[CANCER RESEARCH 28, 2548—2555,December 1968]
The Biochemical
of Patients
Identification
of Blood and Bone Marrow Cells
with Acute Leukemia
JosephF. Seitz and Irma S. Luganova
Biochemical Laboratory of the Leningrad Institute ofHematology
and Blood Transfusion, Leningrad, U.S.S.R.
SUMMARY
Undifferentiated blast white blood cells from 121 cases of
acute leukemia were investigated in relation to respiration,
glycolysis, content and transformation of glycogen, activities
of glycogen-synthesizing enzymes, pyrimidine nucleoside phos
phorylases, glucose-6-phosphate and 6-phosphogluconate de
hydrogenases, and glutathione reductase. The results are coin
pared with the analogous indexes of leukocytes of healthy
persons and of patients with chronic forms of leukemia and
polycythemia.
All types of white blood cells have been divided into two
broad metabolic groups: (a) those exhibiting aerobic glycolysis
under normal conditions of oxygen supply, and (b) those not
undergoing aerobic glycolysis. In the first group we found
granulocytes
of healthy
persons,
leukocytes
of patients
with
polycythemia and chronic myeloid leukemia, and undifferen
tiated white blood cells in the majority (62%) of patients with
acute leukemia, evidently of myeloid origin. In the second
group we found lymphocytes of healthy persons, lymphocytes
of patients with chronic lymphoid leukemia, and undifferen
tiated white blood cells of patients with acute leukemia, evi
dently of lymphoid nature (38%). It has been demonstrated
that the metabolic type of leukocytes depends primarily on
the tissue origin of the cells but not on the degree of maturity
or leukemic transformation. Both groups of blasts of patients
with acute leukemia, just as leukocytes of patients with chron
ic myeloid or lymphoid leukemia, clearly differ from one
another in turnover rate of glycogen, activities of phospho
glucomutase, unidine diphosphate glucose pyrophosphorylase,
unidine diphosphate glucose glycogen glucosyl transferase, and
also in activities
of uridine,
deoxyunidine,
cytidine,
deoxycy
tidine, and thymidine phosphorylases, and glucose-6-phos
phate and 6-phosphogluconate
dehydrogenases. Theoretical
and practical aspects of these observations are considered.
INTRODUCTION
The precise identification of undifferentiated white blood
cells in acute leukemia has great practical significance for ratio
nal therapy,
and vital theoretical
significance
for working
out a
scientifically based classification of cell elements and leuke
mias themselves. At the present time, however, this cannot be
Received
2548
January
24, 1968, accepted
August
26, 1968.
considered a solved problem. The limits of possibilities of tra
ditional morphology in this field at the present level of devel
opment evidently are exhausted. Cytochemistry, despite cer
tam successes (10, 11), still does not guarantee reliable ascer
tainment of the nature of morphologically similar but histo
genetically different blast cells; electron microscopy has
achieved successful results in other fields of biology, but in
this instance has not given decisive information. In this situa
tion, great hopes are associated with the progress of chemical
hematology, the biochemistry of blood cells and blood-form
ing tissue. The dominating tendency is obvious. It appears in
the ever-widening stream of biochemical work on hematologic
problems, particularly on the biochemistry of leukemia cells.
Two factors, however, form a significant obstacle in the path
of progress in this direction. In the first place, more on less
wide metabolic and enzymochemical analysis of white blood
cells demands at least 0.2 to 0.3 ml of cells. Practically, such a
quantity of leukocytes may be obtained from a small sample
of blood only in the case of very high leukocytosis. It is
known that a large, if not dominating, part of acute leukemias
run true to the aleukemia or the subleukemia type. This fact
sharply curtails the possibilities of research. Only patients with
more than 30—50 thousand leukocytes per cu mm can be
subjected to investigation, and even in this case at least 10 ml
of blood is needed. But this is only one of the difficulties
which the investigator faces when studying the biochemistry
of white blood cells. The other is caused by the heterogeneity
of the population of the blood cells during leukemia. Natu
rally, the characterization of one or another type of cell is
conditioned by the homogeneity of the morphologic composi
tion of the given sample. Biochemistry so far does not have
methods at its disposal for selective isolation of different
forms of white blood cells, with the exception of the separa
tion of granulocytes and lymphocytes. A partial way out of
this situation is the selection of patients with the greatest pos
sible uniformity of white blood cells or bone marrow. As is
well known, this is often the case with acute leukemia, where
the more acute the form, the less differentiated and more
uniform is the cell population in the peripheral blood and
bone marrow.
