HORMONAL CYTOLOGY STUDY

HORMONAL CYTOLOGY STUDY DR. ANINDITA SAHA & DR. ANKITA PAUL

Hormones influences the morphology and staining characters of endometrial, endocervical and vaginal cells.

Non- invasive procedure of epithelium for hormonal status

Vaginal epithelium is very sensitive to estrogen and progesterone

Indication of cytological hormonal evaluationAssessment of ovarian functionAfter hysterectomyDuring menstrual cycleIn premature mensesAssessment of abnormal hormonal productionPregnancy , Abortion , Retained placentaVarious endocrine disordersExistence of hormone producing ovarian tumorsAssessment and guidance of hormonal therapy.

For useful interpretation; the following information must be taken in account:-Age of the patientMenstrual history (regular or irregular cycles)Previous past history:Hormonal therapySurgical operations in the genital tractIrradiation

SAMPLE COLLECTIONThe ideal type for sample collection is by: Aspiration of vagino-cervical secretion from the posterior vaginal fornix ORGentle scraping from the lateral mid-third of healthy vaginal wall.

NORMAL CELLS SEENThe normal pap smear shows the following types of squamous epithelial cells. 1)superficial 2)intermediate 3)parabasal

SUPERFICIAL SQUAMOUS CELLS

- -Most mature cells of ectocervix.

-Most abundant during proliferating phase of MC under the influence of estrogen.

-Polygonal with abundant eosinophilic cytoplasm.

-Small pyknotic nucleus(5-6 micron meter)

-Show cytoplasmic keratohyaline granules.

INTERMEDIATE SQUAMOUS CELLS

-Polygonal in shape

-Abundant bluish cytoplasm

-Nucleus is larger with granular chromatin

-Most abundant during secretory phase of MC under the influence of progesterone

PARABASAL CELLS

Immature squamous cell

Round- Oval shape

Nucleus is relatively larger. N/C= 1:2

Nucleus is vesicular with fine reticular chromatin

Cytoplasmic area is smaller

Cytoplasmic texture is granular and dense

BASAL CELLS

Small (8-10 m), round to oval

Dense cyanophilic cytoplasm

Nucleus large, fine reticular chromatin, small nucleoli

ENDOCERVICAL CELLS

-Cells are columnar

-Abundant vaculated cytoplasm

-Eccentrically placed vesicular nucleus , inconspicuous nucleoli

-Cells arranged in strips giving a picket fence appearance or in sheets resembling honey comb

EXFOLIATED ENDOMETRIAL CELLSSmall cells with dark nucleus and scant cytoplasm

Nucleoli is inconspicuous

Arranged in tight balls like 3-D clusters

Seen commonly during 1st 12 days of MC

Background haemorrhage is indication.

Endometrial cells (Exodus ball)

Seen between 6 to10 days of the menstrual cycle.

Last remnants of endometrial shedding and the cells show degenerative changes

Physiology of hormone cycle in womenIn infancy and childhood:Small amount of estrogen without progesterone inactive ovary.

At puberty:1. Follicle Stimulating Hormone : from the pituitary gland --- proliferation of ovarian follicles ---- estrogen secretionMaturation of vaginal epitheliumProliferative phase of endometrium

2. LH (luteinizing hormone): cause maturation of ovarian follicles until rupture and release of ova (ovulation).Maintain corpus luteum and progesterone secretion.Stimulate secretory phase of endometrium

If no pregnancy (no implantation of fertilized ova) --- sudden drop of progesterone and estrogen level ---- menstrual bleeding (shedding of endometrium and basal blood vessels).

If pregnancy occur (implantation of fertilized ovum) --- corpus luteum continuous secret progesterone and gonadotrophic hormones ----- until the third month of gestation.Also; placenta secrete progesterone and gonadotrophic hormones

Normal cyto-hormonal patterns in women Throughout life, women under variations in type and level of hormone, which could be due to some factors such as:-AgePregnancyMenopauseFunction of pituitary ovarian adrenal axis

The general pattern of the smear depends on the level of

gonads hormones, on the vaginal microbiologic factors

and it varies with age.

Hormonal effect Estrogen:Proliferation and maturation of the vaginal squamous epithelial cells, including the superficial cells. Deposition of glycogen within the vaginal epithelium.

Progesterone and androgen:Rapid desquamation of the upper layer of epithelium.Exposed intermediate and parabasal cells to the surface

AT BIRTH

Gonadal hormones are produced in a large amount during pregnancy and pass through the placenta into the fetal circulation.

The squamous epithelium of the cervix and of the vagina of a newborn girl responds to this strong hormonal stimulation.

