Erythrocyte Disorders Part 2

Anemia

General Information

• From the Greek word anaimia, meaning "without blood."

• Anemia is NOT a diagnosis or disease itself, but a manifestation of an underlying condition.

Physiological Effects of Anemia

• Decreased hemoglobin (Hb) oxygen-affinity, resulting in a rightward shift (Shift-R).

• Tissue hypoxia or decreased tissue perfusion in select body areas.

• Increased cardiac output to maintain adequate tissue oxygen tension.

• Increased RBC production by the marrow due to increased erythropoietin (EPO).

  • Bone marrow (BM) compensation occurs 4-5 days after Hb decrease.

  • Increased reticulocyte and nucleated red blood cell (nRBC) values.

Classification of Anemia

• Can be classified morphologically using RBC indices (MCV, MCHC) or based on etiology/cause.

Erythropoiesis

• Effective: Produces functional cells that leave the marrow and supply the blood with adequate cells.

• Ineffective: Produces defective progenitor cells which are destroyed before leaving the marrow.

• Insufficient: Lack of erythroid precursors in the marrow.

Classification Based on MCV and MCHC

• Microcytic Hypochromic:

  • Iron deficiency anemia (IDA)

  • Anemia of chronic disease (ACD)

  • Thalassemia

  • Lead poisoning

  • Inherited Sideroblastic anemia

  • Acquired Sideroblastic anemia

• Normocytic Normochromic:

  • Acute hemorrhage

  • Hemolytic anemias

  • Marrow hypoplasia (aplastic anemia)

  • Renal disease

  • ACD

  • Myelodysplastic syndrome

• Macrocytic Normochromic:

  • Megaloblastic anemia

  • Nonmegaloblastic anemia

  • Megaloblastoid anemia (myelodysplasia)

• Hyperchromic: (No specific anemias listed in this category in the provided text)

MCV-Based Classification

• Microcytic: MCV < 80 fL

  • Iron deficiency anemia

  • Thalassemia

  • Sideroblastic anemia

  • Anemia of inflammation

  • Hb E disease/trait

• Normocytic: MCV 80-100 fL

  • Retic ↑:

    • Hemolytic anemia

      • Intrinsic (Membrane defects, Hemoglobinopathies, Enzyme deficiencies)

      • Extrinsic (Immune causes, Nonimmune RBC injury: Microangiopathic, Macroangiopathic, Infectious agents, Other injury: drugs, chemicals, venoms, extensive burns)

  • Retic N or ↓:

    • Aplastic anemia

    • Anemia of renal disease

    • Myelophthisic anemia

    • Infection (parvovirus B19)

    • Anemia of inflammation

    • Dilutional anemia of pregnancy

• Macrocytic: MCV > 100 fL

  • Megaloblastic:

    • Vitamin B12 deficiency

    • Folate deficiency

    • Myelodysplasia

    • Erythroleukemia

  • Nonmegaloblastic:

    • Aplastic anemia

    • Chronic liver disease

    • Alcoholism

    • Some drugs

Dimorphic Anemias

• Presence of both hypochromic and normochromic cells in the peripheral blood (Anisochromia).

  • Sideroblastic anemia

  • Weeks after iron therapy for IDA

  • Hypochromic anemia after transfusion with normal cells

Pathophysiologic Categories of Anemia

• Impaired RBC Production

  • Heme Disorders: IDA, Sideroblastic Anemia, Porphyrias

  • DNA Disorders: Megaloblastic Anemia, Nonmegaloblastic Anemia

  • Chronic Disease Anemia

  • Anemia of Renal Insufficiency

  • Anemia of Liver Disease

  • Anemia of Endocrine Disease

  • Myelophthisic Anemia

  • Refractory Anemia

  • Aplastic Anemia

  • Pure Red Cell Aplasia

  • Congenital Dyserythropoietic Anemia

• Increased RBC Destruction

  • Globin Chain Disorders Hemoglobinopathies, Thalassemias

  • Hemolytic Anemias

    • Extrinsic hemolysis: CAIHA, WAIHA, PCH, HTR, HDFN, MAHA

    • Intrinsic hemolysis: H. spherocytosis, H. elliptocytosis, H. stomatocytosis, H. pyropoikilocytosis, H. acanthocytosis, PNH

  • Metabolic Disorders G6PD deficiency, PK deficiency

• Anemia of Blood Loss

Anemias of Impaired RBC Production

I. Heme Disorders

1. Iron Deficiency Anemia (IDA)

• Most common form of anemia.

