Erythrocyte and Morphologic Abnormalities of RBC

Maturation of RBC

18 to 21 days

• BFU-E →CFU-E = 7 days

• CFU-E →Pronormoblast = 7 days

• Pronormoblast →Mature RBC = 7 days

Morphologic Trend as Cell Matures (Erythroblast vs. More mature stage)

• Cell size

• N:C ratio

• Nucleoli

• Nuclear chromatin

• Color of cytoplasm

• Cell size - erythroblast larger, mature smaller

• N:C ratio - erythroblast hgher, mature lower

• Nucleoli - erythroblast present, mature absent

• Nuclear chromatin - erythroblast fine, homogenous, mature coarse, clumped, condense

• Color of cytoplasm - erythroblast blue, mature pink

Nomenclature of RBC Maturation series

• First morphologically identifiable RBC precursor

• Contains 1 to 2 nucleoli

• Globin production begins

Pronormoblast

• Intensely basophilic (deeper blue) cytoplasm

• Chromatin begins to condense (resembles a wheel with broad spokes)

• Detectable hemoglobin synthesis occurs

Basophilic normoblast

• First stage in which pink color (due to hemoglobin) is seen

• Hemoglobin synthesis increases

• Last stage capable of mitosis

Polychromatic erythroblast

• Complete condensation of nucleus (pyknotic)

• Hemoglobin production continues on the remaining ribosomes

• Last nucleated stage (nucleus is ejected from the cell later in this stage)

Orthochromic normoblast

• Completion of hemoglobin production

• Residual RNA imparts a bluish tinge to the cytoplasm

• Also known as diffusely basophilic erythrocyte

Polychromatic erythrocyte

Characteristics of Mature RBC

• Size: _ um

• Central pallor: _ of cell size

• Shape: _

• Primary function: _

• Size: 7-8 um

• Central pallor: 1/3 of cell size

• Shape: biconcave/discocyte

• Primary function: deliver oxygen to tissues

RBC membrane consists of:

• _% carbohydrates

• _% lipids

• _% proteins

• 8% carbohydrates

• 40% lipids

• 52% proteins

Lipids in RBC membrane (2)

• Phospholipids - dynamic fluidity to membrane

• Cholesterol - tensile strength to the lipid bilayer

2 types of protein in RBC membrane

• Integral/Transmembrane Protein

• Peripheral/Cytoskeletal Protein

• transport ions, water, and glucose, and anchor cell membrane receptors

• also provides vertical membrane support

Integral/Transmembrane Protein

Example of Integral/Transmembrane Protein

• Aquaporin-1

• Glycophorins (A,B,C)

• certain blood group antigens (Rh)

• provides lateral or horizontal membrane stability

Peripheral protein

Example of Peripheral proteins

• Alpha-spectrin

• Beta-spectrin

• Ankyrin

• Adducin

• Actin

• Protein 4.1 and 4.2

• Tropomodulin

• Tropomyosin

Production of RBCs

• Stimulus:

• Source of EPO:

• Stimulus: Hypoxia

• Source of EPO: Peritubular interstitial cells of the kidney

EPO is a glycoprotein hormone and growth factor that has the following effects (3)

• early release of reticulocytes in BM

• preventing apoptosis of RBC progenitors

• less time of RBC maturation in BM

EPO measurement

• specimen: _

• normal value: _

• EDTA plasma measured by chemiluminescence

• 10 to 30 U/L

Destruction of RBCs

• Extravascular Hemolysis

→%:

→Mechanism:

→Lab findings in excessive hemolysis:

→%: 80% to 90%

→Mechanism: RES macrophage-mediated

→Lab findings in excessive hemolysis: High serum conjugated bilirubin (B1) and urine urobilinogen

Destruction of RBCs

• Extravascular Hemolysis

→%:

→Mechanism:

→Lab findings in excessive hemolysis:

