Hematology Flashcards
RBC Count
Given as 10 ^ 6 / uL or 10 ^ 12 / L
Normal is above 4.75 for males and above 4.18 for females
HGB
Measures the concentration of hemoglobin released by lysed red cells into whole blood
Given as g / DL
Normal is above 14 for males; above 12 for females
HCT
Measures the percentage of whole blood occupied by red cells
Given as a % total blood volume
Normal is above 39 for males, above 35 for females
MCV
Mean corpuscular volume measures the mean size of red cells
Given in femtoliters (10 ^ -15 L)
Normal is 80 - 100
MCH
Mean Corpuscular Hemoglobin measures the mean quantity of hemoglobin in a single red cell
Given in picograms 10 ^ -12g
MCH = HGB / RBC
Low MCH = hypochromatic
High MCH = hyperchromatic
MCHC
Mean Cell Hemoglobin Concentration is the average concentration of hemoglobin in a single red cell - corrects MCH for MCV
MCHC = HGB / HCT
RDW
Red Cell Distribution Width measures the variability in red cell size
Neutrophil
~ 2x size of typical RBC with many fine granules and 2-5 nuclear lobes
~40 - 72% (dif)
The presence of hypersegmented neutrophils (>5 nuclear lobes) is indicative of megaloblastic anemia (B12 or folate deficiency)
Eosinophil
~2x size of RBC with red/orange granules and usually 2 nuclear lobes
0.0% - 6.0% (dif)
Basophil
Contains numerous large, round, purple-black or dark blue cytoplasmic granules
0.0% - 0.2% (dif)
Monocyte
Large, kidney shaped nucleus; pale blue cytoplasm
2.0% - 11.0% (dif)
Lymphocyte
Small with round, dense nucleus and scant blue cytoplasm
20 - 50% (dif)
Spherocytes
Small, spherical RBCs w/o central pallor
Often due to cell membrane defects, i.e. hereditary spherocytosis caused by spectrin mutation and loss of cell membrane
Bite Cells
Caused by removal of Heinz bodies in the spleen;
Suspicious for G6PD Deficiency
Schistocytes
RBC fragments; characteristic of intravascular hemolysis
Target Cells
Suspicious for Thalassemia or HbC Disease
PLT
Platelet count is given in 10^3 / uL or 10^9 / L
Usually 150 - 400 10^9 / L
Microcytosis
MCV < 80
Iron deficiency anemia, thalassemia, lead poisoning
Macrocytosis
MCV > 100
Megaloblastic anemia (B12 or folate deficiency)
Reticulocyte Count
Immature RBCs found in circulation for 1 day prior to maturation
Counted as a percent of RBCs present; normal range is 0.4 - 1.7% (~1% of RBC mass is produced per day) or below 50,000 / uL
Elevated retic count indicative of anemia due to increased RBC destruction or loss
Reticulocyte Index
Provides a ratio of how many fold beyond baseline the normal RBC production is
RI should be between 1 and 2
RI < 1 with anemia indicates decreased production of RBCs
RI > 2 with anemia indicates loss of RBCs (destruction or bleeding) leading to increased compensatory production
Iron absorption
Occurs at the mucosal surface of the duodenum where ferric (3+) iron is reduced to ferrous (2+ iron) which enters the epithelial cell via the action of a divalent metal ion transporter DMT1
Inside the cell some iron is stored in ferritin and some is transported across the basolateral membrane by ferroportin; iron is oxidized to 3+ as it leaves the cell and binds plasma apotransferrin
Hepcidin
A liver peptide produced in response to high iron intake, inflammation, and/or infection; it is a negative regulator of iron absorption
Hepcidin binds ferroportin, down regulating it’s production and resulting in loss of export of iron out of the cell and increased accumulation of iron storage in cellular ferritin
Hepcidin mediates the anemia of chronic infection/inflammation
Iron transport
Iron bound to transferrin moves to the bone marrow where it binds erythroblast surface receptors; the transferrin/transferrin receptor complex is endocytosed and iron is released within the endosome and transported into the cytoplasm by DMT1 to go to sites of ferritin storage
Iron deficiency anemia
Decreased Hgb and Hct
Decreased RBC production characterized by low reticulocyte count and index
Microcytosis (low MCV), Hypochromia (low MCH), and wide range in size of RBCs (high RDW)
Iron overload
Often caused by increased absorption of iron (hemochromatosis)
HLA-H gene codes for a protein in the duodenal cells which acts as a co-factor for absorption; gain-of-function mutation affecting HLA-H may cause increased iron absorption
Clinical consequences: Heart damage (arrhythmia and congestive heart failure), liver damage, endocrine damage
Treatment: Therapeutic phlebotomy, iron chelation
Where does hematopoiesis occur?