• Microcytic, hypochromic.

• Develops when iron loss exceeds iron intake.

• Stages:

  • Iron depletion: Depletion of storage iron, decreased serum ferritin, increased Fe absorption, increased TIBC.

  • Iron-deficient erythropoiesis: Anemia not yet detectable, decreased serum iron, decreased transferrin saturation (<15%), decreased sideroblasts, increased protoporphyrin.

  • Iron deficiency anemia: Anemia is detectable, patient experiences nonspecific symptoms.

• Causes:

  • Chronic blood loss: MOST COMMON cause among adult males and postmenopausal women, excessive menstruation, multiple pregnancies, repeated hemorrhage, hookworm infections, hemorrhagic lesions (associated with benign and malignant tumors), MARCH ANEMIA among soldiers and long-distance runners.

  • Dietary deficiencies and increased demand: Infants, children (6-24 months), pregnant women.

  • Defective iron absorption: Gastrectomy, H2 blocker therapy (drug for peptic ulcer), malabsorption syndromes (rare), sprue.

• Lab features:

  • Microcytic, hypochromic blood picture.

  • Pencil cells.

  • ↓ serum iron, ferritin, hgb/hct, RBC, retics ct.

  • ↓ bone marrow M:E ratio, storage iron.

  • ↑ OFT, RDW, and FEP/ZPP.

  • ↑ TIBC.

  • zinc substitutes for iron in binding protoporphyrin if serum iron is low

• Clinical features:

  • Pica: craving to eat unusual substances such as dirt or ice. Adult Pica: IDA. Childhood Pica: Lead poisoning

  • Glossitis, “Strawberry” tongue.

  • Angular stomatitis.

  • Koilonychia: concave or spoon-shaped nails.

  • Difficulty swallowing due to “webs” of tissue or partial strictures at the junction of the esophagus and hypopharynx.

  • Plummer-Vinson syndrome (combination of glossitis, sore mouth, dysphagia, and iron deficiency).

  • Splenomegaly (uncommon), fatigue, paresthesia, chronic gastritis.

2. Sideroblastic Anemia

• Dimorphic anemia

• Caused by enzymatic defects in the protoporphyrin pathway resulting in defective heme synthesis (↓ ALA synthethase activity) & iron overload

• Excess iron accumulates in the mitochondrial region of the immature erythrocytes in the bone marrow and encircles the nucleus → Ringed Sideroblasts

• Mature erythrocytes in the circulation → Siderocytes

  • Ringed sideroblasts: five or more iron granules encircle at least 1/3 of the nucleus of the cell

• Lab features:

  • ↑ serum Fe, ↑ % saturation of iron-binding protein

  • ↑ storage Fe, ↑ erythroid hyperplasia

  • defective hemoglobinization and ↓ TIBC

• Types:

  • Inherited Sideroblastic Anemia:

    • microcytic, hypochromic

    • X-linked, associated with low levels of δ-ALA enzyme

  • Acquired Sideroblastic Anemia:

    • macrocytic

    • RARS and Secondary Sideroblastic Anemia due to lead poisoning, ethanol, chloramphenicol, copper or pyridoxine deficiency and zinc overload

3. Porphyrias

• Group of inherited and acquired disorders of heme biosynthesis caused by a deficiency of a specific enzyme in the biosynthetic pathway

• Accumulation of porphyrin metabolite in the blood

II. DNA Disorders

1. Nonmegaloblastic Anemia

• Macrocytic anemia with normoblastic marrow.

• Caused by early release of erythrocytes from the marrow, so-called SHIFT RETICULOCYTES.