→%: 10% to 20%

→Mechanism: Fragmentation of RBCs

→Lab findings in excessive hemolysis: low serum haptoglobin and hemopexin, hemoglobinemia, hemoglobinuria

Reference method used for RBC survival studies:

Chromium-51 Radioisotope Labelling

RBC Metabolic Pathways

_______

• main source of ATP (90%)

• glucose is catabolized into _

• produces a net gain of _ molecules of ATP

• important enzyme: _

EMBDEN-MEYERHOF PATHWAY

• main source of ATP (90%)

• glucose is catabolized into PYRUVIC ACID

• produces a net gain of 2 molecules of ATP

• important enzyme: HEXOKINASE, PYRUVATE KINASE, LACTATE DEHYDROGENASE

Embden-Meyerhof Pathway is also called _

anaerobic glycolysis

RBC Metabolic Pathways

_______

• detour of glycolysis (5% to 10%)

• generates _ to reduce glutathione

• detoxifies oxidative compounds and safeguards hemoglobin

• important enzyme: _

RBC Metabolic Pathways

HEXOSE MONOPHOSPHATE PATHWAY

• detour of glycolysis (5% to 10%)

• generates NADPH to reduce glutathione

• detoxifies oxidative compounds and safeguards hemoglobin

• important enzyme: G6PD

Hexose Monophosphate Pathway is also called _

Aerobic glycolysis

RBC Metabolic Pathways

_______

• prevents formation of _

• maintains iron in ferrous state (Fe2+)

• important enzyme: _

RBC Metabolic Pathways

METHEMOGLOBIN REDUCTASE PATHWAY

• prevents formation of METHEMOGLOBIN

• maintains iron in ferrous state (Fe2+)

• important enzyme: CYTOCHROME B5-REDUCTASE

RBC Metabolic Pathways

_______

• generates _

• this binds between globin chains in hemoglobin and enhances delivery of oxygen ro tissues

RBC Metabolic Pathways

RAPOPORT-LUEBERING PATHWAY

• generates 2,3-DIPHOSPHOGLYCERATE

• this binds between globin chains in hemoglobin and enhances delivery of oxygen ro tissues

Main function is to transport O2 from the lungs to tissues and transport CO2 from the tissues to the lungs for exhalation

RBC

A molecule of hemoglobin consists of:

• 2 pairs of polypeptide chains

• 4 heme group

Each heme group is composed of a complex ring structure called _

protoporphyrin IX

Heme consists of:

• 1 atom of ferrous (Fe2+) iron

• 4 pyrrole rings

• Each hemoglobin is capable of binding _ molecules of O2

• 1 g hemoglobin can carry _ of O2

• A small percentage of HbA is glycated. The most characterized of the glycated hemoglobin is _ in which glucose attaches to the -

• Each hemoglobin is capable of binding 4 molecules of O2

• 1 g hemoglobin can carry 1.34 mL of O2

• A small percentage of HbA is glycated. The most characterized of the glycated hemoglobin is HbA1c in which glucose attaches to the N-terminal valine of the beta chain

Heme synthesis occurs in the _ and _ of bone marrow erythroid precursors

• cytoplasm and mitochondria

• The genes that code for the production of globin chains are located on: _

• Translation of mRNA to the globin polypeptide chain occurs on the ribosomes in cytoplasm

• Chromosome 16: alpha, beta

• Chromosome 11: beta, epsilon, delta, gamma

Intrauterine (Embryonic) Hemoglobin

• Gower I: 2 epsilon: 2 zeta

• Gower II: 2 epsilon: 2 alpha

• Portland: 2 zeta: 2 gamma

Newborn hemoglobin

• HbF (60 to 90%)

• HbA (10 to 40%)

2 years to adulthood hemoglobin

• HbA (>95%)

• HbA2 (<3.5%)

• HbF (1 to 2%)

Describes how the affinity of hemoglobin for oxygen relates to the partial pressure of oxygen

