[Physiology][Hematology] General Concepts in Hemolytic Anemias

November 18, 2016 Hematology, Physiology and Pathophysiology No comments , , , , , , , , , , , , , , , , , , , , ,

Hemolysis is the accelerated destruction of red blood cells (RBCs), leading to decreased RBC survival. The bone marrow's response to hemolysis is increased erythropoiesis, reflected by reticulocytosis. If the rate of hemolysis is modest and the bone marrow is able to completely compensate for the decreased RBC life span, the hemoglobin concentration may be normal; this is called fully compensated hemolysis. If the bone marrow is unable to completely compensate for hemolysis, the anemia occurs. This is called incompletely compensated hemolysis.

PS (from wikipedia): The Reticulocyte production index (RPI) is a calculated value used in the diagnosis of anemia. This calculation is necessary because the raw reticulocyte count is misleading in anemic patients. The problem arises because the reticulocyte count is not really a count but rather a percentage: it reports the number of reticulocytes as a percentage of the number of red blood cells. In anemia, the patient's red blood cells are depleted, creating an erroneously elevated reticulocyte count.

Calculation of RPI

Step 1 – Reticulocyte Index is calculated using the formla on the left

Step 2 – The next step is to correct for the longer life span of prematurely released reticulocytes in the blood—a phenomenon of increased red blood cell production. This relies on a table:

Hematocrit (%) Retic survival (days) = maturation correction
36-45 1.0
26-35 1.5
16-25 2.0
15 and below 2.5

Step 3 – The Reticulocyte Production Index is calcualted using the formla below:

Hemolysis can be classified as extravascular or intravascular. Extravascular hemolysis, in which erythrocyte desstruction occurs by macrophages in the liver and spleen, is more common. Intravascular hemolysis refers to RBC destruction occurring primarily within blood vessels. The distinction between intravascular and extravascular hemolysis is not absolute because both occur simultaneously, at least to some degree, in the same patient, and the manifestations of both can overlap. The site of RBC destruction in different conditions can be conceptualized to occur in a spectrum between pure intravascular and pure extravascular hemolysis. Some hemolytic anemias are predominantly intravascular, and some are predominantly extravascular. Others have substantial components of both.

To understand better, the hemolytic anemias can be classified according to whether the cause of hemolysis is intrinsic or extrinsic to the RBC.Intrinsic causes of hemolysis include abnormalities in hemoglobin structure or function, the RBC membrane, or RBC metabolism (cytosolic enzymes). Extrinsic causes may be due to a RBC-directed antibody, a disordered vasculature, or the presence of infecting organisms or toxins. In general, intrinsic causes of hemolysis are inherited and extrinsic causes are acquired, but there are notable exceptions.

Hemolysis Due to Intrinsic Abnormalities of the RBC

screen-shot-2016-11-14-at-9-19-43-pmIntrinsic causes of hemolysis include abnormalities of hemoglobin structure or function, the RBC membrane, or RBC metabolism (cytosolic enzymes). Hemoglobin is the oxygen-carrying protein within RBCs. It is composed of four globular protein subunits, called globins, each with an oxygen-binding heme group. The two main types of globins are the alpha-globins and the beta-globins, which are made in essentially equivalent amount in precursors of RBCs. Normal adult hemoglobin (Hb A) has two alpha-globins and two beta-globins (alpha2beta2).

Abnormalities of Hemoglobin

Disorders of hemoglobin can be classified as qualitative or quantitative disorders. Qualitative abnormalities of hemoglobin arise from mutations that change the amino acid sequence of the globin, thereby producing structrual and functional changes in hemoglobin. There are four ways in which hemoglobin can be qualitatively abnormal: (i) decreased solubility, (ii) instability, (iii) altered oxygen affinity, (iv) altered maintenance of the oxidation state of the heme-coordinated iron. Hemolytic anemia (qualitative abnormalities) result from decreased solubility and instability of hemoglobin. Qualitative hemoglobin disorders often are referred to as hemoglobinopathies, even though the term technicially can apply to both qualitative and quantitative disorders.

Quantitative hemoglobin disorders result from the decreased and imbalanced production of generally structurally normal globins. For example, if beta-globin production is diminished by a mutation, there will be a relative excess of alpha-globins. Such imbalanced production of alpha- and beta-globins damages RBCs and their precursors in the bone marrow. These quantitative hemoglobin disorders are called thalassemias.

Abnormalities of the RBC Membrane

Some hemolytic diseases is characterized by abnormal shape and flexibility of RBCs because of a deficiency or dysfunction of one or more of the membrane proteins, which leads to shortened RBC survival (hemolysis).

