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710 Part VI: The Erythrocyte Chapter 47: Erythrocyte Enzyme Disorders 711

on a lysate, may be more sensitive than the other screening procedures. stippling of erythrocytes is present, examination of the ultraviolet spec-
Prenatal diagnosis of G6PD deficiency is also possible using DNA trum of a perchloric acid extract of the erythrocytes, reflecting the ratio
mutational analyses approach. between pyrimidine and purine nucleotide content, may help to estab-
Testing for red cells enzyme deficiencies is best done in specialized lish the diagnosis of pyrimidine 5′-nucleotidase deficiency. Beyond
598
laboratories. Specimens can be shipped by mail to reference laborato- these relatively simple procedures it is probably rarely useful to pick and
ries. As a rule, whole-blood specimens are suitable and can best be sent choose individual enzyme assays on the basis of family history or clin-
580
at 4°C as some enzymes, notably PFK, are relatively unstable. Blood ical manifestations. Rather, it is usually appropriate to submit a blood
from a healthy volunteer should be shipped with the patient sample to sample to a reference laboratory that has the capability of performing
serve as a shipping control. Exceptions are assays for phosphorylated all the enzyme assays listed in Table 47–3. Preferably, the suspicion of a
sugar intermediates, 2,3-BPG, and nucleotide intermediates, which are specific enzyme disorder causing hereditary nonspherocytic hemolytic
unstable in freshly drawn blood and require immediate deproteiniza- anemia is confirmed by DNA sequence analysis. This also enables pre-
tion in perchloric acid. natal diagnosis which has already been achieved for some of enzymatic
Several aspects should be kept in mind when interpreting test defects. 599–607
results. First, care must be taken to remove leukocytes and platelets in Notably, in an estimated 70 percent of cases of suspected hered-
assays such as for PK, as these cells contain PK activity, obscuring a defi- itary nonspherocytic hemolytic anemia no enzymatic abnormal-
ciency in the red cells. Second, one should be aware of the already men- ity is found. 608,609 Current promising approaches such as red cell
tioned red cell age dependency of, for example, PK, HK, and G6PD. The proteome analysis 610–612 and/or the use of next-generation sequencing
measurement of these enzymes simultaneously can give an idea about technologies may aid in a better and more comprehensive under-
613
red cell age and relative deficiencies. If patients received blood transfu- standing of the etiology of this disorder.
sions, interpreting results from red cell enzyme assays is generally not
possible because the presence of donor erythrocytes will obscure any THERAPY
deficiencies. Some mutant enzymes also display a normal activity in
vitro, whereas in vivo severe hemolysis can occur, reflecting the differ- GLUCOSE-6-PHOSPHATE DEHYDROGENASE
ences between optimal circumstances in vitro and the in vivo cellular
environment. More sophisticated assays to measure, for example, heat DEFICIENCY
instability and kinetics, have to be used in those cases. Interpretation G6PD-deficient individuals should avoid drugs that are predicted to
can be particularly challenging in newborn patients given the differ- induce hemolytic episodes (see Table 47–5). However, it is important
ences in red cell energy metabolism and enzymatic activities between to realize that such patients are able to tolerate most drugs. Unfortu-
adults and newborn infants. 592–596 Molecular diagnosis is now available nately, in the past, a number of case reports incorrectly suggested that
for all red cell enzyme deficiencies. some drugs had hemolytic potential that subsequently were shown to
be safe (see Table 47–5, possible hemolysis). Although it is possible that
DIFFERENTIAL DIAGNOSIS some of these may be hemolytic in some patients or under some cir-
cumstances, this is unlikely, and G6PD-deficient patients should not be
Drug-induced hemolytic anemia resulting from G6PD deficiency deprived of the possible benefit of these drugs.
is similar in its clinical features and in certain laboratory features, to If hemolysis occurs as a result of drug ingestion or infection, par-
drug-induced hemolytic anemia associated with unstable hemoglo- ticularly in the milder A– type of deficiency, transfusion usually is not
bins (Chap. 49). Other enzyme defects affecting the pentose-phosphate required. If, however, the rate of hemolysis is very rapid, as may occur,
shunt, such as a deficiency of GS, also may mimic G6PD deficiency. for example, in favism, transfusions of packed cells may be useful. Good
The diagnosis of hemoglobinopathies can be excluded by performing urine flow should be maintained in patients with hemoglobinuria to
597
a stability test, hemoglobin electrophoresis or DNA sequence analy- avert renal damage. Infants with neonatal jaundice resulting from
sis. These are normal in G6PD deficiency. Some of the screening tests, G6PD deficiency may require phototherapy or exchange transfusion; in
591
particularly the ascorbate cyanide test, may give positive results in the areas in which G6PD deficiency is prevalent, care must be taken not
614
above-named disorders, but a G6PD assay or the fluorescent screen- to give G6PD-deficient blood to such newborns. A single dose of
ing test will be positive only in G6PD deficiency. In addition, defects Sn-mesoporphyrin, a potent inhibitor of heme oxygenase, has been
615
of the erythrocyte membrane should be excluded (Chap. 46), but these advocated to eliminate the need for phototherapy. Patients with hered-
cytoskeletal and other membrane defects are generally associated with itary nonspherocytic hemolytic anemia resulting from G6PD deficiency
characteristic morphologic abnormalities, that makes them easy to dif- usually do not require any therapy. Splenectomy is often ineffective,
ferentiate from hemolysis because of enzyme defects. although some improvement has been reported in a number of cases
Physicians often attempt to establish the cause of hereditary non- following removal of the spleen. 264,616 In most cases, the anemia is not
spherocytic hemolytic anemia on the basis of the appearance of red cells very severe, but in some instances frequent transfusions have been nec-
on a blood film. In reality, red cell morphology is helpful only in the essary. 617,618 The antioxidant properties of vitamin E have been tested in
diagnosis of pyrimidine 5′-nucleotidase deficiency because of the char- G6PD-deficient subjects, and a slight but statistically significant reduc-
acteristic stippling of the red cells that is observed in that disorder. The tion in hemolysis was observed. 619,620 These results could not be con-
appearance of Heinz bodies suggests the possible presence of an unsta- firmed in other studies. 621,622 It has been suggested that desferrioxamine
ble hemoglobin, or defective GSH metabolism. They are more likely to decreases hemolysis. 623,624 Inhibition of histone acetylation by histone
be present after splenectomy. deacetylase inhibitors has been shown to increase G6PD gene transcrip-
Because the laboratory diagnosis of these disorders may entail tion in erythroid progenitor cells and restore G6PD deficiency. 625
considerable expenditure of time and effort, it is prudent to perform
the simplest tests for the most common causes of hereditary non- OTHER ENZYME DEFICIENCIES
spherocytic hemolytic anemia first. Accordingly, it is useful to carry Most patients with hereditary nonspherocytic hemolytic anemia sec-
out screening tests 580,582 for G6PD and PK activity and an isopropanol ondary to red cell enzymopathies do not require therapy, other than
stability test to detect an unstable hemoglobin (Chap. 49). If prominent blood transfusion during hemolytic periods, if the anemia needs

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