Anemia

Dr. Syuhada, Mkes, SpPK

 Anemia
Inability of the blood to supply the tissue with
adequate oxygen for proper metabolic
function.
Determining specific cause of anemia is
important appropriate management.
 Anemia :
Diagnostic criteria of anemia decrease level
of :
Hemoglobin
Hematocrit
RBC count
 Based on Hemoglobin level :
Anemia classified as :
Moderate (Hb 7-10 g/dL)
Severe (Hb < 7 g/dL)
 Clinical symptoms of anemia :
Pallor Syncope (particularly
Weakness following exercise)
Fatigue Dizziness
Lethargy or malaise Headache
Exercise dyspnea Tinnitus or vertigo
Palpitation Irritability
Pica (consumption of
substance such as ice, Difficulty sleeping or
starch, or clay, frequently concentrating
found in IDA) Gastrointestinal symptoms
 Causes of anemia
Blood loss (hemorrhage)
Accelerated destruction of RBCs (immune and nonimmune
hemolytic)
Nutritional deficiency (folate or Vitamin B12)
Bonemarrow replacement (e.g., by cancer)
Infection
Toxicity
Hematopoietic stem cell arrest or damage
Hereditary or acquired defect
Unknown
 Classification of anemia
Hemoglobin and Hematocrit
Red Blood Cell Indices
Red Blood Cell Indices and other tests
 Classification of anemia by RBC Indices

Size (MCV) Hgb content Possible causes

(fL) (MCHC) (%)
Normocytic Normochromic Bonemarrow failure, hemolytic anemia,
(80-100) (32-36) chronic renal disease, leukemia,
metastatic malignancy

Macrocytic Normochromic Megaloblastic and nonmegaloblastic

(>100) (32-36) macrocytic anemias (e.g.,liver disease,
myelodysplasia)

Microcytic Hypochromic Iron deficiency, sideroblastic anemia,

(<80) (<32) thalassemia, lead poisoning, chronic
diseases, chronic infection or
inflammation, unstable hemoglobins
 Laboratory Tests in the Diagnosis of
Anemia
1. Hemoglobin
2. Hematocrit
3. Red Blood Cell Indices
4. Peripheral Blood Smear (PBS)
5. Reticulocyte Count
6. Bone Marrow Smear & Biopsy
 Clinical laboratory

Morphological Classification of
anemia:
Hypochromic and microcytic
Normochromic and normocytic
Macrocytic
 Hemoglobin (Hb)

Is the main component of RBC

Its the physiologic carrier of oxygen to
tissues
 1. Hemoglobin
3 methods for measuring hemoglobin :
Cyanmethemoglobin (*)
Oxyhemoglobin
Measured Iron content
Cyanmethemoglobin method :
Blood is diluted in a solution of Potassium ferricyanide and potassium cyanide,
which oxidizes the hemoglobin to form methemoglobin.
Subsequently, methemoglobin forms cyanmethemoglobin in the presence of
potassium cyanide.
Because the absorption maximum occurs at a wavelength of 540 nm, the
absorbance of solution is read in a spectrophotometer at 540 nm, and
compared with a standard cyanmethemoglobin solution.
The methodes for measuring Hb:

Cyanmethhemoglobin
Oxyhemoglobin
Colorimetry: Sahli, Talqvist
Cyanmethemoglobin
method

K3Fe(CN)6
Blood methemoglobin

KCN
methemoglobin cyanmethemoglobin

Read the solutions

absorbance with
spectrophotometer at 540 nm
 Advantages of Cyanmethemoglobin
method
1. Most forms of hemoglobin are measured
2. Sample can be directly compared with a
standard
3. The solutions are stable
4. Coefficient of variation < 2% at physiologic
ranges
 . THE METHOD OF SAHLI

PRINCIPLE : Hb + HCl HEMATINE ACID (COLOUR BROWN)

METHOD: THE BLOOD IS DILUTED IN AN ACID SOLUTION

THE COLOUR OF COMPOUND IS COMPARED BY
COLOUR STANDARD WITH VISUALIZED

2
3

1
5 6
4

HCl 0,1 N
Automatic Hematology Analyzer
 Errors in hemoglobinometry

Errors inherent in the sample

Errors inherent in the method
Errors inherent in the equipment
Operators errors
 Referenge range of Hb concentration (g/dl)

