AN ADVANCED TUTORIAL ON ERYTHROCYTES (RED BLOOD CELLS)

An advanced tutorial on erythrocytes (red blood cells) based on concepts from embryology, their
physiological function, and associated abnormalities, referencing Guyton and Hall Textbook of Medical
Physiology.

Embryology of Erythrocytes
 Stages of Hematopoiesis
 Erythropoiesis begins early in embryonic development and progresses through different sites:

 Mesoblastic Stage (Yolk Sac Stage):

o Begins around the 3rd week of gestation.
o Primitive erythrocytes are produced in blood islands of the yolk sac. These cells are large
and nucleated.
 Hepatic Stage:
o Starts at 6 weeks and continues until birth.
o The liver becomes the primary site of hematopoiesis. Spleen and lymph nodes contribute
as minor sites.
o Erythrocytes are now smaller, and some become enucleated.
 Medullary Stage (Bone Marrow Stage):
o From the 5th month onward, the bone marrow becomes the predominant site of
erythropoiesis.
o Postnatally, hematopoiesis is restricted to the marrow of axial skeleton bones.
o Developmental Hemoglobins
o Embryonic hemoglobins (e.g., Hb Gower-1, Hb Gower-2, and Hb Portland) dominate
early stages.
o Fetal hemoglobin (HbF) appears around 10–12 weeks of gestation and is replaced by
adult hemoglobin (HbA) after birth.

Physiological Function of Erythrocytes

 Erythrocytes are specialized for oxygen and carbon dioxide transport, facilitated by their unique
structure and hemoglobin content.
 Structure and Adaptation
o Biconcave Shape:
o Increases surface area for gas exchange.
o Flexible to pass through narrow capillaries.
o Lack of Organelles:
o No nucleus or mitochondria, optimizing space for hemoglobin.
o Energy is derived anaerobically via glycolysis, preventing oxygen consumption by the
cell itself.

 Hemoglobin Function
o Each erythrocyte contains ~270 million hemoglobin molecules.
o Oxygen Transport:
o Oxygen binds to the iron in heme groups within hemoglobin.
o High affinity in the lungs; low affinity in tissues, allowing efficient unloading.
o Carbon Dioxide Transport:
o Dissolved in plasma (10%), bound to hemoglobin as carbaminohemoglobin (20%), or as
bicarbonate ions (70%).
 Lifespan and Turnover
o Lifespan: ~120 days.
o Senescent erythrocytes are phagocytosed by macrophages in the spleen, liver, and bone
marrow.
o Hemoglobin is broken down into heme (converted to bilirubin) and globin (recycled as
amino acids).
 Abnormalities of Erythrocytes
 Erythrocyte abnormalities are categorized into:
o Quantitative Abnormalities (e.g., anemia and polycythemia).
o Qualitative Abnormalities (e.g., structural or functional defects).
 Quantitative Abnormalities
o Anemia:
o Decrease in erythrocyte count or hemoglobin concentration.
o Causes:
 Decreased production: Iron deficiency, vitamin B12/folate deficiency, bone
marrow failure.
 Increased destruction: Hemolytic anemia (e.g., sickle cell disease, hereditary
spherocytosis).
 Blood loss: Acute or chronic hemorrhage.
 Clinical features: Fatigue, pallor, tachycardia, dyspnea.
o Polycythemia:
o Increase in erythrocyte count.
o Causes:
 Primary: Polycythemia vera (a myeloproliferative disorder).
 Secondary: Chronic hypoxia (e.g., high altitude, COPD) or erythropoietin-
secreting tumors.
 Qualitative Abnormalities
 Hemoglobinopathies:
o Genetic disorders affecting hemoglobin structure or production.
o Examples:
o Sickle Cell Disease: Mutation in the β-globin gene causes hemoglobin
polymerization under hypoxia.
o Thalassemia: Imbalanced globin chain production.
 Membrane Defects:
o Affect erythrocyte flexibility and survival.
o Examples:
 Hereditary Spherocytosis: Defect in ankyrin or spectrin, leading to spherical,
fragile cells.
 Hereditary Elliptocytosis: Defective spectrin assembly.
 Metabolic Abnormalities:
o Affect glycolysis or antioxidant defenses.
o Examples:
 Glucose-6-Phosphate Dehydrogenase (G6PD) Deficiency: Reduces NADPH
production, causing oxidative damage.
 Pyruvate Kinase Deficiency: Impairs ATP generation, leading to hemolysis.
 Regulation of Erythropoiesis
 o Erythropoietin (EPO):
o A glycoprotein hormone primarily produced by renal peritubular cells in response to
hypoxia.
o Stimulates erythroid progenitors in the bone marrow.

