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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

ENDO#6 - 13/10/23
Reproductive endocrinology of the female
Prof. Persani – 13/10/23 - Author: Axel Duhamel – Reviser: Grazia Negrini

1. Gonadal axis

This is a schematic representation of the gonadal axis. GNRh neurons are hypothalamic neurons in the
medial eminence that originate from the olfactory plateau (with the olfactory neurons) and migrate in the
brain to reach the hypothalamus. Diseases of GnRH neurons are associated with anosmia.
GnRH will stimulate in the pituitary gland in a pulsatile fashion LH and FSH.
LH and FSH in the female stimaulte 2 different cells in the ovary

- LH stimulates thecal cells, external layer of the follicles

- FSH stimulates granulosa cells inside the follicles

Thecal cells produce androgens and granulosa cells express aromatase allowing the formation of estrogen
from testosterone.
Estrogen has a negative feedback on the production of LH and GnRH and has effects on target tissues.
Granulosa cells also produce peptide hormones such as inhibin B allowing the negative regulation of the
pituitary gland.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

Importantly, there are other factors that influence the GnRH pulsatility, LH and FSH secretion. The main one
is Leptin, a hormone important for the regulation of satiety and appetite. It is also an important message for
the GnRH neurons, because it informs the central regulator of the reproductive axis about the energetic
reserves of this organ. If there is a deficiency in the nutritional status and in the amount of adipocytes in this
lumen, there is a decrease in Leptin signaling and this is reflected by a reduction in GnRH pulsatility. So,
Leptin is important to sustain an adequate GnRH pulsatility to avoid the possibility of pregnancy in
conditions of insufficient nutritional supply.
The aromatisation allows the transformation of testosterone into estradiol through the aromatization of the
first ring of the steroid structure. Estradiol is the result of the aromatization of ring A, that becomes an
aromatic ring and this provides the estrogenic activity

2. Follicular dynamics

At birth, the ovary contains 7 million potential germ cells within the primordial follicle. They will undergo
atresia, leading to the presence of 1 million of follicles at birth. Only 300 thousand are present at puberty and
only 450 are ovulated in the fertile period of a woman.

The formation of a viable germ cell is a difficult and energy consuming process due to the amount of
potential errors occurring in the development of germ cells and granulosa cells.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

From primordial cells develop primary follicles, producing the marker AMH. The ovary and primary
follicles produce anti-mullerian follicle and is the marker of residual fertility in a female. In each cycle, there
is a recruitment of antral follicles, forming pre-antral and antral follicles.
All these processes are regulated by the interaction between the oocyte and the surrounding cells (granulosa
+ techa). This information is provided by a series of GF and initiation factors of the TGF family, through a
paracrine action. Through the differentiation of the occyte, the granulosa responds by expressing other
factors that influence the maturation and meiosis of the occyte.
If there is a disorder in the oocyte not producing correctly BMP15 or GDF-9, then the surrounding cells do
not support it and let it apoptose.

There are therefore 2 phases of folliculogenesis:

- gonadotropin independent phase, regulated on these factors deriving from the TFG family through
a paracrine factors.

- gonadotropin (FSH) dependant phase, in which the cells (at the pre-antral stage) start to express FSH
receptors.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

3. Variations of hormones in the menstrual cycle

The variations have several consequences on certain tissues.

4. Menstrual cycles and their alterations

a. Terms and definition
In physiological conditions, the interval between menstruations can vary between 23-35 days, with a
duration of bleeding of 6-7 days.

- oligomenorrhea – intervals > 35 days but < 3 months

- amenorrhea – absence of menstruations for > 3 months. Most frequent cause is pregnancy.

- hypomenorrhea – menstruation of poor volume (< 20 mL; 1 day), usually spotting

- polymenorrhea – interval shorter than 23 days. Mainly due to lutheal defect (defectuous production
of LH by corpus lutheum or defect of progesterone production.

- hypermenorrhea – menstruation of huge volume, often associated with menorragies –

menstruation of long duration, last > 7 days
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

- metrorragies – intermenstrual bleedings can be spotting or largers in the presence of coagulation

defects or fibromas

- dysmenorrhea – painful menstruations (can be associated with endometriosis)

Several endocrine diseases can be associated with these different menstrual cycle disturbances.
b. Endocrine conditions associated with oligomenorrhea or amenorrhea

By measuring gonadotropin, we ewill divide the menstrual cycle problems into:

- Normo or hypogonadotropic state – central defect, diminished stimulation of the ovary.

