INTRODUCTION
 The epithelia of the female genital tract are
sensitive to estrogen , progesterone , androgens &
various additional steroid hormone derivatives &
metabolites.
 In general estrogen promotes & progesterone
inhibits squamous cell maturation.
 Hormones may influence the morphology staining
characteristics of the cervical , endometrial , vaginal
, urethral & bladder mucosa cell.
Hormonal evaluation is based on the degree
of maturation & glycogen storage of the
exfoliated squamous cells.

For useful cytologic evaluation , some
information about the age , menstrual history &
previous hormonal , surgical or treatment of the
patient should accompany the smear.

INDICATIONS FOR CYTOLOGIC HORMONAL
EVALUATION
1)Assessment of ovarian function.
During puberty & reproductive , menopausal &
post menopausal years after hysterectomy.
During menstrual disorders.
In premature menses (childhood).
To determine ovulation time.
2)Assessment of abnormal hormonal production
(placenta, ovaries & other endocrine organs).
 Before , during & after a pregnancy (fertility
study , threatened abortion , retained placenta ).
 Existence of functioning (hormone producing)
tumors.
 Various endocrine disorders.
 Assessment & guidance for hormonal therapy.
NORMAL HORMONAL PATTERNS
 ESTROGEN
 It increases cellular maturation & proliferation
of all layers of squamous epithelium mostly
Superficial cells and Intermediate cells.
DECREASING LEVEL
Deficiency of estrogen increases parabasal cells.
The mucosa becomes thin & superficial layer is
absent in deficient estrogen stimulation.
INCREASING LEVEL
Increase in estrogen level enlarges the parabasal
cells to intermediate .this continue to enlarge &
mature into keratinized superficial cells.
Increases the amount of intra cellular glycogen
( by protein & nucleic acid synthesis )
High estrogenic effect in a vaginal smear is
recognized by a clean background & increased
eosinophilic & karyopyknotic indices.
 PROGESTERONE
 It produce proliferation of intermediate
squamous epithelium at the time of exfoliation.
INCREASING LEVEL
 Clusters of intermediate cells dominate the
smear.
 The amount of intracellular glycogen-increases
 No. of leukocytes –decreases
 ANDROGEN
 Male sex hormone.
 Small amount is present in female.
INCREASING LEVEL
It increases the no. of exfoliated large parabasal cells
with central , almost pyknotic nuclei & dense , glycogen
containing cytoplasm.
GUIDELINES FOR SPECIMEN COLLECTION &
SUITABLE FOR HORMONAL ASSESSMENT
1)SPECIMEN COLLECTION GUIDELINES:
• Smear taken from the lateral vaginal wall
• Smear taken from the junction of the middle & upper 1/3rd
of the vagina.
2)FACTORS THAT MAKE A SMEAR UNSUITABLE FOR
HORMONAL ASSESSMENT:
• Material not collected properly.
• Adequate clinical information not provided (age ,
menstrual status, treatment etc)
• Evidence of an infection.
• Inflammation.
• Air-drying artifact.
HORMONAL PATTERN AT
DIFFERENT STAGES OF LIFE
LIFE STAGE
HORMONAL PATTERN
Newborn
Sterile
No inflammation
Intermediate cells
superficial cells
1st - 2 week of life
Acquires bacteria
Neutrophils appear
Atrophic cells
Childhood
Maturation & cyclic alternations occur months
(or)years before menarche, due to cycles that
are initially anovulatory
Cyclic women , proliferative
phase
Increasing squamous cell maturation
Cyclic women , post
ovulatory phase
Decreasing squamous cell maturation
Pregnancy
Navicular cells may be numerous
Lactation
Atrophy persists but some maturation
may appear slowly .
Post partum
Atrophy
Menopause
Atrophy (or) intermediate cell
predominate
CELLS
1)
SUPERFICIAL CELLS
 Cell – flat , delicate & transparent.
 Size – 35-45 micro meter in diameter.
 Nuclei – small , dark & pyknotic (nuclear material
become condensed & shrunken.