 Nutritional Requirements:
o Iron: Essential for hemoglobin synthesis.
o Vitamin B12 and Folate: Required for DNA synthesis during erythropoiesis.
 Feedback Mechanism
o Hypoxia → Increased EPO → Stimulated erythropoiesis → Restoration of oxygen
delivery.
 Key Laboratory Investigations
 Complete Blood Count (CBC):
o Hemoglobin concentration, hematocrit, RBC count, and indices (MCV, MCH, MCHC).
o Peripheral Blood Smear:
o Morphology of erythrocytes (e.g., microcytic, macrocytic, hypochromic).
 Hemoglobin Electrophoresis:
o Identifies abnormal hemoglobins.
 Bone Marrow Aspiration:
o Evaluates marrow cellularity and erythroid precursors.

\Let’s dive into advanced erythrocyte-related calculations and concepts with detailed step-by-step
examples, including oxygen content, oxygen saturation curves, and related physiological parameters.

1. Oxygen Content of Blood

The oxygen content of blood (CO2C_O2CO2) is calculated using the following formula:

CO2=(1.34×[Hb]×SaO2)+(0.003×PaO2)C_O2 = (1.34 \times [Hb] \times SaO2) + (0.003 \times

PaO2)CO2=(1.34×[Hb]×SaO2)+(0.003×PaO2)

Where:

 CO2C_O2CO2: Oxygen content in mL O2 per 100 mL blood

 1.341.341.34: Oxygen-binding capacity of hemoglobin (mL O2 per gram of Hb)
 [Hb][Hb][Hb]: Hemoglobin concentration (g/dL)
 SaO2SaO2SaO2: Arterial oxygen saturation (percentage as a decimal)
  0.0030.0030.003: Solubility coefficient of oxygen in plasma (mL O2 per mmHg per 100 mL
blood)
 PaO2PaO2PaO2: Partial pressure of oxygen in arterial blood (mmHg)

Example Problem:
A patient has the following values:

 Hemoglobin ([Hb][Hb][Hb]): 15 g/dL

 Arterial oxygen saturation (SaO2SaO2SaO2): 98% (0.98 as a decimal)
 Arterial oxygen partial pressure (PaO2PaO2PaO2): 100 mmHg

Calculate the total oxygen content of the blood.

Solution:

1. Oxygen bound to hemoglobin:

1.34×[Hb]×SaO2=1.34×15×0.98=19.71 mL O2/100 mL blood1.34 \times [Hb] \times SaO2 =

1.34 \times 15 \times 0.98 = 19.71 \, \text{mL O2/100 mL
blood}1.34×[Hb]×SaO2=1.34×15×0.98=19.71mL O2/100 mL blood

2. Oxygen dissolved in plasma:

0.003×PaO2=0.003×100=0.3 mL O2/100 mL blood0.003 \times PaO2 = 0.003 \times 100 =

