- Cervical cytology, also known as the Pap test, is a screening test used to detect cervical cancer and precancerous conditions. It was introduced in the 1940s by Dr. George Papanicolaou. - Screening guidelines recommend starting Pap tests at age 21, with testing every 3 years from ages 21-29 and every 5 years with co-testing from ages 30-65. Screening can stop at age 65 if previous results were normal. - The Bethesda System provides a standardized terminology for reporting cervical cytology results and categorizes findings as negative, atypical squamous cells, low-grade or high-grade squamous intraepithelial lesions, or cancer. Management depends on

1. CERVICAL CYTOLOGY
Dr. Rajesh Deo
Resident
Department of Pathology

2. • A screening test for detection of cervical cancer and
precancerous conditions.
• Dr. George Papanicolaou- introduced cervical
cytology in clinical practice (1940)

3. Whom to screen
• Should start at the age of 21 regardless of the onset of
sexual activity
• Women (21-29 yrs) should have pap test every 3 yrs
• Women (30-65 yrs) should have co-test (Pap test and HPV
test) every 5 yrs or pap test alone every 3 yrs.

4. When to stop routine screening
• Age 65 and above if
- no history of dysplasia or cancer
- 3 negative pap test in a row or 2 negative co-tests in a
row in past 10 years (most recent tests -within 5 years)
• Total hysterectomy (No CIN II/III, Ca)

5. Not applicable to:
• History of cervical cancer
• HIV infected, with weak immune system
• DES exposure in utero

6. NHSCSP guidelines
• Under 24.5 no invitation.
• 24.51st invitation.
• 25 to 49every 3 yrs
• 50 to 64every 5 yrs
• 65+for those who had abnormal tests.

8. THE BETHESDA SYSTEM OF
REPORTING CERVICAL CYTOLOGY
• Started 3 decades back in 1988
• Profound effect on cervical cytology for pathologists and
clinicians.
• Updated in 2014

9. TBS: 2001
SPECIMEN TYPE: Conventional smear vs LBC
 SPECIMEN ADEQUACY
-Satisfactory for evaluation
-Unsatisfactory for evaluation

10.  INTERPRETATION/RESULT
1.NEGATIVE FOR INTRAEPITHELIAL LESION OR
MALIGNANCY-No cellular evidence of neoplasia
Organisms
•Trichomonas vaginalis
•Fungal organisms morphologically consistent
with Candida spp
•Shift in flora suggestive of bacterial vaginosis
•Bacteria morphologically consistent
with Actinomyces spp
•Cellular changes consistent with Herpes simplex virus

11. Non-neoplastic findings (Optional to report)
- Reactive cellular changes associated with:
Inflammation and Radiation
- Glandular cells status post hysterectomy
- Atrophy

12. 2. OTHER
• Endometrial cells
• >= 40 years of age
• Negative for squamous intraepithelial lesion should be
specified.

13. 3.EPITHELIAL CELLABNORMALITIES
• Squamous cell
•Atypical squamous cells
-Of undetermined significance (ASC-US)
-Cannot exclude HSIL (ASC-H)
•Low grade squamous intraepithelial lesion
(LSIL)
•High grade squamous intraepithelial lesion
(HSIL)
•Squamous cell carcinoma

14. • Glandular cell
•Atypical
-Endocervical cells
-Endometrial cells
• Adenocarcinoma in situ
•Adenocarcinoma
-Endocervical
-Endometrial
-Extrauterine
-NOS

15. • OTHER MALIGNANT NEOPLASMS (specify)
• ANCILLARY TESTING
• AUTOMATED REVIEW
• EDUCATIONAL NOTES AND
SUGGESTIONS (optional)

16. SPECIMEN ADEQUACY
Satisfactory for evaluation
1. Presence /absence of
endocervical/transformation zone
component should be mentioned.
2. At least 10 well preserved endocervical
or squamous metaplastic cells singly or in
clusters
3. Other quality indicators/qualifiers-
partially obscuring blood,
inflammation)
4.Specimen with abnormal cells ( ASC-
US, AGC)

17. Unsatisfactory for
evaluation(processed/unprocessed)
Scenarios:
1. No identification
2. Broken slide
3. More than 75 % of squamous cells obscured-no abnormal
cells are identified

18. Organisms
Trichomonas vaginalis:
• Pear- shaped, oval or round
cyanophilic organisms ranging from
15-30 micron
• Nucleus is pale, vesicular,and
eccentrically located.
• Flagella seen in liquid based
preparation.
• Leptothrix may be seen in association
with the organism.

19. Fungal organisms morphologically consistent with Candida spp
• Budding yeasts
• Pseudohyphae

20. Shift in flora suggestive of bacterial vaginosis
• Individual squamous cells -covered by a layer of bacteria
that obscures the cell membrane- clue cells.
• Conspicuous absence of lactobacilli.

