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Formulas and Conversion Factors

• Rule of Three: This rule is used to check the accuracy of hemoglobin and hematocrit results. The formula is: $(Hgb \times 3) = Hct \pm 3 (0.03 L/L)$

• Hemoglobin (HGB) Conversion Factors

  • $1 g/L = 10 g/dL$
  • $1 mm^3 = 10^{12} g/dL$

Hematocrit Determination

• Hematocrit Formula:

  • $Hct\% = \frac{[RBC \times MCV]}{10}$

  • Where:

    • Hct is Hematocrit.
    • RBC is Red Blood Cell count.
    • MCV is Mean Corpuscular Volume.

• Hematocrit (HCT) Conversion Factors:

  • Convert to percentage by multiplying the L/L value by 100%.

Red Blood Cell Indices

• MCV (Mean Corpuscular Volume): Represents the average volume of a red blood cell, expressed in femtoliters (fL).

  • $MCV = \frac{Hct\% \times 10}{RBC \ count} = fL$

    • Reference values for MCV (size):
      • Microcytic: < 80 fL
      • Macrocytic: > 100 fL
      • Normocytic: 80 – 100 fL

• MCH (Mean Corpuscular Hemoglobin): Represents the average weight of hemoglobin per red blood cell, expressed in picograms (pg).

  • $MCH = \frac{Hgb (g/dL) \times 10}{RBC \ count} = pg$
  • Reference values for MCH (size-color):
    • Microcytic, hypochromic: < 27 pg
    • Macrocytic, hyperchromic: > 31 pg
    • Normochromic: 27 – 31 pg

• MCHC (Mean Corpuscular Hemoglobin Concentration): Represents the average concentration of hemoglobin within red blood cells, expressed in g/dL.

  • $MCHC = \frac{Hgb (g/dL) \times 100}{Hct} = g/dL$
  • Reference values for MCHC (color):
    • Hypochromic: < 31 g/dL
    • Hyperchromic: > 36 g/dL
    • Normochromic: 31 – 36 g/dL

Other Indices

• Color Index (C.I.):

  • An approximation of MCH.
  • $C.I. = \frac{(Hemoglobin \ g/dL \times 6.9)}{(RBC \ count \ (in \ millions/\mu L) \times 20)}$
  • Normal Value (N.V.): 0.9-1.1

• Volume Index (V.I.):

  • An approximation of MCV.
  • $V.I. = \frac{MCV \ of \ patient}{Average \ Normal \ MCV}$
  • $V.I. = \frac{(Hematocrit \times 2.3)}{(RBC \ count \ (in \ millions/\mu L) \times 20)}$
  • Normal Value (N.V.): 0.9-1.1

• Saturation Index (S.I.):

  • An approximation of MCHC.
  • $S.I. = \frac{C.I.}{V.I.}$
  • Normal Value: 0.80-1.20

• Mean Corpuscular Average Thickness (MCAT):

  • $MCAT = \frac{MCV}{\pi (MCD/2)^2}$
  • Normal Value: 1.7 to 2.5 microns.

Cell Counts

• Total Cell Count Formula:

  • Total \ cell \ count = # \ cells \ counted \times DF \times VF

• RBC Count:

  • Dilution Factor (DF) = $\frac{101 - 1}{0.5} = 200$
  • Volume Factor (VF) = $\frac{1}{5 (0.2 mm \times 0.2 mm \times 0.1 mm)} = 50$
  • Reference Values:
    • Men: 4.20 x 10^6 to 6 x 10^6 / μL
    • Women: 3.80 x 10^6 to 5.20 x 10^6 / μL

• WBC Count:

  • Dilution Factor (DF) = $\frac{11 - 1}{0.5} = 20$
  • Volume Factor (VF) = $\frac{1}{4 (1 mm \times 1 mm \times 0.1 mm)} = 2.5$

• Corrected WBC Count:

  • Corrected \ WBC = \frac{Uncorrected \ WBC \times 100}{# \ NRBC + 100}
  • Reference Values:
    • Adults: 3.60 x 10^6 to 10.6 x 10^6 / μL

• Eosinophil Count:

  • Total \ cell \ count = \frac{# \ cells \ counted \times DF}{Area \ (36 mm^3) \times depth \ (0.1 mm)}

• Reticulocyte Count:

  • \ RRRR = \frac{# \ of \ reticulocytes}{1000 \ RBC} \times 100
  • Reference Values: 0.5 to 2.5%

• Corrected Reticulocyte Count (CRC):

  • $CRC = (Reticulocyte\%) \times \frac{(Patient's \ Hct\%)}{45\%}$
  • Reference Values: 3 to 5%

• Reticulocyte Production Index (RPI):

  • $RPI = \frac{CRC}{Maturation \ time}$
  • Reference values: In anemic patients, RPI should be >3

• Absolute Reticulocyte Count (ARC):

  • $ARC = \frac{Reticulocyte\%}{100} \times RBC \ count$
  • Reference Values: 20 to 115 x 10^9/L

Terminologies

• Hemogram:

  • A consultative report, the same as a CBC (Complete Blood Count) report.

• MCV: Represents the volume of red cells; a decrease in MCV also means a decrease in RBC.

• MCH: Reflects the hemoglobin content of an individual red cell.

• Normal Red Cell Regulation: Under normal circumstances, red cell production and the circulating red cell mass (RCM) remain at a constant level regulated by erythropoiesis, which functions to meet the body’s oxygen requirement.

• Anemia:

  • Associated with decreased red cells.

• Erythrocytosis and Polycythemia: Designate conditions involving the presence of too many red cells in the circulation.

• Goal in Hematology Lab: Prompt detection and recognition of anemic or polycythemia states so that a definitive diagnosis can be sought and proper treatment will be given.

Absolute vs. Relative Anemia/Polycythemia

Anemia

• Absolute Anemia: There is a true decrease in the RCM (Red Cell Mass).
• Relative Anemia:
  • Means that there is a fluid shift from the extravascular to the intravascular compartment leading to expansion of plasma volume and diluting RCM.
  • Related conditions: pregnancy and diseases associated with hyperproteinemia.

Polycythemia

• Absolute Polycythemia: True increase in the RCM.
• Relative Polycythemia:
  • Decrease in the plasma volume in which the RCM is normal.
  • Encountered with dehydration.

Note: Relative anemia or erythrocytosis is not true hematologic disorders; they must be differentiated from conditions involving an absolute change in the RCM.

Anemia and Polycythemia

Anemia

• Decreased hemoglobin levels.
• Physiologic consequences and symptoms are the direct result of the decreased oxygen-carrying capacity of the blood.
• Hematocrit (packed red cell volume) outside the normal range.

Polycythemia

• Primary consequences include hypervolemia and hyperviscosity.

• Involves an increased number of circulating red cells, and thus an increased RCM is described as erythrocytosis or polycythemia.

• Erythrocytosis related to increased red cell number, hemoglobin content, hematocrit, or some combination thereof.

  Hematocrit greater than 0.52 L/L in men and 0.50 L/L in women is used as a criterion.

  • Relative Erythrocytosis:

    • There is not a true increase in RCM, but rather an apparent increase attributable to a decrease in plasma volume.

  • Absolute Erythrocytosis:

    • Refers to a true increase in RCM and is associated with various causes.

    • Relative erythrocytosis is caused by a decline in plasma volume that can mimic the appearance of an elevated RCM in relation to total blood volume rather than a true increase in RCM.

Disorders Characterized by Increased Red Cell Concentration

Symptoms of Erythrocytosis Include:

• General malaise
• Dizziness and headache
• Full feeling in the head
• Tinnitus
• Bleeding
• Thrombosis

Types of Erythrocytosis

1. Relative Erythrocytosis

• Decreased plasma volume causing relative erythrocytosis may be the result of dehydration secondary to:
  • Diarrhea
  • Vomiting
  • Excessive sweating
  • Increased vascular permeability
  • The use of diuretics
• Also seen in individuals with anxiety or stress, which is called stress syndrome.
• Also seen in spurious erythrocytosis or Gaisbock syndrome.
• Associated with smoking, recognized as tobacco polycythemia.

2. Absolute Primary Erythrocytosis

• Absolute increase in RCM resulting from a clonal pluripotent stem cell disorder seen in polycythemia vera.
• Polycythemia vera:
  • One of a group of chronic myeloproliferative disorders
  • Uncontrolled proliferation of bone marrow elements.
  • Erythrocyte production is not controlled by EPO levels, as reflected by the fact that EPO is decreased in Polycythemia Vera.

