Systematic Approach in Anemia Evaluation and Review

of Peripheral Smears

Jun W. Kim, MDFamily Medicine Residency

Dewitt Army Community Hospital

Objective

Recognize abnormal peripheral blood smear

Review differentials through systematic approach

Approach to Dx

Hx- age, duration, onset, subjacent illness, blood loss (GI, menstruation, surgery…), diet, medications, toxic exposure, occupation, Family Hx, Social Hx

PE- complete exam including skin (jaundice, petechiae), HEENT, Abdomen (hepatosplenomegally), lymphatics, rectal, and pelvic

Basic Labs to Start

Repeat CBC w/ manual differential(WBC, RBC, HCT, HGB, PLT, indices- MCH,

MCHC, MCV, RDW)Peripheral SmearReticulocyte count

Reticulocyte count Retic count = % immature RBC Normal 0.5-1.5% (for non-anemic) <1% Inadequate production >=1% Increased production (? adequacy)

Reticulocyte Correction

%Retic count frequently overestimates Retic count should be compared to non-

anemic RBC count to assess adequacy of response

Corrected Retic count = %Retic X HCT/45

Reticulocyte Production Index

Correction for left shift – Retic lifespan is increased in blood

RPI = % Retic X Hct/45 X 1/CF Hct Correction factor (CF) 40-45 1.0 35-39 1.5 25-34 2.0 15-24 2.5 Normal RPI = 1 (for non-anemic pt) RPI < 2 : hypoproliferative RPI >=2 : hyperproliferative

Retic Production Index

Hypoproliferative

- Iron def. anemia

- B12/folate def.

- Chronic disease

- Sideroblastic anemia

- Aplastic anemia

- Myeloproliferative

Hyperproliferative

- Hemolytic disease- Hemoglobinopathy

(including thalassemia)

Peripheral smear

Optimal area for reviewRBC morphology, WBC differential, PLT

(clumping?)

RBC morphology

7-9 m with 1/3 central palor Lifespan of 110-120 days About the size of nucleus of normal

lymphocyte Poikilocytosis & Anisocytosis

Basophilic stippling

Precipitated RNA lead or heavy

metal poisoning ETOH abuse Hemolytic anemia

Burr cells

Altered lipid in cell membrane

artifact Uremia Renal failure gastric CA transfused old blood

Elliptocytes/ovalocytes

Abnormal cytoskeletal proteins

Hereditary elliptocytosis

Howell Jolly body

Nuclear remnant - DNA

hemolytic anemia absent or

hypofunction spleen

Schistocyte/helmet cells

Fragmented (mechanical or phagocytosis)

DIC TTP HUS Vasculitis prosthetic heart valve severe burns

Sickle cells

Molecular aggregation of Hgb-S

SS, SC, S-thal rarely S-trait

NRBC

Common in newborn severe degree of

hemolysis

Spherocyte

Absent central palor

look smaller Hereditary

spherocytosis immune hemolytic

anemia

Stomatocyte

Mouth like Membrane defect Smear artifact Hereditary

stomatocytosis Liver disease

Target cells

Increased redundancy of membrane

hemoglobinopathies thalassemia liver disease

Tear drop cells

Distorted drop shaped Smear artifact myelofibrosis promyeloblastic

leukemia space occupying

lesions of marrow

Differentials

H&PIndices (MCV, MCHC, RDW)RBC MorphologyRetic responseOther labs as needed

MCV/smear

Micro Normo Macro

Iron panel Retic

Low

Retic

High

High

Low

Iron/B12/Folate

*Occult Blood Loss

Yes

No

Coombs (+)

Coombs (-)

Go to *Occult Blood Loss

B12/Folate

B12 Low

Folate/Low

Normal

MMA/Homocysteine

MMA high – B12 Low

Homocysteine high – Folate Low

Normal – Go to **

**Normal

Bone Marrow Bx

Anemia of Chronic Dis.

