MLAB 1415- Hematology Keri Brophy-Martinez Hemolytic Anemia: Immune Anemias

Classification of Immune Hemolytic Anemia’s Autoimmune hemolytic anemia Drug-induced hemolytic anemia Alloimmune hemolytic anemia

Role of Complement Group of serum proteins that interact with each other to bring about complement-dependent, cell-mediated lysis. Can be activated by two pathways:  classical and alternate. Classical pathway activated by antigen-antibody complexes. Alternate pathway triggered by certain microorganisms, polysaccharides, liposaccharides, and cells or particles even in absence of antibody.  Still results in cell lysis. WILL LEARN ABOUT COMPLEMENT IN IMMUNOLOGY.  DO NOT MEMORIZE PATHWAYS NOW!

Mechanisms of Hemolysis Intravascular Results from activation of the classic complement pathway via immunoglobulin IgG or IgM antibodies. IgM more efficient activator of complement Antibodies bind to antigens on red cells and activate complement resulting in lysis of the cell. Extravascular Most common with immune-mediated hemolysis Usually involves IgG Antibody coated or complement coated red cells are fully or partially phagocytized by cells in the reticuloendothelial system (RES), particularly in the spleen and liver. Partially phagocytized cells are seen as spherocytes on the peripheral smear.

Laboratory Findings in Intravascular Hemolysis Hemoglobinemia Hemoglobinuria Haptoglobin Other test results include elevated lactate dehydrogenase (LDH) levels – LDH is enzyme of red cell metabolism. May or may not see reticulocytosis.

Laboratory Findings in Extravascular Hemolysis Will see spherocytes on peripheral blood smear. Serum bilirubin (indirect) may be elevated and urobilinogen concentration may be increased in both urine and stools. Direct Antiglobulin Test (DAT) may be positive, as well as indirect antiglobulin test (antibody screening test).

Laboratory identification of sensitized red cells Two major techniques Agglutination in saline Detects IgM class of antibodies Antihuman globulin(AHG) or Coombs test(DAT) Detects IgG class of antibodies and/or complement Difference in the ability of IgG and IgM to cause agglutination in saline is based on the difference in size of the two antibodies in relation to the zeta potential

RBC Zeta Potential Caused by a difference in the charge density of the inner and outer layers of the ionic cloud around rbcs when they are suspended in saline. The force keeps rbcs about 25nm apart in solution. So any antibody that causes agglutination must be > 25nm to span the gap between cells

RBC Zeta Potential IgM can span 35nm to overcome forces, causing agglutination IgG spans about 14nm, so it can not reach antigens on separate cells to cause agglutination. Therefore, another technique must be used.

Autoimmune Hemolytic Anemias

Autoimmune Hemolytic Anemia (AIHA) Caused by an altered immune response resulting in production of antibody against the patient’s own red cells, with subsequent hemolysis These autoantibodies are directed against high-incidence antigens Mechanism of antibody formation unknown

Warm AIHA (WAIHA) Most common form of AIHA ~70% Reacts best at body temperature (37oC) Can occur at any age, but incidence increases over the age of 40 Cases can be idiopathic or secondary to another disease IgG coats red cells with or without complement fixation Antibody usually directed against antigens of the Rh system

Laboratory findings of WAIHA Common Findings Positive DAT Normocytic, normochromic anemia Increased reticulocytes Spherocytes Presence of autoantibody in serum Increased bilirubin Decreased serum haptoglobin

Cold Agglutinin Disease (CAD) Optimal thermal activity below 30oC Cases can be idiopathic or secondary to another disease Often seen secondary to Mycoplasma pneumoniae (anti-I) or infectious mononucleosis Mechanism of action IgM antibody attaches to erythrocyte in the cold and fixes complement After warming, the antibody dissociates from the cell, but complement remains, causing lysis or initiating extravascular hemolysis Side note: Almost all adults have cold antibodies in low quantity which cause no problems because they react in the temperature range of 4̊ to 22̊ C. The antibody is usually anti-I.

Laboratory findings of CAD Common Findings Positive DAT Normocytic, normochromic anemia False increase in MCV, MCH, MCHC False decrease in RBC count Increased reticulocytes Spherocytes Agglutinated RBCs, rouleaux, nRBCS Cold agglutinin titer high at 4oc Increased bilirubin Decreased serum haptoglobin

Paroxysmal Cold Hemoglobinurea (PCH) Cold hemolytic anemia which can be severe Common in children under age 5 Seen following respiratory viral infections Caused by a bi-phasic complement-fixing IgG antibody( Donath-Landsteiner) Antibody reacts with RBCs in the capillaries at temperatures < 20oC and avidly binds early complement. Upon warming to 37oC , the antibody molecule disperses from the cell, but the membrane attack complement components are activated on the cell membrane causing lysis Treatment is to avoid the cold

Laboratory findings of PCH Common Findings Decreased hgb Hemoglobinemia Hemoglobinuria Spherocytes Increased bilirubin Decreased serum haptoglobin

Drug-Induced Hemolytic Anemias

Drug-Induced Hemolytic Anemia Drugs that attach to the RBC membrane or alter it in some way Methyldopa/ procainamide, penicillin, cephalosporins Patient produces antibodies directed against a particular drug, its metabolites or the RBC coated with drug Acquired The drug itself does not cause the RBC injury, and not all people taking the drug are affected

Alloimmune Hemolytic Anemia

Alloimmune Hemolytic Anemia A foreign antigen stimulates the host’s immune system to produce a corresponding antibody called an alloantibody Two main types Hemolytic transfusion reactions Hemolytic Disease of the Fetus and Newborn(HDFN)

Types of Alloimmune Hemolytic Anemia Hemolytic transfusion reactions Acute transfusion reaction Occurs within minutes- hours after transfusion The wrong ABO type of blood is transfused. If a patient develops symptoms such as fever, shaking, chills, pain or burning at the site of the iv, the transfusion should be stopped immediately. Delayed transfusion reaction Occurs days – weeks after transfusion Sensitization to non-ABO blood groups occurs during a primary transfusion. When re-exposed during a second transfusion, antibodies build up and a delayed reaction occurs. Symptoms are usually mild and non-specific.

Types of Alloimmune Hemolytic Anemia Hemolytic Disease of the Fetus and Newborn (HDFN) This is caused when the mother’s and baby’s blood groups are incompatible and there is an exchange of fetal-maternal blood bfore or during delivery. The mother builds antibodies against the fetal red cells which then become coated with antibody and are destroyed in the baby’s RES. Associated with ABO or Rh incompatibility. It can be severe and treatment would consist of an exchange transfusion of the newborn.

Referenes Harmening, D. M. (2009). Clinical Hematology and Fundamentals of hemostasis (5th ed.). Philadelphia, PA: F.A. Davis Company. McKenzie, S. B. (2010). Clinical Laboratory Hematology (2nd ed.). Upper Saddle River, NJ: Pearson Education, Inc. Rodak, B. F. (2002). Hematology Clinical Principles and Applications (3rd ed.). St. Louis: Saunders Elsevier.