0.3 \, \text{mL O2/100 mL blood}0.003×PaO2=0.003×100=0.3mL O2/100 mL blood

3. Total oxygen content:

CO2=19.71+0.3=20.01 mL O2/100 mL bloodC_O2 = 19.71 + 0.3 = 20.01 \, \text{mL O2/100 mL

blood}CO2=19.71+0.3=20.01mL O2/100 mL blood

Answer: Total oxygen content of the blood = 20.01 mL O2/100 mL blood

2. Oxygen Delivery (DO2)

Oxygen delivery (DO2DO2DO2) is calculated as:

DO2=CO2×CO×10DO2 = C_O2 \times CO \times 10DO2=CO2×CO×10

Where:

 CO2C_O2CO2: Oxygen content (mL O2/100 mL blood)

 COCOCO: Cardiac output (L/min)
 The factor 10 converts mL O2/100 mL to mL O2/L.
Example Problem:
A patient has the following data:

 Oxygen content (CO2C_O2CO2): 20 mL O2/100 mL blood

 Cardiac output (COCOCO): 5 L/min

Calculate the oxygen delivery.

Solution:

DO2=20×5×10=1000 mL O2/minDO2 = 20 \times 5 \times 10 = 1000 \, \text{mL

O2/min}DO2=20×5×10=1000mL O2/min

Answer: Oxygen delivery = 1000 mL O2/min

3. Oxygen Saturation Curve

The oxygen saturation curve (oxyhemoglobin dissociation curve) shows the relationship between
SaO2SaO2SaO2 and PaO2PaO2PaO2. Key points on the curve:

 P50 Value: The PaO2PaO2PaO2 at which hemoglobin is 50% saturated with oxygen (normal =
~27 mmHg).
 The curve shifts right (reduced affinity) or left (increased affinity) based on physiological
factors.

Factors that Shift the Curve:

Right Shift (↓ Affinity) Left Shift (↑ Affinity)

Increased temperature Decreased temperature

Increased CO2 (Bohr effect) Decreased CO2

Decreased pH (acidosis) Increased pH (alkalosis)

Increased 2,3-BPG Decreased 2,3-BPG

Exercise Fetal hemoglobin

Clinical Insight:

 A right shift facilitates oxygen unloading in tissues (e.g., during exercise).

 A left shift promotes oxygen binding (e.g., in fetal hemoglobin or hypothermia).
Example Problem:
A patient in septic shock has the following:

 PaO2PaO2PaO2: 60 mmHg
 SaO2SaO2SaO2: 90%
 Curve analysis suggests a right shift due to lactic acidosis.
 What is the effect on oxygen unloading in peripheral tissues?

Answer: A right shift enhances oxygen unloading in peripheral tissues, ensuring that despite lower
oxygen levels, tissues still receive adequate oxygen supply.

4. Hematocrit and Blood Viscosity

Hematocrit (HctHctHct) is the proportion of blood volume occupied by RBCs. It impacts oxygen
transport and blood viscosity.

Example Problem:
A patient with polycythemia vera has:

 HctHctHct: 60% (normal: ~45%)

 Cardiac output is reduced due to increased blood viscosity.

What is the effect on oxygen delivery (DO2DO2DO2)?

Answer:
While high HctHctHct increases oxygen-carrying capacity, excessive viscosity reduces cardiac output,
potentially compromising oxygen delivery despite higher CO2C_O2CO2.

5. Bohr and Haldane Effects

1. Bohr Effect:
Describes how pH and CO2 influence hemoglobin's oxygen affinity:
o At low pH (high CO2), oxygen affinity decreases, promoting oxygen unloading in
tissues.
o In lungs (high pH, low CO2), oxygen affinity increases, facilitating oxygen loading.

2. Haldane Effect:
Explains how oxygen levels affect CO2 transport:
o Deoxygenated blood binds more CO2 (e.g., in tissues).
o Oxygenated blood releases CO2 (e.g., in lungs).
Example Problem:
Explain why hyperventilation reduces tissue oxygen delivery.

Answer: Hyperventilation causes respiratory alkalosis (high pH), shifting the oxygen saturation curve
left. This increases oxygen affinity, reducing oxygen unloading in tissues.