Anemia Differential Dx by Flow Chart

First use size (MCV) to sort the Differential Dx

MCV

Micro Normo Macro

Microcytic anemia

Get Iron panel- serum iron, TIBC, ferritin

Iron def. anemia

dec inc dec

Siderobla -stic anemia

inc dec inc

Thalasse mia -mia

inc/nl dec/nl inc/nl

Chronic disease

dec dec inc

Iron def. Anemia

Low Retic count High RDW Due to chronic blood

loss Diet deficiency

Thalassemia Normal to inc. RPI Normal RDW Target cells Mentzer index <13

=MCV/RBC Youden’s index - using

RDW & Mentzer index - sensitivity = 82%

- specificity = 80%

confirm w/ Hgb electrophoresis

Thalassemia continues

Alpha-thalassemia SE Asia & Africa aaaa - normal aaaa^ - silent carrier aaa^a^ - trait (mild) aa^a^a^ - HbH (Bart)

hemolytic disease a^a^a^a^ - hydrops

fetalis (stillborn)

Beta-thalassemia Mediterranean Beta-thal minor

one beta gene,

increased HbA2/HbF Beta-thal major

2 beta genes,

severe, failure to thrive, sig HbF

Sideroblastic anemia

Accumulation of mitochondrial iron in erythroblasts     

Hereditary Drugs - INH, lead,

zinc, alcohol, chloramphenicol, cycloserine, plavix

Hypothermia Confirm w/ BM Bx

Sample question #1

Anemia of chronic disease is due to inadequate production of, or poor response to, which one of the following?

A. Iron B. Folate C. Erythropoietin D. Ferritin E. Hemosiderin

AFP, Nov. 15, 2000

Anemia of chronic disease

Infections: TB, SBE, osteo, chronic UTI or pyelo, fungal

Malignancy: mets, leukemia, lymphoma, myeloma

Chronic inflammatory disorders: RA, SLE, Sarcoid, collagen vascular disease, polymyalgia rheumatica, chronic hepatitis, decubitus ulcer

Macrocytic anemia

Macro

RPI >= 2 RPI < 2

Check Occult Blood Loss Check B12 and folate

No

Yes

Coombs’ test

Macrocytic: RPI < 2

B12/Folate

B12 Low Normal Folate Low

MMA HighMMA

HomocysteineNormal

HomocysteineHigh

ConsiderLiver, Renal,

Thyroid,Alcohol,

Chronic dis.

ConsiderBone Marrow

Bx

Macrocytic: RPI < 2Megaloblastic Anemia

B12 Inadequate absorption Synthesized by

bacteria Meat, fish, dairy (strict

vegans) Absorbed as B12-IF

complex in ileum (gastrectomy)

Ca++ and pH dependant (PPI)

Folate Inadequate intake Synthesized by plants

and micro-organism Green leafy vege’s Fruits Absorbed in jejunum

Sample question #2

Which of the following tests can be useful in determining if an elderly patient has folate deficiency?

A. RBC folate concentration

B. Serum homocysteine level

C. Serum ferritin level

D. Serum methylmalonic acid level

AFP, Oct. 1, 2000

Macrocytic: RPI < 2Megaloblastic Anemia

Smear Macro-ovalocytic Polychromasia Hypersegmented neutrophil

Other Labs Homocysteine – Folate def. Methylmalonic acid – B12 def. Intrinsic Factor Ab test – very

specific for pernicious anemia but only 50% sensitive

Parietal cell AB test – quite sensitive (90%) but not specific

Schilling test

Macrocytic: RPI < 2Non-megaloblastic

Consider Liver, Renal, Endocrine (thyroid), alcohol, drugs

Consider anemia of chronic disease

Get Bone Marrow Biopsy

Myelodysplastic Myeloproliferative -

Leukemia, Lymphoma, Multiple Myeloma

Macrocytic: RPI < 2 continuesAplastic Anemia

Fanconi anemia – congenital Direct stem cell destruction – external

radiation Drugs - chloramphenicol, gold, sulfonamides, felbamate

Other Toxins - Solvents, degreasing agents, pesticides Viral infection - parvovirus B19, HIV, other Idiopathic

Macrocytic: RPI >= 2

Occult Blood Loss?

Yes No

Investigate source

Coombs’ (DAT)

Check for Hemolysis Peripheral smear

Sample question #3

Of the following laboratory results, which one does not occur in hemolytic anemia?