• Occurs in response to:

  • liver disease

  • acute blood loss

  • hemolysis

  • bone marrow infiltration

  • ↑ EPO → bone marrow failure diseases

  • hypothyroidism

  • excessive alcohol intake

2. Megaloblastic Anemia

• Macroovalocytes present

• Giant hypersegmented neutrophils present

• Characterized by maturation arrest or NUCLEAR-CYTOPLASMIC ASYNCHRONY

• characterized by enlargement of ALL rapidly proliferating cells of the body, including marrow cells

• Major abnormality: diminished capacity for DNA synthesis

• Cobalamin (Vitamin B12) Deficiency

  • Intrinsic factor: needed to bind vitamin B12 for absorption into the intestines; secreted by parietal cells

  • Pernicious anemia : caused by deficiency of intrinsic factor, antibodies to intrinsic factor, or antibodies to parietal cells

  • Characterized by achlorhydria and atrophy of gastric parietal cells

  • Vitamin B12 def takes 3-6 years to develop because of high body stores.

  • Causes include:

    • Malabsorption syndromes

    • Diphyllobotrium latum infection

    • Total gastrectomy

    • Total vegetarian diet

  • Clinical features:

    • jaundice, weakness, and CNS problems

• Folic Acid Deficiency

  • Folic acid has low body stores

  • causes a megaloblastic anemia with a blood picture and clinical symptoms similar to vitamin B12 except that there is NO CNS INVOLVEMENT

  • Causes include:

    • Poor diet

    • Pregnancy

    • Chemotherapeutic anti-folic acid drugs such as methotrexate

III. Chronic Disease Anemia

• Microcytic, hypochromic

• associated with chronic nonhematologic disorders leading to defective iron utilization

• causes impaired release of storage iron associated with increased hepcidin levels during inflammation

• Hepcidin: negatively controls the release of iron from cells

• Chronic disease → inflammation → Hepcidin level increases → decreased iron release from macrophage and enterocytes → low serum iron levels → anemia

• SECOND only to IDA as a common cause of anemia

• Lab features:

  • ↑ ESR: due to increased APRs

  • normal to high ferritin

  • ↓ serum iron and TIBC

  • Normal RDW and IBC

  • Hypoproliferative marrow

IV. Anemia of Renal Insufficiency

• Normocytic, normochromic

• Burr cells present

• Lab features:

  • ↑ BUN and creatinine

    • Uremia → acid-base and electrolyte disturbances → BURR cells

  • ↓ EPO → impairs erythropoiesis & BM response to tissue hypoxia → anemia

V. Anemia of Liver Disease

• Shortened RBC survival and inadequate RBC production that occurs secondary to a chronic liver disease.

• Macrocytic or normocytic

• Lab features:

  • Liver disease → increased surface membrane cholesterol → Target cells or Acanthocytes → causing increased RBC destruction by the spleen

  • anemia → splenomegaly and cirrhosis

  • ↑ Retics

  • Normal or ↓ platelet count

VI. Anemia of Endocrine Disease

• 1. Anemia of hypothyroidism

  • Macrocytic or normocytic, normochromic

  • mild-to-moderate anemia w/ normal retics count

  • ↓ thyroid hormone → ↓ metabolic rate and O2 requirement → ↓ EPO by the kidneys → ↓ Erythropiesis → Anemia

• 2. Anemia of androgen deficiency

  • ↓ testosterone secretion → ↓ Erythropoiesis and a decline of approximately 2 \frac{g}{dL} of Hb.

VII. Myelophthisic Anemia

• an anemia associated with bone marrow infiltration and hyperproliferation by nonerythroid cells

• Normocytic, normochromic

• Accompanied by leukoerythroblastic reaction

• Lab features:

  • Normoblasts present in PB with reticulocytosis

  • ↑ WBC with immature forms

  • Normal or ↓ platelets with abnormal forms

• Causes include:

  • Metastatic carcinoma, leukemia, lymphoma

  • Multiple myeloma

  • Lipidoses or storage disease

VIII. Refractory Anemia

• usually occurs in individuals over the age of 50.