3. Absolute Secondary Erythrocytosis (Appropriate)

• Caused by:

  • High altitude adjustment

    • Low pO2 of the air, which reduces arterial O2 saturation
    • Monge’s disease or chronic mountain sickness seen in individuals who fail to adapt to living at high altitudes

  • Pulmonary disease

  • Cardiovascular disease

  • Alveolar hypoventilation

  • Defective oxygen transport

• Physiologic compensation includes:

  • Increased blood volume
  • Increased O2 carrying capacity of blood
  • Increased levels of 2,3-DPG to reduce hemoglobin affinity for O2 and increased cardiac output
  • Pulmonary functions

• Other conditions associated with erythrocytosis include:

  • Chronic obstructive pulmonary diseases or COPD (occasionally)
  • Cardiac disease (e.g., congenital and acquired) (occasionally)

Absolute Erythrocytosis

• Primary erythrocytosis:
  • True increase in RCM associated with a myeloproliferative disorder known as Polycythemia Vera.
  • Increase in the RCM is the result of unregulated red cell production.
• Secondary erythrocytosis:
  • A. Inappropriate response: Increased generation of EPO is the result of localized renal hypoxia or tumor generation of a substance that mimics the action of EPO.
  • B. Appropriate response: More EPO is generated in an attempt to alleviate hypoxia through stimulation of red cell production.

4. Absolute Secondary Erythrocytosis (Inappropriate)

• Not associated with generalized hypoxia.
• Seen most often in association with a variety of urologic disorders, including:
  • Tumors
  • Renal artery stenosis
  • Pyelonephritis
  • Urethral obstruction
  • Renal cystic disease
• Extra-renal tumors such as tumors in:
  • Brain
  • Liver
  • Ovary
  • Uterus
  • Prostate
  • Thymus
  • Adrenal glands
• Rarely, an idiopathic type or unknown cause can occur.
• Anemias can be categorized into four groups:
  • I. Decreased or ineffective bone marrow erythrocyte production
    • a. hypoproliferative
    • b. maturation disorders
  • II. Increased red cell destruction or blood loss
    • a. hemolytic disorders
    • b. blood loss

Physiologic Responses to Anemia

• These responses can be a result of a decrease in the oxygen-carrying capacity of the blood and subsequent hypoxia. The body normally attempts to compensate.

Chemical and Physical Responses

• There are three compensatory mechanisms:

  • 1. Shift to the right in the oxyhemoglobin dissociation curve and an increase in red cell 2, 3 diphosphoglycerate (2,3-DPG), which increases the release of oxygen to the tissues by hemoglobin.
  • 2. Selective redistribution of blood flow to areas of highest oxygen demand.
  • 3. Increase in cardiac output.

• Mild to Moderate Anemic States: These three mechanisms together are effective in maintaining the oxygen pressure close to normal levels, and the patient remains asymptomatic.

• More Severe Anemia: Leads to increased cardiac output and greater cardiac stress, leading to the manifestation of signs such as tachycardia.

Hematologic Response

• A slower but more effective response to anemia involves the triggering of increased red cell production.
• Tissue hypoxia resulting from anemia normally leads to increased erythropoietic stimulation of bone marrow.
• The kidneys are sensitive to the decreased oxygen tension, thus prompting increased production of erythropoietin.
• Erythropoietin (EPO):
  • Stimulates the bone marrow to increase the production of erythroid precursors.
  • Increases their rate of proliferation and maturation.
  • Accelerates their release from the bone marrow.
• As a result, shift reticulocytes or immature red cells are seen in the peripheral blood, causing an increased reticulocyte count and increased reticulocyte production index (RPI).
  • The RPI indicates whether or not the bone marrow is responding adequately to anemia.
  • A 6-8-fold increase is notable if the bone marrow is capable of responding to anemia; however, it takes at least 1 week for a full response to manifest.
  • If the marrow fails to respond, it might indicate:
    • An intrinsic disease.
    • Lack of essential hematopoietic factors.
    • A failure in the erythropoietic mechanism itself.