The RBC membrane consists of a phospholipid-cholesterol lipid bilayer intercalated by integral membrane proteins such as band 3 (the anion transport channel) and the glycophorins. This relatively fluid layer is stabilized by attachment to a membrane skeleton. Spectrin is the major protein of the skeleton, accounting for approximately 75% of its mass. The skeleton is organized into a hexagonal lattice. The hexagon arms are formed by fiber-like spectrin tetramers, whereas the hexagon corners are composed of small oligomers of actin that, with the aid of other proteins (4.1 and adducin), connect the spectrin tetramers into a two-dimensional lattice. The membrane cytoskeleton and its fixation to the lipid-protein bilayer are the major determinants of the shape, strength, flexibility, and survival of RBCs. When any of these constituents are altered, RBC survival may be shortened.

Abnormalities of RBC Metabolism (cytosolic enzymes)

Normal metabolism of the mature RBC involves two principal pathways of glucose catabolism: the glycolytic pathway and the hexose-monophosphate shunt. The three major functions of the products of glucose catabolism in the erythrocyte are (i) maintenance of protein integrity, cellular deformability, and RBC shape; (ii) preservation of hemoglobin iron in the ferrous form; and (iii) modulation of the oxygen affinity of hemoglobin. These functions are served by the regulation of appropriate production of five specific molecules: ATP, reduced glutathione, reduced NADH, reduced NADPH, and 2,3-BPG. Maintenance of the biochemical and structural integrity of the RBC depends on the normal function of >20 enzymes involved in these pathways as well as the availability of five essential RBC substrates: glucose, glutathione, NAD, NAD phosphate (NADP), and adenosine diphosphate (ADP).

  • ATP

The primary function of the glycolytic pathway is the generation of ATP, which is necessary for the ATPase-linked sodium-potassium and calcium membrane pumps essential for cation homeostasis and the maintenance of erythrocyte deformability.

  • 2,3-BPG

The production of 2,3-BPG is regulated by the Rapoport-Luebering shunt, which is controlled by bisphosphoglyceromutase, the enzyme that converts 1,3-BPG to 2,3-BPG. Concentration of 2,3-BPG in the RBC in the RBC in turn regulates hemoglobin oxygen affinity, thus facilitating the transfer of oxygen from hemoglobin to tissue-binding sites.

  • Reduced gluthione

The major function of the hexose-monophosphate shunt is preservation and regeneration of reduced gluthione, which protects hemoglobin and other intracellular and membrane proteins from oxidant injury.

Abnormalities of the glycolytic pathway

Defects in the glycolytic pathway lead to a decrease in the production of ATP or a change in the concentration of 2,3-BPG.

Deficiencies of erythrocyte hexokinase, glucose phosphate isomerase, phosphofructokinase, and pyruvate kinase (PK) all lead to a decrease in ATP concentration. Although genetic disorders involving nearly all of the enzymes of the glycolytic pathway have been described, PK accounts for >80% of the clinically significant hemolytic anemias from defects in this pathway. With the exception of phosphoglycerate kinase deficiency, which is X-linked, all other glycolytic enzyme defects are autosomal recessive.

PK deficiency is the most common congenital nonspherocytic hemolytic anemia caused by a defect in glycolytic RBC metabolism. The syndrome is both genetically and clinically heterogeneous. Both glucose phosphate isomerase and hexokinase deficiencies produce nonspherocytic hemolytic anemia associated with decreased ATP and 2,3-BPG content.

Abnormalities of the hexose-monophosphate shunt

G6PD deficiency is the most frequently encountered abnormality of RBC metabolism, affecting >200 million people worldwide. The gene for G6PD is carried on the X chromosome and exhibits extensive polymorphism. Enzyme deficiency is observed in males carrying a variant gene. Hemolysis in G6PD-deficient RBCs is due to a failure to generate adequate NADPH, leading to insufficient levels of reduced glutathione. This renders erythrocytes susceptible to oxidation of hemoglobin by oxidant radicals, such as hydrogen peroxide. The resulting denatured hemoglobin aggregates and forms intraerythrocytic Heinz bodies, which bind to membrane cytoskeletal proteins. Membrane proteins are also subject to oxidation, leading to decreased cellular deformability. Cells containing Heinz bodies are entrapped or partially destroyed in the spleen, resulting in loss of cell membranes through pitting of Heinz bodies and leading to hemolysis.

Abnormalities of nucleotide metabolism

Pyrimidine-5'-nucleotidase deficiency is an enzymatic abnormality of pyrimidine metabolism associated with hemolytic anemia. The peripheral blood smear in patients with this defect often shows RBCs containing coarse basophilic stippling. Lead intoxication also inactivates the enzyme, leading to an acquired variant of pyrimidine-5'-nucleotidase deficiency.