• Normocytic or macrocytic

• Lab features:

  • Reticulocytopenia, usually pancytopenia

  • Hypercellular marrow showing erythroid hyperplasia

• Now classified with the myeloproliferative disorders as one of five myelodysplastic syndromes.

• Can develop into an acute leukemia with the presence of blast cells in the peripheral circulation.

IX. Pure Red Cell Aplasia

• an unusual disease characterized by the selective depletion of ONLY the ERYTHROID bone marrow tissue

• 1. Congenital RBC aplasia a.k.a. Blackfan-Diamond anemia

  • Diagnosed between 1-6 years old

  • severe normocytic or slightly macrocytic anemia

  • ↓ retics, ↓ developing erythroid cells EXCEPT PRONORMOBLASTS

  • HbF is elevated as high as 5% to 25%

  • RBC fetal antigen present

• 2. Acquired pure red cell aplasia

  • is rare but occurs more in adults than in children

  • ↓ retics, ↓ developing erythroid cells

  • Up to 50% of patients have an associated cancer of the thymus

  • thought to occur through the production of a cytotoxic autoantibody against erythroid precursors and a plasma inhibitor of heme synthesis.

X. Aplastic Anemia

• a marrow disorder characterized by a reduction in the number or function of multipotential stem cells with a resulting PANCYTOPENIA

• commonly found in adults

• The diagnosis of severe aplastic anemia is made in pancytopenic patients when at least 2 of the following three peripheral blood values are found:

  • WBC count < 500 \frac{cells}{mm^3}

  • Platelet count < 20,000 \frac{cells}{mm^3}

  • Retics count <1%

• Aplastic anemia can be acute and rapidly fatal or it can have a slow onset and a chronic course.

• In 50% of patients, no specific agent can be correlated with the disease. Known causes include:

  • Drugs (33%): Chloramphenicol is the most common cause of drug-induced aplasia (reversible)

  • Chemicals or toxins (4%): mustard compounds, benzene, chemotherapy drugs (e.g., busulfan, urethan), antimetabolite drugs, or ionizing radiation

  • Infections and infectious hepatitis (4%): PNH and pregnancy

  • Miscellaneous causes (59%)

• Clinical features:

  • Bleeding as a result of thrombocytopenia

  • Increased susceptibility to infections due to leukopenia

  • All of the symptoms typical of anemia

  • The absence of splenomegaly

  • Iron overload from repeated transfusions

• The marrow is hypocellular with patchy areas of normal cellularity and increased fat cell infiltration.

• Inherited aplastic anemia a.k.a. Fanconi’s anemia

  • Affects to individuals with a genetic predisposition to bone marrow failure.

  • Detected between 5-10 years old.

  • Often, several family members are affected.

  • Clinical features:

    • Hyperpigmentation and hypogonadism

    • Short stature w/ malformation of the fingers and toes

    • Malformation of the organs

    • Abnormalities of the chromosome pattern of lymphocytes and marrow

XI. Congenital Dyserythropoietic Anemia

• a family of inherited refractory anemias characterized by ineffective erythropoiesis and marrow erythroid multinuclearity.

• 3 Types:

  • CDA-I

  • CDA-II: MORE COMMON

    • distinguished from the others in that it has a POSITIVE acidified serum test and a NEGATIVE sucrose hemolysis test

    • a.k.a. Hereditary Erythroblastic Multinuclearity with Positive Acidified Serum test (HEM-PAS)

  • CDA-III

    • has gigantoblasts (giant erythroid precursors)

Anemias of Increased RBC Destruction

I. Globin Chain Disorders

1. Hemoglobinopathies

• a genetic defect which results in an abnormal structure of one of the globin chains of the hemoglobin molecule

• result from a single amino-acid substitution in one of the polypeptide chains

• affected patients are resistant to malaria

• QUALITATIVE defect of the Hb molecule

  • α-like genes: Short arm of Chromosome 16

  • β-like genes: Short arm of Chromosome 11

• Target cells may be present

• Hb S: Valine replaces glutamic acid at 6th ß chain

  • Hb SS/Sickle cell anemia (Homozygous)

    • Two copies of mutated gene

    • Predominant Hb: Hb S

  • Hb AS/Sickle cell trait (Heterozygous)

    • One copy mutated gene

    • Predominat Hb: Hb A

  • NOTE: Hb S only undergoes polymerization upon deoxygenation!

