Cytopathology of Cytopathology of Cerebrospinal FluidCerebrospinal Fluid

GeneviGenevièève Warner ve Warner LearmonthLearmonth

Histopathologist /Cytopathologist.Histopathologist /Cytopathologist.Royal Free Hospital, LondonRoyal Free Hospital, London

andandGroote Schuur Hospital, Groote Schuur Hospital, University of Cape TownUniversity of Cape Town

Diagnostic CytopathologyDiagnostic Cytopathology• Cytopathology is a Cytopathology is a

complementary complementary diagnostic tool.diagnostic tool.

• Correlation with Correlation with other other other other diagnostic modalities:diagnostic modalities:

1. Clinical information: 1. Clinical information: Age, gender, Age, gender, previous illness, previous illness, check database check database results.results.

2. Ultrasound, MRI, 2. Ultrasound, MRI, CTscan.CTscan.

3. Communicate with 3. Communicate with the clinical team.the clinical team.

Cytopathology Cytopathology LaboratoryLaboratory

• Communicate with Communicate with Laboratory Staff Laboratory Staff

• Check laboratory Check laboratory facilitiesfacilities

• Discuss difficultiesDiscuss difficulties

• Availability of Availability of special stainsspecial stains

• Residual specimen Residual specimen for Flow for Flow

CytometryCytometry

What is Cerebrospinal What is Cerebrospinal Fluid? Fluid?

Where does it come from? Where does it come from? Where does it go?Where does it go?• THE NORMAL CSF obtained at lumbar puncture:THE NORMAL CSF obtained at lumbar puncture:

• The cerebrospinal fluid The cerebrospinal fluid (CSF)(CSF) is clear fluid, produced from is clear fluid, produced from arterial blood by the choroid plexuses of the lateral and fourth arterial blood by the choroid plexuses of the lateral and fourth ventricles by a combined process of diffusion, active transfer. ventricles by a combined process of diffusion, active transfer.

• The The choroid plexuschoroid plexus consists of tufts of capillaries with thin consists of tufts of capillaries with thin fenestrated endothelial cells. fenestrated endothelial cells.

• VOLUME: 140 ml. The volume of the ventricles is about 25 ml. VOLUME: 140 ml. The volume of the ventricles is about 25 ml. CSF is produced at a rate of 0.2 - 0.7 ml per minute or 600-700 CSF is produced at a rate of 0.2 - 0.7 ml per minute or 600-700 ml per day. ml per day.

• CirculationCirculation of CSF is aided by the pulsations of the CSF acts as of CSF is aided by the pulsations of the CSF acts as a cushion that protects the brain from shocks and supports the a cushion that protects the brain from shocks and supports the venous sinuses.venous sinuses.

• ChemistryChemistry :CSF from the lumbar region contains :CSF from the lumbar region contains 15 to 45 15 to 45 mg/dl protein mg/dl protein and and 50-80 mg/dl glucose50-80 mg/dl glucose (two-thirds of blood (two-thirds of blood glucose). glucose).

• Cellularity:Cellularity: Normal CSF contains Normal CSF contains 0-5 mononuclear cells0-5 mononuclear cells. . • The CSF pressure: The CSF pressure: lumbar puncture (LP), 8-15 mm Hg with the lumbar puncture (LP), 8-15 mm Hg with the

patient lying on the side, twice that value with the patient sitting patient lying on the side, twice that value with the patient sitting up.up.

Safety procedures Safety procedures Infective specimensInfective specimens

• Tuberculosis Tuberculosis • HIVHIV• HepatitisHepatitis• Septic Septic

meningitismeningitis• Viral Viral

meningitismeningitis

Processing Cerebrospinal Fluid Processing Cerebrospinal Fluid Specimen --- Cytospin Specimen --- Cytospin

TechniqueTechniqueIs there enough fluid for Is there enough fluid for processing?processing?2-10ml usually received2-10ml usually receivedColour of fluid ? Colour of fluid ? /clear/cloudy/bloodstained /clear/cloudy/bloodstained Xanthochromia”( blonde Xanthochromia”( blonde colour) seen in SAHcolour) seen in SAHConsistency? (Mucoid)Consistency? (Mucoid)

Processing Cerebrospinal fluidProcessing Cerebrospinal fluidCytospin methodCytospin method

Staining the cellsStaining the cells

May Grunwald May Grunwald GiemsaGiemsa ( ( MGG)MGG) air dried air dried

Papanicolaou stain Papanicolaou stain alcohol fixed alcohol fixed

Check the origin of “CSF “ Check the origin of “CSF “ specimenspecimen

• Is it really lumbar puncture CSF?Is it really lumbar puncture CSF?

• Is it from an ventricular drain ?Is it from an ventricular drain ? ( hydrocephalic patient)( hydrocephalic patient)

Is it from a cystic brain lesion?Is it from a cystic brain lesion?

• Is it an intra –operative fluid specimen?Is it an intra –operative fluid specimen?• Is it a smear from an intra operative Is it a smear from an intra operative

procedure? procedure?

