RBCs Disorders 2 - Faculty of Pharmacy and Biotechnologypbt.guc.edu.eg/Download.ashx?id=559&file=Lecture 12-RBCs Disorders2_559.pdf · β Thalassemia Pathogenesis: • Persons inheriting

RBCs Disorders 2

Dr. Nabila Hamdi

MD, PhD

ILOs • Discuss the classification of anemia into hypochromic-microcytic, normochromic-

normocytic and macrocytic.

• Categorize laboratory test procedures used in the diagnosis of anemia, outlining the basic workup of a patient who presents with anemia.

• Understand the utilization of peripheral blood and bone marrow smears to assess the deviations from normal marrow response which occur in different types of anemia.

• Compare and contrast anemia secondary to acute vs. chronic blood loss.

• Discuss the different types of hemolytic anemia in terms of: genetics - molecular changes, etiology, pathogenesis, morphology, laboratory diagnosis and clinical features and course.

• Compare and contrast warm vs. cold antibody immunohemolytic anemias.

• Compare and contrast intravascular vs. extravascular hemolysis.

• Discuss and contrast the different types of anemia of diminished erythropoesis in terms of etiology and pathogenesis, marrow and peripheral blood morphology, laboratory diagnostic criteria and clinical features and course.

Outline

I. OVERVIEW

II. ANEMIA OF BLOOD LOSS: HEMORRHAGE

III. HEMOLYTIC ANEMIAS 1. Hereditary Spherocytosis

2. Sickle Cell Anemia

3. Thalassemia

4. Glucose-6-Phosphate Dehydrogenase Deficiency

5. Immunohemolytic Anemia

6. Mechanical Trauma to Red Cells

IV. ANEMIAS OF DIMINISHED ERYTHROPOIESIS 1. Iron Deficiency Anemia

2. Megaloblastic Anemias

3. Aplastic Anemia

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Thalassemia

Overview: • Inherited disorders caused by mutations/deletions that decrease

the synthesis of α- or β-globin chains.

• There is a deficiency of Hb and additional RBC changes due to the relative excess of the unaffected globin chain.

• The mutations that cause thalassemia are particularly common among populations in Mediterranean, African, and Asian regions in which malaria is endemic.

• As with HbS, it is hypothesized that globin mutations associated with thalassemia are protective against plasmodium falciparum of malaria.

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Thalassemia

Overview: • A diverse collection of α-globin and β-globin

mutations underlies the thalassemias, which are autosomal codominant conditions.

• The α chains are encoded by two α-globin genes, which lie in tandem on chromosome 16, while the β chains are encoded by a single β-globin gene located on chromosome 11.

• The clinical features vary widely depending on the specific combination of mutated alleles that are inherited by the patient.

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http://www.chem.purdue.edu/courses/chm333/

β Thalassemia

Pathogenesis:

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β0: no β-globin chains are produced. β+: reduced (but detectable) β-globin synthesis

β Thalassemia

Pathogenesis: • Persons inheriting one abnormal allele have β-thalassemia minor

(also known as β-thalassemia trait), which is asymptomatic or mildly symptomatic.

• Most people inheriting any two β0 and β+ alleles have β-thalassemia major.

• Sequencing of β-thalassemia genes has revealed more than 100 different causative mutations, a majority consisting of single-base changes.

• Gene deletions rarely underlie β-thalassemias.

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β Thalassemia

Two mechanisms contribute to the anemia in β-thalassemia: • The reduced synthesis of β-globin leads to inadequate HbA

formation and results in microcytic hypochromic anemia.

• The excess of unpaired α chains that aggregate into insoluble precipitates, which bind and severely damage the membranes of both red cells and erythroid precursors.

• A high fraction of the damaged erythroid precursors die by apoptosis, a phenomenon termed ineffective erythropoiesis.

• The few RBCs that are produced have a shortened life span due to extravascular hemolysis.

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β Thalassemia

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Inappropriate increase in the absorption of dietary iron, which without medical intervention inevitably leads to iron overload. This is caused by inappropriately low levels of hepcidin, which is a negative regulator of iron absorption.

Pathogenesis of β-thalassemia major

Rund D, Rachmilewitz E. N Engl J Med 2005;353:1135-1146.

Management of Thalassemia and Treatment-Related Complications.