In just such a fashion, by selecting cases of acute leukemia
with large numbers of leukocytes in the blood (or in the bone
marrow) and with maximum uniformity of cell composition,
we have been able in the last ten years to investigate within
rather wide limits the chemistry and the metabolism of undif
ferentiated blast forms of different variants of acute leukemia.
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Biochemicalldentification
Luekocytes or myelokaryocytes or both of 121 patients with
acute leukemia were investigated. Results of this investigation
are given below.
MATERIALS
AND
METHODS
The patients were men and women from 17 to 60 years of
age. Primary morphologic analysis showed that, among acute
leukemias of myeloid origin, the most widespread were myelo
blastic forms (49 cases out of 121), and the number of undif
ferentiated cells varied from 75 to 98%; less frequent were the
closely related biochemically histomonocytic forms (23 out of
121), in which reticular monocytic cells predominated, coin
prising 70—90%; acute leukemia with promyelocytic cell ele
ments dominating in the blood or bone marrow was rarely
met. In 46 cases of acute leukemia (out of 121) undifferen
tiated cells of lymphoid nature dominated (lymphoblasts,
lymphoreticular, plasmatic cells, and in a number of cases,
evidently hemocytoblasts and morphologically unidentified
forms). True, the conclusion concerning the lymphoid nature
of these cells was made not only on the basis of morphologic
analysis but also on biochemical investigation, which will be
treated more fully below. The results of biochemical survey
did not always agree with the results of biochemical tests; this
is understandable if we consider the great difficulties of mor
phologic determination of undifferentiated
blast forms of
myeloid and lymphoid origin, which are outwardly extremely
alike.
It is common knowledge that many outstanding hematolo
gists have considered and still consider this problem practically
unsolvable (e.g., 5, 7, 23). As will be seen from the following
material, biochemical investigation gives additional informa
tion which ensures a much more precise identification of Un
differentiated elements. Leukocytes have been isolated from
the blood with the aid of a gelatin-citrate solution (16, 22);
lymphocytes have been obtained from the blood of healthy
persons by the method of Coulson and Chalmers (6). In these
investigations of metabolism and activities of enzymes, the
same methods were used as in our previous works (17, 20, 32).
The respiration of cells was determined manometrically in a
mixture of Krebs-Ringer phosphate, pH 7.3, and donor's blood
serum of Group IV (AB) (1 : 1), with a concentration of 0.03 5
ml of cells per 1 ml of sample. Glycolysis was determined by
the method of Barker and Summerson (4). Acetone powders
were prepared from cells by processing at —12to —15°C
with
a twentieth part of acetone and then with dry ethyl ether, free
from peroxide. The extracts were obtained by means of short
termed homogenization of acetone powders in a 0.05 M Tris
buffer, pH 7.4, with 10 mg to 1 ml of buffer and incubation
for 20 mm at 0°C.Activity of the enzymes was determined in
the supernatant after centrifugation for 10 minutes at 5000
rpm.
Activities of glucose-6-phosphate dehydrogenase and 6-phos
phogluconate dehydrogenase were determined according to
Kornberg and Horecker (1 2), glutathione reductase—according
to Racker (26), phosphoglucomutase—spectrophotometrically
in the presence of mcotinamide adenine dinucleotide phos
phate and an excess glucose-6-phosphate dehydrogenases (32),
ur@dine diphosphate
glucose (UDPG)-pyrophosphorylase
ofBlood
Cells
according to Munch-Petersen (25), and UDPG-glycogen glu
cosyl-transferase—according to Luck (1 5). UDPG-' 4C was de
termined by the method of Anderson et al. (1) with minor
modifications (32).