- Hypergonadotropic state – elevated gonadotropins, primary defect (so a defect that is affecting
directly the ovary and ovarian follicle)) associated also with polyglandular autoimmune diseases.
Normo/-hypogonadotrope – GnRH pulsatility issue or inablility of producing bioactive products (?)

 Hyperprolactinemia – the elevation of prolactin in blood can be due to the presence of a pituitary
adenoma secreting prolactin (prolactinoma) or sometimes to the administration of anti-dopaminergic
drugs favoring an increase of prolactin in blood. Indeed prolactin secretion is mainly controlled
through the negative dopaminergic tone by specific dopamine-producing hypothalamic neurons.

 Obesity/insuline resistance - found in obesity and polycystic ovarian syndrome (PCOS). Associated
generally with oligomenorrhea.

 Underweight/anorexia nervosa – defect in leptin secretion is associated with a diminished pulsatility

of GnRH, and so the appearance of oligomenorrhea or amenorrhea.

 Overt hyperthyroidism (or hypothyroidism) can be associated with oligomenorrhea. There are also
high levels of prolactin, a reduction of the pulsatility and thus elongated cycle

 Hypopituitarism

 Hypercortisolism, ex. Cushing syndrome, can be associated with diminished pulsatility of GnRH.
Hypergonadotrope – ovarian problem

 Reduced ovarian reserve – can occur in pre-menopause (physiological condition occurring in women
around 50y) or in primary ovarian insufficiency (POI), aka premature reduction of the ovarian
reserve
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

 Hypocortisolism (Addison disease) – in this case the mechanism is an autoimmune mechanism, this
autoimmune process affects also other steroidogenic cells, such as the ones present in the ovary.

 Gonadotropinomas or tumors of the pituitary releasing gonadotropins – rare condition.

Gonadotropins can be normal or high, but they cannot induce the normal ovulation because there are
not the normal oscillation levels of gonadotropins that are required for ovulation and the recruitment
of the dominant follicle.

 Pseudohypoparathyroidism 1a – rare condition. It refers to a general hormonal resistance, so also a

resistance to gonadotropins, that is due to the presence of a mutation in the Gs troponin. This
transduces the stimulatory signal of the gonadotropins inside follicular cells.

 Drugs
o anti-dopaminergic -> hyperPRL production – as antipsycotic/antidepressants/anti-hemetic,

o chemotherapy -> POI

c. Endocrine conditions that cause poly- or menorragies (normogonadotropic state,

hypergonadotropic state)

Endocrine conditions

 Mild hypothyroidism: this will produce a progesterone insufficiency

 Pre-menopause, causing initial progesterone insufficiency, then resulting in reduced ovarian reserve

Non-endocrine conditions

 Fibromas of uterus – myometrial fibromas or endometrial polyps. Can be associated with an

increased bleeding

 Drugs (anticoagulants)
All of these cases are associated with iron loss, leading to a risk of anemia.

5. Puberty

Puberty is a developmental stage of the organism, characterized by an acceleration of somatic growth and
sexual maturation and differentiation. It is the acquiring the possibility to reproduce.

In the last century, there is a trend of anticipation of the starting age of puberty that occurs in association
with the improvement of the socio-economic conditions and nutrition. This allows the leptin message to
enter the hypothalamus and initiate the processes.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

For females, puberty is if it occurs between 8-13 years of age. The start is associated with a nipple bud
(image B2) because the breast is the organ most sensitive to estrogen action (there are modifications also in
uterus). So, this is the first visible change in the body of a young girl. A precautious puberty is defined as an
initiation of puberty before 8. Menarche, which is the first menstruation, occurs between 12-14 years.

There is about one year of delay in the start of puberty in the male sex, beginning at 9-14 years. This is
associated with an increase in the testicular volume > 8mL or 4 mL per testes. These are the signs that
puberty has started.

There are specific changes of pubertal development that are categorized according to the growth of the
mammary gland.

There initiation of puberty (activation of GnRH pulses) is an equilibrium between environmental signals and
peripheral signals (leptin) that influence a family predisposition (genetics) for GnRH pulse initiation
following a Gaussian curve.
If the peripheral and environment signals are stronger than usual, the timing of puberty is delayed/anticipated
and follows a non-gaussian curve.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

The “GnRH pulse generator”, that is the one that starts firing with higher frequency and higher amplitude at
the moment of puberty, mainly during sleep hours. Sleep is very important to promote the maturation of the
sexes.