6. Reticulocyte Production Index (RPI)

RPI assesses bone marrow response in anemia:

RPI=Reticulocyte count (%)×Hematocrit (Hct)Normal Hct÷Maturation factorRPI = \frac{\

text{Reticulocyte count (\%)} \times \text{Hematocrit (Hct)}}{\text{Normal Hct}} \div \text{Maturation
factor}RPI=Normal HctReticulocyte count (%)×Hematocrit (Hct)÷Maturation factor

Example Problem:
A patient has:

 Reticulocyte count: 5%
 Hematocrit: 30% (normal: 45%)
 Maturation factor: 2 (for Hct = 30%).

Calculate the RPI.

Solution:

RPI=5×3045×2=1.67RPI = \frac{5 \times 30}{45 \times 2} = 1.67RPI=45×25×30=1.67

Interpretation: An RPI < 2 indicates inadequate bone marrow response (e.g., hypoproliferative anemia).
 Here’s the answer key to the quiz questions:
 Basic Anatomy and Embryology
 At which embryonic stage does the liver become the primary site of erythropoiesis?
 The hepatic stage, starting at around 6 weeks of gestation.
 Name two hemoglobins present during embryonic development.
 Hb Gower-1 and Hb Portland.

 What is the role of the yolk sac in erythropoiesis?

 It produces the first primitive nucleated erythrocytes during the mesoblastic stage.
 Erythrocyte Physiology
 Why do erythrocytes lack mitochondria?
 To prevent oxygen consumption and maximize oxygen transport efficiency.
 Describe the biconcave shape of erythrocytes and its functional significance.
 Increases surface area for gas exchange and allows flexibility to pass through capillaries.
 What enzyme is responsible for converting CO₂ into bicarbonate within erythrocytes?
 Carbonic anhydrase.
 Pathophysiology
 Explain the mechanism of microcytic anemia in iron deficiency.
 Insufficient iron limits hemoglobin synthesis, leading to smaller and less hemoglobin-rich RBCs.
 What is the genetic mutation responsible for sickle cell disease?
 A substitution of valine for glutamic acid at position 6 of the β-globin gene.
 How does hereditary spherocytosis affect erythrocyte morphology?
 RBCs lose their biconcave shape, becoming spherical and more prone to hemolysis.
 Clinical Presentations
 What are the hallmark features of megaloblastic anemia on a blood smear?
 Macrocytic RBCs, hypersegmented neutrophils, and anisopoikilocytosis.
 Name three triggers for hemolysis in G6PD deficiency.
 Fava beans, sulfa drugs, and infections.
 What are the clinical features of polycythemia vera?
 Plethora, pruritus, splenomegaly, and increased risk of thrombosis.
 Laboratory Diagnostics
 What does a high TIBC indicate in the context of anemia?
 Iron deficiency anemia; it reflects an increased capacity to bind iron.
 How does hemoglobin electrophoresis differentiate between HbA, HbS, and HbF?
 By separating hemoglobins based on their charge; HbS and HbF migrate differently from HbA.
 Which test is used to confirm hereditary spherocytosis?
 Osmotic fragility test.
 Treatment
 What is the mechanism of action of hydroxyurea in sickle cell disease?
 Increases production of HbF, which reduces HbS polymerization and sickling.
 Why is folic acid contraindicated in the presence of untreated vitamin B12 deficiency?
 It can worsen neurological symptoms by bypassing hematologic effects of B12 deficiency.
 Describe the management of acute hemolytic crisis in G6PD deficiency.
 Avoid triggers, provide supportive care (e.g., hydration, oxygen), and, if severe, blood
transfusion.

 Applied Clinical Scenarios

 A patient presents with a hemoglobin of 7 g/dL and glossitis. What two deficiencies should be
considered?
 Vitamin B12 deficiency and folate deficiency.

 Explain how chronic hypoxia can lead to secondary polycythemia.

