Hereditary Glucosephosphate Isomerase
Deficiency
A Review
D O N A L D E. PAGLIA, M.D.,
AND W I L L I A M N. V A L E N T I N E ,
M.D.
From the Division of Surgical Pathology and the Department of Medicine, University of California
School of Medicine, Los Angeles, California 90024, and the Wadsworth General Hospital,
Veterans Administration Center, Los Angeles, California 90073
ABSTRACT
Paglia, Donald E., and Valentine, William N.: Hereditary glucosephosphate
isomerase deficiency: A review. Am. J. Clin. Pathol. 62: 7 4 0 - 7 5 1 , 1974. T h e
clinical, hematologic and biochemical characteristics of the known cases of
glucosephosphate deficiency are reviewed. This hereditary disorder of
erythrocyte metabolism is the third most common enzyme defect resulting in
congenital nonspherocytic hemolytic anemia. Its unique features relative to
other erythroenzymopathies are emphasized. (Key words: Erythrocyte
metabolism; Erythroenzymopathy; Nonspherocytic hemolytic anemia;
Glucosephosphate isomerase deficiency.)
human erythrocyte endures
metabolic restraints not imposed on its
reticulated or nucleated precursors. Incapable of oxidative phosphorylation, it
fills its energy reservoir with ATP derived
almost exclusively from
anaerobic
glycolysis. Thus it logically follows that
shortened cell life span and consequent
hemolytic anemia may be induced by
defective glucose catabolism. Hereditary
deficiencies of 15 specific enzymes have
now been defined in association with
hemolytic syndromes. In most instances,
these have involved glycolytic enzymes, 42
predominantly glucose-6-phosphate dehydrogenase (G6PD) 7 and pyruvate kinase
(PK),39 but they also include enzymes
concerned with glutathione 7,21 and nucleotide metabolism. 46 Pertinent metabolic
pathways a n d r e p o r t e d
enzymatic
deficiencies are summarized in Figure 1.
T H E MATURE
Address reprint requests to Dr. Paglia.
Received J u n e 10, 1974; accepted for publication
July 2, 1974.
740
Recent studies indicate that defective
glucosephosphate isomerase (GPI) (Dglucose-6-phosphate ketol isomerase,
E.C.5.3.I.9.), first reported in 1968 5 and
thought to be quite rare, is probably the
third most commonly occurring erythroenzymopathy. It has features in common with G6PD and PK deficiencies, but
certain dissimilarities are also emerging.
On the basis of broadening experience
with this syndrome and its frequency of
occurrence, it seems appropriate to review
its characteristics for the benefit of those
concerned with laboratory diagnostics.
This enzyme holds additional interest
from a genetic vantage because of its
phenotypic heterogeneity and from a clinical vantage because of its specific location
in the Embden-Meyerhof pathway which
allows, theoretically at least, a potential
for therapeutic intervention.
No u n i q u e clinical or
routine
hematologic findings serve to distinguish
GPI deficiency from other erythroen-
December
1974
741
GLUCOSEPHOSPHATE ISOMERASE DEFICIENCY
Glucose
• GLUCOSE-6PHOSPHATE
DEHYDROGENASE
ATP
• HEXOKINASE
•4
ADP •
6-6-P
PHOSPHOGLUCONATE
DEHYDROGENASE
T^T
• GLUCOSEPHOSPHATE
ISOMERASE
*
6-PG
R-5-P
F-6-P
• PHOSPHOFRUCTOKINASE
FIG. 1. Summary of
glycolytic and some ancillary metabolic pathways
available
to
mature
human erythrocytes. Pentosephosphate shunt activity generates fructose-6phosphate (F6P) and glyceraldehyde - 3 - phosphate
(G3P) t h r o u g h transketolase and transaldolase
(not shown), and these int e r m e d i a t e s a r e subsequently metabolized via
the E m b d e n - M e y e r h o f
pathway (dashed boxes).
T h e solid diamonds designate enzymes which
have been identified as
defective
in
various
hereditary disorders of
erythrocyte metabolism.