Clinical Manifestations of Anemia

• Mild anemic states often do not show any physical manifestation because of the compensatory mechanism of the body; but during extraneous physical activities, it manifests as dyspnea and palpitations.
• In severe conditions, anemia may cause tachycardia and shortness of breath and headaches.
• Other manifestations include pallor, leg cramps, dizziness, fatigue, and insomnia (which are all common as anemia progresses to secondary tissue hypoxia).
• In the most severe form, it may lead to death or coma.

Classification of Anemias

• A. Etiologic Classification:
  • Pertains to the principal underlying pathophysiologic mechanisms of the disorders leading to anemia.
  • Rather difficult, however, since anemias are often caused by several factors.
• B. Physiologic Classification:
  • Ability of the bone marrow to respond to anemia with increased erythropoiesis.
  • Mostly involves the assessment of erythrocyte production using the reticulocyte count and calculated RPI.
  • If the bone marrow is capable of responding to anemia, an increase in shift reticulocytes can be observed.
  • Shift reticulocytes are young polychromatophilic red cells released prematurely from the marrow because of EPO stimulation; therefore, it is a term reflecting their shift from the bone marrow to the peripheral blood.
  • An RPI greater than 3.0 indicates an effective bone marrow response, characteristic of hemolytic anemias and anemias secondary to blood loss.
  • RPI of less than 2.0 suggests an ineffective bone marrow response, associated with hypo proliferative anemias and anemias resulting from maturation disorders.
• C. Morphologic Classification:
  • Established using red cell indices and microscopic examination of red cell morphology.
  • The indices Mean Corpuscular/Cell Value (MCV), Mean Corpuscular/Cell Hemoglobin (MCH), and Mean Corpuscular/Cell Hemoglobin Concentration (MCHC).

Anemia of Bone Marrow Failure and Systemic Disorders

• A result of an absolute failure of the marrow to replace those erythrocytes that are normally destroyed after 120 days or those that are prematurely destroyed, such as by hemolysis.
• The failure may be the result of a primary defect in the marrow itself, which occurs in aplastic anemia and pure red cell aplasia.
• Normal bone marrow myeloid cells may also be replaced by metastatic tumor, resulting in myelophthisic anemia with altered hematopoiesis.
• There are also a number of systemic diseases, including those that affect the renal and endocrine systems, that can result directly in a secondary decrease in the absolute number of erythroid precursors in the marrow.
  • A factor usually has some erythropoietic stimulatory effect.
  • The bone marrow in these cases is functionally normal; however, anemia may develop.

Aplastic Anemia

• A condition presents with pancytopenia.
• Pancytopenia: Defined as a decrease in all cellular constituents in the bone marrow.
• The bone marrow is severely hypoplastic or aplastic.
• The name of this disorder is misleading, as it implies that anemia is the primary problem experienced by these patients; however, their most serious clinical problems relate to neutropenia and thrombocytopenia.

Age, Incidence and Demographics

• Incidence of aplastic anemia is low before the age of 1 year, then increases at an intermediate rate until the age of 50, after which the incidence is highest.
• Marrow aplasia is two to five times more frequent in the far East than in either North America or Europe.

Classification

• Can be categorized into primary and secondary types.
• The primary type has no known precipitating factor, which includes congenital or hereditary Fanconi anemia or idiopathic aplastic anemia.
• Secondary aplastic anemias have a number of identified causative factors and agents.

Primary Aplastic Anemia: Fanconi Anemia (Congenital Aplastic Anemia)

• Rare inherited form of aplastic anemia, first reported in three brothers by Fanconi, after whom the disease was named.
• Autosomal recessive.
• Characterized as a pancytopenic disorder with a hypoplastic to aplastic bone marrow.
• Clinical Presentation:
  • Prominent congenital abnormalities include:
    • Microencephaly
    • Brown skin pigmentation
    • Short stature
    • Malformations of the thumbs
    • Internal strabismus
    • Malformations of the kidney
    • Genital hypoplasia
    • Mental retardation
• Laboratory Findings:
  • Bone marrow is considered hypoplastic. In the blood picture or blood smear, anisopoikilocytosis (the combination of both abnormal variation in shape and size) is seen, as well as relative reticulocytosis, which is increased in the number of reticulocytes.
  • Manifests leukoerythroblastosis, which is pathognomonic for Fanconi anemia.
  • Increased number of immature WBC.
  • Occurrence of nucleated RBC.
  • Other laboratory tests include:
    • Elevated Osmotic Fragility Test (OFT)
    • Elevated EST
    • Elevated Hgb F (persistent because Hgb A is not synthesized)