MicroscopyMicroscopy• Cytopathologist Cytopathologist

needs the needs the best best quality microscope.quality microscope.

• No oil immersion No oil immersion used!used!

• Examine at low Examine at low power first. x40, power first. x40, x100, x400x100, x400

• Check the Check the backgroundbackground

• Before issuing report Before issuing report Check all clinical Check all clinical details again.details again.

“ “ Normal CSF” and Normal CSF” and “Abnormal CSF”“Abnormal CSF”

• NORMALNORMAL• Acellular ---- check Acellular ---- check

that this is a that this is a clearclear fluid specimenfluid specimen

• ?Problems with ?Problems with preparationpreparation

• Scanty cellsScanty cells

Abnormal CSFAbnormal CSF• ABNORMALABNORMAL• Very cellularVery cellular• Mixed Inflammatory Mixed Inflammatory

exudateexudate• Lymphocytic cellsLymphocytic cells• Clusters of cellsClusters of cells• Abnormal cellsAbnormal cells• BackgroundBackground• BacteriaBacteria• Pigment Pigment • FungiFungi• ““mucoid” materialmucoid” material• ContaminantsContaminants

Background -- very very Background -- very very importantimportant

Children on a safe Children on a safe happy holiday in happy holiday in NamibiaNamibia

Bloodstained fluid –intact erythrocytes

–

Specimens from Neurology/ Specimens from Neurology/ Neurosurgery / HIV/AIDS Neurosurgery / HIV/AIDS

Clinic, A&EClinic, A&E• Neurology:Neurology: often non specific symptoms and often non specific symptoms and

signs ---- MS, ME, sarcoidosis, Parkinsons, signs ---- MS, ME, sarcoidosis, Parkinsons, Alzheimers.Alzheimers.

• Lymphoid cells, plasma cells, macrophagesLymphoid cells, plasma cells, macrophages• NeurosurgeryNeurosurgery---- surgery for subarachnoid ---- surgery for subarachnoid

haemorrhage, tumours of brain tissue, haemorrhage, tumours of brain tissue, pituitary tissue, cystic lesions of brain, pituitary tissue, cystic lesions of brain, metastatic neoplasms.metastatic neoplasms.

• Macrophages, Neoplastic cells, “foreign cells, Macrophages, Neoplastic cells, “foreign cells, • HIV/AIDS HIV/AIDS --- Cerebral deterioration, ? --- Cerebral deterioration, ?

lymphoma ? Tuberculosis ? Fungal infectionlymphoma ? Tuberculosis ? Fungal infection• A & EA & E ---- ---- acute meningitis, meningococcal, acute meningitis, meningococcal,

streptococcal, streptococcal, “stroke”, SAH“stroke”, SAH

Acute Inflammation Acute Inflammation MeningitisMeningitis

Lymphoid CellsLymphoid Cells

Mixed Inflammatory Mixed Inflammatory InfiltrateInfiltrate

Camouflage ! Camouflage !

What are these small round What are these small round structures?structures?

Mixed Plasma cells and Mixed Plasma cells and Lymphocytes and small round Lymphocytes and small round

structures with concentric ringsstructures with concentric rings

Special stain – Special stain – mucicarmine.mucicarmine.

( PAS for glycogen stains cells and talc granules too)( PAS for glycogen stains cells and talc granules too)

Sputum from same patient Sputum from same patient when reviewed showed when reviewed showed

CryptococciCryptococci

This patient was a fit healthy man.This patient was a fit healthy man.Admitted with severe dyspnoea.Admitted with severe dyspnoea.

Sputum: ?abnormal cells, TB bacilli, Sputum: ?abnormal cells, TB bacilli, Sputum NAD Sputum NAD

TB culture negative.TB culture negative.Patient developed severe headachesPatient developed severe headaches

? Miliary Tuberculosis? Miliary TuberculosisCSF showed cryptococci.CSF showed cryptococci. Hobby: exploring caves. Hobby: exploring caves.

Cryptococcal infection is usually an Cryptococcal infection is usually an opportunistic organism in patients with opportunistic organism in patients with

HIV/AIDS or immunosuppressionHIV/AIDS or immunosuppression..

Tuberculosis in CSFTuberculosis in CSF

• Cytological appearances are Cytological appearances are notnot specific, specific, maybe lymphocytic or mixed maybe lymphocytic or mixed

inflammatory cellsinflammatory cells

• Maybe cloudy fluid, with necrotic Maybe cloudy fluid, with necrotic backgroundbackground

• Area of endemic TB/ “at risk” personArea of endemic TB/ “at risk” person• Clinical SuspicionClinical Suspicion• HIV/AIDSHIV/AIDS• ImmunosuppressionImmunosuppression• History of previous/current TBHistory of previous/current TB

Tuberculosis and HIV/AIDSTuberculosis and HIV/AIDSThe “Terrible Twins”The “Terrible Twins”

•Ziehl Neehlsen stain for acid fast Ziehl Neehlsen stain for acid fast TB bacilliTB bacilli

Examine Papanicolaou stained Examine Papanicolaou stained slide under Fluorescent light at slide under Fluorescent light at

450 450 μμmmTB bacilli are fluorescent !TB bacilli are fluorescent !

