Anatomy and Process of Endometrial Sampling

CASE PROBLEM
A 24-year old woman came to the office for a follow-up of irregular menstrual
period. She has a period every 10-90 days that lasts 2-10 days and ranges from spotting to
quarter-sized clots. The patient has been trying to conceive, but it has been difficult to time
intercourse given her erratic menstrual cycles. She has been exercising and has lost weight
and now has a BMI of 30 kg/m2.

Gynecologic examination shows a small mobile uterus, normal external genitalia, and
normal adnexa. Endometrial sampling reveals tortuous spiral arteries, coiled glands filled
with carbohydrate-rich mucus, and edematous stroma.

At which time point of the menstrual cycle is the endometrial sampling most likely obtained?

PROBLEMS IDENTIFIED
● Irregular menstrual period
○ every 10-90 days and lasts 2-10 days
○ Spotting to quarter-sized clots
● Difficulty conceiving
● BMI of 30 kg/m2
Gynecologic findings:
● Normal uterus, external genitalia, adnexa
● Tortuous spiral arteries, coiled glands with carbohydrate-rich mucus, and edematous
stroma in the endometrium

The organs of the Female reproductive system and their functions

Female reproductive system:

● Ovaries
○ Female gonads which are homologous to the testes.The ovaries are small,
oval-shaped glands located on either side of the uterus. They function to

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 produce and store eggs (ovum) and also produce hormones that control one’s
menstrual cycle and pregnancy.
○
● Fallopian tubes
○ One of two long uterine (fallopian) tubes that extend laterally from the
uterus. They provide a route for sperm to reach an ovum and transport
secondary oocytes.
○
● Uterus
○ The uterus is the size and shape of an inverted pear. It serves as a pathway for
sperm that are deposited in the vagina to reach the fallopian tubes. It is also
the site of implantation of a fertilized ovum, development of the fetus during
pregnancy, and labor. During reproductive cycles when implantation does not
occur, the uterus is the source of menstrual flow.

○ Anteflexion: normal position or conformation of the uterus; projects

anteriorly and superiorly over the urinary bladder

○ Retroflexion: posterior tilting of the uterus w/c may occur after childbirth;
harmless malposition of the uterus

○ Anatomical subdivisions of the Uterus:

i) Fundus - dome-shaped portion superior to the uterine tubes

ii) Body - tapering central portion

iii) Cervix - inferior narrow portion that opens into the vagina

iv) Isthmus - subdivision between the body of the uterus and the cervix

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 v) Uterine Cavity - interior of the body of the uterus.

vi) Cervical Canal - interior of the cervix

→ Internal os: opening of the cervical canal to the uterine cavity

→ External os: opening of the cervical canal to the vagina

● Vagina
○ The vagina is a tubular long fibromuscular canal lined with mucous
membrane which extends from the exterior of the body to the uterine cervix.
And has 3 main functions:
● provides a passageway for blood and mucosal tissue from the uterus
during a woman’s monthly period
● receives the penis during sexual intercourse and holds the sperm until
they pass into the uterus
● provides a passageway for childbirth

● Mammary Glands
○ a highly evolved and specialized organ present in pairs, one on each side of
the anterior chest wall. The organ's primary function is to secrete milk.
● Vulva
○ vulva , or pudendum, refers to the external genitals of the female and consists
of:
■ Labia – labia majora; covered by pubic hair and contain an abundance
of adipose tissue, sebaceous (oil) glands, and apocrine sudoriferous
(sweat) glands & labia minora; devoid of pubic hair and fat and have
few sudoriferous glands, but they do contain many sebaceous glands.
Both function generally to protect the vagina.

■ Clitoris - a small cylindrical mass composed of two small erectile

bodies, the corpora cavernosa, and numerous nerves and blood
vessels. Functions to experience sexual pleasure.

■ Vestibule – region between the labor minora:

● vaginal orifice - the opening of the vagina to the exterior,

occupies the greater portion of the vestibule and is bordered
by the hymen.