• FRUCTOSEDIPHOSPHATE
ALDOLASE
DHAP
Glycine
• TRIOSEPHOSPHATE
ISOMERASE
GLYCERALDEHYDE3-PHOSPHATE
DEHYDROGENASE
•
•PHOSPHOGLYCERATE
KINASE
DIPHOSPHOGLYCERATE
PHOSPHATASE
PHOSPHOGLYCEROMUTASE
•y-GLUTAMYLCYSTEINE
SYNTHETASE
Glutamic Acid
+
Cysteine
2-PG
PHOSPHOPYRUVATE
HYDRATASE
t
I
PEP
• PYRUVATE
KINASE
\j- ADP
[
•ADENYLATE
2 ADP <
r» ATP
Pyruvate
LACTATE
DEHYDROGENASE
«• ATP + AMP
t - » "AOH
k » NAD
Lactate
zymopathies. Patients may present any of
the nonspecific features generally associated with chronic hemolytic anemia,
such as icterus, reticulocytosis, indirect
hyperbilirubinemia, decreased haptoglobin, etc. Clinical severity ranges from
well-compensated chronic hemolysis to
severe fluctuating anemia necessitating
frequent transfusions. T h e associated
laboratory findings are similarly variable.
Since the latter are nonspecific, the diagnosis is derived virtually by exclusion after
failure to demonstrate antiglobulin reactions, hemoglobinopathies or membrane
defects.
Incidence and Distribution
Defective GPI has been implicated in 20
individual cases of hemolytic anemia occ u r r i n g in 16 apparently unrelated
families (Table 1). Of these, eleven were
identified as male, seven as female, and
the r e m a i n d e r unspecified. Afflicted
families have been largely of European
ancestry, but cases have also been observed in Japanese 27,28 and Mexicans.6,11 •24,33
Eleven resided in the Northwestern Hemisphere, seven in Europe.
Gene frequency remains undefined.
Lohr and colleagues 24 found no deficien-
742
PAGLIA AND VALENTINE
A.J.C.P.—Vol.
62
Table 1. Proposed Designations and Characteristics
Patient Data
References
Age
(Yr.),
Sex
5,18
15, M
GPI-Whitley County*
32
13, F
12, M
VA, M
(None)
14
Designalion
GPI-Seattle»
GPI-Espeln
(None)
Hemoglobin
(Gm. per 100 n 1.)
Reticulocytes
French/Irish
9.8
28
Anglo-Saxon
8.0
7.6
6.0
72
42
71
12.3
11
Ancestry
" C r T 'A
(Days)
4.5 (39-44)t
5
1,2.3.24,40
24, M
21, M
German
23
10, F
?, F
Sardinian
(None)
(%)
12
(30-40)
20. M
(None)
(None)
31
12, M
GPI-Los Angeles
11,24
1,M
GPI-Winnipeg
11,24
26, F
7.4
34
2.5
Portuguese •Canadian
11.1
33
3.9
7-9
16
7.3
10
Mexican-American
GPl-Recklinghausen
19,24
9, F
German
GPl-Narita
28,30
2, M
Japanese
GPl-Matsumoto
27,30
24, F
Japanese
8.6
7
Mexican
11.0
25
10.0
20
Mexican
9-10
15-20
(None)
6, M
6
(None)
16
9, M
CPl-Valle Hermoso
33
14, F
* Proposed in retrospect.
t Range of normal controls
t Classification according to Detter et at
2.7
5
(25-32)
10.5 (25-32)
4
18
cies in an unselected series of 350,
whereas we have observed two incidental
heterozygotes in approximately 1,000.45
Detter and associates 18 discovered 20 electrophoretic variants of ten different types
in random surveys of 3,397 individuals,
but activity measurements were not
routinely performed. In a similar survey
of 1,650 persons, Fitch and colleagues 20
found five variant patterns in nine subjects. In both studies, Asiatic indians had
more than a 1% incidence of specific
variants; otherwise, no phyletic distributions have been reported.
