procedure# indications # anatomy and normal lining cells# acellular material#abnormal bronchial cells # atypical cancer cell#lung carcinoma cytology

1. BRONCHIAL BRUSHINGS AND
WASHINGS IN PULMONARY DISEASES
DR Y L S

2. ANATOMY
• The respiratory system is broadly divided into the upper
respiratory tract, lower respiratory tract and lungs
• Upper respiratory tract: It extends from nose to larynx
Lower respiratory tract: It extends from the trachea to the lungs
and is comprised of trachea, bronchi and bronchioles
• Lungs: It is made of respiratory bronchioles, alveolar ductsalveolar
sacs and alveoli.

3. • Epithelium shows mucosal lining from the nose to the alveoli.
• Most of the portion of the respiratory tract from the nose to the bronchi is lined by
ciliated columnar epithelium except
• vestibules of the nose
• vocal cords
• superior surfaces of the epiglottis, which are lined by non-keratinizing stratified
squamous epithelium.
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5. • Parts Mucosal lining
• Nasal vestibules -Stratified squamous epithelium
• Nasal sinuses -Ciliated columnar epithelium
• Nasopharynx -Ciliated columnar epithelium
• Larynx Mostly : Ciliated columnar epithelium
• Vocal cords and superior surfaces of the
• epiglottis: Stratified squamous epithelium
• Trachea and bronchi- Ciliated columnar epithelium, goblet
• cells, neuroendocrine cells
• Terminal bronchioles -Simple columnar to cuboidal epithelium
• Alveoli- Type I and II pneumocytes and alveolar
• macrophages

6. Bronchial Specimens
• Bronchial Aspirate and Wash
• Introduction of flexible fiber optic bronchoscope enables the
clinician to procure samples from the peripheral part of the
lower respiratory tract.
• With the help of bronchoscope, the bronchial secretion is
aspirated directly.
• In case of bronchial washing technique, 3–5 mL of balanced salt
solution or normal saline is introduced through the
bronchoscope and the fluid is re-aspirated.
The direct smear or multiple smears from the centrifuged fluid
sample are made for staining.

7.  Bronchial Brush
• The lesion is directly visualized by the fiber optic bronchoscope.
• The brush is applied to the lesion through the instrument.
• Multiple smears are made and fixed in 95% ethanol or the brush
 is rinsed in a collection solution and sent to the laboratory.
• The main advantage of the brush is that direct visualization of
 the lesion is possible and therefore diagnostic cellular yield is
 excellent.
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8. FLEXIBLE FIBER OPTIC ENDOSCOPE
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9. BRONCHIAL BRUSHES
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10. Bronchoalveolar Lavage
• Bronchoalveolar lavage (BAL) is a powerful investigative
technique in the field of pulmonary medicine.
• Procedure: BAL should be done before performing any other
techniques such as bronchial wash/brush or biopsy.
•
• Under local anesthesia, fiber optic bronchoscope is wedged
in a desired sub segmental bronchus at the desired location.
• At first, 20 mL of normal saline is flushed through bronchoscope
in the desired segment of the lung and gentle suction is done to
collect the lavage specimen.

11.  The whole procedure is done for five times to get 40-50ml solution and should be processed as
early as possible.
 Indications: The predominant clinical indications of BAL are:
 • Any case of non resolving pneumonia
 • Interstitial lung disease
 • To diagnose opportunistic infection in an immunocompromised
 host
 • Suspected case of peripheral lung carcinoma.
 Other than diagnosis, BAL is also useful for therapy of alveolar proteinosis, cystic fibrosis and
pulmonary alveolar microlithiasis
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12. INSTRUMENTS USED FOR BAL
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14. Adequacy of BAL Sample
• There are no absolute indicators of adequacy of the BAL sample.
• However, it is considered that cell counts more than 2 X 10^6,
 or more than 10 macrophages per high power field are the markers
of adequacy of the sample.