6. Pubertal delay or hypogonadism in females

This is an extremely heterogenous condition.

- The primary defect is associated with a condition of high circulating gonadotropin

(hypergonadotrope), the most frequent condition associated with this state is the Turner Syndrome.

- Central defects are associated with either a state of hypo- or normogonadotropic hypogonadism:

o organic lesions (ex. pituitary or hypothalamus)

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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

o or genetic defects (ex. Kallmann syndrome; less frequent in females)

o or the so called functional hypothalamic defects (ex. anorexia nervosa, stress, excessive
exercise; more frequent in females).
7. Turner syndrome

TS is caused by numerical or structural anomalies of the X chromosome and represents one of the main
chromosomal anomalies, with a prevalence of 1/2000 female newborns.

- About 50% of the cases are associated with a karyotype 45X,

- 25% are associated with structural defect of one of the two X chromosomes,

- 25% that are associated with the mosaic between cells of the organism with a normal karyotype of
46XX and cells within the same organism, with an abnormal karyotype 45X.

This variable karyotype, especially in the mosaic group, can account for the large phenotypic variability of
individuals with this syndrome. So, the majority of turners are 45,X, then we have ¼ that are mosaic and ¼
that are either i(Xq),ringX or del Xq/Xp.

The most frequently associated anomaly is short stature (the double X is important to have a normal
height), all TS patients are invariably short.

Ovarian insufficiency is present in all patients at puberty but a fraction (20%) of patients have a
spontaneous puberty and can conceive for a short period of time before they undergo very rapidly a
premature menopause after puberty around 25yo.

There are other abnormalities seen here that help with the differential diagnosis: the most evident are
Epicanthus palpebralis, neck lymphedema, lateralized lymph nodes and short fingers.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

Here the evolution of antral follicles (red) and germ cells (blue) in a normal person (straight line) and a TS
patient (dotted line).
In the normal condition, there are 7 milllion follicles during gestation and a progressive decay to 1 million,
then the appearance of other follicles at puberty, at around 50y/o menopause (no more follicles).
In TS, the ovary has a normal population of germ cells but the astresia of germ cells is very fast. Only 20%
keep ovulatory activity and are able to conceive.

In the graph, it is possible to see that FSH levels are at levels close to menopausal range in 45X patients at
birth. This is rarer in patients who are mosaics: the proportion of patients with a spontaneous puberty is
higher among the mosaics.

8. Menopause and aging

Organs at term during growth and the aging process

• Placenta: adaptation to autonomous extrauterine life

• Infancy teeth: adaptation to cranio-facial growth

• Thymus: immune system maturation and tolerance

• Ovary: preservation from risk of late reproduction

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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

The ovary is one of the organs that has a terminated function during the aging process, which is to preserve
from the risk of a late reproduction.

Menopause occurs physiologically according to the distribution seen above. It occurs mainly between 50-54
years at a median being 51. There is a steep slope just after 50 years.
Those with late menopause are probably women with a strong “organism” that can afford a late pregnancy,
as menopause is very closely related to the aging potential of an organism.
You can also see that about 10% of women have menopause before 45y. This is a very early age, because we
have a delayed/elongated life-expectancy. So, there is a prolonged time that is conducted in the absence of
estrogen activity. This can affect several tissues and have severe consequences on the well-being later on.
There is a correlation in between the age of menopause and the survival  the later you finish menstruating
the more your body is strong.
9. Primary ovarian insufficiency:
Primary ovarian insufficiency is defined as menopause occurring before 40 years of age. The prevalence of
this condition is 1% of women. The frequency of the disease decreases by 1 log every 10 years (so at 30y,
the prevalence is of 1:1000 ; at primary amenorrhea in 46XX, the prevalence is of 1:100000).
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

10% of women have an early menopause (EM) between 40-45yo. This is becoming an increasing problem
due to the median age of conception increasing (30yo now) and the increasing life expectancy (longer time
with on estrogen). This results in an anticipated aging: skin, bones, CV system, …

These conditions need to be diagnosed and treated with a supplementation of estrogen, at least until the
normal age of menopause (51).
In addition, due to the linger life expectancy, most women undergoing physiological menopause should
receive supplemental estrogen. Risks associated with hormone replacement therapy can be breast cancer (but
today, diagnosis is rather easy).