Embryonic stage (0 - 3 months) - yolk sac
Fetal stage (3 - 7 months) - liver and spleen
Birth and early childhood - most of the marrow cavity
Adulthood - axial skeleton (vertebrae, pelvis, sternum, ribs, skull)
Hematopoietic Stem Cells (HSCs)
Multipotential stem cells that can give rise to all blood cells (both lymphoid and myeloid lineages) via assymetric cell division
Oligopotent Hematopoietic Stem Cells
Common progenitor cells of the lymphoid line and myeloid line
CFU-L - gives rise to all lymphoid cells
CFU-GEMM - gives rise to all non-lymphoid blood cells (granulocyte, erythroid, monocyte, megakaryocyte)
Erythropoietin (EPO)
Made by kidney cells in response to hypoxia; promotes erythrypoiesis, hemoglobin production, and causes increased release of reticulocytes into the peripheral blood
Erythrocyte maturation
CFU-ME BFU-E Pronormoblast Basophilic Normoblast Polychromatophilic Normoblast Orthochromatic Normoblast (last nucleated stage) Reticulocyte Erythrocyte
Granulocyte maturation
Myeloblast - common progenitor to all 3 granulocyte lineages Promyelocyte Myelocyte - secondary granules appear Metamyelocyte Band Segmented Neutrophil (Seg) --> Neutrophil, Eosinophil, Basophil
Megakaryocyte Maturation
Megakaryoblast
Promegakaryocyte
Megakaryocyte
Platelet
Monocyte Maturation
Monoblast
Promonocyte
Monocyte
Marrow cellularity
Refers to the portion of the marrow that is hematopoietically active
Marrow may be hypercellular (signaling increased proliferation of one or more lineage) or hypocellular (signaling attack on marrow cells)
Factors affecting hemoglobin’s oxygen affinity
Bohr Effect - CO2 produced in the tissues as a byproduct of metabolism equilibrates in the blood to form carbonic acid; Hemoglobin has higher O2 affnity at higher pH (i.e. lungs) and lower O2 affinity at lower pH (i.e. tissues)
Temperature - metabolic rates are higher at increased temperatures (exercise, fever); hemoglobin has decreased O2 affinity at higher temperatures
2-3, BPG - a byproduct of anaerobic glycolysis present in RBCs at higher levels during conditions of hypoxia & anemia; 2-3, BPG binds to deoxyhemoglobin, stabilizing the T configuration and decreasing O2 affinity
Hemoglobin Variants
Embryonic - Hemoglobin Gower 1, Hemoglobin Gower II, Hemoglobin Portland
Fetal Hemoglobin (a2y2) - predominates after 8 weeks, higher O2 affinity than HbA
HbA (a2B2) - 97% of adult hemoglobin
HbA2 (a2d2) - 3% of adult hemoglobin, more in B-thalassemia
Physically unstable hemoglobins
Can lead to hemolytic anemia, a.k.a. “Heinz Body anemia” due to presence of Heinz bodies - precipitated, denatured hemoglobin within the cell
I.e. Hb Koln, Hb Poole
High affinity hemoglobins
May lead to erythrocytosis because O2 delivery to the tissues is reduced, leading to increased release of erythropoietin and stimulated RBC production
i.e. Hb Chesapeake
Low affinity hemoglobins
May be associated with cyanosis (more deoxygenated hemoglobin circulating, < 85% oxygen saturation) often with mild anemia (fewer RBCs needed)
Methemoglobinemia
Overproduction of methemoglobin, hemoglobin in which iron is bound to the heme group in its ferric (3+) form leading to decreased ability of hemoglobin to unload oxygen
Acquired - oxidation of the heme group by free radicals
Genetic - homozygous deficiency of cytochrome b5 reductase
Treatment: methylene blue
Factors affecting hemoglobin’s oxygen affinity
Bohr Effect - CO2 produced in the tissues as a byproduct of metabolism equilibrates in the blood to form carbonic acid; Hemoglobin has higher O2 affnity at higher pH (i.e. lungs) and lower O2 affinity at lower pH (i.e. tissues)
Temperature - metabolic rates are higher at increased temperatures (exercise, fever); hemoglobin has decreased O2 affinity at higher temperatures
2-3, BPG - a byproduct of anaerobic glycolysis present in RBCs at higher levels during conditions of hypoxia & anemia; 2-3, BPG binds to deoxyhemoglobin, stabilizing the T configuration and decreasing O2 affinity
Hemoglobin Variants
