Wednesday, September 17, 2008

Glucose-6-phosphate dehydrogenase (G6PD) deficiency

Note the RBC's with Heinz Bodies present (Nile Blue Sulfate stain)

Glucose-6-phosphate dehydrogenase (G6PD) deficiency (a defect in the hexose monophosphate shunt) was first noted in the time of Hippocrates when Pythagoras warned against eating fava beans. It is a sex-linked trait. The disorder is fully expressed in the hemizygous male and the homozygous recessive female (both alleles abnormal). All RBC’s of homozygous recessive men and women will have a G6PD deficiency. The heterozygote female (one normal and abnormal allele each) will present with two populations of RBC’s: one with G6PD and the other without G6PD. This group will be clinically normal, but with about 50% of the level of G6PD in normal individuals. This disorder is found world-wide and is the most common RBC enzyme deficiency associated with hemolysis.

The World Health Organization has divided G6PD deficiency into five classes: Class I is the most severe of hemolytic expressions. Class II hemolytic episodes are mild. Class III presents with mild hemolytic episodes. Class IV does not manifest hemolytic episodes. Class V does not manifest hemolytic episodes. Each of these classes are characterized by different genetic expressions, levels of G6PD enzyme activity and deficiency, and electrophoretic test patterns.

G6PD is an important enzyme in the chemistry of the body. It purpose is to generate NADPH (the reduced form of Nicotinamide Adenine Dinucleotide Phosphate) which is necessary to rejuvenate oxidized glutathione to its reduced form. The reduced glutathione then protects hemoglobin from oxidative denaturation processes by reducing free radicals and H2O2. The reaction process to generate NADPH in the RBC is as follows:

NADPH is a H+ donor. It prevents oxidative stress by providing hydrogen ions to the hemoglobin preventing oxidation to functionless methemoglobin. If methemoglobin is allowed to build up, then it will precipitate out as Heinz bodies. In this state the RBC is vulnerable to macrophage attack where the Heinz body will be removed resulting in pitting, bite cells, and/or blister cells. In this RBC, the spectrin cross-linking increases, producing a more rigid cell which becomes trapped for longer intervals in the spleen. This means a higher rate of RBC destruction.

In the normal physiological activity of the RBC the following occurs:

If oxidation of heme iron occurs by any of the following:
  • » Auto-oxidation of long standing,
  • » Ferricyanide reaction,
  • » Nitrates or nitrites,
  • » Sulfonamides,

Then this creates a ferric state of heme iron (Fe+++) and it cannot bind to nor transport O2.This new state of hemoglobin is not methemoglobin. As the RBC ages, the level of G6PD decrease to deficient levels. This reduction occurs over 120 day period, at which time the aging RBC’s are removed from circulation. If there is a deficiency or absence of G6PD, the RBC aging process is faster.

Clinical symptoms for the most individual with variants of G6PD deficiency are absent. These lead normal lives, experiencing mild to moderate anemia conditions are generally asymptomatic. The variants of G6PD that are defective and subject to significant hemolytic episodes, the patient will experience: headaches, dizziness, palpations, dyspnea, abdominal pain, and back pain. Expected clinical laboratory findings include:
  • [1] Hemoglobinuria (with brown to black colored urine).
    [2] If severe episode, a hemoglobin drop of 3 to 4 gm/dL.
    [3] Hemoglobinemia (with pink to brown plasma).
    [4] Reticulocyte count = 8 to 12 %
    [5] Bilirubin = >1.5 mg/dL
    [6] Haptoglobin = < ldh =" ">200 IU/L with LDH-1 and LDH-2 as the predominant isoenzyme forms
    [8] Peripheral blood smear will contain [1] bites cells, [b] spherocytes, [c] cell fragments, [d] blister cells, [e] Heinz bodies, [f] polychromasia, and [g] poikilocytosis
    [9] Urobilinogen = increased

Specific hematological tests useful in diagnosing G6PD deficiency are:
  • [1] Ascorbate Cyanide Test. This is a sensitive test, but not very specific. Its principle is based upon the absence of G6PD in the RBC and the failure to reduce H2O2.
  • [2] Fluorescent Spot Test. This is a testing principle in which glucose-6-phosphate, NADP, and saponin is added to whole blood. If G6PD is present, then NADP is converted to NADPH. NADPH will fluoresce if present.
  • [3] Methemoglobin Reductase Test. This is a screening test that uses fresh whole blood, NADPH, and methylene blue. The principle is based up the reduction of methemoglobin to hemoglobin by NADPH. G6PD deficient RBC lack this ability. Caution. This test needs to be performed immediately after collecting the blood. If blood is allowed stand for too long a period of time, a false positive may result.

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