Clinical Features
There is considerable variation in clinical expression. Insidious onset of chronic
hemolysis and its sequelae are usually
noted in infancy or early childhood, but
the disease may present as neonatal jaundice, 27,28,32 sometimes necessitating exchange transfusion. 32 Slight to moderate
splenomegaly has been observed alone
o r in c o m b i n a t i o n
aly 5,6,27,28,32,35,37
with
hepatomeg-
Variable severities of anemia are
reflected by the wide spectrum of mean
values for hemoglobin and reticulocytes
December 1974
743
GLUCOSEPHOSPHATE ISOMERASE DEFICIENCY
of Hereditary GPI Deficiencies
GPI Activity
Erylhroc ytes
GIM Michaelis Constant
Leukocytes
Normal Mean)
Other
<%of
19
23
16
7
14
10
PH
Optimum
Erythrocytes
(mM RiP)
Plasma = 0
1Leukocytes
{mM F6P)
Normal
Normal
0.06
0.06
Thermal
Sensitivity
Electrophoresis and
Connneiits
Pill 0-10$ (last major
band)
Slow major, absent minor
bands
Parents normal
Normal
25
23
73
Fibroblasts = 53
Platelets = 27
Normal
8.7
8.4
Very labile
PHI 9 - 9
Coincidental elliplocylosis
50
50
Resembled Pill 9. Coincidental G6PD deficiency
Coincidental G6PD
deficiency
10
10
14
22
Spherocytic
29
Platelets = 33
8.5
0.10
Very labile
Slow; one parent normal
8.5
0.11
Labile
Slightly slower
8.3
0.05
Labile
Moderately slower
15
35
40
Low
Plastna = Normal
Normal
0.22
Ubile
Fast: 4 bands: parents resemble PHI 7 - 1
38
<I0
Plasma = < 30
Acid shift
0.18
Very labile
Normal
23
Very labile
15
25
Normal
Plasma = Normal
i.0-8.5
recorded in Table 1 and by considerable
variance in transfusion requirements.
Some patients require monthly transfusions for considerable periods, 32 whereas
others remain well-compensated without
any. T h e clinical c o u r s e may be
punctuated by acute hemolytic crises,
which are often associated with intercurrent infections.8'19'32-33,37 Exposure to certain drugs or chemicals was suspected to
have initiated an acute episode in at least
one case,33 but this has been documented
with certainty only when there was coexistent G6PD deficiency.37
0.07
l-abile
Normal
Hematologic Features
A broad spectrum of cytologic alterations
of affected erythrocytes has been described.
Almost invariably, anisopoikilocytosis and
polychromatophilia are prominent. Nucleated erythrocytes, target and burr cells,
basophilic stippling, Pappenheimer and
Howell-Jolly bodies, and dense spiculated
microspherocytes may also occur in relatively small numbers. These findings are
common to most erythroenzymopathies
and are not specifically indicative of GPI
deficiency.
744
PAGLIA AND VALENTINE
Autohemolysis during sterile incubation
at 37 C , when tested, has conformed to
the type I category of Selwyn and Dacie,38
with slight to moderate hemolysis partially
corrected by glucose. 5,6,32 ' 33,36,40 An entirely normal autohemolysis test has been
reported. 16
Osmotic fragility is usually unaltered
but may be slightly increased after 24-hr.
incubation at 37 C. Oski and Fuller reported an affected individual with increased spherocytes and fragility whose
parents were heterozygous for GPI deficiency but devoid of other hematologic
alterations. 31 Elliptocytosis with decreased
fragility has been found in combination
with partial (50%) deficiency of GPI in a
young girl and her mother, both of whom
had hemolytic anemia. 23 Other family
members with either trait alone were
hematologically normal.
As shown in Table 1, erythrocyte survival times are markedly shortened, and
splenic s e q u e s t r a t i o n is d e m o n s t r a ble. 12,24,25
Despite the observation that GPI deficiency also occurs to varying extents in the
leukocytes, significant alterations in
leukocyte counts, distribution, morphol-
F6P
GPI
NADP
Some procedures additionally include
6-phosphogluconate
dehydrogenase
(6PGD) to double the sensitivity of the
assay, but this is unnecessary in view of
the relatively high GPI activity in blood
cells, and may be undesirable because
some commercial preparations of 6PGD
contain traces of GPI, a problem also
encountered with some lots of G6PD. The
small amounts of endogenous 6PGD in
hemolysates result in observed activities
which are slightly higher than actuality,
but this is not of sufficient magnitude to
alter identification of heterozygous or
62
ogy or function have not been reported.
