peripheral smear basics (part 1) - hemepathreviewhemepathreview.com/heme-review/part1-1-pb.pdf ·...
TRANSCRIPT
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Peripheral
Smear Basics
(Part 1)
Amy Sanchez, M.D.
11-16-2005
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Peripheral Smear
• Inexpensive, rapid, reliable
• Window into the functional status of
the bone marrow
– especially cytopenic states
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Vs. automated machines
• Able to weigh importance of observed findings and assess their importance within context of clinical data
• Machines tend to generalize a wide array of morphologic abnormalities
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Systematic approach
• Slide preparation
• Optimal area of review
• RBCs – Size
– Shape
– Pallor
– Premature forms
– Inclusions
• White cells – Lymphocytes
– Neutrophils
– Eosinophils
– Basophils
– Monocytes
– Blasts
– Organisms
• Platelets
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Slide preparation
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Optimal area of review
Ideal thickness
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RBCs
SIZE
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RBC Size
• Normal RBC should be the size of
a mature lymphocyte nucleus
• Measured by the MCV
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Elevated MCV = Macrocytosis
MCV > 100um3
• B12/Folate deficiency, aplastic anemia
• Autoimmune hemolytic anemia
• Liver and thyroid disease, alcoholism
• Cold agglutinin disease
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Decreased MCV = Microcytosis
MCV < 80um3
• Iron deficiency
• Thalassemias
• Anemia of chronic
disease
• Hemoglobinopathies
– C, E, S
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SHAPE
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Pathologic Red Blood Cells in
Peripheral Blood Smears
Type of Cell Underlying Change Disease States
Acanthocyte (spur cell) Altered cell membrane lipids Abetalipoproteinemia, liver
disease, postsplenectomy
Bite Cell (degmacyte) Heinz body pitting by spleen G6PD deficiency, drug-induced
oxidant hemolysis
Ovalocyte (elliptocyte) Abnormal cytoskeletal proteins Hereditary elliptocytosis
Rouleaux Circulating paraprotein Paraproteinemia
Schistocyte (helmet cell) Mechanical destruction in
microvasculature
TTP, HUS, prosthetic heart
valves
Spherocyte Decreased membrane
redundancy
Hereditary sphereocytosis,
immunohemolytic anemia
Stomatocyte Membrane defect with
abnormal cation perm
Hereditary stomatocytosis,
immunohemolytic anemia
Target Cell (codocyte) Increased redundancy of cell
membrane
Liver disease, thalassemia
postsplenectomy,HgbC dz, Fe
deficiency
Burr Cell (echinocyte) Altered membrane lipids Usually artifactual but maybe
uremia
Tear Drop Cell (dacrocyte) Myelofibrosis
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•Irregular, long, sharply pointed and bent
spicules
•Absence of central pallor
•Most commonly seen in liver disease
Acanthocytes (Spur cells)
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Sphingomyelin
Absence of apolipoprotein B results in inability to transport triglycerides in the blood
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McLeod phenotype/syndrome
• Absence of erythrocyte surface Kx
antigen (part of Kell antigen group)
• Acanthocytes with chronic but well
compensated hemolytic anemia
• Disease processes include muscular
dystrophy, cardiomyopathy, and
“neuroacanthosis”
• May be associated with chronic
granulomatous disease
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Tear Drop Cells (Dacrocytes)
Myelofibrosis or bone marrow infiltrate
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Bite Cells
• G6PD deficiency
G6PD deficiency
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STOMATOCYTES
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RBC with slit-like or
rectangular area of central
pallor, a mouth
Most often seen in liver
disease
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Burr Cells (Echinocytes)
Projections- smaller
more regular than
acanthocytes
Often Artifactual but
seen in UREMIA
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Spherocytes
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Hereditary spherocytosis
• Northern European ancestry
• Spectrin, ankyrin or band 3 or 4.1 deficiency
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• Defects in vertical stabilization of
the phospholipid bilayer of the
RBC membrane cause separation
of the spectrin - phospholipid
bilayer
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Hereditary spherocytosis
• Normal biconcave red cell loses membrane fragments and adopts a spherical shape
• Inflexible cells are trapped in the splenic cords, phagocytosed by macrophages
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Elliptocytes
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Hereditary elliptocytosis
• Autosomal dominant trait
• Spectrin abnormality or deficiency of protein 4.1
• Asymptomatic without anemia and usually with no splenomegaly and only mild hemolysis
• RBC hemolysis occurs in the spleen, thus splenectomy corrects the hemolysis, but not the RBC membrane defect.