21. Bacteria morphologically consistent with Actinomyces spp
• Tangled clumps of filamentous
organisms recognizable as “ cotton
ball” clusters
• Associated with IUCD use

22. Cellular changes consistent with Herpes simplex virus
• Nuclei have a “ ground glass”
appearance
• Large multinucleated epithelial cells
with molded nuclei

23. Other non neoplastic findings
Reactive cellular changes
associated with inflammation
• Nuclear enlargement (1/1.5-2 times
more)
• Occasional binucleation or
multinucleation
• Nuclear outlines smooth, round and
uniform, vesicular nuclei
• Prominent single or multiple nucleoli
• Cytoplasm : polychromasia,
vacuolization, perinuclear halos
without peripheral thickening

24. Reactive cellular changes associated with radiation:
• Cell size is markedly increased
without increase in N:C ratio i.e
cytoplasm n nucleus is
proportionately increased
• Prominent single or multiple
nucleoli is coexisting repair
• Cytoplasmic vacuolization and/or
polychromatic staining
• Bizzare cell shape may occur

25. Reactive cellular changes associated with
intrauterine contraceptive device
• May be present singly or in small
clusters, mimicking a signet ring
appearance

26. Atrophy:
• Flat, monolayer sheets or dispersed parabasal cells
• Blue blobs
• Multinucleated giant cells

27. Endometrial cells
• Exfoliated endometrial cells in ball like
clusters
• Normal during first half of menstrual cycle
• Endometrial cells (>= 40years of age)

28. Epithelial cell abnormalities
Squamous cell:
Atypical squamous cells(ASC)
• of undetermined significance(ASC-US)
• cannot exclude HSIL(ASC-H)

29. Atypical squamous cells
• Refer to cytological changes, suggestive of SIL which are
qualitatively or quantitatively insufficient for definitive
interpretation.
• Doesnot represent a single biological entity
• 50% with ASC are infected with HPV
• Remaining may mimic numerous non neoplastic condition-
inflammation, air drying artefacts, atrophy with degeneration,
radiation

30. ASCUS
• > 90 % of ASC
• s/o LSIL but are quanititatively or
qualitatively insufficient for definite
interpretation.
• Cells resemble superficial or intermediate
cells
• 2.5-3 times increase in nuclear size with
minimal nuclear irregularities
• 2 times size of squamous metaplastic cells
• Repeat after 6 months or HPV test(LBC)
or colposcopy.

31. ASC-H
• Cells resemble parabasal and basal
cells
• nuclei 1.5 to 2.5 larger than normal
• Hyperchromatic nuclei with
irregularities
• Management - colposcopy

32. Squamous Intraepithelial lesion ( SIL)
•Low grade squamous intraepithelial lesion (LSIL)
HPV(low risk)/mild dysplasia/CIN 1
•High grade squamous intraepithelial lesion (HSIL)
Moderate and severe dysplasia/CIS/CIN 2 & 3
/HPV(high risk)

33. HPV
• Low risk HPV: HPV 6, 11
• High risk HPV: HPV 16( most common), 18, 31, 33
• HPV are associated with both low and high grade SIL.
• LSIL & HSIL -2 different biological entities
• (LSIL)Transient infections: regress over a period of 1 to 2
years
• (HSIL)Persistent infections associated with increased risk of
precancerous lesions/ invasive carcinoma

34. HPV
• 15 high risk HPVs currently identified,
• HPV-16 60% of cervical cancer cases, and
• HPV-18 accounts for another 10% of cases
• On average, 50% of HPV infections are cleared within 8
months,
• and 90% of infections are cleared within 2 years.
• HPVs infect immature basal cells of the squamous
epithelium in areas of epithelial breaks,
• or immature metaplastic squamous cells present at the
squamocolumnar junction.

35. PATHOGENESIS
• viral proteins E6 and E7 INHIBITS activity of tumor
suppressor proteins that regulate cell growth and
survival.
• Viral E7 binds RB (hypophosphorylated
form)degradation by proteasome pathway
• Binds and inhibits p21 and p27(CDKI) enhances cell
cycle progression and inhibit DNA damage repair
.

36. PATHOGENESIS..contd
• viral E6 proteins of high-risk HPV subtypes binds to the
tumor suppressor protein p53 and promote its
degradation by the proteasome.
• In addition, E6 up-regulates the expression of
telomerase, which leads to cellular immortalization.
The net effect is increased proliferation of cells that are
prone to acquire additional mutations that may lead to
cancer development.