Nature did not consider menopause. Menopause is also seen in Rhesus macaques. However, in the wild,
these animals usually die before reaching menopause due to other diseases (live to < 20y). In captivity,
menopause is observed at around age 27 in these animals.

a. Clinical presentation and treatment

Issues Consequences Notes

Fertility Possible natural conception before
defects complete follicle depletion
Interventions to rescue fertility
after POI diagnosis
Psychological distress
Oocyte cryopreservation before
follicle depletion
Auto-transplantation of
cryopreserved ovarian tissue
Oocyte donation
Hormonal Flushes, sweating, vaginal dryness, Consequences at several tissue levels
defects urogenital manifestations,
irritability/anxiety, sleep disorders prolongation of life expectancy, but
Sexual dysfunction
Impaired performance (physical
and mental)
Bone disease (osteoporosis)
Neurodegenerative diseases
Cardiovascular/metabolic diseases
Increased Autoimmune risk; increased
morbidity/ mortality for all causes of death
mortality and cancer specific
Risk to conceive a male with Fragile X
syndrome
Frequently ineffective
Severe impact for affected woman and
family
These interventions are rarely
supported by NHS
Concerns on long term conservation of
ovarian samples
Ethic concerns: parental consent for
minors
have increasing impact due to the
may be reduced by appropriate HRT
(estrogens and/or androgens).
Elevated costs for examinations and
therapeutics.
Controversial protection on breast
cancer
Risks not influenced by HRT?
Role of overweight?
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

The clinical relevance of POI has two orders, fertility defects and hormonal defect and acquired importance
due to the increasing age of conception and increasing life expectancy.
Fertility defect are important and are targeted through interventions to rescue fertility and through oocyte
cryopreservation before follicle depletion. The cryopreservation is usually complicated and oocyte donation
(not available in all countries and regions) is the preferred route. Earlier the diagnosis and prediction of POI,
earlier and better the fertility can be saved.
The hormonal defect is associated with manifestations typical of menopause: hot flashes, sweating, vaginal
dryness, urogenital manifestations, irritability/anxiety, sleep disorders, sexual dysfunction, bone disease,
neurodegenerative disease (Alzheimer), and cardiovascular metabolic diseases. It is associated with an
increased mortality in cancer and an increased autoimmune risk. These are all consequences of the increased
time spent without estrogens.
The defect is targeted through hormonal replacement therapy: estrogen and in some cases low amounts of
androgens. The only contraindication is the genetic predisposition to breast cancer.
Treatment of women with POI needs a multidisciplinary approach in order evaluate

 Ca/bone metabolism – make sure to have an adequate Ca intake and Ca metabolism; by evaluating
PTH, Ca/P, 25OH-vitD and bone mineral density (BMD); be sure to have an adequate vit D level
(supplementation).

 Oncologic evaluation/prevention – using new generation highly sensitive imaging for breast cancer
screening

 Psychological support – especially for younger women that are without children

 Gynecological evaluation – ovary/uterus US, PAP test.

 Metabolic and dietary evaluation and education of how to avoid overweight – ex. being overweight
can be dangerous for the bones and because this disease can increase the risk of metabolic diseases.

 Cardiovascular risk – by evaluating coagulation parameters, folate, vit B12, homocysteine, carotid
artery doppler
b. Diagnosis
Fertility and subfertility:

- Fertility is optimal before 30y.

- From 30, subfertility is present.

- Sterility (loss of fertility) occurs at 10 years before menopause

- Oligomenorrhea occurs 5 years before menopause.

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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

The early signs of menopause occur therefore after the loss of fertility. The diagnosis of POI should therefore
be made early enough in order to preserve fertility (ie: diagnosis should be made during normal menses and
normal ovarian reserve).
Instead, the guideline show that the diagnosis is made with at an overt clinical state and with an elevated
FSH: POI is suspected after amenorrhea for 4 months and two separate FSH > 25 U/L at 1-month interval.