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Target Cells
Characteristic of: – Liver disease
– Post-splenectomy
– Hemoglobinopathies
• Thalassemia
• Hb C, D and E
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Canoe cells (aka Taco
cells, folded cells)
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HgbSC disease with canoe and sickle cells
“Washington Monument crystals”
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Schistocytes
Microangiopathic
hemolytic anemia
DIC, TTP, HUS
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Rouleoux and Clumping
Rouleaux
RBC Clumping
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Quiz!
• 1. 60 yo woman with a history of treated
breast cancer now presents with anemia
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Most Likely Diagnosis?
• Tear drop cells 2o to bone marrow
infiltration with tumor
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2. 45 yo man with macrocytic anemia
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Most Likely Diagnosis?
• Liver disease
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PALLOR
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Hypochromic anemia
MCH < 27 pg
• Disorders of globin synthesis – Thalassemic syndromes
• α-Thalassemia
• Β-Thalassemia
• Disorders of heme synthesis – Sideroblastic anemias
• Hereditary (X-linked auto. Dominant)
• Acquired idiopathic
• Acquired toxic
• Disorders of Fe metabolism – Fe deficiency
– Chronic disease
– Neoplasia
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Iron Deficiency
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Iron deficiency s/p
transfusion
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PREMATURE
RBCs
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Reticulocytes
• Decreased cell survival – Blood loss
– Autoimmune HA
– Nonimmune HA
• TTP, HUS, DIC
• Spherocytes
• G6PD
• PNH
• Hemoglobinopathy
• Thalassemia
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Nucleated RBCs
• Not normally present in
adult patients PS
• Present in:
– Severe hemolysis
– Profound stress or
hypoxemia
– Myelophthisic condition
– Leukoerythroblastic
smear
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INCLUSIONS
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Basophilic Stippling
Precipitated ribosomes (RNA)
Fine – variety of anemias
Siderblastic, sickle cell,
megaloblastic
Coarse – Pb intoxication,
thalassemia
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Heinz Bodies
• Precipitated denatured Hgb
• Seen in G6PD deficiency
• Seen with supravital staining – Crystal violet
– Brilliant cresyl blue
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Howell Jolly Bodies
Dense,usually
single
Nuclear remnant
Seen in:
- Postsplenectomy
- Hemolytic anemia
- Megaloblastic
anemia
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Pappenheimer bodies
Small, dense basophilic granules
Fe-containing mitochondrial remnant or sidersome
Seen in:
- Sideroblastic anemia
(Hereditary, idiopathic or secondary)
- Post-splenectomy
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LEUKOCYTES
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WBC
• Neutrophil
• Eosinophil
• Basophil
• Lymphocyte
• Monocyte
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Polymorhponuclear Neutrophils
(PMNs)
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PMNs
• Normal range ANC: 1.5-7.0 103/mm3
• Reflects only the CIRCULATING PMNs
• Does not include marginated PMNs or
stored PMNs in the bone marrow
• Mechanisms for neutrophilia:
– Demargination
– Release of bone marrow component
– Increased production
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Mechanisms Causing
Nonneoplastic Neutrophilia
Mechanism Time Course Causes
Demargination Minutes Epinephrine release,
acute stress, exercise
Mobilization of
bone marrow
Hours Corticosteroids,
infections,
inflammation
Increased
production
Days Sustained infection,
chronic inflammation,
CSF-producing
tumors, CSF therapy
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Distinguishing Between Reactive
Changes and Neoplasia
Parameter Reactive Neoplastic
WBC <30 x 103/mm3 >50 x 103/mm3
Toxic Neutrophils Present Absent*
Left Shift Includes myelocytes Includes blasts
Basophilia Absent Present
Platelet count Variable, decreased in
sepsis
Increased
Platelet Morphology Unremarkable Abnormal
Nucleated RBC Absent Present
*Except in patients with infection
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Reactive Neutrophil
3 Features:
Toxic granulations
Dohle bodies
Cytoplasmic vaculoes
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Immature Granulocyte