• 2 The BAL sample is labeled unsatisfactory when it shows excessive blood,
degeneration and squamous cells.
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15. OTHER SAMPLING TECHNIQUES INCLUDE :
• TRANS BRONCHIAL NEEDLE ASPIRATION
• TRANS OESOPHAGEAL FINE NEEDLE ASPIRATION
• PERCUTANEOUS FINE NEEDLE ASPIRATION
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16. NORMAL CONSTITUENTS OF LUNG SAMPLES
 Lining cells
• Ciliated columnar cells
• Goblet cells
• Alveolar macrophages
• Squamous epithelial cells
 Associated cells
 • Polymorphs
 • Lymphocytes
 histiocytes

17.  Acellular material
 • Curschmann’s spirals
 • Ferruginous (asbestos) bodies
 • Charcot-Leyden crystals
 • Carbon particles
 • Hemosiderin
 Contaminants
 • Pollen grains
 • Vegetable materials
 • Contaminants from food particles
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18. NORMAL CYTOLOGY
• The normal cellular components of the respiratory tract are
• Ciliated columnar cells
• Goblet cells
• Basal cells
• Squamous cells
• Clara cells
• Neuroendocrine cells
• Type 1 and 2 pneumocytes
• Alveolar macrophages.
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19. Ciliated Columnar Cells
 the predominant lining cells of the lower respiratory tract.
• They are mainly noted in washing/brushing and BAL specimens.
• The cells are columnar in appearance with moderate cytoplasm. Nuclei are located on one side
of the cell.
• Cilia come out from the terminal plate of the columnar cells .
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20. Basal or reserve cells
• Present mostly in bronchi
• Short cells with little cytoplasm and oriented mostly towards the basement membrane.
• cytology –small round to cuboidal cells with thin rim of cytoplasm with high N/C ratio with dark
chromatin .
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21. Goblet Cells
• Goblet cells are present mainly in bronchial washing/brushing or BAL
samples
• The individual cells have a moderate amount of vacuolated
 cytoplasm filled with mucin .
• The nuclei are round and monomorphic.
• Goblet cells are seen in abundant numbers in
 chronic tracheobronchial irritation and asthma.
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22. Alveolar Macrophages
• The alveolar macrophages show moderate to abundant cytoplasm filled with phagocytosed
brownish to blackish dust particles .
• The nuclei a Centrally placed kidney-shaped nuclei.
• Bi- and multinucleation present.
• Fine chromatin.
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23. Squamous Epithelial Cells
 The stratified squamous cells may occur as an abnormal reaction replacing normal pseudo
stratified columnar epithelium.
 They are polygonal cells with abundant transparent cytoplasm with distinct cell borders and
small nucleus with granular chromatin .
.
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24. Other cells such as
• neuroendocrine cells
• clara cells
• type 1 alveolar pneumocytes
• are present in lower respiratory tract but not seen in cytological routine
preparations
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25.  NONCELLULAR COMPONENTS
 1)Mucus
 These are acellular homogenous substances. The largest amount
 of mucus is usually present in bronchial obstruction and
 bronchioloalveolar carcinoma.
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26.  2)Curschmann’s Spirals
 Curschmann’s Spirals are corkscrew-shaped long inspissated
 mucus cast produced in the small bronchioles due to excessive
 mucus production. seen in chronic bronchitis, asthma and in smokers.
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27.  3)Ferruginous Bodies
 The ferruginous (asbestos) bodies are approximately 50–200
 micron long rod-shaped golden-brown structures with two
 terminal bulbous tips.
 These are coated with protein and iron indicates asbestos exposure.
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28.  4)Charcot-Leyden Crystals
 These are bi pyramidal needle-shaped crystals with two pointed
 ends.
 Charcot-Leyden crystals are seen frequently in asthma patients,
 eosinophilic pneumonia and allergic bronchopulmonary
 Aspergillosis.
