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==Approach to diagnosis of hemolytic anemia==
==Approach to diagnosis of hemolytic anemia==
===Laboratory findings===
* '''Total bilirubin''':
* '''LDH''':
* '''Haptoglobin''':
* '''Reticulocyte count''':
* '''Direct antiglobulin test (DAT)''':
 
* Elevated corrected reticulocyte count will be elevated (>2%), LDH, indirect bilirubin, and low haptoglobin. Haptoglobin is produced in the liver and bind free hemoglobin. During hemolysis, haptoglobin decreases as it is consumed at a faster rate compared to the rate of production by the liver.
====Specific findings for intravascular hemolysis====
* Urine hemosiderin and urine hemoglobin, and microangiopathic blood smear.
====Specific findings for extravascular hemolysis====
* Spherocytic red cell morphology
* Negative urine hemosiderin and urine hemoglobin.
===Peripheral smear===
===Peripheral smear===
Findings on peripheral smears can guide towards diagnosis. Some important findings are:
Findings on peripheral smears can guide towards diagnosis. Some important findings are:
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====Sickle cells====
====Sickle cells====
* Do [[hemoglobulin]] [[electrophoresis]] for [[sickle cell disease]].
* Do [[hemoglobulin]] [[electrophoresis]] for [[sickle cell disease]].
====Bite cells====
* Do G6PD levels
* If G6PD levels decreased then G6PD anemia present. With normal levels diagnosis of unstable hemoglobinopathy should be considered.
====Target cell====
* Order hemoglobulin electrophoresis.
* Electrophoresis helps to differentiate between thalassemia and liver diseases (elevated liver enzymes), and other hemoglobinopathies.
====Schistocytes====
====Schistocytes====
* [[HUS]]-[[TTP]], [[DIC]], [[Prosthetic heart valve]], [[malignant hypertension]]
* [[HUS]]-[[TTP]], [[DIC]], [[Prosthetic heart valve]], [[malignant hypertension]]
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==Laboratory Findings==
==Laboratory Findings==
===Peripheral blood smear===
* Elevated '''total bilirubin''': Elevated bilirubin, or hyperbilirubinemia, is a hallmark of hemolytic anemia
* Fragments of the red blood cells ("[[schistocyte]]s") can be present.
* Elevated '''LDH''':
* Some red blood cells may appear smaller and rounder than usual ([[spherocyte]]s).
* Low '''haptoglobin''': Haptoglobin is produced in the liver and bind free hemoglobin. During hemolysis, haptoglobin decreases as it is consumed at a faster rate compared to the rate of production by the liver.
* [[Reticulocytosis|Reticulocytes]] are present in elevated numbers. This may be overlooked if a special [[staining (biology)|stain]] is not used
* Elevated '''reticulocyte count''':
* The reticulocytes response act as an important indicator for the severity of the hemolytic anemia. The different methods used to judge the correct response are, absolute reticulocyte count, corrected reticulocyte count, and reticulocyte production index.
* Positive '''direct antiglobulin test (DAT)''':
===Reticulocyte count===
 
====Absolute reticulocyte count====
===Specific findings for intravascular hemolysis===
The normal values are:
*Elevated urine hemosiderin
* RBC count - 5 million/microLitre
*Elevated urine hemoglobin
* Reticulocyte count - 0.5 -1.5 %
*Microangiopathic blood smear
* Absolute reticulocyte count - 25,000 to 75,000/microLitre
====Reticulocyte production index====
* [[Reticulocyte]]s are newly-produced red blood cells. They are slightly larger than totally mature red blood cells, and have some residual ribosomal [[RNA]]. The presence of RNA allows a visible blue stain to bind or, in the case of fluorescent dye, result in a different brightness. This allows them to be detected and counted as a distinct population.
* The '''Reticulocyte production index''' (RPI, also called a ''corrected reticulocyte count'') is a calculated value used in the diagnosis of [[anemia]].  
* The corrected absolute reticulocyte count = absolute reticulocyte count / reticulocyte maturation time (in days).
* 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.
* The idea of the RPI is to assess whether the [[bone marrow]] is producing an appropriate response to an anemic state.
* Reticulocyte production should increase in response to any loss of red blood cells.
* It should increase within 2-3 days of a major acute [[hemorrhage]], for instance, and reach its peak in 6-10 days.
* If reticulocyte production is not raised in response to anemia, then the anemia may be due to an acute cause with insufficient time to compensate, or there is a defect with red blood cell production in the bone marrow.
* Marrow defects include nutritional deficiencies (i.e. iron, folate, or B12) or insufficient [[erythropoietin]], the stimulus for  red blood cell production.


