1) The document describes the Bethesda System for reporting cervical cytology results, which standardizes terminology for specimen adequacy, interpretation, and epithelial cell abnormalities. 2) Key cell types of the cervix are described along with non-neoplastic and reactive findings like organisms, inflammation, and hormonal changes. 3) Epithelial cell abnormalities are classified as either squamous or glandular, ranging from atypical/precancerous lesions to carcinoma. Nuclear and architectural criteria are provided to distinguish between low and high-grade squamous intraepithelial lesions.

1. CERVIX-CYTOLOGY
BETHESDA SYSTEM
Dr. Sobia Khalid

2. Papanicolaou (Pap) stain
 Alcohol dried; better for nuclear detail
 Stains ribosomes blue green, particularly in parabasal
cells, mesothelial cells and metaplastic squamous
cells
 Stains metabolically inactive cells pink, such as
superficial cells
 Stains keratinized cells or thick specimens orange
(benign or malignant)

3. Papanicolaou (Pap) stain

4. Liquid based cytology
 Head of spatula, where cells are lodged, is broken off
into small glass vial containing preservative fluid, or
rinsed directly into preservative fluid
 Sample is sent to lab, then spun and treated to
remove mucus, pus or other obscuring material
 Random sample of remaining cells is taken and
deposited onto a slide
 Reduces number of inadequate smears and need for
repeat smears

5. Liquid based cytology

6. Diff-Quik stain
 An air dried, Giemsa-type stain
 Better for background material or to assess adequacy
of endocervical smears to detect C. trachomatis
 Used for fine needle aspirates, not for cervical smears

7. CERVIX - Normal cells
 Basal cells
 Parabasal cells
 Intermediate cells
 Superficial cells
 Endocervical cells

8. Basal Cells
 Small undifferentiated cells
 Seldom identified in pap smears

9. Parabasal cells
 Cytoplasm is round, dense, basophilic
 Nucleus is vesicular, central, round and relatively
large (50μm²)
 May see naked nuclei
 Higher N/C ratio and smaller size than intermediate
cells

10. Parabasal cells

11. Intermediate Cells
 Cytoplasm is polygonal, transparent, basophilic
 Nucleus is about the size of a red blood cell (35 μm²),
is vesicular, round/oval
 Nuclear texture and size is reference for dysplasia

12. Intermediate Cells

13. Superficial cells
 Cytoplasm is polygonal, transparent, eosinophilic
 Nucleus is pyknotic, round/oval

14. Superficial cells

15. Endocervical cells
 Columnar cells
 Vacuolated or granular cytoplasm
 Prominent cell borders
 Basal nuclei with fine granular chromatin
 Occasional nucleoli
 Honeycomb appearance
 Ciliated if tubal metaplasia

16. Endocervical cells

17. Squamous Metaplasia
 Single cells, loosely cohesive groups or sheets
 Variable cell size and shape
 Cell borders are more defined with maturity
 Cytoplasm is variable with occasional vacuoles
 Nuclei is slightly larger than intermediate cell nuclei,
has finely granular and evenly distributed chromatin,
nuclear membrane is smooth, no prominent nucleoli

18. Squamous Metaplasia

19. BETHESDA
SYSTEM

20. BETHESDA SYSTEM
 Uniform terminology introduced in 1988 and revised
in 1991 and 2001
 To establish uniform terminology and standardize
diagnostic reports.
 In addition, it introduced a standardized approach
for reporting if an individual specimen is adequate
for evaluation.

21. BETHESDA SYSTEM, 2001
 Specimen adequacy
 General categorization (optional)
 Interpretation/result
 Automated review and ancillary testing (include if
appropriate)
 Educational notes and suggestions (optional)

22. Adequacy
 Satisfactory for evaluation (describe presence or
absence of endocervical/transformation zone
component and all other quality indicators, such as
partially obscuring blood, inflammation, etc.)
 Unsatisfactory for evaluation (indicate reason)
 Specimen rejected/not processed (indicate reason)