Activities of cytidine and deoxycytidine phosphorylases
were measured in samples of the following content : cytidine
(deoxycytidine), 4 j@moles;phosphate buffer 0.1M , pH 7.4,40
zmoles; extract (1 :100), 0.2 ml; total volume of the sample,
0.7 ml. The free ribose and deoxyribose which appeared was
determined in the deproteimzed supernatant after the nucleo
sides were removed with charcoal (2).
The activities of uridine, deoxyuridine, and thymidine phos
phorylases were determined spectrophotometrically,
generally
according in principle to Yamada (36) and Friedkin and
Roberts (8). Protein was determined according to Lowry et a!.
(14). Here it is worthwhile mentioning that numerous experi
ments have shown the absence of statistically significant dif
ferences between the investigated biochemical indexes of
blasts of the peripheral blood and those of the bone marrow in
the same patient with acute leukemia. For this reason, the
calculation of mean values was taken on the basis of the total
number of experiments with blasts, regardless of where they
were isolated, from the blood or from the bone marrow.
RESULTS
Two Metabolic Types of Blood Cells
The most fundamental metabolic indicator enabling us to
distinguish between myeloid and lymphoid cell elements, re
gardless of the degree of their differentiation from mature
forms to blasts, has been found to be aerobic glycolysis. As
seen in Table 1, all forms of leukocytes may be divided into
two groups according to whether they have the faculty for
aerobic glycolysis. According to our data, normal leukocytes
(in a sampleof cellsafterisolationfromthe donor'sblood,
there were usually about 90% granulocytes and 10% mono
nuclear cells) from patients with chronic myeloid leukemia
and undifferentiated white blood cells of more than half of all
cases of acute leukemia accumulate lactic acid in the presence
of air, providing that glucose is present in the medium. On the
other hand, lymphocytes of normal human blood, lympho
cytes of patients with chronic lymphoid leukemia, and acute
leukentic blast cells of lymphoid origin do not have this abil
ity. We designated leukocytes of the first group (with aerobic
glycolysis) as cells of Metabolic Type I, and leukocytes of the
second group as cells of Metabolic Type II. We consider that
among blood cells the faculty for aerobic splitting of glucose
into lactic acid is one of the most characteristic properties of
myeloid elements, whereas purely oxidative metabolism in
conditions of normal air supply is inherent in lymphoid cells.
The fact that undifferentiated leukocytes in acute leukemia
divide themselves into these two metabolic types indicates, in
our opinion, the existence of two large groups of acute leu
kemia—myeloid and lymphoid—according to their nature. It is
worth mentioning that both metabolic groups are hetero
geneous morphologically and biochemically. In particular,
Metabolic Type I was divided into two clearly distinguishable
subgroups (Tables la and lb. Included in Subgroup Ia are
DECEMBER 1968
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2549
Joseph F. Seitz and Irma S. Luganova
1Leukocytes02
Table
consumption
cells)With
(pinoles/hr/ml of cells)Glycolysis
AnaerobicHealthy
glucose
±2.3
persons47.6
(pinoles lactic acid/hr/mi of
Without
glucoseAerobic
62.6
±4.4
±8.9
(20)Persons
(20)
with chronic myeloid leukemia44.4
(15)Blasts
±2.9
(15)
56.6 ±3.4
(15)111.1
(15)
±2.3
(32)
61.5 ±3.9
(18)
±5.0
(44)
45.8 ±1.7
58.7 ±2.5
118.9 ±6.3
(19)
49.7 ±3.1
(11)90.0
(12)
(23)
0
(11)
of patients with acute leukemia, Type I:
Ia
lb
Lymphocytes ofhealthy
(9)Lymphocytes
persons49.7
ofpatients
(27)Blasts leukemia52.2
lymphoid
with chronic
of patients with acute leukemia, Type II55.7
(20)156.6
257.8
±8.9
205.5
±6.8
(20)
±6.3
136.7 ±8.9
(30)
156.7
181.0
±3.1
(22)
50.9 ±5.4
(8)0
(27)
±3.4
(21)
53.6 ±5.8
(7)0
(46)
±10.4
(17)
180.0 ±17.0
±6.6
126.6 ±8.9
(19)
Respiration and glycolysis in human white blood cells in the normal state and in leukemia. Numbers in parentheses indicate
number of persons.