37. LSIL
• Cells occur singly and in sheets
• Cytological changes confined to cells
with “ mature” intermediate or superficial
type cytoplasm.
• >3x intermediate nuclei with slightly
increased N:C ratio
• Hyperchromatic to normochromatic
• Smooth to irregular nuclear membrane
• Nucleoli absent or inconspicuous
• Koilocytosis: cells with enlarged
hyperchromatic nuclei, irregular contour
with perinuclear halo

38. • LSIL does not progress directly to invasive carcinoma and
in fact most cases regress spontaneously; only a small
percentage progress to HSIL.
• LSIL is not treated like a premalignant lesion.
• LSILs are ten times more common than HSILs.
• More than 80% of LSILs are a/w HPV.

39. MANAGEMENT OF LSIL
• <25yrs and LSIL+ve follow up with cytology at
12months
• >25 yr, HPV –vefollow up after 3yrs
• >25yr, HPV+ve colposcopy
• >25yrs, HPV unknown Repeat cytology in
12months

40. Koiocyte

41. HSIL
• Cytological changes involve smaller and
less mature cells than LSIL.
• Cells occur singly or in syncytial like
aggregates
• Nuclear hyperchromasia with greater
nuclear irregularities
• N:C ratio higher than LSIL
• Chromatin fine or coarsely granular
• Nucleoli absent but occasionally seen
• Cytoplasm is immature, lacy and delicate
or densely metaplastic or occasionally
mature and densely keratinized
• Immediate LEEP or colposcopy

42. • HSIL, there is a progressive deregulation of the cell cycle
by HPV increased cellular proliferation, decreased or
arrested epithelial maturation, and a lower rate of viral
replication, as compared with LSIL.
• Derangement of the cell cycle in HSIL may become
irreversible and lead to a fully transformed malignant
phenotype, and thus all HSILS are considered to be at
high risk for progression to carcinoma.
• More than 100% HSIL a/w HPV.

43. MANAGEMENT OF HSIL
• >25YRS and +ve HSILImmediate excisional
procedure at the time of colposcopy if lesion is
identified.
• If biopsy confirmed HSIL not identified at the
time of colposcopyp16 IHC to be done.

47. Squamous cell carcinoma
• Malignant tumor showing differentiation towards
squamous cells
• Does not subdivide squamous cell carcinoma
• Tadpole cells, tumor diathesis

48. KERATINIZING SCC
• Mostly singly and less commonly cellular
• Variation in shape and size caudate to spindle
• Dense orangeophilic cytoplasm
• Dense opaque nuclei with irrregular nuclear membrane
• Coarsely granular to irregular with chromatin clearing
• Less commonly tumor diathesis

49. NON-KERATNIZING SCC
• Occur singly or in clusters
• Smaller than HSIL
• Coarsely clumped chromatin with chromatin clearing
• Prominent nucleoli
• Tumor diathesis common necrotic debris and broken
blood elements

51. Bethesda system 2014: changes
• increased use of liquid-based preparations
• addition of co-testing (Pap and HPV testing)
• primary HPV testing as additional screening options
• further insights into HPV biology
• approval and implementation of prophylactic HPV
vaccines and
• updated guidelines for cervical cancer screening and
clinical management.

52. What has changed?
1.Reporting of benign-appearing endometrial cells is now
recommended for women aged ≥45 years.
- to increase the predictive value of this category.

53. 2. No new category was created for squamous lesions with
LSIL and few cells s/o concurrent HSIL.
LSIL in addition to ASC-H(LSIL is present with possibility of
HSIL.)

54. LIQUID BASED CYTOLOGY
• Is a monolayer slide preparation technology introduced to
overcome shortcomings of conventional pap smear
• Produces a sample fully representative of the material

55. Advantages of LBC over conventional pap
smear
Provides a more representative cervical sampling
Reduces obscuring factors
Lower unsatisfactory rates
Less screening time
Fewer artefacts in cellular morphology
Cells are deposited on slide in a monolayer
Provides representative residual material that can be used for
additional test like HPV test
Immunohistochemical analysis

56. FDA approved
1.Surepath
• Centrifugation and sedimentation through a density
gradient
2.ThinPrep
• Filtration and collection of vacuum-packed cells on a
membrane and transferring to the glass slide

58. Processing: SurePath
• Works on principle of density gradient.
• The vials are vortex mixed to re-suspend cells.
• An aliquot is treated with a density reagent and
centrifuged.
• This produces a concentrated pellet of cells

59. • Cells sediment to form a thin layer and excess fluid is
discarded.
• Staining is an integrated part of the process

60. Disadvantages of LBC
• Cost
• Not suitable for smaller centers dealing with fewer
samples
• Some studies show equal specificity in detecting HSIL and
carcinoma as conventional

61. HPVVACCINE
• Currently used in UK is Gardasil.
• Protects against 4 types of HPV (16,18,6 and 11)
• 99% effective against cancer caused by HPV 16 and 18.
• 1st dose12 -13 yrs older
• 2nd dose12 months or 6months gap between the doses.
• For aged>15yrs or older who have not vaccinated at 12-13
yrs 3 doses to be taken.

62. Thank you!! 