 Hypoxia stimulates increased erythropoietin (EPO) production, promoting erythrocyte
production.
 Why does carbon monoxide poisoning cause hypoxia despite normal oxygen levels?

 CO binds to hemoglobin with high affinity, reducing oxygen transport capacity.

 Advanced Topics
 How does erythropoietin regulate erythropoiesis in response to hypoxia?
 Hypoxia stimulates renal peritubular cells to release EPO, which promotes erythroid progenitor
proliferation.
 Compare and contrast the oxygen dissociation curves of HbA and HbF.
 HbF has a left-shifted curve compared to HbA, indicating higher oxygen affinity.
 Explain the physiological basis for the cyanosis observed in methemoglobinemia.
 Methemoglobin cannot bind oxygen, leading to reduced oxygen delivery and tissue hypoxia.

 Quiz Continuation (25–50)

 What role does 2,3-bisphosphoglycerate (2,3-BPG) play in oxygen release by hemoglobin?

 It decreases hemoglobin’s oxygen affinity, facilitating oxygen release to tissues.

 Name one drug that causes oxidative stress in G6PD deficiency.

 Primaquine.
 What is the primary diagnostic marker for iron-deficiency anemia?
 Low serum ferritin.
 Describe the role of macrophages in the clearance of senescent RBCs.
 Macrophages in the spleen phagocytose old RBCs and recycle hemoglobin.
 How does high altitude affect erythrocyte production?

 Hypoxia induces erythropoietin release, increasing RBC production.

 Differentiate between absolute and relative polycythemia.
 Absolute: Increased RBC mass; Relative: Normal RBC mass but decreased plasma volume.
 Why is bilirubin elevated in hemolytic anemia?
 Accelerated breakdown of RBCs increases unconjugated bilirubin levels.
 How do reticulocyte counts help differentiate between anemia causes?
 High reticulocyte counts indicate bone marrow compensation for hemolysis or blood loss.

 What is the significance of basophilic stippling?

 Indicates impaired RNA degradation, seen in lead poisoning or thalassemia.
 Describe one therapeutic indication for methylene blue.
 Treating methemoglobinemia.
 Why is a splenectomy performed in hereditary spherocytosis?
 To reduce RBC destruction and anemia severity.
 What is the lifespan of an erythrocyte, and why is this limited?
 ~120 days; lack of organelles limits repair mechanisms.
 How does vitamin C enhance non-heme iron absorption?
 It reduces ferric iron (Fe³⁺) to ferrous iron (Fe²⁺), which is more absorbable.
 8. Explain the role of transferrin in iron metabolism.
 Transports iron in the blood to tissues.
 What is the functional consequence of ankyrin mutation in erythrocyte membranes?
 Loss of membrane stability, leading to hemolysis.
 Define the term "anisocytosis."
 Variation in RBC size.
 Why do patients with thalassemia major often require chelation therapy?
 To manage iron overload from frequent transfusions.
 What causes pruritus in polycythemia vera?
 Increased histamine release due to basophil activation.
 How is erythropoietin synthesized, and which organ regulates its production?
 Synthesized in renal peritubular cells in response to hypoxia.
 What causes macrocytosis in alcoholism?
 Folate deficiency and direct alcohol effects on the marrow.
 Why does carboxyhemoglobin impair oxygen transport?
 It prevents oxygen binding to hemoglobin.
 How is the MCV used to classify anemia types?
 Microcytic (<80 fL), normocytic (80–100 fL), macrocytic (>100 fL).
 Why does bone marrow expand in β-thalassemia?
 To compensate for ineffective erythropoiesis.
 What is the purpose of a Coombs test in hemolytic anemia?
 Detects antibodies against RBCs.
 How do prosthetic heart valves cause hemolytic anemia?
 Shear stress damages RBCs.
 What morphological changes are seen in erythrocytes during severe sepsis?
 Anisocytosis, schistocytes, and possible spherocytes.