α Thalassemia

Pathogenesis:

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Unlike β-thalassemia, α-thalassemia is caused mainly by deletions involving one or more of the α-globin genes

www.fpnotebook.com

α Thalassemia

Pathogenesis: • The severity of the disease is proportional to the number of α-globin

genes that are missing

• With loss of three α-globin genes there is a relative excess of β-globin or (early in life) γ-globin chains.

• Excess β-globin and γ-globin chains form relatively stable β4 and γ4 tetramers known as HbH and Hb Bart, respectively, which cause less membrane damage than the free α-globin chains that are found in β-thalassemia.

• As a result, ineffective erythropoiesis is less pronounced in α-thalassemia.

• Unfortunately, both HbH and Hb Bart have an abnormally high affinity for oxygen, which renders them ineffective at delivering oxygen to the tissues.

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Thalassemia Morphology:

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β-thalassemia major β-thalassemia minor and α-thalassemia

Poikilocytosis (variation in cell size), and anisocytosis (variation in cell shape).

RBCs are regular in shape

Peripheral blood smears: microcytic hypochromic anemia

Thalassemia

Morphology: • The ineffective erythropoiesis and hemolysis result in a striking

hyperplasia of erythroid progenitors, with a shift toward early forms. • The expanded erythropoietic marrow may completely fill the

intramedullary space of the skeleton, invade the bony cortex, impair bone growth, and produce skeletal deformities.

• Extramedullary hematopoiesis and hyperplasia of mononuclear phagocytes result in prominent splenomegaly, hepatomegaly, and lymphadenopathy.

• The ineffective erythropoietic precursors consume nutrients and produce growth retardation and a degree of cachexia reminiscent of that seen in cancer patients.

• Unless steps are taken to prevent iron overload, over the span of years severe hemosiderosis develops.

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Thalassemia Clinical Course: β-Thalassemia minor and α-thalassemia trait are often asymptomatic. There

is usually only a mild microcytic hypochromic anemia, these patients have a normal life expectancy (differential diagnosis with iron deficiency!)

β-Thalassemia major manifests postnatally as HbF synthesis diminishes. Affected children suffer from growth retardation that commences in infancy.

• Repeated blood transfusions improve the anemia and reduce the skeletal

deformities associated with excessive erythropoiesis.

• With transfusions alone, survival into the second or third decade is possible, but systemic iron overload gradually develops.

• Unless patients are treated aggressively with iron chelators, cardiac dysfunction from secondary hemochromatosis inevitably develops and often is fatal in the second or third decade of life.

• When feasible, bone marrow transplantation at an early age is the treatment of choice. 15

Thalassemia Diagnosis: β-thalassemia major can be strongly suspected on clinical

grounds.

Prenatal diagnosis of β-thalassemia is challenging due to the diversity of causative mutations, but can be made in specialized centers by DNA analysis.

Hb electrophoresis, reveals reduced level of HbA (α2β2). HbA2 (α2δ2) level may be normal or increased. HbF level might increase.

HbH disease can be diagnosed by detection of β4 tetramers by electrophoresis.

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G6PD Deficiency

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G6PD Deficiency • Abnormalities affecting the enzymes responsible for the synthesis of reduced

glutathione (GSH) leave RBCs vulnerable to oxidative injury and lead to hemolytic anemias.

• By far the most common of these anemias is that caused by Glucose-6-Phosphate Dehydrogenase (G6PD) deficiency.

• G6PD gene is on the X chromosome. More than 400 G6PD variants have been identified, but only a few are associated with disease.

• One of the most important variants is G6PD A−, which is carried by approximately 10% of black males in the US.

• G6PD A− has a normal enzymatic activity but a decreased half-life.RBC become progressively deficient in enzyme activity and the reduced form of glutathione. This in turn renders older RBCs more sensitive to oxidant stress.

• In other variants such as G6PD Mediterranean, found mainly in the Middle East, the enzyme deficiency is more severe.

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G6PD Deficiency

Pathogenesis:

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G6PD deficiency produces no symptoms until the patient is exposed to an environmental factor that produce oxidants.

Drugs Infections

•Antimalarials (primaquine) •Sulfonamides •Nitrofurantoin • Phenacetin •Aspirin (in large doses) •Vitamin K derivatives

Generation of oxidants

(Hydrogen peroxide)

GSH oxidized glutathione (GSSG) X

Oxidants attack globin chains

G6PD Deficiency

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Pathogenesis: Oxidized hemoglobin denatures and

precipitates, forming intracellular inclusions called Heinz bodies, which can damage the cell membrane sufficiently to cause intravascular hemolysis.