Embryonic - Hemoglobin Gower 1, Hemoglobin Gower II, Hemoglobin Portland
Fetal Hemoglobin (a2y2) - predominates after 8 weeks, higher O2 affinity than HbA
HbA (a2B2) - 97% of adult hemoglobin
HbA2 (a2d2) - 3% of adult hemoglobin, more in B-thalassemia
Physically unstable hemoglobins
Can lead to hemolytic anemia, a.k.a. “Heinz Body anemia” due to presence of Heinz bodies - precipitated, denatured hemoglobin within the cell
I.e. Hb Koln, Hb Poole
High affinity hemoglobins
May lead to erythrocytosis because O2 delivery to the tissues is reduced, leading to increased release of erythropoietin and stimulated RBC production
i.e. Hb Chesapeake
Low affinity hemoglobins
May be associated with cyanosis (more deoxygenated hemoglobin circulating, < 85% oxygen saturation) often with mild anemia (fewer RBCs needed)
Methemoglobinemia
Overproduction of methemoglobin, hemoglobin in which iron is bound to the heme group in its ferric (3+) form leading to decreased ability of hemoglobin to unload oxygen
Acquired - oxidation of the heme group by free radicals
Genetic - homozygous deficiency of cytochrome b5 reductase
Treatment: methylene blue
White Blood Cell Count and Differential
Normal adult WBC Count: 4,500 - 10,500 / uL or 4.5 - 10.5 x 10^9 / L
Neutrophils: 40 - 60% Lymphocytes: 20 - 40% Monocytes: 2 - 8% Eosinophils: 1 - 4% Basophils: .5 - 1%
Causes of underproduction anemia
Iron-deficiency Chronic infection / inflammation Malignant disease Renal insufficiency Endocrine disorders Lead intoxication Vitamin B12 / Folate deficiency
Pathophysiology and lab findings of lead intoxication
Lead inhibits the synthesis of protoporphyrin as well as the enzyme that attaches iron to the porphyrin ring, leading to decreased Hb production
Characterized by mild to moderate anemia (Hgb 8 - 12 g/dL), decreased reticulocyte count, microcytosis with mild hypochromia, basophilic stippling, increased protoporphyrin
Sideroblastic anemia
Results from impaired production of protoporphyrin or incorporation of iron into porphyrin ring - iron accumulates in mitochondria
Inherited and acquired forms
Pathophysiology of B12 / folate deficiency
Folic acid and B12 are co-factors for hematopoiesis; deficiency causes cells to increase in size, arrest in S phase of mitosis, and undergo destruction
B12 absorption, transport, and deficiency
B12 binds to intrinsic factor (IF) in the stomach and is absorbed in the terminal ileum; after absorption, B12 is bound to transcobalamin binding protein II (TcII) and transported to the liver for storage or to the bone marrow for use
B12 deficiency may result from auto-immune causes (anti-intrinsic factor antibodies produced), IF deficiency 2/2 gastritis, or defector transport / storage (TCII deficiency)
B12 deficiency - laboratory findings & clinical consequences, treatment
B12 deficiency causes megaloblastic anemia with an onset on the order of months
Characterized by macrocytosis (MCV > 97), decreased reticulocyte count, RI < 1, increased bilirubin due to intramedullary destruction of RBCs
Clinical presentation includes neurologic features; treatment with large doses of folic acid may exacerbate neurologic damage
Treatment: IM or SC injections of B12 daily for 2 weeks, then weekly until HCT is normal, then monthly for life
Folate - absorption, transport
Characteristic anemia & treatment
Folate is absorbed in the jejunum and stored in the liver where it is secreted in the bile and reabsorbed (enterohepatic circulation)
Most common causes of deficiency are dietary insufficiency, malabsorption, and increased demand
Characteristic anemia is megaloblastic with an acute onset on the order of weeks; MCV > 97, decreased reticulocyte count, RI < 1, increased bilirubin due to intramedullary destruction of RBCs
Treatment: 1 mg/day orally
Differential Diagnosis for microcytic, hypochromic anemia
Iron deficiency, lead poisoning, occult GI bleed, decreased production (primary marrow disease) peripheral destruction (hemolysis), anemia of chronic disease