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 ● external urethral orifice - the opening of the urethra to the
exterior.

● paraurethral (Skene’s) glands - mucus-secreting glands are

embedded in the wall of the urethra. The paraurethral glands
are homologous to the prostate.

● greater vestibular (Bartholin’s) glands - which open by ducts

into a groove between the hymen and labia minora. They
produce a small quantity of mucus during sexual arousal and
intercourse that adds to cervical mucus and provides
lubrication.

● Mons Pubis - functions as a source of cushioning during sexual

intercourse. The mons pubis also contains sebaceous glands
that secrete pheromones to induce sexual attraction.

● Bulb of the vestibule - consists of two elongated masses of

erectile tissue just deep to the labia on either side of the
vaginal orifice. The bulb of the vestibule becomes engorged
with blood during sexual arousal, narrowing the vaginal orifice
and placing pressure on the penis during intercourse.
● Perineum
○ the diamond-shaped area medial to the thighs and buttocks of both males
and females. It contains the external genitals and anus, and functions to help
strengthen the pelvic floor.

The Histology and the Blood Supply of the Uterus

The uterus is composed of three layers of tissue, namely the perimetrium,
myometrium, and endometrium.

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 a. Perimetrium
i. The outer layer of the uterus, also known as serosa. It is a part of the
visceral peritoneum and is composed of simple squamous epithelium
and areolar connective tissue.

ii. Laterally, it becomes the broad ligament which attaches the uterus to
either side of the pelvic cavity and which contains the blood vessels to
the ovaries, fallopian tubes, and uterus.

iii. Anteriorly, it covers the urinary bladder and forms the shallow,
vesicouterine pouch which prevents the sliding of the urinary bladder
past the uterus.

iv.Posteriorly, it covers the rectum and forms the deep, rectouterine

pouch or pouch of Douglas, situated between the uterus and urinary
bladder and the most inferior point in the pelvic cavity.
b. Myometrium
i. The middle layer of the uterus consists of three layers of smooth
muscle fibers that are thickest in the fundus and thinnest in the cervix.
The inner and outer layers are longitudinal/oblique, while the middle
layer is circular.

ii. Functions in expelling the fetus from the uterus during labor and
childbirth through coordinated contractions of this layer in response
to oxytocin.

c. Endometrium
i. The highly vascularized inner layer of the uterus which has three
components.

(1) An innermost layer that lines the lumen is composed of simple

columnar epithelium which are ciliated and secretory cells.

(2) An underlying endometrial stroma which is a very thick region

of lamina propria made of areolar connective tissue, and;

(3) Endometrial (uterine) glands that develop as invaginations of

the luminal epithelium and which extend almost to the
myometrium

ii. Two Layers of Endometrium

(1) Stratum functionalis - functional layer; lines the uterine cavity

and sloughs off during menstruation

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 (2) Stratum basalis - basal layer; the deeper layer that is
permanent and which gives rise to a new stratum functionalis
after each menstruation.
● Blood Supply of the Uterus

The uterine arteries which are branches of the internal iliac artery are the
blood vessels that supply blood to the uterus.

a) Arcuate arteries - branches of uterine arteries that are arranged in a

circular fashion in the myometrium.
b) Radial arteries - branches of arcuate arteries that penetrate deeply
into the myometrium.

Before the branches of radial arteries enter the endometrium,

they divide into two kinds of arterioles:
i) Straight arterioles - supply the stratum basalis with
materials for the regeneration of the functional layer (stratum
functionalis)

ii) Spiral arterioles - supply the stratum functionalis; change

markedly during the menstrual cycle

Opposite to the uterine arteries is the uterine veins which drains the blood
leaving the uterus into the internal iliac veins. Blood supply in the uterus is
essential as it supports regrowth of a new stratum functionalis after
menstruation, implantation of a fertilized ovum, and for development of the
placenta.