Hb-O2 Dissociation Curve

Hb-O2 Dissociation Curve

• Shape:_

• PO2 of approximately _ results in 50% saturation of hemoglobin molecule (P50 value)

• sigmoidal

• 27 mmHg

Hb-O2 Dissociation Curve

• Shift to the left

• HIGH oxygen affinity

• LOW oxygen release to tissues

• LOW PCO2

• LOW temperature

• LOW H+ ions/alkalosis

• LOW 2,3-DPG

Hb-O2 Dissociation Curve

• Shift to the right

• LOW oxygen affinity

• INCREASED release of oxygen to tissue

• HIGH PCO2

• HIGH TEMP

• HIGH H+ ions / acidosis

• HIGH 2,3-DPG

• dysfunctional hemoglobins that cannot transport oxygen

Dyshemoglobin

• Methemoglobin

• Carboxyhemoglobin

• Sulfhemoglobin

Methemoglobin (Hi)

• Hb iron is in _

• Acquired/inherited

• Causes (2)

• associated with _ blood color

• Treatment: _

• Hb iron is in FERRIC STATE (Fe3+)

• Acquired/inherited

• Causes CYANOSIS, DYSPNEA

• associated with CHOCOLATE BROWN blood color

• Treatment: METHYLENE BLUE / ASCORBIC ACID

Carboxyhemoglobin (HbCO)

• _ times greater affinity to Hb than O2

• Acquired (smoking, car exhaust)

• Toxic effects (_% levels)

• assocaited with _ colored blood

• Treatment: _

• 240 times greater affinity to Hb than O2

• Acquired (smoking, car exhaust)

• Toxic effects (20-30% levels)

• assocaited with CHERRY RED colored blood

• Treatment: REMOVE CO SOURCE, O2 THERAPHY

Sulfhemoglobin (SHb)

• sulfur atom at heme pyrrole ring

• acquired (drugs/sulfur chemicals)

• cannot be converted to _

• Associated with _ color

• Treatment: _

• sulfur atom at heme pyrrole ring

• acquired (drugs/sulfur chemicals)