23. Assessment of adequacy
 Adequate number of squamous cells (conventional
smear should have 8000-12000 cells, liquid-based
preparation should have 5000 cells)
 The presence or absence of endocervical cells should
be reported; an adequate number of endocervical
cells (at least 10 well-preserved endocervical or
metaplastic cells) confirms sampling of transition zone
 Specimen with more than 75% of cells obscured by
inflammation and bacteria is unsatisfactory (however,
should still report presence of abnormal cells)

24. Unsatisfactory: scant
squamous cellularity

25. Adequate

26. Interpretation/result
 Negative for Intraepithelial Lesion or Malignancy
(NILM)
 Epithelial Cell Abnormality
 Other malignant neoplasm (Specify)

27. Negative for Intraepithelial
Lesion or Malignancy (NILM)
 Organisms
 Trichomonas vaginalis
 Fungal organisms morphologically
consistent with Candida species
 Shift in flora suggestive of bacterial
vaginosis
 Bacteria morphologically consistent with
Actinomyces species
 Cellular changes associated with Herpes
simplex virus

28. Trichomonas vaginalis
(pear shaped)

29. Candida (Branching
pseudohyphae and spores)

30. Gardenella Vaginalis
(Clue cells)

31. Actinomyces
(Dust bunnies)

32. Herpes Simplex

33.  Other non-neoplastic findings (optional to
report, list is not inclusive)
 Reactive cellular changes associated with:
• inflammation (includes typical repair)
• irradiation
• Intrauterine contraceptive device (IUD)
 Glandular cells status post hysterectomy
 Atrophy

34. Reactive cellular changes,
Repair
 Flat monolayer
sheets with
distinct
cytoplasmic
outlines,
streaming
nuclear
polarity,
prominent
nucleolus in
almost every
cell.

35. Reactive cellular changes,
Radiation
 Enlarged nuclei
with abundant
polychromatic
cytoplasm with
vacuolization.
Mild nuclear
hyperchromasia
without coarse
chromatin,
prominent
nucleoli

36. Reactive cellular changes,
IUD
Glandular cells in
small clusters with
increased N/C
ratio and
cytoplasmic
vacuoles. Nuclear
degeneration and
prominent nucleoli
present.

37. Glandular cells status post
hysterectomy
 Goblet cell
metaplasia
and bland
cellular
features.

38. Atrophy
 Sheets of
uniform orderly
parabasal
cells. Some
nuclei may
show grooves,
but chromatin
pattern is fine.
Atrophic cells
may have
nucleoli.

39.  Other
 Endometrial cells (in a woman greater than
or equal to 40 years of age; specify if
“negative for squamous intraepithelial
lesion”)

40. Endometrial cells
Cells occur in small
clusters. Small,
round nuclei similar
in size to a normal
intermediate cell
nucleus.
Inconspicuous or
absent nucleoli.
Cytoplasm is scant,
basophilic and
sometimes
vacuolated. Cell
borders are ill-
defined.

41. Epithelial Cell Abnormality
 SQUAMOUS CELL
 Atypical squamous cells
 - of undetermined significance (ASC-US)
 - cannot exclude HSIL (ASC-H)
 Low grade squamous intraepithelial lesion (LSIL)
 - encompassing HPV/mild dysplasia/CIN I
 High grade squamous intraepithelial lesion (HSIL)
 - encompassing: moderate and severe
dysplasia/CIN2/CIN3/CIS
 - with features suspicious for invasion (if invasion
suspected)
 Squamous cell carcinoma

42. Atypical squamous cells of
undetermined significance (ASC-
US)
 May be neoplastic (HPV related, LSIL, HSIL) or
reactive
 Nuclear changes more marked than reactive, less
than LSIL; nucleus is 2.5-3x size of intermediate cell
nucleus
 In perimenopausal women (40-55 years), cells with
bland nuclear enlargement (2-3x size of intermediate
cell nuclei), smooth nuclear membranes and fine
chromatin are likely to be negative for SIL/malignancy

43. Atypical squamous cells - of
undetermined significance
(ASC-US)

44. Atypical squamous cells -
cannot exclude HSIL (ASC-H)

45. Squamous Intraepithelial
Lesions

46. Low grade squamous
intraepithelial lesion (LSIL)
 HPV/mild dysplasia/CIN I
 usually transparent cytoplasm
 enlarged nuclei at least 3x size of nucleus of
intermediate cell, but N/C ratio is less than 1/3
 hyperchromasia (darker than intermediate cell)
 cell may be enlarged (largest atypical cells in
gynecologic cytology are LSIL), but still has
elevated N/C
 no nucleoli
 binucleation is more common than multinucleation
 koilocytes may be present