A smear , obtaining with a thin cotton applicator , contain a clear predominance of superficial cells

IN CHILDHOOD

After a few days after birth the maternal hormones are eliminated.

The smears contain mostly parabasal cells, reflecting the absence of gonadal hormones

AT PUBERTY

Even before the first menstrual period occur ,the vaginal smear begins to change;

intermediate cells replace the parabasal cells and

a few superficial cells reflect the onset of estrogen production in the ovaries.

DURING THE REPRODUCTIVE YEAR

DAY 1 of the cycle is the first day of menstruation. DAY 1 DAY 5(onset of menstruation) smear shows 1) blood 2) desquamated endometrial cells in singles and clusters 3) polymorphonuclear leukocytes. 4) squamous cells predominatly intermediate type. Such cells form clumps and their cytoplasm is folded and degenerated. On the 4th or 5th day, the squamous cells begin to show less clumping and a better cytoplasmic preservation.

DAY 6 DAY 13/14 (proliferative/preovulatory) increase in estrogen

1)Endometrial cells in clusters2) squamous cells predominatly intermediate type later replaced by superficial type(12th to 14th day)3) thick cervical mucus forms fern-like crystalline structures that vanish just prior to ovulation4) Small macrophages5) small nipple-like nuclear protrusions may occasionally be seen in the endocervical cells

A cluster of endometrial glandular cells observed on day 7 of menstrual cycle.

Day 11 of menstrual cycle. The smear contains a mixture of intermediate and superficial cells.

Mid cycle - fernlike structure.

DAY 14-DAY 28(secretory/post ovulatory/luteal phase) increase progesterone intermediate squamous cells with few superficial squamous cells with cytoplasmic foldings.

As the time of menstrual bleeding approachesintermediate cells form clusters or clumps.Marked increase in lactobacilli, resulting in cytolysis of the intermediate cells. The cytolysis results in moth-eaten cell cytoplasm, nuclei stripped of cytoplasm (naked nuclei) in a smear with a background of cytoplasmic debris (dirty type of smear) This appearance of the smear persists until the new cycle begins with the onset of the menstrual bleeding.

Luteal phase intermediate cells

Cytolysis by Doderlein lactobacilli

DURING PREGNANCY Vaginal smears reflect the balance of hormones during pregnancy. Generally the high level of progesterone (placental) do not allow the complete maturation of the squamous epithelium. 1)clustering of intermediate squamous cells 2) predominance of navicular cells, defined by yellow cytoplasmic deposits of glycogen, displacing the nuclei to the periphery, and sharply defined, accentuated borders (presence of navicular cells is not diagnostic of pregnancy) 3) In the later stages of pregnancy, extensive cytolysis of squamous cell cytoplasm by lactobacilli is not uncommon 4)Endocervical cells increase in number. appearance of significant number of parabasal cells indicates fetal death.

MENOPAUSEEarly Menopause: Slight Deficiency of Estrogens 1)predominantly of dispersed intermediate cells occasionally showing cytolysis 2)some large parabasal cells 3) reduction in the proportion of superficial squamous cells

Crowded Menopause: Moderate Deficiency of Estrogens: 1)thick, crowded clusters of intermediate and large parabasal cells. The cytoplasm frequently contains deposits of glycogen in the form of yellow deposits, similar to navicular cells observed in pregnancy

Atrophic or Advanced Menopause: 1) relatively few cells 2)dominant squamous cells are of the parabasal type. 3) Blue blobs are sometimes noted, these being interpreted by some as mucin by others as degenerate cells. 4)granular debris in background.

Menopausal smear: parabasal cells and naked nuclei

Atrophic smear

Maturation IndexIt is the percentage study of the parabasal, intermediate, and superficial squamous cells\100 cells counted from exfoliated epithelial cells of healthy vaginal smear.It is determined by morphology of the nucleus and thickness of cytoplasm of epithelial cells.

Reading of the maturation index Shift to the right: indicate an increase number of superficial cell (maturation) i.e. 0\0\100 under the effect of increase estrogen like effect

Shift to the left: indicate an atrophic effect e.g. post menopause women i.e 100\0\0 with no effect of estrogen.

Shift to the mid-zone: means progesterone like effect e.g. secretory phase of endometrium i.e. 0\100\0

Effect of extrinsic hormones on vaginal cytologyEstrogen: MI=0\10\90Increase cell maturation Proliferation of all layers of epitheliumProgesterone: MI= 0\90\10Proliferation of intermediate cellsDecrease superficial cell maturationAndrogen like H. (testosterone) MI= 20\80\0 Increase number of parabasal and intermediate cellsNo superficial cells.

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