Suggesting Neoplasia
Myeloblast
Promyelocyte
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The Malignant Mimicker:
Leukemoid Reaction
• All precursor granulocytes in the PBS
• WBC in the range up to 100K
• Response to severe stress or infection
• Other signs of malignancy not present
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Neutrophil Disorders with
Abnormal Morphology
• Pelger-Huet anomaly – Bilobed or nonsegmented nucleus
– asymptomatic
• May-Hegglin anomaly – Cytoplasmic inclusions resembling Dohle bodies
– Many asymptomatic
• Chediak-Higashi syndrome – Giant cytoplasmic granules in all granulocytes
– Immunodeficiency
• Hypersegmentation – B12/Folate deficiency, myelodysplasia, myeloid
leukemia, chemotherapy, or renal failure
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Pelger-Huet Anomaly
• Inherited, AD or
acquired
• Acquired =
“pseudo” Pelger-
Huet as in MDS
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Chediak-Higashi Syndrome
• Autosomal
recessive
• Giant granules
• Severe
immunodeficiency
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Hypersegmentation
• Normal to have 4-5 lobes
• Seen most commonly in B12/folate def
• Uremia
• Chemotherapy
• Also seen in MDS and other myeloid neoplasms
• Can be inherited
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Eosinophilia
• Allergic/hypersensitivity reactions
• Drug allergies
• Parasitic infections
• Connective tissue/collagen vascular disease
• Neoplasms
– T-cell lymphoma
– Hodgkin lymphoma
• Sarcoidosis
• Hypereosinophilic syndrome/Chronic eosinophilic leukemia
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Basophilia
• Much more common in
malignancies like CML
vs. reactive
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Reactive Lymphocytosis
• Diseases with nonreactive
morphology
– Infectious lymphocytosis (Whooping
cough)
– Transient stress lymphocytosis
• Diseases with reactive morphology
– EBV, IM, CMV, Toxo, adenovirus, HHV-
6, viral hepatitis
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Atypical/reactive lymphocytes
Plasmacytoid
lymphocyte
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Features of Leukemias/Lymphomas
• T-cell large granular lymphocyte
leukemia/NK Cell Leukemia
• Blasts with and without Auer rods
• Hairy cells
• Cleaved cells
• Smudge cells
• “Clover cells”
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T-cell LGL Leukemia/NK
Cell Leukemia
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Small Cleaved Cells
Follicular lymphoma
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Smudge Cells
CLL
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Hairy Cell Leukemia
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Blasts with Auer Rods
(Myeloblasts)
AML
Auer Rod=
Fused
lysosomal
granules
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Blasts without Auer Rods
Lymphoblasts (ALL) vs. Atypical lymphs
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Clover Cells
Adult T-Cell Leukemia/Lymphoma
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ORGANISMS
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Ehrlichiosis
• Found in the SE and S. Central
US
• Transmitted by ticks
Found in the SE and S. Central US
Transmitted by ticks
Rickettsial organism
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Histoplasma
<>
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Babesiosis
• Protozoa
• Endemic in the NE US
• Transmitted by the Ixodidae tick
• Similar to Malaria
• Tetrad form is pathonogmonic
• Risk Factors: – Post-splenectomy
– Immunocompromised
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Malaria
• Ringed stage
(trophozoite)
• Can see other
stages within
RBCs
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PLATELETS
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Estimate platelet count
on PBS
• 100x oil immersion
• Minimum of 5 fields
• Average # platelets, then X by
20,000
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Increased Normal Decreased
Over 20
platelets per
HPF
> 400,000
7 – 20
platelets per
HPF
Under 7
platelets per
HPF
< 140,000
Platelet Count
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Giant Platelets
• Size of an RBC
• Usually indicates a hyperreactive bone marrow 2o to underlying condition – ITP, TTP, DIC
• Can be inherited in the form of Bernard-Soulier syndrome, platelet dysfunction
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Platelet Clumping and
Satellitelosis
• Causes artificially low platelet
counts
• 2o EDTA used in collection tubes
Solution is to use
sodium citrate instead
of EDTA
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Quiz!