undifferentiated white blood cells of patients with acute mye
loblastosis, morphologically described as myeloblasts; in Sub
group lb were included cases of acute histomonocytic leuke
mia with a predominance in the blood or bone marrow of
undifferentiated monocytic cells.
Metabolically, there is complete analogy of Type I and Type
II of acute leukemia with chronic myeloid and chronic
lymphoid leukemia. Leukocytes of patients with chronic mye
losis and acute myeloid leukemia (metabolic Type I) have the
potential for aerobic glycolysis. As a result of this quality,
both categories of cells have the inverse Pasteur reaction (Crab
tree effect); respiration decreases when glucose is added. We
have noticed this connection of aerobic glycolysis with the
inverse Pasteur reaction many times in various cell types, and
this was interpreted as the result of competition of respiration
and glycolysis for adenylic acceptor and inorganic phosphate
(27, 29—31,33).
The picture is different in cells of lymphoid origin, regardless
of the degree of their differentiation. Mature lymphocytes of a
healthy person, mature and relatively mature lymphocytes of
patients of chronic lymphadenosis, undifferentiated blasts of
patients with acute lymphadenosis, acute plasma cell leukemia,
and acute lymphoreticulosis do not have aerobic glycolysis
and, correspondingly, do not have the inverse Pasteur reaction
(Crabtree effect).
From these results we may draw several conclusions of major
significance. In the first place, aerobic glycolysis is not a sign
of malignancy inasmuch as normal granulocytes have this fac
ulty, but leukemic lymphocytes are devoid of it. It is interest
ing that the presence of aerobic glycolysis in myeloid cells and
the absence of it in lymphoid ones is so fundamental a prop
erty of each of these lines of cells that the processes of malig
nant transformation or cell differentiation do not affect it.
2550
Indeed, aerobic glycolysis is observed in normal as well as in
leukemic granulocytes and in mature polymorphonuclear leu
kocytes as well as in undifferentiated blasts. In principle, the
mechanisms of regulation of respiration and glycolysis in nor
mal and leukemic cells are identical : the Pasteur effect and the
inverse Pasteur reaction in leukemia qualitatively do not
change, though quantitatively a slight weakening of the Pas
teur effect is observed in blasts in acute leukemia. This is not
observed in leukocytes of patients with chronic myeloid leuke
mia when compared with normal leukocytes.
We should mention also a noticeable decrease in the glyco
lytic activity of blasts, in comparison with normal leukocytes
with a practically invariable level of respiratory activity. How
ever, among cells of different variants of acute leukemia of
Type I there is no essential difference in respiration or glycoly
sis.
The discovery of aerobic glycolysis in leukocytes of myeloid
origin and purely oxidative metabolism in lymphoid cells has
great theoretical significance, inasmuch as it exposes the link
of these energy-giving processes with certain classes of tissue,
but not with specifically pathologic (cancer, leukemia) or nor
mal physiologic (differentiation) processes. Moreover, the exis
tence of different types of metabolism in the very youngest
representatives of the myeloid and lymphoid series indicates
the relative autonomy of these two systems and, perhaps, of
the remoteness of their common precursor. On the other hand,
in a practical sense the test for aerobic glycolysis allows us to
distinguish between outwardly very similar blast forms of
lymphoid and myeloid origin in acute leukemia, which en
hances the possibility of rational use of one or another type of
medical treatment.