• cannot be converted to NORMA HbA

• Associated with MAUVE LAVENDER/GREENISH PIGMENT color

• Treatment: AVOIDANCE OF THE OFFENDING AGENT

Abnormal variation in RBC volume or diameter

Anisocytosis

• Normocytic: 7 to 8 um

• Microcytic: <6 um

• Macrocytic: >8 um

Variation in hemoglobin content of RBCs

Anisochromia

• Normochromic: 1/3 central pallor, MCHC = 32 to 36 g/dL

• Hypochromic: <1/3 central pallor, MCHC<32 g/dL

• Hyperchromic: >1/3 central pallor, MCHC>36 g/dL

Abnormal variation in RBC shape

Poikilocytosis

RBC Morphologic Abnormalities

• deep staining large oval red cell

Macroovalocyte

RBC Morphologic Abnormalities

• Macroovalocyte

Megaloblastic anemia

RBC Morphologic Abnormalities

• small,dense RBC

• no central pallor

• low surface area:volume ratio

Spherocyte

RBC Morphologic Abnormalities

• Spherocyte

• Heriditary spherocytosis

• AIHA

• Severe burns

RBC Morphologic Abnormalities

• Few irregularly spaced projections of varying length and width

Acanthocyte/ Thorn cell

RBC Morphologic Abnormalities

• Acanthocyte/Thorn cell

• severe liver disease

• abetalipoproteinemia

• McLeod syndrome

RBC Morphologic Abnormalities

• RBC with blunt or pointed, short projections that are evenly spaced

Burr cells / Echinocyte

RBC Morphologic Abnormalities

• Burr cell / Echinocyte

• Uremia

• Pyruvate kinase deficiency

RBC Morphologic Abnormalities

• Hb concentrated in the center and around the periphery

Target cell / codocyte

RBC Morphologic Abnormalities

• Target cell / Codocyte

• Liver disease

• Obstructive jaundice

• Thalassemia

• Hemoglobin C Disease

RBC Morphologic Abnormalities

• slit like (mouth like) area of central pallor

• Hereditary stomatocytosis

• Acute alcoholism

• Rh null syndrome

RBC Morphologic Abnormalities

• narrow oval cells

• egg-shaped or cigar shaped

Elliptocyte

RBC Morphologic Abnormalities

• Elliptocyte

• Hereditary elliptocytosis

• IDA

• Thalassemia major

RBC Morphologic Abnormalities

• Fragmented RBCs caused by rupture in peripheral circulation

Schistocyte

RBC Morphologic Abnormalities

• Schistocyte

• Disseminated Intravascular Coagulation

• Hemolytic Uremic Syndrome

• Microangiopathic Hemolytic Anemia

• Thrombotic Thrombocytopenic Purpura

RBC Morphologic Abnormalities

• has single pointed extension resembling tear drop or pear

Dacryocyte

RBC Morphologic Abnormalities

• Dacryocyte

• Primary Myelofibrosis

• Thalassemia

• Myelopthisic Anemia

• Megaloblastic anemia

RBC Morphologic Abnormalities

• Crescent-shaped cell

• Caused by crystalization of HbS due to low oxygen tension

Sickle cell / Drepanocyte

RBC Morphologic Abnormalities

• Sickle cell/Drepanocyte

• Sickle cell anemia

• Sickle cell-beta thalassemia

RBC inclusions

• Bluish tinge throughout cytoplasm

• Also called polychromasia

• Contect: RNA

Diffuse basophilia

RBC inclusions

• Diffuse basophilia

• Hemolytic anemia

• After treatment for iron, vitamin B12 or folate deficiency

RBC inclusions

• dark blue-purple, punctuate granules

• Content: precipitated RNA

Coarse basophilic stippling

RBC Inclusions

• Coarse Basophili Stippling

• Lead poisoning

• Megaloblastic anemia

• Thalassemia

• Hemoglobinopathies

• Myelodysplastic Syndrome

RBC Inclusions

• Dark blue-purple round granule

• Content: DNA remnants

Howell-Jolly bodies

RBC Inclusions

• Howell-Jolly bodies

• Post-splenectomy

• Megaloblastic anemia

• Thalassemia

• Myelodysplastic syndrome

RBC Inclusions

• Not visible in Wight's staine

• In supravital stain: dark blue-pyrple granule near RBC membrane

• Content: Denatured hemoglobin

Heinz bodies

RBC Inclusions

• Heinz bodies

• G6PD deficiency

• Unstable hemoglobin

• Oxidant drugs/chemicals

RBC Inclusions

• Irregular clusters of light to dark blue granules near periphery

• Content: Non-heme iron

Pappenheimer bodies

RBC Inclusions

• Blue rings or figure-8 inclusion

• Content: mitotic spindle remnant

• Megaloblastic anemia

• Myelodysplastic syndrome

RBC Inclusions

• Not visible in wright's stain

• Supravital stain: Fine, evenly dispersed dark blue granules

• “golf-ball RBCs”, “raspberry”

• Content: precipitated beta-globin chains

Hemoglobin H inclusion

RBC Inclusions

• Hemoglobin H inclusion

Hemoglobin H disease

RBC Inclusions

• hexagonal, rod-shaped crystals

• bar of gold appearance

Hemoglobin C crystal

RBC Inclusions

• finger-like or quartz like crystal of dense hemolgobin protruidng from RBC membrane

Hemoglobin SC crystals

RBC Inclusions

• Hemoglobin SC crystals

Hemoglobin SC disease

RBC Inclusions

• NRBCs in bone marrow with dots surrounding th nucleus

• Content: Non-heme iron

Ringed sideroblast

RBC Inclusions

• Ringed sideroblast

• Sideroblastic anemia

• Myelodysplastic syndrome