Biochemical Features
GPI Assay
Diagnosis ultimately relies upon identification of the specific biochemical lesion. Screening tests for deficiencies of
G6PD and PK are now p e r f o r m e d
routinely in most clinical laboratories, and
a similar screening procedure for GPI
deficiency has been described by Blume
and Beutler. 10 In addition, relatively simple quantitative assays for GPI activity are
well within the capability of any laboratory equipped to perform ultraviolet spectrophotometry.
Most quantitative assay systems are similar to our modification 5 of that described
by Chapman and associates.15 Substrate
(F6P) is provided in a medium buffered
to about pH 8. Purified yeast G6PD and
NADP serve as an indicator system, and
the reaction is initiated by addition of cell
lysate. Rate of F6P isomerization to G6P is
measured by monitoring absorbance increases at 340 nm. as NADP is reduced to
NADPH in the subsequent oxidation of
G6P:
G6PD
G6P
A.J.C.P.—Vol.
6-phosphogluconate
NADPH
homozygous deficiency states. Errors due
to contamination of F6P with small
amounts of G6P may be eliminated by
preincubation of the entire system prior
to addition of lysate.8 Corrections for the
GPI contributions by leukocytes contaminating erythrocyte preparations are
sometimes necessary when sedimentation
technics are used to isolate the formed
elements. Much purer erythrocyte suspensions are attainable utilizing the
sulfoethyl-cellulose filtration technic described by Nakao and associates,29 which
December 1974
GLUCOSEPHOSPHATE ISOMERASE DEFICIENCY
has the further advantage of preserving
reticulocytes and macrocytes in greater
numbers.
Any laboratory wishing to expand its
diagnostic armamentarium to encompass
assays of GPI or any other blood cell
enzyme is referred to Beutler's concise yet
comprehensive monograph on pertinent
methodology. 8
Variations in GPI Activity
Reported values for GPI activities vary
considerably among different laboratories, largely due to technical variations and the potential errors cited in the
previous section. Nevertheless, once a
normal range is established for a given
procedure, the distinction between normals and inherited deficiencies is usually
quite clear. In the u n c o m p l i c a t e d
heterozygous deficiency state, which clinically is asymptomatic, assays reveal approximately half-normal GPI activities,
almost invariably falling within the range
of 40 to 60% of normal laboratory mean.
As shown in Table 1, anemic subjects,
who are either homozygous or doubly
heterozygous for GPI deficiency, possess
erythrocytic values approximating 25% of
normal means. The latter may be deceptively high because of the accompanying
reticulocytosis and because the cell population available for study is quite young, as
evidenced by the shortened chromium
survival times. It is well established that
most erythrocytic enzymes are considerably more active in reticulocytes and young
erythrocytes, and that these activities
decay at individually characteristic rates as
the cells age. It is more appropriate,
therefore, to compare measurements with
those obtained from individuals with
comparable reticulocytosis due to other
causes. When such comparisons are made,
anemic patients may have less than 10%
of expected activities in their erythrocytes.
Erythrocytic GPI activities which are
significantly elevated above those ex-
745
pected on a cell-age basis may be encountered in cord blood specimens 22 and in a
variety of dyserythropoietic disorders, 43,44
preleukemic states, 44 acquired refractory
anemias, 44 and folate deficiency.44
On an individual cell basis, GPI activity
in leukocytes is greater than that in
erythrocytes by approximately two orders
of magnitude. Normal control values for
leukocytic GPI are subject to greater
variation, but deficiencies are still easily
identified. Significant deficiencies have
existed in all but one 33 of the reported
cases appropriately studied (Table 1). The
one exception, GPI-Valle Hermoso, discussed in the section, Genetic Polymorphism and Isozymes, also had normal GPI
activities in plasma.