Below we shall see that the division of white blood cells into
two metabolic types is not just a formal measure, but reflects
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Biochemical Identification
ofBlood
Cells
Table 2
(mm)(@zgJml
transferaseHealthypersons6160±550
LeukocytesGiycogen@tmoies/gm
(6)Patients
withpolycythemia9610
(16)Patients
with chronic myeloid leukemia4540
(5)Blasts
of cells)Specific
protein
activity
in cpm/mgPhosphoglucomutaseUDPG-pyrophosphorylaseUDPG-glycogen
(15)1098±168
(15)80±7
(20)52±5
±450
(12)1646
±185
(12)96
(16)72
±260
(15)16128
±852
(15)64
(17)48
(17)2.8
(14)7745±1010
(14)36±6
(14)13±2
(14)2.7±0.5
±72
(13)21196
±2554
(13)60
±6
(18)15
±1
(18)3.8
±1
±1
(20)4.7±0.3
±4.0
±11
±4
±4
±0.3
of patients with acute leukemia
Type I:
Ia940±112
(4)lb711
(9)Lymphocytes
ofhealthy
persons933
±111
(5)2127
(4)Lymphocytes
of patients with chronic
lymphoid leukemia1600
(5)Blasts
of patients with acute leukemia,
Type II662
±700
±0.7
(4)6
(5)9
±160
(14)2020
±314
(14)10
±1
(13)10
±1
(13)1.2
±0.2
±51
(17)6008
±644
(17)9
±1
(21)4.0
±0.5
(21)0.9
±0.2
(8)
Glycogen content, its turnover rate and the activities of enzymes of the biosynthesis of glycogen in normal and leukemic human Ieukocytes. UDPG, un
dine diphosphate glucose. Numbers in parentheses indicate number of persons.
deep enzymochemical and functional differences in myeloid
and lymphoid tissues. This is demonstrated especially clearly
in the case of the glycogen system.
Glycogen and Glycogen Synthesizing Enzymes in
Leukocytes in Acute Leukemia
In a healthy person, the glycogen content in granulocytes is
much greater than its level in lymphocytes. In polycythemia
and chronic myelosis, the glycogen content changes in oppo
site directions: in the first case it sharply increases, in the
second, it decreases (18, 28—30,34, 35).
Along with the activity of alkaline phosphatase, the changes
in the glycogen level in leukocytes, registered biochemically or
cytochemically, may serve as a reliable means of demarcation
in some complicated cases of polycythemia and chronic mye
losis. Still more striking are the data on incorporation of glu
cose-14C label in the glycogen of leukocytes (18, 28—30). In
standard experimental conditions (18), specific radioactivity
of glycogen in chronic myeloid leukemia exceeds the normal
by 15 times but in polycythemia by only 1.5 times (Table 2).
Leukocytes of metabolic Types I and II of acute leukemia
retain differences also in their indexes of glycogen turnover.
As seen in Table 2, specific radioactivity of polysaccharide in
blasts with aerobic glycolysis (Type I) is higher than in cells of
Type II and, in subgroup Ib, exceeds the corresponding index
in blasts of Type II by 3.5 times and is close to the index of
leukocytes of patients with chronic myeloid leukemia.
