Erythropoiesis and Erythrocyte Physiology Notes

Erythropoiesis and Erythrocyte Physiology

Erythropoiesis is the specific process through which erythroid precursor cells undergo differentiation to eventually become mature red blood cells (RBCs). This process is regulated and stimulated by the hormone Erythropoietin.

The timeline of development involves several distinct phases:

The transition from a pronormoblast to an orthochromic normoblast typically takes between $3$ to $5$ days.
After approximately $1$ day as an orthochromic normoblast, the cell becomes a reticulocyte.
Reticulocytes remain in the bone marrow for $2$ to $3$ days before being released into the systemic circulation.
Once in circulation, the reticulocyte continues its maturation for one additional day before becoming a mature erythrocyte.

Hypoxia serves as the primary stimulus for the production of erythropoietin, which subsequently triggers the erythropoiesis process.

Stages of Erythroid Development

The development of red blood cells follows a specific sequence of stages, often referred to by two different nomenclature systems (Normoblast and Rubriblast systems):

Pronormoblast / Rubriblast
Basophilic Normoblast / Prorubricyte
Polychromatophilic Normoblast / Rubricyte
Orthochromic Normoblast / Metarubricyte
Reticulocyte / Diffusely Basophilic Erythrocyte / Polychromatophilic Erythrocyte
Mature Red Blood Cell / Discocyte

Cellular Characteristics of Erythroid Precursors

Pronormoblast / Rubriblast

Size: $14 - 20\,mm$
Nucleus: Contains fine chromatin and usually has $1$ to $2$ nucleoli.
Cytoplasm: Characterized by a deeply basophilic, nongranular appearance.
N/C Ratio: $8:1$

Basophilic Normoblast / Prorubricyte

Size: $12 - 17\,mm$
Nucleus: Nucleoli are typically no longer visible; the chromatin is slightly coarse.
Cytoplasm: Exhibits intensely basophilic properties.
N/C Ratio: $6:1$

Polychromatophilic Normoblast / Rubricyte

Size: $10 - 15\,mm$
Nucleus: The nuclear volume occupies only half of the total cell area. The chromatin is intensely staining and moderately condensed.
Cytoplasm: Appears blue-gray to pink-gray.
N/C Ratio: $4:1$

Orthochromic Normoblast / Metarubricyte

Size: $7 - 12\,mm$
Nucleus: Contains a small, pyknotic nucleus that is dark, structureless, and nonfunctional.
Cytoplasm: Appears pink.
N/C Ratio: $1:2$

Reticulocyte / Diffusely Basophilic Erythrocyte / Polychromatophilic Erythrocyte

Size: $7 - 10\,mm$
Nucleus: Non-nucleated.
Cytoplasm: Pink to slightly pinkish-gray. It contains a fine basophilic reticulum of RNA, which is only visible when using a Supravital stain such as New Methylene Blue.

Mature Red Blood Cell / Discocyte

Size: $6 - 8\,\mu m$
Nucleus: Non-nucleated.
Cytoplasm: Pink.

Red Blood Cell Disorders: Size and Variation

Anisocytosis refers to a variation in the size of red blood cells. The Red Cell Distribution Width (RDW) is the numerical expression correlating with the degree of anisocytosis.

Normal Value (NV) for RDW: $11.5 - 14.5\%$

Classification by Mean Corpuscular Volume (MCV)

Normocytic: Normal MCV, ranging from $80 - 100\,fL$ .
Microcytic: MCV is less than $80\,fL$ .
Macrocytic: MCV is greater than $100\,fL$ .

Clinical Conditions Associated with Size

Normocytic: Observed in Acute Post Hemorrhagic Anemia, Hemolytic Anemia, and Aplastic Anemia.
Microcytic: Observed in Chronic disease, Thalassemia (which typically presents with a normal RDW), and Iron Deficiency Anemia (associated with an increased RDW).
Macrocytic: Observed in Chemotherapy, Liver disease, Alcoholism, and Megaloblastic anemia (caused by Vitamin $B_{12}$ or Folic acid deficiency).