A. Reticulocytosis

B. Increased unconjugated bilirubin

C. Increased haptoglobin

D. Increased LDH

E. Hemosiderinuria

AFP, June 1, 2004

Macrocytic: RPI >= 2 Hemolytic Anemia

Other Lab Characteristics

RBC morphology Serum haptoglobin Serum LDH Unconjugated bilirubin Hemoglobinuria Hemosiderinuria

Macrocytic: RPI >= 2 Hemolytic Anemia

Coombs’ (DAT)

Positive Negative

Immune HemolysisDrug related HemolysisTransfusion, Infection, Cancer

Hemoglobinopathy, G6PD, PK, Spherocytosis, Eliptocytosis, PNH, TTP, DIC

Coombs’ positive with SpherocytesAutoimmune hemolytic anemia

Warm AIHA Abrupt onset IgG Anti-Rh, e, C, c, LW, U Jaundice Splenomegaly SLE, CLL, Lymphoma Drugs: methyl-dopa,

mefenamic acid, cimetidine, cefazolin

Cold AIHA Insidious onset IgM, complement Anti-I, I, Pr Cold agglutinin titer Absent jaundice Mycoplasma Virus

Coombs’ positive with SpherocytesOther immune hemolytic anemia

Alloantibody hemolytic anemia Transfusion reaction Feto-maternal incompatibility (Kleihauer-Betke test)

Drug related Hemolytic anemia Toxic immune complex (drug+Ab+C3)

- Quinine, Quinidine, Rifampin, INH, Sulfonamides,

Tetracyclin Hapten formation (anti-IgG)

- PCN, methicillin, ampicillin

Coombs’ Negative Hemolytic anemia

Episodic - G6PD def., PNH Hemoglobinopathy

- Sickle, crystals or target cells Elliptocytosis Spherocytosis DIC, TTP

Coombs’ Negative Hemolytic AnemiaMembrane Defects

Spherocytosis Common among

Northern European Autosomal dominant Decreased spectrin Osmotic fragility test Autohemolysis test

Elliptocytosis 90% with no clinically

significant hemolysis Abnormal membrane

protein

Coombs’ Negative Hemolytic AnemiaDeficiency of RBC Enzymes

Pyruvate Kinase Def.

Severe anemia in newborns

Adults symptomatic Jaundice Splenomegaly Fluorescent screening

test Quantitative test

G6PD Def.

X-linked Mediterranean,

African American, and Asian

Oxidant drugs – ASA, quinine, primaquine, chloroquine, sulfacetamide, sulfamethoxazole, nitrofurantoin, chloramphenicol, procainamide, quinidine

Infections Quantitative test

Coombs’ Negative Hemolytic AnemiaHemoglobinopathy

HbS disease Valine substitution for

Glutamic acid at the 6th position of b-chain

Sickle crises Severe anemia Screening test - Na

Metabisulfite solubility

Hgb electrophoresis

Coombs’ Negative Hemolytic AnemiaHemoglobinopathy continues

HbC disease Mild hemolysis Splenomegaly Lysine substitution HbC crystals “bar of gold” Hgb electrophoresis

HbSC disease Sickle and SC crystals

“Washington monument” Less crises More retinopathy/aseptic

necrosis

Coombs’ Negative Hemolytic AnemiaParoxysmal Nocturnal Hemoglobinuria

Rare chronic condition Recurrent abdominal pain, vomiting,

headaches, eye pain, thrombophlebitis Episodic Hgb in urine, Hemosiderinuria Abnormal cell membrane - increased lysis

by complement Screening - Sucrose hemolysis test Confirm - Acid hemolysis test (Ham’s test)

Coombs’ Negative Hemolytic AnemiaFragmented RBC’s & Thrombocytopenia

TTP-HUS Thrombocytopenia Microangiopathic

hemolytic anemia Neurologic symptoms and

signs Renal failure Fever Idiopathic - 37 % Drug-associated - 13 % Autoimmune disease - 13 % Sepsis - 9 % Pregnancy - 7 % Bloody diarrhea - 6 % Hematopoietic cell transplantation - 4 %

DIC Depletion of clotting factor (TTP – normal) Thrombocytopenia Bleeding (64%) Renal dysfunction (25%) Hepatic dysfunction (19%) Respiratory dysfunction

(16%) Shock (14%) Thromboemboli (7%) Central nervous system

involvement (2%) Sepsis, trauma, malignancy

TTP-HUS / DIC

Normocytic AnemiaHyperproliferative (RPI >= 2)

Use same flow chart as macrocytic hyperproliferative

Occult Blood Loss?

Yes No

Investigate source

Coombs’ (DAT)

Check for Hemolysis Peripheral smear

Normocytic AnemiaHypoproliferative (RPI < 2)

1. Get iron panel (ferritin)/B12/folate - some clue from RBC indices to check early

disease, high RDW, peripheral smear.

2. Consider liver, renal, drugs, toxin, endocrine (thyroid), and anemia of chronic disease.

3. Get BM bx - Leukopenia, thrombocytopenia, CRI < 0.1 - Aplastic anemia/pancytopenia - Abnormal (immature) cells on smear

Questions?