* Reticulocyte Production Index is calculated as follows:
===Specific findings for extravascular hemolysis===
* The degree of anemia (done by normalizing hematocrit of 45%)
*Spherocytic red cell morphology
* Reticulocyte maturation time (RMT)
*Negative urine hemosiderin
* The Reticulocyte maturation time is 1.0 days for a hematocrit of 45 % to 2.5 days for a hematocrit of 15 %:
*Negative urine hemoglobin
* Reticulocyte production index = Reticulocytes %  x  (HCT / 45)  x  (1 / RMT)
* The reticulocyte index (RI) should be between 1.0 and 2.0 for a healthy individual.
* RI < 2 with anemia indicates decreased production of reticulocytes and therefore red blood cells.
* RI > 2 with anemia indicates loss of red blood cells (destruction, bleeding, etc) leading to increased compensatory production of reticulocytes to replace the lost red blood cells.
** Value of 45 is usually used as a normal hematocrit.
** So,in a person whose reticulocyte count is 5%, hemoglobin 7.5 g/dL, hematocrit 25%, the RPI would be:


'''5 x [corrected retic count]/[maturation correction] = 5 x (25/45) /2 = 1.4'''
===Peripheral blood smear===
*'''Schistocytes''': These are fragments of red blood cells. They are also known as helmet cells. Presence of schistocytes is not specific for hemolysis but is specific for a microangiopathic process, which can be associated with hemolysis.
*'''Spherocytes''': These are red blood cells that contain a high volume to surface area ratio. These cells have a relative lack of membrane and thus appear as spheres rather than the typical biconcave shape of normal red blood cells. They appear smaller and rounder than normal red blood cells.
*'''Bite cells''': These are also known as Heinz bodies. They are typically seen in patients with hemolysis from G6PD deficiency. Heinz bodies comprise denatured hemoglobin.
*'''Target cells''': These are also known as codocytes. They are not specific for hemolysis, as they can be found in liver disease, thalassemia, hemoglobin E, and hemoglobin C. Electrophoresis helps to differentiate between thalassemia and liver diseases (elevated liver enzymes), and other hemoglobinopathies.
*'''[[Reticulocytes]]''': These are primitive or immature red blood cells. They contain residual ribosomes and RNA. The presence of RNA allows a visible blue stain to bind or, in the case of fluorescent dye, result in a different brightness. This allows them to be detected and counted as a distinct population. When reticulocytes are present in elevated numbers, this suggests an adequate bone marrow response to anemia. The reticulocytes response act as an important indicator for the severity of the hemolytic anemia. The normal reticulocyte count is 0.5-1.5%. In the case of hemolysis, the reticulocyte count can increase to beyond 10%. The different methods used to assess the appropriate response are:
**Absolute reticulocyte count: This is measured in number of cells per microliter. The normal range for absolute reticulocyte count is 25,000 to 75,000 per microliter.
**Corrected reticulocyte count: This is calculated as absolute reticulocyte count divided by reticulocyte maturation time (in days).
**Reticulocyte production index: This is calculated as reticulocytes %  x (hematocrit / 45) x  (1 / reticulocyte maturation time). The reticulocyte index (RI) should be between 1.0 and 2.0 for a healthy patient.
***RI < 2 with anemia indicates decreased production of reticulocytes and therefore red blood cells.
***RI > 2 with anemia indicates loss of red blood cells (destruction, bleeding, etc) leading to increased compensatory production of reticulocytes to replace the lost red blood cells.


===Other lab tests===
* Elevated [[lactate dehydrogenase]] (LDH)
* Elevated [[haptoglobin]]
* The direct [[Coombs test]] is positive if hemolysis is caused by an immune process
===Liver function test===
* The level of unconjugated [[bilirubin]] in the blood is elevated. This may lead to [[jaundice]].
===Urinalysis===
* Hemosiderin in the urine indicates chronic intravascular hemolysis.
* There is also [[urobilinogen]] in the urine.