 Cytoplasm – delicate pink (eosinophilic / acidophilic)
2)
INTERMEDIATE SQAMOUS CELLS
• Cell – flat
• Size – 35-45 micro meter
• Nucleus – spherical / oval
-vesicular
• Cytoplasm – basophilic
• A variant of intermediate cells –boat shaped navicular
cells.
Navicular cells – common in early menopause &
pregnancy.
• Oval shaped cells store glycogen in the form of
cytoplasmic deposits – stains yellow in PAP stain
3) PARABASAL CELLS
 Size – 12 -30 micro meter.
 Nuclei – vesicular.
 Shape – round / oval.
 Cytoplasm – basophilic.
4) BASAL CELLS
 Shape – round / oval.
 Cytoplasm – scanty , basophilic.
 Nuclei – vesicular
-fine chromatic granules
-occasionally tiny round nucleoli
CYTOLOGICAL INDICES
The degree of proliferation , maturation &
desquamation of vaginal cells is influenced by
various hormones.
The cytological assessment of these hormones is
given by different types of indices . All are based
on recognition & exact typing of the epithelial cells
exfoliated from the surface of the stratified
squamous vaginal mucosa .
 If the smear is to inflammatory & a repeat
specimen is not possible after the proper
treatment , the exfoliated urethral cells in the
first portion of a voided urine specimen can be
examined for this evaluation.
 The terminal part of the urethral mucosa is
almost as sensitive to hormonal changes as in
the vaginal mucosa but more difficult to
evaluate.
TYPES OF INDICES
Maturation index (MI)
 It is determined by the percentage of the basal ,
intermediate & superficial cells as presented as 3 part
ratio with the basal cells started 1st , intermediate cells
2nd & superficial cells 3rd.
Example : MI = 80 /20 / 0
Indicates :
80 % - Parabasal cells
20 % - Intermediate cells
0 % - Superficial cells
RESULTS READ AS:
Basal
Intermediate
0
0
0
100
100
0
superficial
100
Shift - left
0
Shift -middle
0
Shift – right.
RESULT READ AS
Shift to left – atrophic effect.
Shift to right – increased estrogen like effect.
Shift to midzone – progesterone like effect
2)KARYOPYKNOTIC (KPI) / CORNIFICATION
INDEX
The percentage of squamous cell with sharp
squared cytoplasmic edges with pyknotic nuclei
compared with other mature squamous cells with
vesicular nucleus.
Parabasal cells are not counted.
3)MATURATION VALUE (MV)
This method is useful for providing hormonal
evaluation data to a computer.
Each parabasal cell is counted as – 0
Intermediate cell as – 0.5
superficial cell as - 1.0
The addition of all the values given to the first
100 epithelial cells is recorded.
Example : MI =0 / 35 / 65
0 x 0 =0
35x 0.5 = 17.5
65x1.0 = 65.0
MV = 82.5
MV of 100 indicates – pure population of superficial
squamous cells
MV of 0 indicates – pure population of parabasal cells.
STAGES
Newborn
MI
MV
VARIATION
0 / 90 / 10
55
+/ - 10
80 / 20 / 0
10
+ / - 20
Pre ovulatory
0 / 40 / 60
80
+ / - 10
Post ovulatory
0 / 70 / 30
65
+ / - 15
Menopause
0 / 80 / 20
60
+ / - 20
50 / 50 / 0
25
+ / - 40
Estrogen therapy
0 / 10 / 90
95
+ / - 10
Progesterone therapy
0 / 90 / 10
55
+ / - 10
Androgen therapy
20 / 80 / 0
40
+ / - 10
Infancy
Post menopause
4)EOSINOPHILIC INDEX (EI)
The no. of mature squamous cells with
eosinophilic (pink) cytoplasm , nuclear
appearance is compared with the no. of other
mature squamous cells with basophilic (blue)
cytoplasm.
The result is given in ratio.
Parabasal cells not counted.
