RBCs Disorders 2 Dr. Nabila Hamdi MD, PhD
ILOs Discuss the classification of anemia into hypochromic-microcytic, normochromicnormocytic 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 3
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 (protective against malaria) Autosomal codominant conditions. 4
β-thalassemia Genetics: β0: no β-globin chains are produced. β+: reduced (but detectable) β-globin synthesis β chains are encoded by a single gene (2 alleles) on chromosome 11 5
Pathogenesis: β-thalassemia 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. 6
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. Excess of unpaired α chains aggregate into insoluble precipitates, which bind and severely damage the membranes of both RBCs 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. 7
β Thalassemia Microcytic hypochromic anemia 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 8
Management of Thalassemia and Treatment-Related Complications. Rund D, Rachmilewitz E. N Engl J Med 2005;353:1135-1146.
α Thalassemia Genetics: α chains are encoded by two genes (4 alleles) on chromosome 16 Unlike β-thalassemia, α-thalassemia is caused mainly by deletions involving one or more of the α-globin genes www.fpnotebook.com Hydrops fetalis 10
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. 11
Morphology: Thalassemia β-thalassemia major Poikilocytosis (variation in cell size), and anisocytosis (variation in cell shape). β-thalassemia minor and α-thalassemia trait RBCs are regular in shape Peripheral blood smears: microcytic hypochromic anemia For β-thalassemia intermedia and HBH disease, findings lie between these two extremes 12
Thalassemia Morphology: Bone marrow: Hyperplasia of erythroid progenitors due to ineffective erythropoiesis and hemolysis, with a shift toward early forms. Bone complications: the expanded erythropoiesis may completely fill the intramedullary space of the skeleton, invade cortex, impair bone growth, and produce skeletal deformities. Extramedullary hematopoiesis and hyperplasia of macrophages result in prominent splenomegaly, hepatomegaly, and lymphadenopathy. Growth retardation & Cachexia: due to nutrient s consumption by the ineffective erythropoietic precursors. Hemosiderosis and secondary hemochromatosis develop over the span of years, if steps are not taken to prevent iron overload (chelation therapy) 13
Thalassemia Clinical Course: β-thalassemia minor and α-thalassemia trait Often asymptomatic There is usually only a mild microcytic hypochromic anemia (differential diagnosis with iron deficiency!) Normal life expectancy No blood transfusions required β-thalassemia major manifests postnatally as HbF synthesis diminishes. Affected children suffer from growth retardation 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. Systemic iron overload gradually develops due to tranfusion and increased absorption of dietary iron. 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. 14
Diagnosis: Thalassemia 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: Reduced level of HbA (α2β2) HbA2 (α2δ2) level may be normal or increased HbF level might increase β4 tetramers in HbH disease 15
G6PD Deficiency 16
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. 17
G6PD Deficiency Pathogenesis: G6PD deficiency produces no symptoms until the patient is exposed to an environmental factor that produce oxidants. Drugs Antimalarials (primaquine) Sulfonamides Aspirin (in large doses) Vitamin K derivatives Infections Generation of oxidants (Hydrogen peroxide) GSH X GSSG Intravascular and extravascular hemolysis Oxidants attack globin chains 18
G6PD Deficiency Pathogenesis: Oxidized hemoglobin denatures and precipitates, forming intracellular inclusions called Heinz bodies, which can damage the cell membrane sufficiently to cause intravascular hemolysis. Heinz bodies Bite cells 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). 19
G6PD Deficiency 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. 20
Immunohemolytic Anemia Immunohemolytic anemias are uncommon and classified on the basis of (1) the nature of the antibody and (2) the presence of predisposing conditions. 21
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. 22
Immunohemolytic Anemias Warm Antibody Immunohemolytic Anemias Caused by IgG that are active at 37 C. More than 60% of cases are idiopathic (primary). 25% are secondary to an underlying disease of the immune system or are induced by drugs (α-methyldopa, penicillin) Opsonization of RBC by autoantibodies leads to erythrophagocytosis in the spleen. 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. 23
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. When these cells travel to warmer areas, the weakly bound IgM antibody is released, but the coating of C3b remains and cells are phagocytosed by macrophages, mainly in the spleen and liver; hence, the hemolysis is extravascular. 24
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. 25
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). While microangiopathic hemolysis is not usually in and of itself a major clinical problem, it often points to a serious underlying condition. 26
Mechanical Trauma to RBCs Morphology: The morphologic alterations in the injured red cells (schistocytes) are striking and quite characteristic; burr cells, helmet cells, and triangle cells may be seen 27
28
References ROBBINS Basic Pathology 9 th Edition Source of the cover: http://kidney2.blogspot.com/2012_07_01_archive.html 29
Thank you 30