultrasound: which done in the day 2 of the cycle, basal

scanning to assess the uterus (as regards the endometrial thickness), the
ovaries (as regards the ovarian volume, the antral follicular count and
the size of the follicles) and to exclude the presence of any ovarian or
adnexal pathology.

Estimation of the Serum level of antimullrian hormone done by ELIZA

and total testosterone hormone done by ELIZA technique, it will be
estimated day 3 -5 of the cycle.

A standard dose of tamoxifen (NOLVADEX 10mg) twenty milligram
per day (two tablets), starting on day 2 of the cycle for 5 days is to be
given.

Follow up by transvaginal ultrasound every 3 days for the following

two weeks will be done to evaluate the ovarian response by defining the

56
 Summary

number of growing follicles in both ovaries and detection of the

ovulation by either disappearance of the mature follicle, Corpus luteum
formation, trilaminar endometrium and minimal fluid collection in the
doglus pouch.

The level of the antimullerian hormone and the total testosterone

hormone at which the ovaries respond by ovulation were evaluated.

The result was:

1. The number of ovulated cases was 23 (28.8%) and the number of non-
ovulated cases was 57 (71.3%).

2. The mean level of AMH was 9 3.97 1.51 and 5.15 2.07) with the
median 4.0 and 4.9 for ovulated and non- ovulated groups respectively.

3. The mean leve of Total testosterone level was (0.72 0.43 and 0.70
0.41) with the median 0.60 and 0.59 for ovulated and non-ovulated
groups respectively.

4. There were statistically significant differences between the two studied

groups regarding AMH, but there were no statistically significant
differences between the two studied groups regarding total testosterone
levels.

Our results concluded that:

AMH and serum testosterone cannot be used as predictive parameters to

predict response to ovulation induction by tamoxifen.

57
 Conclusion

CONCLUSION
1) AMH is significantly higher in PCOS patients who didn't ovulate in
response to the standard dose of tamoxifen than those who ovulated.

2) The total testosterone level was comparable between both PCOS patients
who didn't ovulate in response to the standard dose of tamoxifen than
those who ovulated.

3) No definite cut off value has been detected to discriminate cases who
responded to treatment from those who did not respond.

58
 Recommendations

RECOMMENDATIONS
AMH and serum testosterone cannot be used as predictive parameters to
predict response to ovulation induction by tamoxifen.

59
 References

REFERENCES
1. Stein IF, Leventhal NL. Amenorrhea associated with
bilateral polycystic ovaries. Am J Obstet Gynecol 1935;
29:18191.

2. Lenart-Lipiska M, Matyjaszek-Matuszek B, Woniakowska

E, Solski J, Tarach JS, Paszkowski T. Polycystic ovary
syndrome: clinical implication in perimenopause. Prz
Menopauzalny 2014; 13(6):348-51.

3. Zawadzki JK, Dunaif A. Diagnostic criteria for polycystic

ovary syndrome: towards a rational approach. In: Dunaif
A, Givens JR, Haseltine FP, Merriam GR, editors. Polycystic
Ovary Syndrome. Boston: Blackwell Scientific
Publications; 1992. P. 377-84.

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