Adenosine deaminase (ADA) excess is an unusual abnormality. It is caused by a genetically determined increase in the activity of a normal erythrocyte enzyme. The excessive deaminase activity prevents normal salvage of adenosine and causes subsequent depletion of ATP and hemolysis.

ASH Guideline for RBC Transfusion

July 13, 2016 Critical Care, Hematology, Transfusion No comments , , , , , ,

The Guideline

The development of clinical practice guidelines for RBC transfusion has been challenged by a limited availability of high-quanlity evidence to support practice recommendations. There is general agreement that RBC transfusion is typically not indicated for hemoglobin (Hgb) levels of >10 g/dL and that transfusion of RBCs should be considered when Hb is <7 to 8 g/dL depending on patient characteristics. The decision to transfuse RBCs should be based on a clinical assessment of the patient that weighs the risks associated with transfusion aganist the anticipated benefit. As more studies addressing RBC transfusion become available, it becomes increasely clear that liberal transfusion strategies are not necessarily associated with superior outcomes and may expose patients to unnecessary risks.

The most recently published guidelines from the AABB (formerly the American Association of Blood Bank) are based on a systematic review of randomized, controlled trials evaluating transfusion thresholds. These guidelines recommend adhering to a restrictive transfusion stratety and consider transfusion when Hb is 7 to 8 g/dL in hospitalized, stable patients. This strong recommendation is based on high-quality evidence from clinical trials comparing outcomes in liberal versus restrictive transfusion strategies in this patient population. A restrictive transfusion strategy is also recommended for patients with preexisting cardiovascular disease. In this population, transfusion should be considered when Hb levels are <8 g/dL or for symptoms such as chest pain, orthostatic hypotension, tachycardia unresponsive to fluid resuscitation, or congestive heart failure. This weak recommendation is based on moderate-quality evidence due to limited clinical trial data directly addressing this population of patients. Additional clinical practice guidelines exist that specify Hb targets for critical care patients with conditions including sepsis, ischemic stroke, and acute coronary syndrome.

RBC transfusion is indicated in patients who are actively bleeding and should be based on clinical assessment of the patient in addition to laboratory testing. Much remains to be learned about the optimal resuscitation of the bleeding patient. However, a recent study examining transfusion in patients with active upper gastrointestinal bleeding showed superior outcomes in patients treated with a restrictive transfusion strategy (<7 g/dL).

The Physiologic Response to Anemia

The initial response to anemia is a shift in the oxygen dissociation curve to the right as modulated by an increase in production of 2,3-DPG in RBCs. This shift allows for the unloading of oxygen to the tissues at higher partial pressures of oxygen, ensuring adequate oxygen delivery despite the reduction in RBC mass.

As anemia progresses, the cardiac output will increase by an increase in the heart rate to preserve the delivery of oxygen in the setting of decreased oxygen content. As RBC mass is reduced in anemia, the viscosity of the blood decreases. This reduction in viscosity leads to an increase in regional blood flow at the tissue and organ level, driving up local perfusion area and pressures leading to increased oxygen extraction. While a change in viscosity may be the trigger for increased regional blood flow, there has been suggestion that local blood vessel dilatation may be mediated by the release of nitric oxide (NO) from the RBCs. In order for these mechanisms to work properly, the patient must be at or near a euvolemic state. In considering these regulatory mechanisms, it is important to understand that the transfusion of RBCs will incease viscosity by adding stored RBCs that may not have the same vasoactive capabilities of native RBCs. As such, a transfusion of RBCs may inhibit compensatory mechanisms for low oxygen states, without significiantly increasing oxygen delivery.

There is evidence that low levels of Hb can be tolerated in healthy subjects. Hematocrits of 10% to 20% have been achieved in experimental studies using normovolemic hemodilution without untoward effects. Weiskopf and colleagues studied patients who underwent isovolemic reduction of Hb to 7, 6, and 5 g/dL. No evidence of reduced oxygen delivery was detected at any of the tested values of Hb; however, there was a subtle reversible reduction in reaction time and impaired immediate and delayed memory observed at Hb below 6 g/dL. An important source of data regarding the impact of anemia on surgical outcome comes from studies of Jehovah's Witness patients. Carson has demonstrated that the risk of death in these pateints at Hb between 7 and 8 g/dL is low. However, the odds of death increase by 2.5 for each gram decrease in Hb below 8 g/dL. The mortality is very high at Hb levels below 5 g/dL. It should be noted that these data are from patients who refuse all RBC transfusions. There is time to intervene between a low Hb and resulting morbidity or mortality in most patients.