(Images shown below are courtesy of Melih Aktan MD, Istanbul Medical Faculty - Turkey, and Hospital Universitario La Fe Servicio Hematologia)
(Images shown below are courtesy of Melih Aktan MD, Istanbul Medical Faculty - Turkey, and Hospital Universitario La Fe Servicio Hematologia)

Revision as of 06:24, 17 October 2017

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Editor-In-Chief: C. Michael Gibson, M.S., M.D. [1] Shyam Patel [2]

Overview

Hemolytic anemia is anemia caused secondary to shortened survival of circulating red blood cells. The normal life span of RBCs is 110 to 120 days. RBC destruction before that time is defined as hemolytic anemia. As opposed to the normal senecence of RBC, the random hemolysis (premature RBC death) is increased in hemolytic anemia.

Approach to diagnosis of hemolytic anemia

Peripheral smear

Findings on peripheral smears can guide towards diagnosis. Some important findings are:

Spherocytes

Sickle cells

Schistocytes

Acanthocytes

  • Liver disease

Parasitic inclusion

Thick and thin smear

Direct and Indirect antiglobulin test

  • Direct antiglobulin tests test for antibody coating red blood cells.
  • Done by mixing the patient's erythrocytes with anti-human globulin (IgG & C3).
  • If agglutination occurs, the test is positive.
  • 95% sensitive.
  • The indirect antiglobulin (indirect Coombs') is done to detect antibodies present in the patient's serum but not coating their red cells.
  • Helps in detecting alloantibodies that were introduced by prior transfusion or by fetal transfer of red blood cells to the mother.

Laboratory Findings

  • Elevated total bilirubin: Elevated bilirubin, or hyperbilirubinemia, is a hallmark of hemolytic anemia
  • Elevated LDH:
  • Low haptoglobin: Haptoglobin is produced in the liver and bind free hemoglobin. During hemolysis, haptoglobin decreases as it is consumed at a faster rate compared to the rate of production by the liver.
  • Elevated reticulocyte count:
  • Positive direct antiglobulin test (DAT):

Specific findings for intravascular hemolysis

  • Elevated urine hemosiderin
  • Elevated urine hemoglobin
  • Microangiopathic blood smear

Specific findings for extravascular hemolysis

  • Spherocytic red cell morphology
  • Negative urine hemosiderin
  • Negative urine hemoglobin

Peripheral blood smear

  • Schistocytes: These are fragments of red blood cells. They are also known as helmet cells. Presence of schistocytes is not specific for hemolysis but is specific for a microangiopathic process, which can be associated with hemolysis.
  • Spherocytes: These are red blood cells that contain a high volume to surface area ratio. These cells have a relative lack of membrane and thus appear as spheres rather than the typical biconcave shape of normal red blood cells. They appear smaller and rounder than normal red blood cells.
  • Bite cells: These are also known as Heinz bodies. They are typically seen in patients with hemolysis from G6PD deficiency. Heinz bodies comprise denatured hemoglobin.
  • Target cells: These are also known as codocytes. They are not specific for hemolysis, as they can be found in liver disease, thalassemia, hemoglobin E, and hemoglobin C. Electrophoresis helps to differentiate between thalassemia and liver diseases (elevated liver enzymes), and other hemoglobinopathies.
  • Reticulocytes: These are primitive or immature red blood cells. They contain residual ribosomes and RNA. The presence of RNA allows a visible blue stain to bind or, in the case of fluorescent dye, result in a different brightness. This allows them to be detected and counted as a distinct population. When reticulocytes are present in elevated numbers, this suggests an adequate bone marrow response to anemia. The reticulocytes response act as an important indicator for the severity of the hemolytic anemia. The normal reticulocyte count is 0.5-1.5%. In the case of hemolysis, the reticulocyte count can increase to beyond 10%. The different methods used to assess the appropriate response are:
    • Absolute reticulocyte count: This is measured in number of cells per microliter. The normal range for absolute reticulocyte count is 25,000 to 75,000 per microliter.
    • Corrected reticulocyte count: This is calculated as absolute reticulocyte count divided by reticulocyte maturation time (in days).
    • Reticulocyte production index: This is calculated as reticulocytes % x (hematocrit / 45) x (1 / reticulocyte maturation time). The reticulocyte index (RI) should be between 1.0 and 2.0 for a healthy patient.
      • RI < 2 with anemia indicates decreased production of reticulocytes and therefore red blood cells.
      • RI > 2 with anemia indicates loss of red blood cells (destruction, bleeding, etc) leading to increased compensatory production of reticulocytes to replace the lost red blood cells.


(Images shown below are courtesy of Melih Aktan MD, Istanbul Medical Faculty - Turkey, and Hospital Universitario La Fe Servicio Hematologia)

References

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