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 ● The Cervical Mucus
The cervical mucus is secreted by the secretory cells of the cervical mucosa. It is
composed of two main fractions which includes an insoluble gel (mucin) and an
aqueous phase with soluble components (lipids; fatty acids; prostaglandins; trace
metals; proteins; enzyme inhibitors, and immunoglobulins) (Daunter & Councilman,
1980). It becomes more favorable to sperm as it is more alkalinic (pH = 8.5) during
ovulation. It supplements the energy needs of the sperm and protects it from
phagocytes and hostile environment of the vagina and uterus. It also plays a role in
sperm capacitation.

Composition:
mucin (insoluble gel) – lubricates, lessen friction
aqueous phase (lipids, fatty acids, prostaglandins, trace metals, proteins,
enzyme inhibitors, and immunoglobulins)

Functions:
Immunological: Protects from phagocytes and hostile environment of the
vagina and uterus
Immune system components found in mucus include immunoglobins,
complement, antibodies, cytokines, antimicrobial proteins and immune cells
Fertility: Sperm capacitation
Alkalinic during ovulation, favoring environment for sperm
Supplement energy needs of the sperm
Antisperm antibodies – infertility ,, but functions in fertility are poorly defined

● The Hormonal Regulation of the Female Reproductive System

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 a. Gonadotropin-releasing hormone (GnRH)
- Secreted by the hypothalamus, controls the ovarian and uterine
cycles, and stimulates the release of FSH and LH.

b. Follicle-stimulating hormone (FSH)

- From the anterior pituitary; initiates follicular growth

- Stimulates secretion of estrogen by ovarian follicles

- Influence take-up of androgens by the granulosa cells

c. Luteinizing hormone (LH)

- From the anterior pituitary; stimulates further development of the
ovarian follicles

- Stimulates secretion of estrogen by ovarian follicles.

- Stimulates the theca cells of a developing follicle to produce

androgens.

- Triggers ovulation at midcycle and promotes formation of the corpus

luteum which produces and secretes estrogens, progesterone, relaxin,
and inhibin.

d. Estrogen
- Six types are present in women; three of which are present in
significant quantities: (1) beta β-estradiol; (2) estrone; (3) estriol.

i.) beta β-estradiol: most abundant in a non-pregnant woman;

synthesized from cholesterol in the ovaries.

- Promote development of female reproductive structures, secondary

sex characteristics, and breasts.

- Increase protein anabolism and lower blood cholesterol level;

synergistic with human growth hormone (hGH)

- Inhibit release of GnRH bh=y hypothalamus, and secretion of LH and

FSH by anterior pituitary, when it is in moderate levels in the blood.

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 e. Progesterone
- Secreted by cells of corpus luteum

- Prepare and maintain the endometrium for implantation of a fertilized

ovum, together with estrogens

- Prepare mammary glands for milk secretion, and in high levels inhibit
secretion of GnRH and LH

f. Relaxin
- Produced by corpus luteum each monthly cycle

- Inhibits contractions of the myometrium, thus relaxing the uterus

- Functions in implantation of fertilized ovum as it occurs more readily

in a relaxed (’quiet’) uterus.

- Increases flexibility of pubic symphysis and helps dilate the uterine

cervix, easing the delivery of the baby.

g. Inhibin
- Secreted by granulosa cells of growing follicles and by corpus luteum
after ovulation

- Inhibits secretion of FSH, as well as LH, albeit to a lesser extent.