The source of plasma GPI is unknown,
but it too is usually low in severe deficiency states and presumably reflects deficiencies in many other organ systems.5'27,33
Plasma activities in one case investigated by Miwa and associates28 were normal, but leukocytic activities were low.
Fibroblasts and platelets have been shown
to be affected as well, 119 adding strength
to the postulate that a multi-system disorder is involved. The lack of apparent
significant consequences to leukocytes or
to other tissue cells possessing nuclei and
organelles is presumably because such
cells, unlike mature erythrocytes, possess
alternate metabolic pathways and a potential capacity to replenish depleted enzymes by continued protein synthesis.
Qualitative Alterations in
GPI Characteristics
Kinetics and Molecular Features
Unlike erythrocytic PK anomalies, the
polymorphism of GPI defects is not commonly manifested by altered enzyme kinetics. Apparent Michaelis-Menten constants
(Km) of residual GPI in either erythrocytes
or leukocytes have never been reported to
diverge significantly from normal controls
746
PAGLIA AND VALENTINE
(Table 1). This has been true whether
crude hemolysates or partially purified
preparations were examined. Most reported Km(F6P) values approximate 0.10
mM F6P, with no apparent difference
between values for erythrocytes and
leukocytes. Purification procedures per se
do not substantially alter Km (F6P) values
from those obtained with hemolysates. 4
Similarly, most cases have a common
pH optimum for maximum in vitro activity, approximately pH 8.5 (Table 1). Only
Miwa and associates 27 have reported an
a b n o r m a l p H - o p t i m u m curve, with
GPI-Matsumoto manifesting a slight
acidic shift. Variably abnormal isoelectric
points have been measured in GPI from
four different cases.24 Molecular weight
determinations on purified preparations
have not yielded significant deviations
from normality. 1,11,19,24,40
A.J.C.P.—Vol.
62
between normals and homozygotes 30 ' 33,40
or indistinguishable from normals. 11
T o summarize, most persons who have
GPI deficiency share several qualitative
characteristics of their residual erythrocytic enzyme: normal kinetics for F6P, normal pH optima and molecular weights,
and pronounced thermolability.
Electrophoretic Properties
The existence of molecular heterogeneity in this disorder is best demonstrated by the wide disparity in
electrophoretic migration rates among
reported cases. T h e extensive work of
Detter and associates18 established patterns of GPI migration on starch gel both
for the wild enzyme (designated PHI 1)
and for ten unusual isozymes, one of
which was found in the first known case
of GPI-deficiency hemolytic anemia. Most
variants, as well as PHI 1, contained three
Thermostability
major c o m p o n e n t s which m i g r a t e d
cathodally
at slightly alkaline pH. Their
Susceptibility to heat inactivation is
studies
supported
the hypothesis that the
another feature common to GPI deficienwild
enzyme
from
erythrocytes existed as
cies. When normal hemolysates are incua
dimer
of
identical
subunits, while mubated at 45 to 48 C in controlled media,
tant
forms
apparently
combined a normal
GPI activity remains relatively stable, ususubunit
with
one
somehow
altered to
ally registering more than 80% of initial
11,40
effect
differing
mobilities
(designated
levels after 1 hour.
By contrast, all
eight variants studied by this technic were PHI 2-1 to PHI 10-1). In the initial case of
moderately to markedly labile, frequently GPI deficiency, the proband possessed no
falling to less than 50% of initial activities component identifiable with normal PHI
within 10 to 20 min. (Table 1). Curves 1, apparently combining the rare alleles
describing enzymic activity as a function from his mother (PHI 9-1) and father
of incubation periods may be linear, 11,24,30 (PHI 10-1) to yield phenotype PHI 9-10.