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29.  5)Psammoma Body
 These are darkly stained concentric lamellated structures
 made up of calcium. Psammoma bodies are usually seen in
 Bronchio alveolar carcinoma.
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30. Other noncellular components
include:
• Corpora amylacea-seen in chronic bronchitis heart failure emphysema
• Calcium oxalate crystals-seen in aspergillous infection
• amyloid-amylodosis
• Schaumann and asteroid bodies seen within multinucleated giant cells in
granuloma conditions.
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31. Abnormalities of
Bronchial Epithelial Cells
Ciliocytophoria
• The cells in the presence of cilia degenerate and may show
 only terminal plate.
• Infection particularly viral infection of the bronchial lining epithelial cells
may cause detachment of cilia from the cell. This condition is known as
ciliocytophoria.
• cytology smear shows detached fragment of cilia or cells with
 cytoplasm and nucleus without any cilia.
• There is no association of malignancy with ciliocytophoria
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32. Bronchial cell hyperplasia (Creola body)
 Commonly seen
 • Chronic bronchitis,asthma and TB
 Morphology
 • Large three-dimensional papillary clusters of the bronchial
 cells
 • Smooth peripheral outline
 • Focal areas of the surface cells show cilia
 • Outer cells in palisade arrangement
 • Cells are small with monomorphic nuclei
 • Fine nuclear chromatin
 • Occasional prominent nucleoli
 Differential diagnosis: Adenocarcinoma
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33. nuclear enlargement and pleomorphism in the reactive
bronchial
hyperplasia
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34. Reserve cell hyperplasia
 Causes: Infection, injury
 Morphology
 • Tight cohesive small clusters
 • Small cells
 • High N/C ratio
 • Scanty cytoplasm
 • Hyperchromatic nuclei
 • Nuclear molding
 D/D: Small cell carcinoma
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35. Goblet cell hyperplasia
• Occur in response to acute injury, chronic bronchitis, bronchiectasis, asthma
and chronic infections
• It shows hyper distended vacuole pushing the nucleus towards basement
membrane.
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36. Hyperplastic type II pneumocytes
 Causes: Tuberculosis, pulmonary infraction, radiation injury,
 chemotherapeutic drugs, pneumonia, interstitial lung disease
 Morphology
 • Tight papillary clusters
 • Abundant vacuolated cytoplasm
 • Enlarged nuclei
 • Fine chromatin
 • Prominent nucleoli
 D/D: Adenocarcinoma
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37. INFECTIONS
 Cytological examination of the respiratory samples play good
 role in the diagnosis of various infective processes.
 Bacterial Infections
 Large varieties of Gram-positive and Gram-negative organisms
 cause bacterial pneumonia. The common organisms causing
 bacterial pneumonia are Staphylococcus aureus, pneumococci,
 Streptococcus, Klebsiella pneumoniae, Pseudomonas, Haemophilus
 influenzae, etc.
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38. Mycobacterial infection
 • Cytology:
 – Multiple epithelioid cell granulomas
 – Multinucleated Langhans type giant cells
 – Necrosis
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39. Actinomycosis
• Actinomyces species are commonly present in tonsillar crypts
and gingival crevices .
• the organisms are cotton ball-like floppy hemtoxylin-stained tangled filamentous material with
associated squamous cells.
• Background of the smear showing polymophs.
• Nocardia
• They infect lung in the immunocompromised patient.
• Nocardia is slender, filamentous, branching organisms.
• They are Gram-positive and weak acid-fast positive organism.
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41. Fungal Infections
 Aspergillus: Septate, uniform acute angle branching, 30
 micron width hyphae, septate
 Special stain: PAS, mucicarmine, methanamine silver
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42.  • Zygomycetes (mucor): Broad, nonseptate, wide-angle branching
 Special stain: PAS, mucicarmine
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43.  • Cryptococcus: 5–10 μ diameter round, thick outer capsule,
 narrow-based budding
 Special stain: India ink
 Histoplasma: Small round (2–5 μ), narrow budding, usually
 inside the macrophages
 Special stain: PAS, Z-N stain
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44.  • Candida: Thin slender pesudohyphae with budding yeast
 Special stain: PAS
 Abbreviations: PAS—periodic acid-Schiff; Z-N—Ziehl–Neelsen
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45. VIRAL INFECTIONS
 Herpes Virus
• Shows many multinucleated giant cells.