Other, less severely damaged cells lose their deformability and suffer further injury when splenic phagocytes attempt to “pluck out” the Heinz bodies, creating so-called bite cells. Such cells become trapped upon recirculation to the spleen and are destroyed by phagocytes (extravascular hemolysis).

Heinz bodies

Bite cells

G6PD Deficiency

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Clinical Features: • Drug-induced hemolysis is acute and of variable severity. Typically, patients

develop hemolysis after a lag of 2 or 3 days.

• Since G6PD is X-linked, the red cells of affected males are uniformly deficient and vulnerable to oxidant injury.

• In the case of the G6PD A−variant, it is mainly older RBCs that are susceptible to lysis. Since the marrow compensates for the anemia by producing new resistant red cells, the hemolysis abates even if the drug exposure continues.

• In G6PD Mediterranean, the enzyme deficiency and the hemolysis that occur on exposure to oxidants are more severe.

Immunohemolytic Anemias

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Immunohemolytic anemias are uncommon and classified on the basis of (1) the nature of the antibody and (2) the

presence of predisposing conditions.

Immunohemolytic Anemias

• The diagnosis of immunohemolytic anemias depends on the detection of antibodies and/or complement on red cells.

Direct Coombs antiglobulin test: the patient’s red cells are incubated with antibodies against human immunoglobulin or complement. In a positive test result, these antibodies cause the patient’s red cells to clump (agglutinate).

The indirect Coombs test assesses the ability of the patient’s serum to agglutinate test red cells.

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Immunohemolytic Anemias

Warm Antibody Immunohemolytic Anemias Caused by IgG or, rarely, IgA antibodies that are active at 37°C.

More than 60% of cases are idiopathic (primary).

25% are secondary to an underlying disease affecting the immune system (systemic lupus erythematosus) or are induced by drugs (α-methyldopa, penicillin)

The hemolysis usually results from the opsonization of red cells by the autoantibodies, which leads to erythrophagocytosis in the spleen and elsewhere.

Incomplete consumption (“nibbling”) of antibody-coated RBCs by macrophages leads to membrane loss and formation of spherocytes, which are rapidly destroyed in the spleen.

Most patients have chronic mild anemia with moderate splenomegaly and require no treatment.

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Immunohemolytic Anemias

Cold Antibody Immunohemolytic Anemias Usually caused by low-affinity IgM antibodies that bind to RBC membranes

only at temperatures below 30°C, such as occur in distal parts of the body (ears, hands, and toes) in cold weather.

Although bound IgM fixes complement well, the latter steps of the complement fixation cascade occur inefficiently at temperatures lower than 37°C.

As a result, most cells with bound IgM pick up some C3b but are not lysed intravascularly. When these cells travel to warmer areas, the weakly bound IgM antibody is released, but the coating of C3b remains.

Because C3b is an opsonin the cells are phagocytosed by macrophages, mainly in the spleen and liver; hence, the hemolysis is extravascular.

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Immunohemolytic Anemias

Cold Antibody Immunohemolytic Anemias • Binding of pentavalent IgM also cross-links red cells and causes them to

clump (agglutinate).

• Slugging of blood in capillaries due to agglutination often produces Raynaud phenomenon in the extremities of affected individuals.

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Mechanical Trauma to RBCs

Pathogenesis: • Abnormal mechanical forces result in red cell hemolysis in a variety of

circumstances.

• Mechanical hemolysis is sometimes produced by defective cardiac valve prostheses (the blender effect), which can create sufficiently turbulent blood low to shear red cells.

• Microangiopathic hemolytic anemia is observed in pathologic states in which small vessels become partially obstructed or narrowed by lesions that predispose RBCs to mechanical damage (DIC, malignant hypertension, SLE, disseminated cancer).

• While microangiopathic hemolysis is not usually in and of itself a major clinical problem, it often points to a serious underlying condition.

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Mechanical Trauma to RBCs

Morphology:

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The morphologic alterations in the injured red cells (schistocytes) are striking and quite characteristic; “burr cells,” “helmet cells,” and

“triangle cells” may be seen

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References

ROBBINS Basic Pathology 9th Edition Source of the cover: http://kidney2.blogspot.com/2012_07_01_archive.html

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Thank you…

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