but in many instances are bimodal, with a
Investigations of other cases of GPI
very rapid decline in the first 10 to 20 deficiency have shown electrophoretic abmin. followed abruptly by a normal decay normalities in which the major composlope. 3,19,24,30,33 Such biphasic patterns are nents were also similar to type 9,37,40 but
highly suggestive of two GPI components, some patterns bore no resemblance to any
one of which may be catalytically more previously described. 11,24,32 In one famactive but also much more susceptible to ily,32 parents and clinically unaffected sibheat inactivation. Hemolysates from lings possessed zymograms which were
heterozygotes have yielded either similar indistinguishable from normal, while procurves with a more subtle bimodality 30 or band patterns showed decreased migralinear curves with slopes intermediate tion rates and virtual absence of minor
December 1974
GLUCOSEPHOSPHATE ISOMERASE DEFICIENCY
bands. It was presumed that the proband
was homozygous for mutant parental
genes which coded for a subunit with
decreased activity and only slightly altered
migration. T h r e e variants, GPI-Matsumoto, 3 0 GPI-Valle Hermoso, 3 3 and
that in a case reported by Carrier, 14 have
entirely normal electrophoretic patterns.
While most studies have reported three
primary bands, as few as one major
component 24,32 and as many as four 28 have
been observed. Some minor bands are
probably products of enzyme oxidation
and do not truly represent djstinct subunits. 34 Studies with purified GPI extracted from human erythrocytes and
skeletal muscle strongly support the dimeric s t r u c t u r e suggested by electrophoretic data and genetic considerations. 13,41
Genetic Polymorphism and Isozymes
Genealogic studies uniformly have demonstrated an autosomal recessive pattern of genetic transmission. Clinically
normal heterozygotes inherit a normal
GPI-determinant gene from one parent
and a mutant allele from the other,
resulting in approximately half the usual
erythrocytic GPI activity. Clinically affected individuals with markedly impaired
GPI activity acquire abnormal genes from
both p a r e n t s . T h e y may be either
homozygous for a common aberrant gene
or doubly heterozygous for dissimilar
mutant alleles. The former alternative is
obviously more probable when consanguinity exists, as it did in at least five of
the 16 affected families.6,14,27,28,33
As in PK deficiency, some mutant genes
appear to code for the production of
catalytically inactive or quantitatively
deficient enzyme, while others govern
structural alterations that may adversely
affect enzyme stability. Numerous combinations of mutant alleles are thus possible,
accounting for the diversity of variants
747
demonstrated by electrophoresis. GPISeattle, 5,18 for example, resulted from
simultaneous inheritance of two rare alleles at the GPI locus, as noted previously.
GPI-Los Angeles, 11,24 on the other hand,
appeared to result from a paternal gene
coding for highly labile enzyme in combination with a maternal gene coding for
inactive or absent enzyme, genotype
GPI-Los Angeles/GPI ( - ) .
Tissue-specific isozymes of GPI apparently do not exist in either normal or
deficient states. Similar molecular weights
and isoelectric points have been obtained
from purified enzyme extracted from
normal human erythrocytes and skeletal
muscle. 13,41 Electrophoretic studies by Detter and associates 18 demonstrated virtual
identity of GPI among various blood cells,
plasma and other tissues. These findings
were supported and expanded by Payne
and colleagues, 34 who reported identical
electrophoretic and isoelectric properties
of GPI from normal erythrocytes, heart,
liver, kidney and brain. The electrophoretic abnormality of GPI-Narita was shown
by Nakashima and associates30 to be identical to that of the GPI present in leukocytes and plasma and in spleen, liver and
skeletal muscle extracts. These observations strongly support a common structural identity of GPI among these various
tissues and thus, by inference, a single
genetic determinant.
In only one instance, GPI-Valle Hermoso, 33 has the deficiency appeared to be
isolated to erythrocytes, with normal activities in leukocytes and plasma and
normal electrophoretic patterns of residual enzyme. It was hypothesized that
the proband was homozygous for a single
mutant gene at the GPI locus. The resultant structural alteration apparently induced severe instability of the enzymic
protein with relatively inconsequential effects on catalytic effectiveness or electrophoretic properties. Unlike mature
748
PAGLIA AND VALENTINE
erythrocytes, tissues capable of continued
protein synthesis could constantly replenish a diminishing complement of unstable enzyme and thereby show normal
activities by in vitro assay. Such a situation
has a counterpart in the highly unstable
A( —) variant of G6PD, which is less active
but electrophoretically indistinguishable
from the A isozyme.
A,J.C.P.—Vol.