Red Blood Cell Disorders: Hemoglobin Content

Anisochromia refers to variations in the hemoglobin content or staining of RBCs.

Classification by Mean Corpuscular Hemoglobin Concentration (MCHC)

Normochromic: Normal MCHC, between $31 - 36\%$ .
Hypochromic: MCHC is less than $31\%$ .
Hyperchromic: MCHC is greater than $36\%$ .

Clinical Conditions Associated with Hemoglobin Content

Normochromic: Found in Acute Post Hemorrhagic Anemia, Hemolytic Anemia, and Aplastic Anemia.
Hypochromic: Found in Iron Deficiency Anemia (IDA), Thalassemia, and Chronic Post Hemorrhagic Anemia.
Hyperchromic: Characteristic of Spherocytosis.

Polychromasia or Polychromatophilia refers to a blue-gray and pink coloration of the RBCs, indicating early release from the bone marrow.

Poikilocytosis: Red Blood Cell Shape Abnormalities

Poikilocytosis is the presence of abnormally shaped red blood cells. These are categorized based on the underlying cause.

Poikilocytes Secondary to Developmental Defects

Megalocytes / Macroovalocytes / Macrocytes: Associated with Megaloblastic anemia.

Poikilocytes Secondary to Membrane Abnormalities

Acanthocyte / Spur cell / Thorn cell
Echinocyte / Burr’s cell / Crenated RBC / Sea Urchin cell
Codocyte / Target cell / Mexican Hat cell
Leptocyte: A thinner variant of the Codocyte.
Spherocyte / Bronze cell
Stomatocyte
Elliptocyte
Ovalocyte

Poikilocytes Secondary to Trauma

Schistocyte / Schizocyte / Keratocyte / Helmet cell / Bite cell
Dacryocyte: Teardrop-shaped cell.
Microspherocyte / Pyropoikilocyte
Semilunar bodies / Half-moon / Crescent cell
Blister cells
Knizocyte / Pinch cell

Poikilocytes Secondary to Abnormal Hemoglobin Content

Drepanocyte / Sickle cell

Detailed Comparison of Anemia Types

Type	Cause	Explanation	Characteristics	Example
MEGALOBLASTIC	Vitamin $B_{12}$ or Folic acid deficiency	Nuclear maturation lags behind cytoplasmic maturation. Cells grow larger without dividing.	Oval macrocytes, hypersegmented neutrophils, hyperchromic, macrocytic, hyporegenerative, RBCs with Howell-Jolly bodies	Pernicious anemia
IRON DEFICIENCY	Deficiency of iron	Cytoplasmic maturation lags behind nuclear maturation due to deficiency of iron needed for hemoglobin synthesis.	Microcytic, hypochromic RBCs	Iron deficiency anemia

Identification and Clinical Significance of Poikilocytes

Acanthocyte (Spur cell / Thorn cell)

Associated with:

Abetalipoproteinemia
Pyruvate kinase deficiency
Alcoholic cirrhosis with hemolytic anemia
Malabsorption states
Post splenectomy states
Hepatitis of the newborn
Severe hemolytic anemia associated with cirrhosis and metastatic liver disease

Echinocyte (Burr's cell / Crenated RBC)

Typically associated with anemia related to renal insufficiency.

Codocyte (Target cell / Mexican Hat cell)

Associated with:

Hemoglobinopathies (SS, CC, DD, EE)
Thalassemia
Obstructive liver disease
Post splenectomy states
Iron deficiency anemia

Spherocyte (Bronze cell)

Associated with Hereditary Spherocytosis (HS), Isoimmune and Autoimmune hemolytic anemia, severe burns, and banked blood stored for long periods.

Distinction between HS and Autoimmune/Isoimmune anemias:

Hereditary Spherocytosis: Increased Osmotic Fragility Test (OFT), Increased MCHC, Negative (--) Direct Antiglobulin Test (DAT).
Isoimmune & Autoimmune Hemolytic Anemia: Increased OFT, Increased MCHC, Positive (+) DAT.

Stomatocyte

Associated with Hereditary stomatocytosis, Rh Null disease, Alcoholism, Cirrhosis, and Obstructive liver disease.