● The Phases of the Female Reproductive Cycle

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 h. Menstrual Phase (1-5 day cycle)
i. Ovarian Cycle
1. From the preceding menstrual cycle, the declining steroid
production by the corpus luteum alongside the sudden decline
of inhibin results in the FSH levels to rise. Beyond that,
increased FSH levels may also be attributed to the increasing
GnRH due to the decrease in estrogen and progesterone levels
(Reed & Carr 2000). The elevated levels of FSH during the first
days of the menstrual cycle allows several primordial follicles
to be recruited to develop into primary follicles and into
secondary follicles (Tortora & Derrickson 2009).
ii. Uterine Cycle
1. Menstrual flow occurs from the uterus which releases 50–150
mL of blood, tissue fluid, mucus, and epithelial cells shed from
the endometrium. The discharge is attributed to the decline in
progesterone and estrogen levels which causes the release of
prostaglandins that cause the uterine spiral arterioles to
constrict. The result of which causes the cells of stratum
functionalis to die due to the deprivation of oxygen from the
reduction of blood supply. After the degeneration of the
stratum functionalis, it will then separate away from the living
stratum basalis and will be part of the menstrual flow (Tortora
& Derrickson 2009).

i. Preovulatory Phase (6-13 day cycle)

i. Ovarian Cycle
1. Estrogen will increase due to the conversion of
androstenedione released by theca cells into estrogen through
interaction with aromatase released by granulosa cells of some
secondary follicles (Reed & Carr 2000).
2. This increase in estrogen alongside inhibin will cause FSH levels
to decline as a form of negative feedback. Following the
decline of FSH, a dominant follicle with the most FSH receptors
will be selected among the cohort of follicles. Those that are
not selected or less dominant follicles will not receive
sufficient FSH. This insufficiency will lead to the follicle to
cease growing and undergo atresia (Tortora & Derrickson
2009). This suppression of less dominant follicles is further
influenced by the dominant follicle as it exerts its dominance
by secreting increasing estrogen and inhibin as it grows in
follicular size (Reed & Carr 2000).

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 3. In the presence of estrogen, FSH will also stimulate the
formation of LH receptors in granulosa cells. The formation of
LH receptors in the granulosa cells will subsequently help in
the positive feedback on the pituitary gland for LH secretion
(Reed & Carr 2000).
ii. Uterine Cycle
1. The increase in circulating estrogen in the blood due to
developing follicles will stimulate the repair of the
endometrium. Specifically, the cells of stratum basalis will
undergo mitosis to form a new stratum functionalis. As the
endometrium thickens, straight endometrial glands develop,
and the arterioles coil and lengthen as they penetrate the
stratum functionalis (Tortora & Derrickson 2009).
j. Ovulation (14 day cycle)
i. Ovarian Cycle
1. High levels of estrogen will result in positive feedback wherein
it results in the increase in the release of GnRH and LH (Tortora
& Derrickson 2009). Specifically, estrogen stimulates the
frequency of GnRH in the hypothalamus which consequently
stimulates the release of LH in the anterior pituitary gland
(Reed & Carr 2000).
2. The LH surge will stimulate the luteinization of the granulosa
cells in the mature oocyte (Reed & Carr 2000). After the
rupture of the mature follicle, the ovulated oocyte and its
corona radiata will be swept into the uterine tube (Tortora &
Derrickson 2009).

k. Postovulatory Phase (15-28 day cycle)

i. Ovarian Cycle
1. After ovulation, a blood clot formation forms from the minor
bleeding of the rupture of the follicle which subsequently
forms the corpus hemorraghicum (Tortora & Derrickson 2009).
Under the effects of LH, the luteinized granulosa cells combine
with the theca lutein cells to form the corpus luteum. The main
function of the corpus luteum is to secrete progesterone which
helps encourage the preparation of the estrogen primed
endometrium for implantation of the fertilized ovum (Reed &
Carr 2000). It is also worth noting that corpus luteum also
secretes estrogen, inhibin and relaxin (Tortora & Derrickson
2009).