Secondary Aplastic Anemia: Etiology and Pathophysiology

• Drugs and chemicals
• Radiation
• Abnormal immune mechanisms
• Various other factors

A. Drugs and Chemicals

• Chloramphenicol is a drug associated with marrow aplasia.
• Transient marrow hypoplasia after treatment with chloramphenicol is fairly common; associated with the appearance of vacuolated cells in the bone marrow, especially among the erythroid series.
• Sometimes, a more serious persistent marrow aplasia follows Chloramphenicol therapy.
• Other drugs include:
  • Benzene and benzene derivatives
  • Hydantoins
  • Sulfonamides
  • Gold preparations
• Insecticides
  • Chlordane
  • Chlorophenotane
  • Gamma benzene hexachloride

B. Radiation

• Long-term low-dose irradiation has an increased incidence of aplastic anemia.
• Causes damage to stem cells or the hematopoietic microenvironment.
• It is likely that damage to the microenvironment is reversible, as indicated by successful marrow transplants in aplastic patients.
• High-dose radiation may have unsuccessful bone marrow transplants, which is associated with acute exposure such as radiotherapy, radioactive isotope administration, and work in unsafe nuclear power plants.

C. Immune Mechanism

• Some support for an autoimmune mechanism as the cause of aplasia, either directly by lymphocytes or by some humoral factor.
• Human natural killer (HNK) cells inhibit growth of hematopoietic cells in vitro; therefore, lymphocytes from the patients no longer suppress in vitro.
  • Are responsible for lymphocyte-mediated aplasia.

D. Miscellaneous Etiologies

• Infections, most commonly non-A, non-B hepatitis, cause aplastic anemia.
• These patients usually have not had severe hepatitis, and the infection has usually resolved by the time the aplasia develops several months later.
• Other reported cases have been associated with military tuberculosis, brucellosis, and parasitic infestation.
• Aplastic anemia can also be seen in paroxysmal nocturnal hemoglobinuria (PNH).
• Clinical Presentation:
  • Symptoms are directly related to pancytopenia.
  • If severe, the typical symptoms occur.
  • Decreased neutrophils result in an increased incidence of bacterial infections;
  • Hemorrhage may be seen as a consequence of thrombocytopenia.
• Laboratory and Correlations with Diseases:
  • The hallmark for secondary aplastic anemia is decreased blood cell counts.
  • Peripheral blood smear (PBS) is decreased with notable reticulocytosis.
  • Reticulocyte production index is also decreased.
  • LAP score or alkaline phosphatase is increased
  • Serum iron is elevated because there is no cell production.
  • Erythropoietin is in the normal to elevated range.
  • Hams Acidified Serum (HAS) test is positive
    • 1.Principle: complement is activated by the alternate pathway, binds to RBC’s and lyses PNH cells which are sensitive to complement activation.
    • 2.Positive HAS test also occurs in the rare congenital dyserythropoeitic anemia type II, also known as Hereditary Erythroblastic Multinuclearity with Positive Acidified Serum lysis test (HEMPAS).

Pure Red Cell Aplasia (PRCA)

• Rare condition that may be inherited or acquired as a primary or secondary disorder.
• The inherited form is known as Diamond Black Fan anemia.
• The acquired form is seen primarily in individuals older than 40 years of age, and is characterized by a severe anemia and normal to slightly decreased peripheral blood leukocyte and platelet counts.

Pathophysiology

• Idiopathic or an immune mechanism.
• Some patients have an immunoglobulin inhibitor of erythroid precursor such that in vitro incubation of a patient’s own serum and marrow cells inhibits erythroid growth.
  • In the absence of the patient’s serum, the patient’s erythroid cells do grow.
• Patients have increased erythropoietin levels.
• Much less common with respect to an immune etiology is an inhibitor of erythropoietin.
• It can also be secondary to a number of agents like benign thymomas.

Clinical Presentation

• Extreme pallor
• Splenomegaly and hepatomegaly
• Anemia develops insidiously, and the onset is so gradual that the patient effectively compensates; thus, by the time a patient presents with symptoms, the anemia is usually severe.