Predictive biomarkers of early menopause is crucial in order to preserve fertility in women with POI. One of
the most promising parameters is AMH (Anti-Mullerian hormone – produced by antral and pre-antral
follicles, ie: follicles that are supposed to ovulate). Therefore depending on the AMH produced by these
follicles after birth, it is possible to predict through an algorithm the average age of menopause.
An AMH in 31 (ie the max amount of AMH) at 5µg/L, it is possible to predict a menopause around 50.
AHM tends to increase until 25y, so it is not possible to diagnose before that.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

c. Etiology
There are several environmental factors that play an important role (the professor only mentioned briefly
chemo, radio, smoking and infections, but I added this from the past year notes). For example,

- chemo- or radiotherapy for tumors at early ages, can induce POI. Note that there is an increase in
cancer prevalence at younger ages, but reduced mortality. However, these women will have risk of
infertility later in life. For this reason, some hospitals offer assisted reproductive technologies for
young cancer patients who wish to, for example, store their gametes.
There is, of course, different risk for different types of malignancy, as well as regarding the different types of
treatment. The cytotoxic agents are divided into high, intermediate and low/no risk. This allows for the
prevention of infertility in some cases, where another treatment can be chosen.

- Other environmental factors include exposure to toxins/disruptors (ex. heavy and/or long/lasting
smoking; > 20 sig/day: +35% risk; alcohol, phthalates, etc.) and, also, certain infections.

Genetic predisposition to POI:

- Familiarity to POI

- Turner syndrome

- Monogenic forms of POI

- Autoimmunity
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

Autoimmune POI:
Unfortunatly, there are no biomarkers for autoimmune POI. They represent 15-20% of all POI. It is usually
associated with polyglandular autoimmune syndrome type I (PGA type 1).
PGA type 1 is a rare disease that is associated with mutations in the AIRE gene (involved in T cell
maturation in the thymus). It is characterized by:

- anti-21-hydroxylase antibodies, frequently found in patients with also Addison disease.

- Anti-TPO antibodies have no role in the disease but are found in 15-20% of POI patients (can be a
surrogate marker).
PGA is associated with candidiasis, hypoparathyroidism, Addison disease, and oophoritis in 60% of the
cases

It is therefore necessary to screen for AMH, FSH, anti-21-hydroxylase antibodies, anti-TPO antibodies in
women with a personal history organ-specific and systemic autoimmune diseases. Women with POI are at an
increased risk for Hashimoto’s thyroiditis.
Family history is very important: Women reporting a mother with an early age at menopause had a 6-fold
increased risk of POI. In a study in Milan, we saw that 37% of women with POF responded that they have
someone in their family with early menopause.
d. POI and fragile X syndrome
Fragile X syndrome is a mental retardation syndrome that mainly affects males. These males have an
expansion of triplets in a particular region of the FMR1 gene (CGG-triplets), requires about 200 triplet
expansion.
Women that are carrying permutational genes (CGG triplets over 50-60) start to have an increasing risk of
POI and EM. POI before 40 years of age, in the presence of a premutation (meaning an expansion of
triplets between 55-100), is associated with a 20% risk of POI and 30% risk of EM.

- Around 55 triplet expansion, there is an increased risk of POI from 1% to 2%.

- Then, the progressive triplet expansion from 55 to 70 is associated with a risk going from 2% to 5%.

- And it will hit a peak risk at 80-90 expansion.

- If triplet expands to full 200, the risk of POI is close to 100%.

So, a woman with 80-90 triplets expansion is highly likely to have a male child with more than 200 triplets
and with mental retardation.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

This a family free: the females had a pre-mutated gene and had menopause at 38 (mother), 26 and 25
(daughter) and the grandson has a full mutation.

Such an premutation is not rare in the population: : 1 out of 157 in populations of Mediterranean origin,
lower in Asian populations, 1 out of 259 in general population. Therefore, FMR1 expansion gene analysis is
needs to be performed in women with EM.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

There are several genes associated, so nowadays it is very difficult to perform analysis of all the candidate
genes by Sanger sequencing because you will never succeed. However, there are particular phenotypes that
can suggest you to perform direct genetic investigations, ex. drooping eyelids caused by mutation in FOXL2,
which causes Blepharophimosis, Ptosis, and Epicanthus inversus Syndrome (BPES), and is associated with
POI at a young age.
Using thus genetic analysis it is therefore possible to predict future symptoms to come and the possibility of
a preservation of fertility.