62
however, remains unimpaired in GPIdeficient cells,3,33 this because glucose and
mannose are both phosphorylated by
hexokinase but are converted to F6P by
separate isomerases. 9
Partial blockage of G6P isomerization
and its consequent increased intracellular
concentration leads to accelerated shunting through the pentosephosphate pathway. T h e technic described by Davidson
Metabolic Consequences of GPI
and Tanaka 1 7 measures this by monitorDeficiency
ing 1 4 C0 2 evolution from glucose labeled
Even though GPI deficiency in other in the first-carbon position. In severe GPI
tissues appears to induce no significant deficiency, the shunt may be operating at
metabolic impairment, deleterious effects or near maximum capacity, so little or
on the erythrocyte population are clearly none of the expected increase in shunt
demonstrable. Intracellular concentra- activity occurs in5 response to methylene
tions of glycolytic intermediates and blue stimulation.
adenine nucleotides may be variably disEvolution of 1 4 C0 2 from glucose labeled
14
turbed but have also appeared normal.
in the second-carbon position depends
Erythrocyte concentrations of G6P are upon recycling of pentose through the
frequently increased, 2,19,27,28,31,33,36,37 some- shunt following its conversion to F6P by
times to as much as three to five times transketolase and transaldolase (Fig. 1).
normal levels, and may affect overall Since F6P must traverse the GPI step to
glycolysis by feedback inhibition of glu- re-enter the shunt, a markedly decreased
cose phosphorylation, the latter despite recycling capacity results from GPI defithe highly increased hexokinase activities ciency, as low as 0.2% 31 to 10% 5,33 of
characteristic of young cells. Metabolites normal. Intermediate impairment may be
immediately distal to the enzymatic block, encountered with the partially deficient
such as F6P and FDP, are often de- cells of heterozygotes. 32,33
creased, 31,33,36,37 although they may appear
Even with apparently normal in-vitro
normal 28 or elevated27 unless compared with glycolysis, there are obviously severe inblood of comparably young mean cell age. vivo effects of GPI deficiency. Cell life
T h e latter is also true of ATP/ADP span is drastically shortened as measured
concentrations, which have been reported by radiochromium survival times (Table
as normal, 14,27,28,33 increased, 2,5 or de- 1), and there is evidence of splenic sequescreased. 31,36,37 Concentrations of 2,3-DPG tration. 12,24,25 These effects may ultimately
have been observed to be normal 16 or be a consequence of altered membrane
elevated. 33,36 Such obvious alterations of deformability, especially under conditions
i n t e r m e d i a t e s a n d nucleotides are of relative acidosis as demonstrated by
confined to cells with severe deficiencies; Chilcote and Baehner, 16 resulting in -.elecheterozygotes show either normal cellular tive reticulocyte destruction by splenic
concentrations or only subtle deviations.
macrophages, as shown by Matsumoto
and
associates.25
In vitro rates of glucose consumption
and lactate production are quite variable
Splenectomy often results in clinical
but may be decreased, especially in com- arid hematologic improvement, this deparison with other young cell popula- spite the apparent paradox of further
tions. 31,33 Conversion of mannose to lactate, shortening of radiochromium half-lives
December 1974
GLUCOSEPHOSPHATE ISOMERASE DEFICIENCY
following splenectomy. T h e
same
phenomenon has been observed in PK
deficiency and presumably reflects the
labeling and rapid demise of markedly
deficient cells which ordinarily would not
have survived long enough to be present
in a presplenectomy specimen. 26 As in PK
deficiency, variably deficient erythrocytes
appear to be selected for destruction at
different rates.
Although heterozygous deficiency is not
accompanied by clinical symptoms or obvious hematologic alterations, when it
occurs in association with other cellular
abnormalities the combined effects may
result in overt hematologic manifestations. T h e family described by Leger and
colleagues 23 is a case in point. Both proband and mother manifested elliptocytosis
with 50% of normal erythrocytic GPI
activity levels, and both had hemolytic
anemia. A sister with the same GPI deficiency but without elliptocytosis was clinically normal. The father and brother
were devoid of either trait. Such findings
are similar to observations in partial PK
deficiency combined with hemoglobinopathies or hereditary spherocytosis. 45 In contrast, individuals with coexistent partial
deficiencies of GPI and G6PD have been
reported to be phenotypically normal. 36,37
Schroter and associates37 suggested that
G6PD deficiency might even be of benefit in
this circumstance by providing elevated G6P
concentrations to assist in overcoming the
partial GPI block.