Laboratory Findings and Correlations with Disease

• Severe normocytic normochromic anemia in which the red cells came from the liver and spleen and not in the bone marrow.
• Reticulocyte is severely decreased and can reach as low as zero.
• Bone marrow is normal
• Other hemocyte count is normal.

Diamond Black Fan Anemia (Congenital Pure Red Cell Aplasia)

• Rare congenital disorder
• Normochromic normocytic anemia
• Normal leukocyte and platelet count
• A marked decrease in marrow erythroblasts.

Etiology

• Unknown
• Clinical Presentation: Patients less than 14 years old presents with mental and sexual retardation.
• Laboratory Findings and Correlations with Disease:
  • Hemoglobin values are extremely low, ranging from 1.7-9.4 g/dl.
  • Normocytic normochromic with no reticulocyte production and no nucleated red cell
  • Reticulocyte production index is decreased.
  • Hb F is elevated
  • Bone marrow is cellular
  • Erythropoietin is elevated.

Acute Acquired Erythropoietic Hypoplasia

• Caused by Parvovirus B19 infection, the manifestation of which is the absence of cell production for 1-2 weeks.
• Self-limiting condition, allowing normal erythropoiesis to reoccur after the infection.

Chronic Acquired Erythropoietic Hypoplasia

• Otherwise known as Chronic acquired erythroblastophthisis.
• Caused by thymoma and autoimmune etiology, which implicates the T cells in causing aplastic anemia.

Congenital Dyserythropoeitic Anemias (CDA)

• Rare, hereditary disorders characterized by refractory anemia that varies in severity and abnormalities of bone marrow erythrocyte precursors, including nuclear abnormalities such as karyorrhexis, multinuclearity, and other bizarre changes.
• 1. Type I CDA
  • This may cause neonatal jaundice, but patients have a normal life span.
  • Clinically patients may be diagnosed or present with splenomegaly.
  • Morphologic findings include mild macrocytosis, mild anisocytosis, and mild poikilocytosis.
  • Bone marrow demonstrates binucleated cells, cells with incompletely separated or multilobulated nuclei, megaloblastic nuclear chromatin structure, and erythrophagocytosis.
• A diagnostic feature is the finding of thin, Feulgen-positive, internuclear chromatic bridges joining two normoblasts.
• 2. TYPE II CDA
  • Hereditary erythroblast multi-nuclearity with positive acidified serum test (HEMPAS).
  • Most common CDA.
  • The major laboratory diagnostic findings include abnormal erythrocyte sensitivity to acidified normal serum (similar to that seen in PNH).
  • Clinical findings include splenomegaly and varying degrees of jaundice.
  • Laboratory findings include binuclearity and multinuclearity of 10-40% of erythroid precursors.
  • Negative sucrose hemolysis test (differentiate PNH from HEMPAS).
  • Red cells react with the anti-I and anti-i (I and i antigen is known as HEMPAS antigen).
  • In the PBS normocytic cells are seen with significant diagnostic feature of red cell membrane doubling – they present as ghost cells with double membrane.
• 3. TYPE III CDA
  • Has pronounced multinuclearity of bone marrow normoblasts so that the size is comparable to that of a megakaryocyte.
  • Autosomal dominant pattern.

Hemolytic Anemias

• Anemias may result from either decreased erythrocyte production or increased erythrocyte loss or destruction.
• When the rate of destruction exceeds the bone marrow’s capacity to produce red cells, hemolytic anemia results.
• Since a normal bone marrow can increase its rate of production by as much as six to eight times normal, the red cell lifespan must be significantly shortened for anemia to develop.
• Some anemias result from a combination of decreased production and increased destruction. In megaloblastic anemias and thalassemias, decreased production is the major underlying problem.
• In iron deficiency and chronic disease, may also have a somewhat shortened red cell life span.
  • These anemias are said to have a hemolytic component, but because hemolysis is not the primary underlying cause, they should not be called hemolytic anemia.