The preservation of fertility in POI through genetic counselling (NGS Overarray) in families at risk yields a
30-40% positive result and allows:

- Early diagnosis: anticipation of conception, or oocyte cryopreservation before ovarian failure in

young relatives of one POI patient

- Informed risk of X-linked mental retardation (FMR1 premutation)

- Neoplastic risk (meiotic, DNA replication/repair genes)

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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

I. PCOS (polycystic ovary syndrome)

1. Hirsutism
There are also other diseases of the ovaries which can be associated mainly with oligomenorrhea and
androgenization. Hirsutism is the most evident phenomenon of androgenization.
Hirsutism is the excessive growth of terminal hair (thick and pigmented) in areas typical of the male sex,
under androgen stimulation. On the contrary, hypertrichosis is the increased thin hair growth not androgen-
dependent.
There is a hirsutism score called the Ferrimann and Gallwey score. Different androgen-dependent areas of
hair growth are examined, and different stages are defined:
- Mild: 8-15
- Moderate: 15-24
- Severe: >24

9 areas of the body should be evaluated (upper lip, chin, chest, arm, upper abdomen, lower abdomen, upper
back, lower back and thighs), with a score of 1-4, depending on the density of the hair growth.

The typical disease of the ovaries associated with hirsutism is PCOS (polycystic ovary syndrome). Another
cause of hirsutism could be congenital adrenal hyperplasia, in which a defect in steroidogenesis leads to an
increased release of androgens from the adrenal glands.
2. PCOS definition
PCOS is a heterogenous condition, nowadays considered to have a relevant genetic predisposition. The
clinical manifestations include menstrual irregularities (hypo-oligomenorrhea), hyperandrogenism
(hirsutism, acne, alopecia), obesity/insulin resistance and polycystic ovaries. Polycystic ovaries are formed
due to an ovulation defect: the follicles are recruited, and they grow, but there is no ovulation so, they remain
inside the ovaries as a series of follicles.
3. Epidemiology
It is one of the most frequent endocrine anomalies in females, affecting 6.6% of women of reproductive age.
It is particularly frequent in the Mexican and Mediterranean populations.
There is a high PCOS prevalence in various conditions: families with PCOS, premature adrenarche, obesity
and insulin resistance, diabetes mellitus type 1 or type 2 and autoimmune thyroiditis.
4. Diagnostic criteria
There are different definitions of PCOS, which changed over the years. In the past years, the presence of
polycystic ovaries, observed on US, was required for diagnosis but nowadays it is not thanks to the
biochemical diagnosis of hyperandrogenism.
So, clinical and/or biochemical hyperandrogenism (to assess the androgen status) associated with oligo- or
anovulation are enough for diagnosis.
As already said, PCOS is also generally associated with obesity and insulin resistance (probably a
mechanism which also leads to ovarian dysfunction). However, not all PCOS women are overweight.
Moreover, these people have also a higher to develop diabetes Mellitus.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

Clinical hyperandrogenism manifests with hirsutism, acne and alopecia. Rarely it can be associated with
virilization: voice modification, muscle mass increase, and clitoromegaly. Virilization is usually associated
with tumors of adrenals or ovaries which secrete a greater amount of androgens.
Biochemical hyperandrogenism must be evaluated within the first 5 days of menstrual cycle. The
parameters to be measured are:
- Total testosterone and delta4-androstenedione: androgens typically elevated in PCOS
- 17 hydroxylate progesterone and DHEAs: are more likely to be elevated in the case of
hyperandrogenism of adrenal origin, so they are measured to exclude this diagnosis
- LH/FSH ratio: a high LH/FSH ratio (> 1.5-2) is indicative of PCOS. In a normal cycle, FSH is
higher than LH in the follicular phase.
The ovulatory defect (oligo-anovulation) can have clinical manifestations: oligo-amenorrhea, difficult
conception and infertility. If these are not present, to demonstrate the ovulation defect progesterone must be
evaluated between 20-24 days of the menstrual cycle. Higher levels of progesterone are indicative of
luteinization and ovulation (they will remain low if there is no ovulation).

Other instrumental evaluations are US monitoring and variation in basal temperature (rise of body
temperature at ovulation).
5. Differential diagnosis
Similar manifestations are observed also in other diseases, so other diagnoses must be excluded in all women
before making a diagnosis of PCOS.
- Thyroid disease: TSH must be measured
- Prolactin excess: can lead to anovulation and oligomenorrhea
- Non-classical congenital adrenal hyperplasia: it appears in the second decade of life, it is associated
with hirsutism and menstrual irregularities
- Pregnancy: amenorrhea
- Hypothalamic amenorrhea and anorexia nervosa can be associated with hirsutism
Primary ovarian insufficiency: is more associated with menstrual irregularities (amenorrhea)
combined with symptoms of estrogen deficiency including hot flashes and urogenital symptoms
- Androgen-secreting tumor: it is more associated with virilization signs
- Cushing syndrome: many of the signs and symptoms can overlap with PCOS due to the increased
secretion of androgens
- Acromegaly: oligomenorrhea and skin changes (hirsutism) may overlap with PCOS

Q: Can women with PCOS have a normal pregnancy?