• The individual cells show multiple enlarged basophilic nuclei with ground-glass appearance,
slaty gray homogenized chromatin and large intra nuclear eosinophilic inclusions.
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46. Cyto megalo virus
• Smears show markedly enlarged cells with enlarged nuclei.
• Large amphophilic intra nuclear inclusion with a peripheral clear halo
 and margination of chromatin on the inner nuclear membrane.
• Small satellite basophilic cytoplasmic inclusions are seen
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47. Pneuma cyctis cainii infection
 • Associated with immunocompromised patient, particularly
 in AIDS
 • Best sampling method: bronchioloalveolar lavage
 – Frothy appearance
 – Extracellular aggregate
 – Not stained by Papanicolaou’s staining
 – Cup-shaped structure of 6–8 μm diameter
 – One surface flat and a central dark zone
 • Special stain
 – Methanamine silver
 •
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48.  Adenovirus
• The cytology smears show cilliocytophoria affected cells show two types of inclusions.
• The first type shows small reddish well-circumscribed inclusion surrounded by clear halo and
other type shows large basophilic homogenous inclusion that completely fills the entire
nucleus.
• Measles Virus
• Cytology smear shows enormous
• multinucleated giant cells (Warthin-Finkelday cells) with more
 than 100 nuclei containing eosinophilic intracytoplasmic and intranuclear inclusions.
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49. WHO classification of lung
tumors
 Squamous cell carcinoma
 – Papillary
 – Clear
 – Small cell
 – Basaloid
 • Small cell carcinoma
 – Combined small cell carcinoma
 • Adenocarcinoma
 – Acinar
 – Papillary
 – Bronchioloalveolar
 – Solid adenocarcinoma
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50.  Large cell carcinoma
 – Large cell neuroendocrine carcinoma
 – Combined large cell neuroendocrine carcinoma
 – Basaloid carcinoma
 – Lymphoepithelioma-like carcinoma
 – Clear cell carcinoma
 – Large cell carcinoma with rhabdoid phenotype
 • Adenosquamous carcinoma
 • Sarcomatoid carcinoma
 • Carcinoid
 – Typical carcinoid
 – Atypical carcinoid
 • Salivary gland carcinoma
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51. Squamous cell carcinoma
 MCC lung cancer associated with smoking arises centrally from major or segmental bronchi.
 Polyhedral cells
 • Eosinophilic cytoplasm
 • Intracellular keratin (orangeophilic cell)
 • Round nucleus with moderate nuclear pleomorphism
 • Hyperchromatic nucleus, inconspicuous nucleoli
 • Fiber cells and tadpole cells
 • Ghost of squamous cells
 • Background necrosis and granular debris
 • Keratin pearls
 IHC: Positive for CK 5/6, CEA.
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53. oval to polyhedral cells with moderately pleomorphic
nuclei in squamous cell carcinoma of lung
(Papanicolaou’s stain X HP
Polyhedral cells with orangeophilic cytoplasm and
dark hyperchromatic nuclei in squamous cell
carcinoma of lung (Papanicolaou’s stain X HP)
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55.  Differential diagnosis
 1)Squamous metaplasia: Monomorphic small round nuclei
 with homogenous chromatin
 2) Reactive squamous atypia: Monomorphic with regular
 nuclear margin
 3) Small cell carcinoma: Hyperchromatic nuclei, absence of
 nucleoli, crushing artifact, molding
 4)Adenocarcinoma: Cytoplasmic vacuoles, fine nuclear
 chromatin, prominent nucleoli
 5) Metastatic squamous cell carcinoma: Clinical history

56. Adenocarcinoma
 Mcc in females located in peripheral part of lung. subtypes include acinar,
papillary, solid and broncho alveolar types.