• 75 yo asymptomatic man with
WBC 60,000
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Diagnosis?
• CLL (chronic lymphocytic leukemia)
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• 35 yo man with WBC 33,000 and thrombocytopenia
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Diagnosis
• Reactive neutrophilia, sepsis
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Conclusion
• Systematic approach to reviewing a peripheral blood smear
• All cell lines are evaluated
• MUST take into account the clinical history
• Integrate the information to make a differential diagnosis
• Most peripheral smears are non-specific and have a constellation of findings
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White Blood Cell
Morphologic
Abnormalities Related
to Hereditary Disorders
(Part 2)
CP Talk
Jacqueline Nguyen, DO
April 2, 2007 8am
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White Blood Cells
• Qualitative
abnormalities
– Hereditary disorders
• Morphologically
abnormal
– Neutrophil inclusions
– Abnormal neutrophil
nuclei
– Macrophage/histiocytic
abnormalities
http://www.mhhe.com/biosci/ap/histology_mh/wbccomp.jpg
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Neutrophil Inclusions
• Neutrophil inclusions-inherited
– Alder Reilley anomaly
– May Hegglin anomaly
– Chediak-Higashi Syndrome
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Alder Reilley Anomaly
• Resemble the large
primary granules of
promyelocytes
• Large, purple to purplish-
black, coarse azurophilic
granules
• No impaired PMN function
• Inclusion is a
mucopolysaccharide
(PAS+)
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Alder Reilley Anomaly
• Vacuolated/abnor
mally granulated
lymphocytes in
some case
• Eosinophils and
basophils contain
large basophilic
granules
www.academic.marist.edu/.../HematologyI/7-24.jpg
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Alder Reilley Anomaly
• Autosomal recessive
• Associated with
several different types
of genetic
mucopolysaccharide
disorders (Hurler,
Hunter, San Fillipo,
Maroteaux-Lamy, but
not Moriquo) www.academic.marist.edu/.../tn_bloodsmears28.jpg
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Alder Reilley Anomaly
• Not specific for one of the
mucopolysaccharidoses
• First discovered in
Hurler's syndrome
• May be seen following
bone marrow transplants
and chemotherapy
www.med-ed.virginia.edu/.../wcd/qualitative.cfm
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May Hegglin Anomaly
• Large blue cytoplasmic inclusions
resembling giant Döhle bodies
hsc.unm.edu/Pathology/MedLab/images/mhegglin.jpg
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May Hegglin Anomaly
• Thrombocytopenia
• Enlarged platelets
• Variable neutropenia
• Inclusions also seen in
eosinophils, basophils,
and monocytes
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May Hegglin Anomaly
• Autosomal dominant
• Many patients are
asymptomatic
• Non-muscle myosin
heavy chain A (MYH9)
mutation
• No impairment on PMN
function www.bekkoame.ne.jp/.../WBC/photo/MayHeggrin3.jpg
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Chédiak-Higashi
Syndrome
• Large, well-defined,
round to irregular, blue
to green-gray
cytoplasmic granules
(MPO+)
• All granulated cells and
even
lymphocytes/natural
killer cells affected hsc.unm.edu/pathology/MedLab/images/chediak.jpg
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Chédiak-Higashi Syndrome
• Autosomal
recessive
• Cytopenias
• Platelet and NK-
cell dysfunction
www.pathology.ucla.edu/.../case6/image6.gif
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Chédiak-Higashi Syndrome
• Affects many granule-
containing cells
– Melanosomes-partial
occulocutaneous
albinism
– Neurons- neurological
abnormalities
– Impaired PMN function-
severe, recurrent
pyogenic infections
(decrease killing and
chemotaxis) http://www.medscape.com/content/2003/00/46
/65/466530/art-adnc466530.fig8.jpg
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Chédiak-Higashi
Syndrome
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Quiz
• Match the picture:
– May Hegglin?
– Chediak-Higashi?
– Alder Reilley?