Classification

• An intrinsic defect is one in which the patient’s erythrocytes would not survive normally when transfused into a normal recipient; it may affect the cell membrane, metabolic systems, or hemoglobin molecules. Most inherited hemolytic anemias.
• An extrinsic defect is defined as one in which the lifespan of a normal red cell would be shortened if they were transfused into the patient under investigation; extrinsic defects result from abnormal environmental factors that damage normal red cells. Most extrinsic anemias are acquired. Red cells coated with antibodies or complement, or damaged by some abnormal environmental factor or by liver or renal disease, have a shortened lifespan

Intrinsic Hemolytic Anemias

I. Hereditary Anemias of Increased Destruction

• There are three major categories of hereditary disorders causing anemias that are due to abnormal erythrocyte destruction:
  • 1. Intrinsic erythrocyte membrane structural abnormalities (most common and best understood)
  • 2. Erythrocyte enzymopathies (most common and best understood)
  • 3. Extrinsic plasma constituent abnormalities (rare)

A. Hereditary Erythrocyte Enzymopathies

• PYRUVATE KINASE DEFICIENCY (PK)

  • Most common enzyme deficiency of the E-M pathway and is a mild to moderately severe hemolytic anemia.
  • Occurs worldwide and is an autosomal recessive trait.
  • PK mutants are numerous, and most people with hemolytic anemia are doubly heterozygous for two mutant genes
  • Heterozygous PK deficiency generally demonstrates about half the normal PK activity and is not associated with anemia or other hematologic changes.

• Acquired PK deficiency occurs in some cases of dyserythropoietic syndromes.

• Pathophysiology:

  • PK catalyzes the formation of pyruvate from phospho-enol pyruvate, accompanied by the transformation of adenosine diphosphate to ATP.
    • Thus, the PK deficiency leads to a marked reduction in ATP production, which is necessary for the maintenance of the membrane sodium-potassium pump.
  • Irreversible membrane injury resulting from potassium loss and an increase in membrane calcium causes decreased deformability and premature destruction in the spleen.
  • 2,3-diphosphoglycerate accumulates in PK deficiency as a result of the block in glycolysis; the cellular concentration of 2,3-DPG may exceed two to three times normal.
    • This increase is responsible for a shift to the right in the oxyhemoglobin dissociation curve, causing red cells to release oxygen more readily to the tissues to compensate for anemia.

• GLUCOSE 6 PHOSPHATE DEHYDROGENASE DEFICIENCY (G6PD)

  • Most common red cell enzymopathy associated with hemolysis and is inheritance of any one of a large number of abnormal genes that code for the G6PD enzyme.
  • Pathophysiology:
    • Oxidative denaturation of hemoglobin is the major contributor.
    • G6PD is necessary for converting Glucose 6-phosphate to 6-phosphogluconate and for the subsequent production of NADPH and reduced glutathione (GSH).
      • GSH protects enzymes and hemoglobin against oxidation by reducing hydrogen peroxide and free radicals.
    • Hydrogen peroxide is generated in small amounts during normal red cell metabolism and in larger amounts when an oxidant drug interacts with oxyhemoglobin.
    • Normal cells exhibit sufficient G6PD activity to maintain adequate GSH level; when G6PD is deficient, red cells cannot penetrate sufficient GSH to detoxify peroxide.
      • Hemoglobin is then oxidized to methemoglobin.
    • The heme is liberated from globin, and globin denatures, forming Heinz bodies.
      • Heinz bodies attach to membrane sulfhydryl groups, including cell rigidity.
      • Due to cell rigidity, red cells can no longer traverse the splenic microcirculation and lysis occurs.
  • Drugs and chemicals associated with G6PD are primaquine, phenylhydrazine, nitrofurantoin, nalidixic acid, sulfanilamide, methylene blue, and naphthalene.

II. Hereditary Erythrocytes Membrane Abnormalities

• The erythrocyte membrane skeleton is responsible for preserving red blood cell shape and integrity; the normal membrane is composed of equal amounts of lipid and protein.
• Membrane abnormalities that result in hemolytic anemia are due to alterations in structural proteins, caused by a deficiency in proteins or a defective skeletal protein structure.
• Erythrocyte destruction results from loss of membrane surface area, increased rigidity, fragmentation, and deformation.
• Red cells become trapped in the spleen and are prematurely destroyed.
• A. HEREDITARY SPHEROCYTOSIS
  • A hemolytic disorder characterized by numerous microspherocytic erythrocytes on the blood film. Is autosomal dominant, and therefore manifests in heterozygotes.
  • Hereditary spherocytosis is the most common hemolytic anemia in people of Northern European extraction.
  • Pathophysiology:
    • Defect in