• Tests for Hb S

  • 1. Sickling Test - Metabisulfite Slide Test

    • Reagent: Metabisulfite (reducing agent)

    • Principle: Additon of metabisulfite enhances deoxygenation of Hb and sickling of Hb S

    • Disadvantages: (1)Does not differentiate sickle cell trait from sickle cell anemia (2)Other rare sickling Hbs (Hb STravis, Hb C Harlem, Hb I) will produce a positive result)

  • 2. Sickle Solubility Test

    • Reagent: saponin (to lyse the RBC); dithionite/sodium hydrosulfite (reducing agent)

    • Principle: Reduced Hb S will increase TURBIDITY of the solution

    • Disadvantages: (1) Presence of other sickling hemoglobins

• Hb C: Lysine replaces glutamic acid at 6th ß chain

• Hb E: Lysine replaces glutamic acid at 26th ß chain

• Hb OArab: Lysine replaces glutamic acid at 121st ß chain

• Hb DPunjab: Glycine replaces glutamic acid at 121st ß chain

• Hb M: Tyrosine replaces proximal or distal histidine in α or ß chain

  • Associated with methemoglobinemia and cyanosis (responds to methylene blue)

• Hb H: ß 4

• Hb Barts: γ 4

2. Thalassemias

• caused by an abnormally long or short polypeptide resulting from genetic errors

• Leads to deficiency or absence of one of the types of globin chains

• With microcytic, hypochromic anemia

• Decrease in synthesis of one or more of the globin chains

• QUANTITATIVE defect of the Hb molecule

• Target cells present

• Types:

  • A. Alpha thalassemia

    • 4 β → Hgb H (fastest migrating)

      • Adult α0

    • 4 δ → Bart’s Hgb

      • Newborn α0

      • results in still birth

      • incompatible w / life → Hydrops fetalis

    • Predominant in Malaria belt

      • Sub-Saharan Africa

      • Mediterranean Basin

      • Middle East, South Asia, and Southeast Asia

| * B. Beta thalassemia
* Predominant in Mediterranean and African regions
* absence of (β0) or a marked decrease in (β+) β-chain production, there is an excess of α-chains
* Decreased Hb A1, Increased Hb A2 (≥ 3.5%) and Hb F

|

• Must know!

  • Hemoglobin constant spring (Hb CS)

    • an α-chain variant with 31 extra amino acids

    • α-Chains are functionally normal but synthesized more slowly

    • present a clinical picture of thalassemia.

II. Hemolytic Anemias

• occurs whenever there is increased RBC destruction and shortened cell survival

• 2 Categories:

  • A. Intrinsic hemolytic anemias

    • usually hereditary and occur from defects in the RBC membrane, metabolism, or the Hb molecule

  • B. Extrinsic hemolytic anemias

    • represent the RBC survival disorders that are acquired and occur secondary to a primary condition or stimulus

• Lab features:

  • ↓ haptoglobin

  • ↑ serum potassium and LD-1

  • ↑ serum CO and carboxyhemoglobin

  • ↑ B1 (2-2.5 mg/dL)

  • ↑ retics and nRBC may also be present

  • ↑ RDW

  • Normocytic, Normochromic

• A. Intrinsic Hemolytic Anemias

  • a. Hereditary spherocytosis (HS)

    • MOST COMMON membrane defect

    • Caused by a defect in SPECTRIN → increased RBC destruction → Splenomegaly

    • Management: Splenectomy

      • Corrects RBC destructrion caused by splenomegaly, NOT the spherocytosis caused by intrinsic spectrin defect

  • b. Hereditary elliptocytosis (HE)

    • Caused by (1) deficiency of protein 4.1 (2) defective spectrin dimer-dimer interaction (3) defective ankyrin-protein 3 interaction