Cord blood 13,5-20,5

First day of life 15,0-23,0
Children 6 months-6 years 11,0-14,5
Children 6 years-14 years 12,0-15,5
Adult males 13,0-17,0
Adult females 12,0-15,5
Pregnant 11,0-14,0
Mild anemia : 10-11g Hb/dl
Moderate anemia : 7-11
Severe anemia :<7
 Hematocrit
Packed Cell Volume (PCV)
The ratio of the volume of RBCs to the
volume of whole blood (%)
Referenge range for hematocrit:
Man: 42-45%
Woman: 37-47%
 macro

micro
 2. Hematocrit
Problems in measurement :
Incorrect centrifuge calibration
Choice of sample site
Incorrect ratio of anticoagulant to blood
owing to improper amount of blood drawn
Reading error
 Erythrocyte count
Measurement of erythrocyte:
Manually
Otomaticaly
Referenge range for erytrocyte count:
Man: 4,5 6,5 000.000 /mm3
Woman: 4,0 5,6 000.000 /mm3
 Red blood cell indices
The RBCs indices:
Mean Corpuscular Volume (MCV)
Mean Corpuscular Hemoglobin (MCH)
Mean Corpuscular Hemoglobin Concentration
(MCHC)
 MCV
Estimation of average size of the RBC
Calculated:

HCT (%) X 10fl

MCV:
RBC count (millions/ul)

Referenge range: 80-100 fl

< n: microcytic
> n: macrocytic
 MCH
To determine the content of Hb in RBC
Calculated:

Hb (g/dl) X 10pg
MCH:
RBC count (millions/ul)

Referenge range:27-31 pg
<n: hypochromic
 MCHC
To determine the content of Hb concentration in
RBC
Calculated:

Hb (g/dl) X100%
MCHC:
Hct (%)

Referenge range: 32-36 %

<n: hypochromic
 Peripheral blood smear
Stain: Wright and Giemza
Reddish orange appearance, central pallor: 2 -
3 um
Diameter: 6-8 um biconcaf disc, volume: 90 fl
 Alteration in size: anisocytosis
Microcytic
Macrocytic

Alteration in shape: poikilocytosis

Sferocyt
Leptocyt
Skistocyt
Stomatocyt
Pencil cell
Ovalosit
Image of Normal Red Blood Cells
Normochromic and normocytic
Hypochromic and microcytic
macrocytic
Polichromasia
Basophilic stippling
Macrocytic
Aniso-poikilositosis
Aniso-poikilositosis
Stomatocyt
Sickel cell
Roulleaux
 Morphologic classification
of anemia

Hypochromic and microcytic anemia:

Iron deficiency anemia
Thalassemia
Anemia of chronic diseases
Sideroblastik anemia
 Normochromic and normocytic anemia
Acut blood loss
Hemolytic anemia
Aplastic anemia
 Macrocytic anemia:
Megaloblastic anemia:
Non megaloblastic:
 Iron deficiency anemia
Is the most commonly recognized cause of
hypochromic anemia
Its caracterized by decrease in
Hb concentration, hematocrit, MCV
Results after there is total depletion (loss) of
the body iron store
 Iron deficiency anemia
Sequential steps in the development of iron
deficiency anemia:
Depletion (loss) of iron stores
Iron deficiency of erythropoiesis
Iron deficiency anemia
 Iron deficiency anemia
DISTRIBUTION OF IRON IN THE BODY
Total body iron of a healthy adult 2 5 g.
66.4 % : in the Hb of red cells (1 ml red cells contains
about 1 mg of iron)
30 % : ferritin and hemosiderin
3.3 %: Myoglobin of muscle cells and respiratory
enzymes of all cells
0.2% :cytochrome oxidase, catalase, peroxidase
0.1% : Transferrin (1-globulin).
 Iron deficiency anemia
IRON ABSORPTION
Daily intake of iron 0.5-2 mg
Adults with normal iron stores absorps about
5 10 % of total daily intake
Absorption takes place in the duodenum and
proximal portion of jejunum
 Iron deficiency anemia
Factors affecting daily iron requirements:
Growth spurts
Menstruation
Pregnancy
Lactation and breast feeding
Iron deficiency
 Iron deficiency anemia
Hematologic change:
Hb decreases before decrease of red cells
Decrease of MCHC, MCV and MCH resulting in a
hypochromic and microcytic anemia
Morphology of blood film
- hypochromic and microcytic red cell
- pencil cell
- target cell
 Iron deficiency anemia