Example : 2:14
2 – eosinophilic sq. cell
14 – basophilic sq. cells
5)FOLDED CELL INDEX (FCI)
No . of mature sq. cells with folded
cytoplasmic rims are compared with no. of mature
cells with flat non folded cytoplasm.
The folded cells with cytoplasmic glycogen are
considered as less mature than flat cells.
6)SUPERFICIAL CELLS INDEX (SCI)
Superficial sq. cells with any type of nucleus is
compared with any other sq. cell present in the smear.
7)CROWED CELL INDEX (CCI)
The no. of mature sq. cells that are clustered
together in groups of 5 or 6 cells is compared to the no.
of similar cells found singly / less than 3 or 4.
INDICES USED FOR HORMONAL
ASSESSMENT
TYPE OF INDEX
CELL TYPE RATIO COMPARED
Maturation index
Parabasal : intermediate : superficial
Karyopyknotic
index
Eosinophilic index
Superficial : intermediate
Folded cell index
Folded cytoplasm : non folded
cytoplasm
Crowed cell index
Clusters of more than 4 cells : single cell
Eosinophilic : basophilic
NORMAL CYTOHORMONAL AVERAGES
1)NEW BORN (UPTO 8 WEEKS)
MI =0 / 90 / 10
Intermediate cell – increase with glycogen in their
cytoplasm.
- this is due to maternal hormones
in infant’s blood.
1st 4 – 8 days - Very little cellular debris , bacteria ,
leucocytes , lymphocytes (or)mucous can
be found.
2 – 3 weeks – increased no. of leucocytes &
micro organism occurs.
3 – 4 weeks – intermediate cell decreases.
2) INFANCY & NEWBORN (3 WEEKS TO PUBERTY)
MI = 80 / 20/ /0
Vaginal pool smears are scanty in cells & contain
mainly parabasal cells.
Leucocytes – few.
These smears are very similar to the cells found in
the late postmenopausal period except for their
smaller size & the absence of inflammatory
background.
3)MENSTRUAL AGE (REPRODUCTIVE PERIOD)
MI = 0 / 70 / 30
The vaginal cellular changes start to appear several
months before the beginning of menstruation.
A nucleated eosinophilic sq. cells are present.
Basal cells & polymorphonuclear cells are the main
cells found during the reminder of the cycle.
Superficial & intermediate cells – always present.
Parabasal cells-few.
4)DURING PRE OVULATORY TIME.
(Proliferative phase, 3-14 day of the cycle)
MI=0/40/60
8th day of cycle -the cyanophilic intermediate cells
gradually increases in size with
progressive cytoplasmic eosinophilia
& nuclear pyknosis.
- no. of leukocytes – decreases.
mucous – decreases.
5) DURING OVULATION & POST OVULATORY TIME
(Secretory phase 15 – 19th days)
MI = 0 / 70 / 30
Super ficial & eosinophilic cells – decreases
Intermediate cells – increases & the cells show a
tendency to cluster together.
6) DURING MENSTRUAL FLOW.
MI = 0 / 60 / 40
Increase of fresh & old red blood cells , cyanophilic
intermediate cells, histiocytes, mucous & leukocytes.
Grandular & stromal endometrial cells are seen singly in
clusters /in acinic formation.
The evaluation of smears taken at 3 day intervals will
indicate if ovulation has occurred & the level of estrogenic
effect.
7) MENOPAUSE
MI = 0 / 80 / 20
Exfoliated superficial & intermediate sq. cells
become smaller & show some decrease in their
staining capacity.
8) EARLY POST MENOPAUSE
2-6 years cessation of menstruation a decrease in
estrogenic activity occurs.
Parabasal & intermediate cells – increases
Glycogen is found in some of the cells.
9) LATE POST MENOPAUSE
MI = 100 / 0 / 0
Some smear may become completely atrophic.
Atrophic parabasal cells:
size – varies
shape – oval / round with mild irregularities
cytoplasm – cyanophilic
-show degenerative changes in
the form of vacuolization.
No glycogen is present.