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 2. The lifespan of the corpus luteum is dependent on the
continued support of LH (Reed & Carr 2000). If the ovum is not
fertilized, the corpus luteum will last for two weeks as the
secretory activity of corpus luteum declines and corpus luteum
undergoes luteolysis under the influence of estrogen and
prostaglandin to form a scar tissue known as the corpus
albicans (Reed & Carr 2000; Tortora & Derrickson 2009). If the
ovum is fertilized, the corpus luteum will persists longer than
the normal two-week lifespan as the human chorionic
gonadotropin (hCG) released from the chorion of the embryo
will act like LH to continue the secretory activity of the corpus
luteum which consequently stops the corpus luteum from
degenerating (Tortora & Derrickson 2009).
ii. Uterine Cycle
1. Progesterone and estrogen produced by the corpus luteum will
stimulate the growth and coiling of the endometrial glands.
Consequently, endometrial glands will undergo secretory
activity through glycogen secretion for nourishment of the
embryo. If fertilization does not occur, progesterone and
estrogen levels will decline which will result in the start of the
menstruation cycle (Tortora & Derrickson 2009).

Q: At which time point of the menstrual cycle is the endometrial sampling most likely
obtained?
Uterine tissues for endometrial sampling was most likely obtained during the
postovulatory phase in the secretory period. This phase happens for 14 days from
day 15-28 of the reproductive cycle wherein the uterus prepares for fertilization and
implantation. As the name implies, the secretory phase or period involves the
secretory activity of the endometrial glands. Progesterone levels rise due to the
rupturing of the mature follicles. Glycogen, the molecular form of carbohydrates, is
released by the glands. Mucous secretions are enhanced as cervical mucus provides
energy supply to the sperm. Progesterone also stimulates the endometrium to
increase vascular supply by coiling the spiral arteries, thereby increasing surface area
for blood to flow. Lastly, endometrial proliferation is reduced and the uterine lining
thins out (Thiyagarajan & Jeanmonod, 2022).

Diagnostics and Pathophysiology

● Irregular menstrual period
● every 10-90 days and lasts 2-10 days
● Spotting to quarter-sized clots
● Difficulty conceiving

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 ● Gynecologic findings: Normal uterus, external genitalia, adnexa, Tortuous spiral
arteries, coiled glands with carbohydrate-rich mucus, and edematous stroma in the
endometrium

Obese women often present with oligomenorrhoea (infrequent periods), amenorrhoea

(stopped completely) or irregular periods (Seif et al. 2015). The age of onset of obesity and
that of menstrual irregularities are significantly correlated. Obesity is associated with many
co-morbidities. Women who are obese suffer disorders of reproduction including infertility,
polycystic ovarian syndrome (PCOS) and menstrual disorders.

Specifically, women with obesity (BMI ≥30 kg/m2) were twice as likely as normal weight
women to have an irregular menstrual cycle. Furthermore, women with higher waist
circumferences and waist-to-hip ratios were more likely to have long cycles (greater than 35
days) (Itriyeva 2022).

It was reported that the prevalence of menstrual cycle irregularities was 8.4% in women
who were 74% overweight, as opposed to 2.6% in women who were <20% overweight [9]. A
further study documented that being 15% overweight was associated with a significantly
higher chance of having a menstrual cycle longer than 43 days

Range of abnormalities include

● irregularities in amount of menstrual blood loss,
● duration of bleed,
● length and regularity of menstrual cycle.
● heavy menstrual bleeding (HMB)

Pathophysiology
Hormonal effects
- Elevated androgen levels (ovarian and adrenal), thereby production of estrogens
- Relative FSH deficiency
- Excess production of LH, also stimulates production of androgens
- Inhibited production of SHBG
- In PCOS: elevated free and total testosterone and androstenedione levels; low SHBG
levels; , mildly elevated dehydroepiandrosterone sulfate (DHEAS), and occasionally
an elevated LH to FSH ratio

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 Body Mass Index (BMI)
l. Approximate measure of body fat based on height and weight
BMI Weight Status

Below 18.5 Underweight

18.5-24.9 Healthy Weight

25.0-29.9 Overweight

30.0 and above Obesity

Treatment
● Weight management program (weight loss).
● Hormonal treatment (but may be risky considering risk factors associated with
obesity)
○ combined oral contraceptive pills (COCP)
○ progestogen-only pill (POP)
○ subdermal progestogen implants
○ progestogen-releasing intrauterine devices

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