Morphology of bone-marrow film (Giemsa

staining)
- defective erythropoiesis
- normoblastic hyperplasia, decreased
acidophilic normoblasts
- smaller normoblasts due to decrease of
cytoplasm
- ragged cytoplasm of normoblasts with
pycnotic nuclei
- naked nuclei (normoblast without
cytoplasm)
 Iron deficiency anemia
BIOCHEMICAL CHANGES
Decreased serum iron concentration < 40 Ug/dl
Increased Iron Binding Capacity (IBC) > 370 Ug/dl
Decreased transferrin saturation < 15%
Decreased serum ferritin concentration < 12 ng/ml
Increased erythrocyte protoporphyrin.
Increased transferrin receptor concentration

RESPONSE OF IRON TREATMENT

Reticulocytosis
Increased Hb concentration of 2 g/dl in 3 weeks
 4. Peripheral Blood Smear
Much information concerning the cause of anemia can be determined from a
PBS.
Coexistent neutropenia, thrombocytopenia, and anemia :
bone marrow failure
Lack of a nutritional substance to provide adequate bone marrow production
Excess lobulation of PMN (Hypersegmentation) macrocytic anemia
Basophilic stippling in RBC :
increased bone marrow production and reticulocytosis
Remnants of RNA (lead poisoning, malignancy)
 5. Reticulocyte Count
Useful in determining the response and potential of the bone
marrow
Reticulocytes are non-nucleated RBCs that still contain RNA.
Reticulocytes maybe visualized after incubation with
supravital dyes (New Methylen Blue, Brilliant Cresyl Blue)
Normal range : 0.5 2.0 % from RBCs
LEUKOCYTE DISORDER
Nucleus and cytoplasm disorders

GRANULOCYTE :
CYTOPLASM DISORDER :

1. Hypergranulation.
a. Toxic granulation
b. Alder - Reilly Anomaly
2. RNA Inclusion.
a. Dohles Bodies
b. May-Hegglin Anomaly
3. Vacuolisation.
Jordans Anomaly
4. Agranulation.
5. Giant Granulation.
Chediak-Higashi Syndrome
6. Decreasing of Peroxides activities.
a. Leukaemia
b. Alius-Crignashi Anomaly
NUCLUES DISORDER :

1. Hyper segmentation.
a. Acquired
b. Hereditary
2. Segmentation Obstacles.
a. Acquired
b.Hereditary : Pelger-Huet Anomaly
3. Nucleus Protuberance.
Seman Anomaly
EOSINOPHYLE :
o Presently Anomaly :
Hyper segmentation, Hypogranulation
o Homozygous Pelger-Huet :
Segmentation obstacle
o Alder-Reilly Anomaly :
Hypergranulation
o Chediack-Higashi Syndrome :
Giant granulation
o Jordans Anomaly :
Vacuolisation

o Leukaemia
MONOCYTE :

Alder-Reilly Anomaly
Chediack-Higashi Syndrome
Jordans Anomaly
Alius Crignashi Anomaly

LYMPHOCYTE :

NUCLEUS : Circle segmentation

Rieder Cell

CYTOPLASM : Vacuolisation
Leukemia
 Definition
Leukemia is a malignant disease of
hematopoietic tissue, characterized by
replacement of normal bone marrow
elements with abnormal (neoplastic) blood
cells.
 Classification of Leukemia
Acute Myeloid
Acute Lymphoblastic
Chronic Myeloid
Chronic Lymphoid
 Classification of Acute Myeloid
Leukemia