A: Yes, they can. Ovulation normally occurs, with no risk of reduced ovarian reserve so spontaneous
pregnancy is possible. However, if spontaneous pregnancy is not possible and if these women undergo an
assisted hyperstimulation of the ovaries, they are more at risk of developing ovarian hyperstimulation
syndrome which leads to very high estrogen levels, which could cause thromboembolism for example.
(Ovarian hyperstimulation syndrome is an iatrogenic complication of assisted reproduction technology).
These women cannot have a hyperstimulation of the ovaries with also high LH.
In general, after pregnancy, PCOS manifestations decrease so pregnancy can be considered a way to
improve ovarian functions. However, women with PCOS have a higher risk during pregnancy to develop
metabolic syndromes due to insulin resistance, indeed the risk of gestational diabesity is also increased.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

The image below shows the flow chart for differential diagnosis of ovarian dysfunction together or not with
clinical hyperandrogenism. Thyroid functions and prolactin must be evaluated because their alterations
should suggest a differential diagnosis (thyroid disease or hyperPRL). Other values to be measured are LH,
FSH, and estradiol in the first 3-5 days of the cycle. If clinical hyperandrogenism is present, also
testosterone, 17OH-progesterone, androstenedione and AMH must be evaluated. AMH is usually low in POI
and tends to be high in PCOS, because of the accumulating follicles in the ovaries. The different
combinations of the results of this screening can lead to the right diagnosis.

- PCOS: there is an elevation of androgens, LH higher than FSH, normal or high estradiol, AMH
tends to be high and polycystic appearance of the ovaries.

- POI: there is an absence of hyperandrogenism, high FSH compared to LH, low estradiol, low AMH,
and no follicles at US.

- 21OH-ase deficit, an enzymatic defect in the adrenals. There are higher levels of androgens, such as
17OHprog, and both the ratio LH/FHS and AMH are normal.

The clinical manifestations change with the age of the patients. During adolescence, the typical signs are
reproductive abnormalities, clinical hyperandrogenism and overweight/obesity. With menopause, metabolic
abnormalities (type 2 diabetes), postmenopausal hyperandrogenism and risk of developing cardiovascular
diseases are more observed. Insulin sensitizers, drugs used in women with PCOS with cardiovascular risk,
are also useful in reducing ovarian defects and ameliorating ovarian functions. One example of an insulin
sensitizer is Metformin.
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International Medical School – ENDO#6 – prof. Persani – Reproductive endocrinology of the female

6. Treatment
The treatment for PCOS includes a lot of different drugs/lifestyle changes different for each sign/symptom.
To treat oligomenorrhea/amenorrhea in some cases lifestyle changes (5-10% weight loss + structured
exercise) could be sufficient to restore normal ovulation. To restore the androgen levels, combined
contraceptive pills can be used: they set at rest the ovaries, which are not stimulated anymore and androgens
are not produced. Cyclic progestins and Metformin can also be used mainly in women with insulin
resistance.
For hirsutism, cosmetical interventions and lasers are recommended. However, if in the long term the
clinical treatment is effective also the hirsutism decreases.
Pharmacological therapy must be considered if there is a patient concern or if cosmetic treatment is
ineffective/inaccessible/unaffordable. It should be trialled for at least 6 months before making changes in
dose or medication. The primary therapy is COCP (monitor glucose tolerance in those at risk of diabetes). If
3-6 of COCP is ineffective, anti-androgen (such as Spironolactone) is used.
For infertility, physicians advise smoking cessation, optimal weight, exercise, folate supplementation and
advice regarding the age-related decline in infertility to allow optimal timing of family planning. Infertility
therapies may include metformin (helpful in restoring ovulation), clomiphene (anti-estrogen used only for
some days, it favors ovulation), gonadotropins, surgery and in vitro fertilization (as already said women with
PCOS who underwent in vitro fertilization are at higher risk to develop ovarian hyperstimulation syndrome).
For cardiometabolic risk, lifestyle changes in those who are overweight reduce the diabetes risk by 50-60%,
also metformin reduces the risk of diabetes by 50% in adherent high-risk groups.