  • c. Hereditary pyropoikilocytosis (HPP)

    • budding and fragmentation of RBCs at 45°–46° C

    • severe congenital hemolytic anemia, which is characterized by microcytosis, striking micropoikilocytosis and fragmentation

  • d. Hereditary stomatocytosis/hydrocytosis/xerocytosis

    • membrane abnormality results in increased permeability of the membrane to Na+, K+ and water, resulting in hydrated, macrocytic red cells

    • MCV may be as high as 150 fL

  • e. Hereditary acanthocytosis

    • abetalipoproteinemia

  • f. Paroxysmal Nocturnal Hemoglobinuria (PNH)

    • The ONLY ACQUIRED, INTRINSIC RED CELL DEFECT

    • the red cell defect renders them more susceptible to C3 binding → C’-mediated intravascular lysis

    • C’ regulatory proteins are decreased or absent in PNH including:

      • Decay Accelerating Factor (DAF, CD55): a glycoprotein that antagonizes the convertase complexes of complement

  • Membrane Inhibitor of Reactive Lysis(MIRL, CD59): a protein that controls the membrane attack complex, C5b-9. * C8-binding protein: a homologous restriction factor

    • Lab features:

      • Hemosiderinuria is almost always present → serious iron loss.

      • Screening test: Sucrose hemolysis screening test

        • Screening tests

          • POSITIVE Sucrose Hemolysis/ Sugar Water Hemolysis Test

  • Confirmatory Tests:
    * POSITIVE Acidified Serum Test/ Ham test (PAST)
    * Flow Cytometry (CURRENTLY USED)

• B. Extrinsic Hemolytic Anemias

  • a. all cause a normocytic, normochromic anemia and are acquired disorders

  • A. IMMUNE DEFECTS

    • Warm Autoimmune Hemolytic Anemia (WAIHA)

      • RBCs are coated with IgG and/or C’

    • Cold Autoimmune Hemolytic Anemia (CAIHA)

      • RBCs are coated with IgM and C’

    • Paroxysmal Cold Hemoglobinuria (PCH)

      • Donath-Landsteiner antibodies (Autoanti-P; IgG) present with biphasic hemolysis

      • Cold temp (<20°C): fixes complement to RBCs

      • High temp (37°C ): C’-coated RBCs lyse

  • Hemolytic Transfusion Reactions

  • Isoimmune Hemolytic Anemia : Hemolytic Disease of the Newborn and Fetus

  • C. NON-IMMUNE DEFECTS

    • Microangiopathic Hemolytic Anemia (MAHAs)

      • group of anemias due to mechanical destruction of RBCs mainly in bone marrow where they are synthesized

    • March Hemoglobinuria

      • transient hemolytic anemia that occurs after a forceful contact of the body w / hard surfaces

    • Iatrogenic anemia: due to excessive blood loss from blood extraction (usually in small children and neonates)

III. Metabolic Disorders

1. Glucose-6-phosphate dehydrogenase deficiency

• MOST COMMON ENZYME DEFICIENCY in the Hexose Monophosphate Shunt

• leads to accumulation of intracellular oxidants which denatures the proteins to become Heinz bodies

• Heinz bodies and BITE CELLS

2. Pyruvate Kinase (PK) deficiency

• MOST COMMON ENZYME DEFICIENCY in Embden-Meyerhof Pathway

• lack of ATP causes impairment of the cation pump

IV. Anemia of Blood Loss

1. Acute Posthemorrhagic Anemia

• characterized by a SUDDEN loss of blood

• Normocytic, normochromic anemia

2. Chronic Posthemorrhagic Anemia

• characterized by a GRADUAL, LONG-TERM loss of blood

• Normocytic, normochromic anemia or microcytic, hypochromic anemia 2° to depletion of iron stores

Laboratory investigation of thalassemia and hemoglobinopathies

CBC, ferritin, FEP to rule out iron deficiency

HPLC or capillary electrophoresis for Hb A2 and Hb F measurements and detection of Hb variants

Perform alkaline and acid gel electrophoresis when a Hb variant is found in HPLC or CE