Elliptocyte / Ovalocyte

Elliptocyte: Hereditary elliptocytosis. A thinner variant is called a Pencil or Oat cell.
Ovalocyte: Hereditary ovalocytosis, megaloblastic bone marrow, and myelodysplasia.

Schistocyte (Schizocyte / Keratocyte / Helmet / Bite cell)

Associated with Disseminated Intravascular Coagulation (DIC), Thrombocytopenic purpura, burns, and Microangiopathic hemolytic anemia.

Dacryocyte (Teardrop cell)

Associated with Myeloid metaplasia and Hypersplenism.

Microspherocytes / Pyropoikilocytes

Observed in severe burns, Hereditary microspherocytosis, and Hereditary pyropoikilocytosis. A hemolytic blood picture may show polychromasia and microspherocytes.

Semilunar Bodies / Half-Moon / Crescent

Associated with Malaria.

Drepanocyte (Sickle cell)

Associated with Sickle anemia and SC disease.

RBC Morphology Evaluation and Grading

Steininger Sample Criteria (WNL to 4+)

Morphologic Characteristics	WNL	1+	2+	3+	4+
Macrocytes ( $>9\,mm$ )	$0 - 5$	$5 - 10$	$10 - 20$	$20 - 50$	$>50$
Microcytes ( $<9\,mm$ )	$0 - 5$	$5 - 10$	$10 - 20$	$20 - 50$	$>50$
Hypochromia	$0 - 2$	$3 - 10$	$10 - 50$	$50 - 75$	$>75$
Poikilocytosis	$0 - 2$	$3 - 10$	$10 - 20$	$20 - 50$	$>50$
Burr cells	$0 - 2$	$3 - 10$	$10 - 20$	$20 - 50$	$>50$
Acanthocytes	$<1$	$2 - 5$	$5 - 10$	$10 - 20$	$>20$
Schistocytes	$<1$	$2 - 5$	$5 - 10$	$10 - 20$	$>20$
Dacryocytes	$0 - 2$	$2 - 5$	$5 - 10$	$10 - 20$	$>20$
Codocytes	$0 - 2$	$2 - 10$	$10 - 20$	$20 - 50$	$>50$
Spherocytes	$0 - 2$	$2 - 10$	$10 - 20$	$20 - 50$	$>50$
Ovalocytes	$0 - 2$	$2 - 10$	$10 - 20$	$20 - 50$	$>50$
Stomatocytes	$0 - 2$	$2 - 10$	$10 - 20$	$20 - 50$	$>50$
Sickle cells	Absent	Report 1+	--	--	--
Polychromatophilia (Adult)	$<1$	$1 - 6$	$7 - 15$	$15 - 20$	$>20$
Polychromatophilia (Newborn)	$1 - 5$	$5 - 10$	$10 - 20$	$20 - 50$	$>50$
Basophilic stippling	$0 - 1$	$1 - 5$	$5 - 10$	$10 - 20$	$>20$
Howell-Jolly bodies	Absent	$1 - 2$	$3 - 5$	$5 - 10$	$>10$
Siderocytes (Pappenheimer)	Absent	$1 - 2$	$3 - 5$	$5 - 10$	$>10$

Brown RBC Morphology Grading Chart

Graded as 1+ ( $1 - 5$ /field), 2+ ( $6 - 10$ /field), 3+ ( $>10$ /field):

Spherocyte
Teardrop RBC
Acanthocytes
Polychromatophilia
Helmet cells
Schistocytes

Graded as 1+ ( $3 - 10$ /field), 2+ ( $11 - 20$ /field), 3+ ( $>20$ /field):

Stomatocytes
Target cells
Ovalocytes
Poikilocytosis
Burr cells
Elliptocytes
Bizarre-shaped RBC

Graded as Positive only (regardless of count):

Sickle cells
Basophilic stippling
Pappenheimer bodies
Howell-Jolly bodies

Rouleaux Grading

1+: Aggregates of $3$ to $4$ RBCs.
2+: Aggregates of $5$ to $10$ RBCs.
3+: Numerous aggregates with only a few free RBCs.