47. LSIL

48. LSIL

49. HSIL / CIN II / moderate
dysplasia
 N/C ratio is 1/3 to ½
 cell size is same as squamous metaplastic or
parabasal cells
 polygonal shape (like intermediate or superficial cell)
 denser cytoplasm
 enlarged and hyperchromatic nucleus
 nuclear membranes may be irregular (crinkled paper)
 no nucleoli

50. HSIL / CIN II / moderate
dysplasia

51. HSIL / CIN II / moderate
dysplasia

52. HSIL / CIN III / severe
dysplasia
 Usually single, small, round/oval cells
 scant cytoplasm
 nuclei same size as dysplastic cells but increased N/C
ratio (greater than 1/2) since only minimal rim of
cytoplasm
 nuclei vary markedly in contour and have irregular
coarsely clumped chromatin
 no macronucleoli
 no tumor diathesis

53. HSIL / CIN III / severe
dysplasia

54. HSIL / CIN III / severe
dysplasia

55. HSIL-Carcinoma in Situ
 Hyperchromatic crowded groups
 Undifferentiated cytoplasm
 Ill-defined cell borders
 Loss of nuclear polarity

56. Carcinoma in situ

57. Carcinoma in situ

58. Squamous cell carcinoma
 Highly irregular shaped cells (tadpole, caudate)
 Keratinized are orange, often with squamous pearls
 Non-keratinized cells have dense, hard, basophilic
cytoplasm; also cannibalism (tumor cells surround
other cells)
 Tumor diathesis in background (necrosis,
hemorrhage, inflammatory cells) is suggestive of
malignancy

59. Squamous cell carcinoma

60. Squamous cell carcinoma

61. EPITHELIAL ABNORMALITIES
- GLANDULAR
 Atypical
 Endocervical cells, NOS or specify in comments
 Endometrial cells, NOS or specify in comments
 Glandular cells, NOS or specify in comments
 Atypical
 Endocervical cells, favor neoplastic
 Glandular cells, favor neoplastic
 Endocervical adenocarcinoma in situ
 Adenocarcinoma
 Endocervical
 Endometrial
 Extrauterine
 Not otherwise specified (NOS)

62. Atypical Glandular Cells -
AGC
 Morphological changes of glandular cells which are
too pronounced for inflammatory or reactive origin,
but insufficient to diagnose an adenocarcinoma
 More likely to be neoplastic if decreased cytoplasm,
irregular nuclear membranes and nucleoli present

63. Atypical Glandular Cells -
AGC

64. Atypical Glandular Cells -
AGC

65. Adenocarcinoma in situ
(AIS)
 Tightly crowded sheets of malignant cells with
architectural disarray, often with short strips of
pseudostratified columnar cells near edges
 Nuclei may be partially denuded, causing a feathered
appearance
 Nuclei are enlarged, usually oval, and
hyperchromatic
 Often prominent nucleoli
 No tumor diathesis

66. Adenocarcinoma in situ
(AIS)

67. Adenocarcinoma in situ
(AIS)

68. Adenocarcinoma
 Multilayering
 May form glandular structures with central lumina or
acinar formations with peripheral nuclei
 Cells are large or small with pleomorphism; have fluffy
cytoplasm, cytoplasmic vacuolization, loss of nuclear
polarity, true nuclear crowding, nuclei with clumped
chromatin, marked variation of nucleoli, occasional
mitotic figures; invasion is often characterized by
heavy blood with abundant glandular material

69. Adenocarcinoma

70. Adenocarcinoma

71. ANCILLARY TESTING
 Describe briefly the test method(s) and report the result
so that it is easily understood by the clinician

72. AUTOMATED REVIEW
 If case is examined by automated device, specify
the device and result

73. EDUCATIONAL
NOTES/SUGGESTIONS
 If provided, should be concise and consistent with
clinical guidelines published by professional
organizations.

74. THANK YOU