Megaloblastic anemia - etiology, CBC, and smear findings
CBC findings:
Decreased RBC, HGB, HCT
Increased MCV, MCH, RDW
Smear findings:
Sparser number of RBCs, RBCs are large, presence of hypersegmented neutrophils
Etiology: Vitamin B12 / folate deficiency, chemotherapy / radiation causing dysplasia of bone marrow
Intravascular hemolysis - Mechanism
Hemoglobin from lysed RBCs is released into circulation where it dissociates into alpha-beta dimers, which bind to the liver protein haptoglobin; this complex is removed from circulation by the liver
Excess free Hb dimers may be broken down and converted to bilirubin in the liver or may be oxidized to methemoglobin
Extravascular hemolysis - Mechanism
The faulty RBC is ingested by a macrophage in the spleen; iron is removed from hemoglobin and released for storage in transferrib; the porphyrin ring is converted to bilirubin which enters the blood where it is taken up by the liver and conjugated with glucuronic acid, which is secreted into the gut and transformed into fecal urobilinogen
Laboratory findings relevant to hemolysis
Increased retic count / RI (compensatory)
Presence of spherocytes, schistocytes
Hyperbilirubinemia (mostly unconjugated; the hepatic process of conjugation with glucuronic acid is overwhelmed)
Increased serum / urine hemoglobin
Hereditary Spherocytosis
Characterized by variable onset of moderate anemia, jaundice, and splenomegaly
Most commonly caused by spectrin deficiency resulting in loss of plasma membrane and formation of spherocytic RBCs which are less deformable and have greater osmotic fragility
Labs: Increased retic count and RI Decreased MCV Spherocytes on smear Unconjugated Hyperbilirubinemia
G6PD Deficiency
X-linked recessive deficiency of G6PD enzyme, important in the pathway that restores reduced glutathione necessary in protecting the cell against oxidative stress; in the absence of G6PD, oxidative stress causes oxidation of hemoglobin which denatures and attaches to the membrane, damaging spectrin; spectrin damage results in decreased deformability of RBC, splenic trapping, and extravascular hemolysis
Hemolytic crisis may be precipitated by fava beans, aspirin
Pyruvate Kinase Deficiency
Enzyme defect in PK, which converts phosphoenolpyruvate to pyruvate; results in decreased ATP, loss of membrane plasticity and increased membrane rigidity, extravascular hemolysis (in the spleen)
Sickle Cell Disease
Autosomal recessively inherited condition resulting from mutation in the beta globin chain at the 6th AA from glutamate to valine
Characterized by severe anemia (Hb 6-9g/dL), high reticulocyte count, increased RDW, abnormal smear with presence of sickled cells, increased bilirubin, LDH, AST (released from lysed RBCs)
Treatment: bone marrow transplant, hydroxyurea (induces production of HbF), transfusion
Sickle Beta0 Thalassemia
Caused by the presence of one sickle cell gene and one defective beta globin gene
Characterized by severely low Hb (6-9g/dL), elevated retic count, microcytic RBCs
Clinically severe
Sickle Hemoglobin C Disease (SC Disease)
Caused by the presence of one sickle cell gene and one HbC gene (mutation of beta globin AA 6 from glutamate to lysine)
Characterized by moderately low Hb (10-12 mg/dL), slightly elevated retic count
Mild to moderate clinical severity because the presence of HbC interferes with the polymerization of HbS within the RBC
Clinical and lab findings of thalassemia
Decreased MCV, MCHC
Increased retic count
Abnormal peripheral smear with microcytosis, target cells, increased reticulocytes
Increased bilirubin, LDH, AST
Splenomegaly
Extramedullary hematopoiesis (frontal bossing, osteopenia, splenomegaly)
Treatment of thalassemia
Transfusion - indicated in B-thalassemia major; started within the first 2 years of life to maintain Hb values between 8-10 g/dL in order to prevent extramedullary hematopoesis, allowing normal growth and development
Hydroxyurea - increases HbF production
Bone marrow transplant - requires HLA-identical, unaffected sibling