 Discrete, cluster and glandular pattern
 • Sheets of cells with honeycomb pattern
 • Round cells with moderate vacuolated cytoplasm
 • Central to eccentric nucleus
 • Fine chromatin
 • Prominent nucleoli
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57. malignant cells withmoderate amount of cytoplasm and enlarged nuclei with prominent nucleoli
Discrete malignant cells
entangled in mucous in
adenocarcinoma of lung
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58. Small Cell Carcinoma
 20% of ca lung and centrally located. highly aggressive and metastasize easily.
 Dissociated cells
 • Small cells resembling lymphocytes
 • Scanty cytoplasm
 • Hyperchromatic nucleus with inconspicuous nucleoli
 • Nuclear molding
 • Paranuclear blue bodies
 • Crushing artifact and nuclear threading
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59. Malignant cells with scanty cytoplasm and hyperchromatic
nuclei in small cell carcinoma of lung (Papanicolaou’s stain
X HP)
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60.  Immunocytochemistry
 The small cell carcinoma is positive for CD56, chromogranin,
 synaptophysin, cytokeratin and TTF-1.
 Differential Diagnosis
 • Lymphomas: Small round dissociated cells with scanty
 cytoplasm may simulate non-Hodgkin’s lymphoma (NHL).
 The presence of lymphoglandular bodies is characteristic of
 NHL
 • Lymphocytes: Lymphocytes in chronic inflammation may
 be mistaken as small cell carcinoma. Lymphocytes are
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61.  usually dissociated and smaller than the cells of small cell
 carcinoma.
 • Poorly-differentiated carcinoma: Poorly differentiated SQC
 and adenocarcinoma with small round cells may be confused
 with small cell carcinoma. Cellular dissociation, scanty
 cytoplasm and nuclear molding favor the diagnosis of small
 cell carcinoma.
 • Carcinoid tumor: Discrete cells with relatively monomorphic
 nuclei may pose a diagnostic dilemma with carcinoid tumor.
 However, the presence of abundant granular cytoplasm, salt
 and pepper chromatin, and prominent nucleoli favor the
 diagnosis of carcinoid.
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62. Undifferentiated Large Cell
Carcinoma
 10% of ca lung mostly located in periphery.
 Large cell with marked nuclear pleomorphism
 • Moderate to abundant cytoplasm
 • Ill-defined cytoplasm
 • Severely pleomorphic bizarre nuclei
 • Multiple prominent nucleoli
 • Large tumor giant cell
 • Polymorphs sticking to the cells
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63. Discrete malignant cells with enlarged markedly
pleomorphic
nuclei along with many polymorphs
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64. Differential diagnosis of large cell carcinoma
(LCC)
 Sarcomas: Rare; immunostain—vimentin for sarcoma,
 cytokeratin (CK) for LCC
 • Amelanotic melanoma: HMB45 immunostaining.
 • Reactive changes: Regular nuclear margin and no
 significant chromatin abnormality
 • Squamous cell carcinoma: Orangeophilic cells, fiber cells
 and tadpole cells
 • Adenocarcinoma: Gland-like arrangement and background
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65. CARCINOIDS
 Dissociated cells
 • Rosettes
 • Monomorphic cells
 • Moderate to abundant cytoplasm with red granularity
 • Round monomorphic nuclei with salt and pepper
 Chromatin.Mitosis is uncommon
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66. REFERENCES
 1)ORELLS CYTOPATHOLOGY
 2)PRANAB DEY DIAGNOSTIC PATHOLOGY
 3)LIPPINCOAT SUDHA DD of exfoliative and aspiration cytology
 4)KOSS DIAGNOSTIC PATHOLOGY AND ITS HISTOPATHOLOGICAL BASES,5TH
EDITION,2006.
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