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Answer
May Hegglin
Chediak-Higashi
Alder Reilley
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Abnormal Neutrophil
Nuclei
• Pelger-Huët anomaly
• Hereditary hypersegmentation of
neutrophils
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Pelger-Huët Anomaly
• Bilobed (pince nez)
or non-segmented
neutrophil nuclei
seen in most PMNs
• Coarse clumping of
the nuclear
chromatin in
neutrophils,
lymphocytes, and
monocytes http://www.med.unc.edu/medicine/web
/Smearreview/img034.jpg
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Pelger-Huët Anomaly
• Autosomal dominant
• No other lineage
abnormalities
• No functional
abnormalities
• Mutations in lamin ß-
receptor (LBR gene on
Chrom 1)
http://www.bphealthcare.com/healthcare
/galleries/haem/case5-a1.jpg
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Pelger-Huët Anomaly
• Heterozygous in good
health, and their natural
resistance to infection is
unimpaired
• Homozygous PHA is
associated with skeletal
anomalies,
developmental delay,
and seizures
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Hereditary
Hypersegmentation
• AKA Undritz anomaly
• More than 3 cells
having 5 lobes or a
single cell with 6 lobes
found in the course of a
100 cell differential (or
5% with 5 lobes) http://www.med-
ed.virginia.edu/courses/path/
innes/images/wcdjpeg/wcd%
20hyperseg.jpeg
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Hereditary
Hypersegmentation
• Autosomal
dominant
• No other
abnormalities
• No associated
findings
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Histiocyte/Macrophage
Abnormalities (Bone
Marrow Cells)
• Inherited abnormalities
– Gaucher cell
– Niemann Pick cell
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Gaucher Cell
• Crumpled tissue-paper cytoplasm
• Caused by enlarged, elongated lysosomes filled with glucocerebroside
• Accumulation in BM, liver, spleen, and lungs leads to pancytopenia, hepatosplenomegaly, and pulmonary disease
• Infiltration in BM causes thinning of the cortex, pathologic fractures, bone pain, bony infarcts, and osteopenia
http://pathcuric1.swmed.edu/PathDe
mo/gen1/gen130.jpg
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Gaucher Cell
• Seen in
– Gaucher’s
disease
– Myeloproliferative
syndromes
(CML)= “pseudo-
Gaucher” cells
http://www.sfu.ca/biology/faculty/
kermode/laboratory/gaucher-
cells.jpg
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Gaucher Disease
pathology.catholic.ac.kr/.../genetic/ge13.jpg
The enlarged spleen shows effacement of white pulp and massive expansion of pale red pulp.
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Gaucher Cell
• Acid phosphatase +
• PAS+
• Labs
– ↓ in WBC acid ß-
glucosidase
– ↑ in serum acid
phosphaatase
– ↑ in serum ACE
http://www.neuropathologyweb.org/cha
pter10/images10/10-GCl.jpg
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Gaucher Cell
• Enzyme
replacement
therapy now
available
(imiglucerase
[Cerezyme]) http://www.academic.marist.edu/~jzmz/
HematologyI/MicroexamBM25.jpg
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Niemann Pick Cell
• Foamy, vacuolated cytoplasm
• Accumulation of
sphingomyelin
• Seen in Niemann-Pick
disease (sphingomyelinase
deficiency), Wolman disease,
cholesterol ester storage
disease, lipoprotein lipase
deficiency, and, GM1
gangliosidosis type 2 http://www.thecrookstoncollection.
com/Collection/medslides/Slides/
Niemann-pick-cell.jpg
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Niemann Pick Cell
• Weakly PAS+
• Birefringence on
polarized light
• Yellow-green on
UV
http://pathology.catholic.ac.kr/pat
hology/specimen/genetic/ge13.jpg
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Niemann Pick Disease
• Systemic involvement
– Progressive lung disease
– Hepatosplenomegaly
– Short stature
– Pancytopenia
• Lab findings
– ↓ WBC sphingomyelinase
– May see vacuolation of PB lymphocytes and monocytes
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Niemann Pick Disease
• No specific treatment available
• Generally more rapid clinical
course than Gaucher’s disease
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Quiz
• Name that cell
– Gaucher cell?
– Niemann Pick
cell?
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Answer:
Gaucher cell
Niemann Pick cell
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References
• Kjeldsberg, Practical Diagnosis of
Hematologic Disorders
• Osler Notes
• Internet