““CSF” received from a five CSF” received from a five year old sheep farmer’s year old sheep farmer’s

daughter daughter

• CSF was actually CSF was actually from the orbit !from the orbit !

• Refractile hooklets Refractile hooklets from Echinococcus from Echinococcus granulosus granulosus

• Hydatid disease Hydatid disease spread by the spread by the black - backed black - backed jackal to sheep jackal to sheep and sheep dogs.and sheep dogs.

Hydatid disease presents as a cyst in Hydatid disease presents as a cyst in any tissue of the body. Clear fluid is any tissue of the body. Clear fluid is aspirated . Hooklets seen. Sometines aspirated . Hooklets seen. Sometines scolices and the laminated membranescolices and the laminated membrane

Hydatid Disease Hydatid Disease ( Echinococcus granulosus) ( Echinococcus granulosus)

Histology of Cyst wallHistology of Cyst wall

Abnormal Cells in CSFAbnormal Cells in CSF

• Lymphoma Lymphoma /Leukaemia/Leukaemia

• Metastatic tumours Metastatic tumours • Childhood tumoursChildhood tumours• Primary Primary Intracranial Intracranial

“Tumours” /Cysts“Tumours” /Cysts• Primary Primary IntracerebralIntracerebral

TumoursTumours• Patients are often Patients are often

in Oncology Clinicin Oncology Clinic

Lymphoma/ Leukaemia Lymphoma/ Leukaemia

AIDS patients develop cerebral and other extranodal B-cell AIDS patients develop cerebral and other extranodal B-cell

lymphomas due to loss of the surveillance function of T-cellslymphomas due to loss of the surveillance function of T-cells. .

Pap

Metastatic Metastatic Malignant Malignant MelanomaMelanoma

with Macronucleoli --- “Big Mac”with Macronucleoli --- “Big Mac”

Cell types: Small “lymphoid”, Binucleate, Multinucleate, Balloon. **Prominent macronucleoli ( L.Koss)

Metastatic neoplasmsMetastatic neoplasmsBreast, lung, ovary, testis Breast, lung, ovary, testis

Metastatic Metastatic RetinoblastomaRetinoblastoma

MedulloblastomaMedulloblastoma

Retinoblastoma and ?Retinoblastoma and ?pinealoblastomapinealoblastoma

Craniopharyngioma CystCraniopharyngioma CystOr ? squamous carcinomaOr ? squamous carcinoma

Capillary Capillary HaemangionblastomaHaemangionblastoma

Intraoperative smearsIntraoperative smears

Primary Brain TumoursPrimary Brain TumoursIntra operative smearsIntra operative smears

Pitfalls and Mimicry in Pitfalls and Mimicry in CytologyCytology

• Oogpister or Oogpister or “Eye pisser” in “Eye pisser” in the Namib desert the Namib desert mimics a lizardmimics a lizard

Pollen grain which can be mistaken for parasitic remnants

Report on Report on whatwhat you see, you see, not what you expect to see.not what you expect to see.

Scan, look for the “odd man Scan, look for the “odd man out”.out”.

Be aware of odd/ unusual Be aware of odd/ unusual appearances, Avoid appearances, Avoid

complacency !complacency !

This is a giraffe in Etosha, Namibia

White bone in mouth !

Giraffes are herbivorous!

Lack of calcium in leaves of local acacia trees.Giraffes suck bones to avoid osteoporosis !!

Look carefully!

Remember !Remember !There is more to Cytology than meets There is more to Cytology than meets

the Eyethe Eye..

So keep an eye out !So keep an eye out !

Further ReadingFurther Reading

1.1. J. Bell, J.Clin Pathol, 1994;J. Bell, J.Clin Pathol, 1994;4747:573-:573-378 378

Update on Central nervous Update on Central nervous System. Cerebrospinal Fluid.System. Cerebrospinal Fluid.

2.2. Website : Neuropathology, Dimitri Website : Neuropathology, Dimitri P. Agamonolis, Akron Childrens’ P. Agamonolis, Akron Childrens’ Hospital, N.E Ohio, University Hospital, N.E Ohio, University Colleges of Medicine. (Colleges of Medicine. (excellent images, excellent images, videos, discussions).videos, discussions).

DedicationDedication

• This presentation is dedicated to the This presentation is dedicated to the memory of memory of

• The late Mr Patrick Carey , The late Mr Patrick Carey , Neurosurgeon, RCSI, Dublin. Neurosurgeon, RCSI, Dublin.

andand• The late Professor P.J. Bofin, The late Professor P.J. Bofin,

Neuropathologist, RCSI, Dublin.Neuropathologist, RCSI, Dublin.