Major blood group alloantigens
Alloantigens are biochemically distinct differences in polysaccharides on the surface of RBCs that are immunologically different but functionally identical
The major blood alloantigens are A, B, O, and AB, differentiated by different polysccharides / glycoproteins
Rh System
The Rh antigen system consists of 3 pairs of alleles: C/c, D/-, and E/e
RhD is the most clinically significant, referred to as Rh+ or Rh-
85% of US Caucasians are RhD+
Blood components
Whole Blood - indicated in massive transfusions to replace oxygen-carrying capacity and blood volume
Packed Red Blood Cells (PRBCs) - indicated to replace oxygen-carrying capacity (Hct = 70%) in chronic anemia or acute blood loss
Fresh Frozen Plasma (FFP) - indicated to treat coagulopathy related to procoagulatnt deficiency; acellular, contains all essential clotting factors as well as complement factors and other plasma proteins
Donor Platelet Concentrate - platelet function is maintained but concentrates are a poor source of clotting factors; indicated for bleeding associated with thrombocytopenia
Immediate hemolytic transfusion reaction
Results from transfusion with incompatible blood products (usually ABO mismatched); results in activation of complement and intravascular hemolysis leading to shock, acute renal failure, and intravascular coagulation
Treatment: stop the infusion, maintain renal output with IV fluids and diuretics
Howell-Jolly Bodies
Single, dense blue dot inclusion in RBCs, evident on smear; comprised of nuclear DNA
Caused by (real or functional) asplenia, megaloblastic anemia
Heinz Body
Blue dot inclusion in the periphery of RBCs, evident on smear with supravital dye
Comprised of denatured/oxidized hemoglobin attached to the inner cell membrane
Cause: G6PD deficiency
Associated with bite cells
Hypersegmented Neutrophils
More than 5 lobes
Associated with megaloblastic anemia
Total Iron Binding Capacity (TIBC)
Measures the capacity of the blood to bind iron with transferrin
High in iron deficient anemia because the body is trying to maximize capture of all available iron
Low in anemia of chronic disease because the body sequesters iron in intracellular ferritin in order to keep it away from pathogens in circulation
Transferrin Saturation
Gives the percentage of transferrin that is available to bind iron in circulation
Calculcated by serum iron / TIBC x 100
Low in iron deficiency anemia
Low-normal in anemia of chronic disease; seen with increased ferritin
High in hemochromatosis
Hemoglobin E
Caused by a point mutation in in the B-globin gene, creating an abnormally spliced B-globin mRNA and production of small amounts of abnormal B-globin protein that interacts weakly with alpha globin
Homozygotes present with mild Beta Thalassemia; heterozygotes are clinically normal
Hemoglobin C
Caused by a Glutamate –> Lysine point mutation in the hemoglobin beta chain
Homozygotes present with mild hemolytic anemia; heterozygotes are clinically normal
Cold antibody autoimmune hemolytic anemia
Cold antibodies, usually IgG or IgM, transiently bind RBCs in cooler areas of the body; as they move back to central circulation, they activate complement through the C5-C9 MAC; the antibody dissociates because of low affinity at these warmer central temperatures and complement destroys the cell via intravascular hemolysis .
Warm antibody autoimmune hemolytic anemia
Warm antibodies, usually IgG, bind the red cell and trigger splenic macrophage phagocytosis of the red cell via interaction of the Fc domain, leading to extravascular hemolysis
Little or no complement activity
Direct antiglobulin test (DAT) or Coomb’s Test
Evaluates the presence of either IgG or C3d or C4d on the surface of a patient’s RBCs via the addition of Coombs reagent which has antibodies for IgG, C3d and C4d, causing agglutination.
Methylmalonic Acid (MMA)
Specific test for Vitamin B12 deficiency; elevated in B12 deficiency
Homocysteine
Elevated in B12 and folate deficiency