Pathology
The glycolytic dependence of mature
erythrocytes makes it virtually certain that
biochemical lesions such as GPI deficiency
are causally related to premature loss of
affected cells, yet the precise pathogenetic
mechanism of hemolysis has not been
defined unequivocally in any erythroenzymopathy. Studies such as those by Arnold and associates3 indicate that unstable
mutant GPI isozymes may be rapidly
749
inactivated in vivo with progressively more
serious effects on cellular metabolism. The
enzymatic impairment produces alterations of glycolytic intermediates which
may be directly deleterious or may
further block glycolysis by feedback inhibition. Diminished cellular stores of ATP
and appropriate cofactors eventually become insufficient to supply specific energy
needs. Ultimately, when cation gradients
can no longer be maintained or oxidative
stresses effectively countered, membrane
alterations may result in selective destruction by the reticuloendothelial system,
especially in the spleen, where low levels
of glucose and oxygen and decreased pH
further diminish both glycolysis and
oxidative phosphorylation that may be
occurring in reticulocytes. 16,25
Gross and microscopic pathologic
changes again are nonspecific. Splenomegaly with congestion is the most common
finding, but there may 25 or may not 5,32 be
thickening and fibrosis of the cords and
lymphoid hyperplasia. 25 Hemosiderin deposition may be evident. 32 Hepatomegaly
has also been noted clinically.28,37 Selective
splenic phagocytosis of reticulocytes (but
rarely of erythrocytes) has been demonstrated by Matsumoto and colleagues, 25
who noted that erythrophagocytosis did
occur in hepatic reticuloendothelial cells.
Therapy
Theoretically, it is attractive to postulate
that in-vivo stimulation of pentosephosphate pathway activity might result in
prolongation of cell life spans by avoiding
the partial block at the GPI step. While
traversal of the shunt by a molecule of
glucose is only 5/6 as effective in ATP
yield as degradation via the EmbdenMeyerhof pathway, a positive ATP balance would still pertain. Preliminary attempts at such therapy, however, have
been ineffective whether ascorbic acid 5 or
methylene blue 2 was used as the shunt
stimulant. This may be explained by the
750
PAGLIA AND VALENTINE
observation that pentosephosphate pathway activity may already be functioning at
or near maximum capacity in severely
deficient cells.
Stimulation of glycolysis by intravenous
infusion of inorganic phosphate was also
attempted by Arnold and associates2 as a
therapeutic measure. They reported an
increase in hemoglobin from 12.7 to 14.5
Gm. per 100 ml. over a ten-day period
with a concomitant decrease in reticulocytes to about 70% of the initial level.
On the basis of in-vitro incubation
studies, mannose might be expected to
provide an effective alternative substrate
to circumvent the defective enzyme block.
Beutler and Teeple, 9 however, demonstrated that glucose competitively inhibits mannose phosphorylation, which is
also inhibited noncompetitively by its
product, mannose-6-phosphate. T h e latter compound can accumulate because
mannosephosphate isomerase may be
even less active than hexokinase.
Splenectomy currently remains the
most hopeful therapeutic measure. Although not curative, in a number of
instances splenectomy has been followed
by distinct but variable clinical improvement. 5,16,31,32,35 Chronic hemolysis continues, but hemoglobin and reticulocytes
often increase, and transfusion requirements may diminish or disappear. On the
basis of these few cases and the similar but
broader experience gained with PK
deficiencies, there appears to be a reasonable probability that splenectomy will result in clinical improvement. One reported patient, 19 however, failed to respond at all to splenectomy, so a good
prognosis cannot be assured. On the basis
of our current knowledge, patients who
are encumbered by symptoms, transfusion requirements, or frequent or severe
episodes of acute hemolysis are considered
appropriate candidates for splenectomy.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
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