Type of Leukemia Abbrevation FAB* Alternate Names

Acute Myeloblastic Leukemia : AML Acute Nonlympho-
-without cytologic maturation M0 blastic Leukemia
-With minimal maturation M1 (ANLL)
-With maturation M2
Acute Promyelocytic Leukemia APL M3 Hypergranular
promyelocytic
Acute Myelomonocytic Leukemia AMML M4 Naegeli-type
leukemia
Acute Monocytic Leukemia AMoL M5 Schilling-type
leukemia
Erythroleukemia AEL M6 Di Guglielmos
syndrome. Eryhtremic
myelosis
Acute Megakaryoblastic Leukemia AMegL M7
 Classification of Acute
Lymphoblastic Leukemia
Type of Leukemia Abbrevation FAB Alternate Name

Precursor B-cell ALL :

-early-pre-B-cell ALL L1,L2 Common ALL
-Pre-B-cell ALL L1,L2 Common ALL
B-cell ALL L3 Burkitts leukemia

T-cell ALL L1,L2

 Classification of Chronic Myeloid
Leukemia
Type of Leukemia Abbrevation FAB Alternate Names

Chronic Myelogenous Leukemia CML Chronic Granulo-

cytic Leukemia

Chronic Eosinophilic Leukemia CEL

Chronic Basophilic Leukemia CBL

 Classification of Chronic
Lymphocytic Leukemia
Type of leukemia Abbrevation FAB Alternate Names
Chronic Lymphocytic Leukemia CLL
-B-cell CLL
-T-cell CLL

Prolymphocytic Leukemia PLL

Hairy Cell Leukemia HCL Leukemic reticulo-

endotheliosis
Sezary syndrome Mycosis fungoides,
leukemic phase
 Etiology and Risk Factors
Host Factors :
Heredity
Congenital Chromosomal Abnormalities
Immunodeficiency
Chronic Marrow Dysfunction
Environmental Factors :
Ionizing Radiation
Chemicals and Drugs
Viruses
 Incidence
In USA : 8-10 new cases/100.000
individuals/year.
Increases exponentially with age
Ratio adult : children = 10 : 1
Ratio males : females = 1-2 : 1
ALL more common in children, AML more
common in adults
 Comparison of Acute and Chronic
Leukemia
ACUTE CHRONIC
Age All ages Adults

Clinical onset Sudden Insidious

Course (untreated) < 6 mo 2-6 yr

Leukemic cells Immature Mature

Anemia Mild to severe Mild

Thrombocytopenia Mild to severe Mild

White Blood Cell count Variable Increased

Organomegaly Mild Prominent

 Clinical Features of Acute Leukemia
Pathogenesis Clinical Manifestation
Bone Marrow Failure
Anemia Fatigue, malaise, pallor
Thrombocytopenia Bruising, bleeding
Granulocytopenia Fever, infections
Organ Infiltration
Marrow expansion Bone or joint pain
Spleen Splenomegaly
Liver Hepatomegaly
Lymph nodes Lymphadenopathy
Central Nervous system Neurologic symptoms
Gums,mouth Gingival hypertrophy, oral lessions
 Laboratory Evaluation of Acute
Leukemia
1. CBC/peripheral blood smear.
2. Cytochemistry
3. Immunologic marker studies
4. Cytogenetics
5. Molecular genetics
 1. CBC/Peripheral Blood
Smear
Anemia :
Mild to severe
Normochrom normocytic
Platelet : decrease
WBC count :
Highly variable : decrease markedly increase
PBS :
Reveals blast/other immature cells (incl.normoblast)
 2. Cytochemistry
Special stains to identify chemical components of cells
(enzymes, lipids) specific for certain cell lines
Includes :
Myeloperoxidase
Sudan Black B
Specific Esterase
Nonspecific Esterase
Periodeic Acid Schiff (PAS)
 3. Immunologic Marker
Studies
1. Cell surface markers
2. Cytoplasmic markers
3. Terminal deoxynucleotidyl Transferase (TdT)
 Reference Book:
Denise M.Harmening
Clinical Hematology and Fundamentals of
Hemostasis, 4th edition, 2001

Anemia : pp 74-83
Leukemia : pp 272-300
Thank You