DECEMBER
The activities of glycogen-synthesizing enzymes in undif
ferentiated leukocytes of two types of acute leukemia (Table
2) are very characteristic. It is known that the synthesis of
glycogen in animal cells takes place with the participation of
UDPG and three enzymes: phosphoglucomutase, UDPG-pyro
phosphorylase and UDPG-glycogen glucosyl transferase. The
whole process of synthesis of this polysaccharide, beginning
with glucose, may be schematically depicted in the following
manner:
glucose + ATP hexokinas@ glucose-6-phosphate
+ ADP
glucose-6-phosphate
phosphoglucomutase
glucose-i-phosphate
glucose-i-phosphate
+ UT? UDPG-pyrophosphorylase
UDPG
pyrophosphate
UDPG + glycogen(fl@G@Yc0@en
glucosyl-transferas@jr@p
glycogen(@@1)
A study of enzymes participating in the biosynthesis of
glycogen showed that in blasts of Type II, the activity of
phosphoglucomutase was 4 times less and in UDPG-pyrophos
phorylase and UDPG-glycogen transferase was 3 times less
than in myeloblasts belonging to the first metabolic group
(subgroup Ia). Blasts of Type II differ even more strongly
from undifferentiated cells of subgroup lb (monocytic-like un
1968
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2551
Joseph F. Seitz and Irma S. Luganova
@
differentiated cells). This indicates that biologic differences
exist not only between blasts of Type I and II of acute leuke
mia but that they also exist among cells of metabolic Type I of
acute leukemia: practically all the indexes of enzyme activity
and specific radioactivity of glycogen in Subgroup lb are much
higher than in Subgroup la (Table 2).
Worthy of attention is the fact that the activities of glyco
gen-synthesizing enzymes in Type I cells of acute leukemia,
especially subgroup lb are much nearer to those in leukocytes
of patients with chronic myelosis than to blasts of metabolic
Type II, while the latter are very close to lymphocytes of
patients with chronic lymphadenosis and even to lymphocytes
of healthy persons.
We consider these results as still further evidence that undif
ferentiated white blood cells of the first metabolic type have
originated from some primary myeloid elements and the ana
logous cells of Type II from lymphoid ones. At the same time
we must admit that the least differentiated of the investigated
leukocytes, myeloblasts (metabolic Type I) and lymphoblasts
(metabolic Type II), have between them much more enzymo
chemical similarity than the more mature forms of myeloid
and lymphoid series. One might think that even earlier precur
sors of these forms would be less distinguishable, if not identi
cal. The cell in which are blended not only morphologic but
chemical features of two lines of blood cells should be the
primary stem cell. It is possible that its development in one or
another direction is determined by the environment, that is,
the tissue in which it exists.
parison of activities of pyrimidine nucleoside phosphorylases.
This
of enzymes
catalyzes
reversible
uridine + phosphate
phosphorolytic
‘.
uracil
+ ribose-1-phosphate
The activities of uridine, deoxyuridine, cytidine, deoxycyti
dine, and thymidine phosphosylases (Table 3) were tested in
extracts of acetone powder of undifferentiated leukocytes of
patients with acute leukemia.
As a rule, the activities of blasts of metabolic Type I were
higher than blasts of Type II. The difference was especially
noticeable in this respect between Subgroup lb and Group II.
Only deoxycytidine phosphorylase in Subgroup Ia (myelo
blasts) was lower than in blasts of Group II. The other en
zymes in cells with aerobic glycolysis were much more active
than in undifferentiated cells of lymphoid nature. The biggest
differences, as seen in Table 3, were shown by deoxyuridine
phosphorylase, uridine phosphorylase, and thymidine phos
phorylase. Therefore, in practice, it is enough for identifica
tion of blasts to determine the activity of these three enzymes,
or even of one of them, for instance, deoxyuridine phosphory
lase.
Glucose-6-phosphate Dehydrogenase, 6-Phosphogluconate
Dehydrogenase and Glutathione Reductase in Leukocytes
of Patients with Acute Leukemia
The changes
Pyrimidine Nucleoside Phosphorylases in Human
Leukocytes in Acute Leukemia
6-phosphogluconate
of glucose-6-phosphate
dehydrogenase
dehydrogenase
in leukocytes
and
of patients
with acute leukemia are subject to the general rule noted for
other enzymes: their activity is reduced, usually much more
sharply in the case of acute leukemia of the second type.
Further delimitation of leukocytes of patients with acute
leukemia of metabolic Type I or II may be achieved by coin
3LeukocytesPhosphorylases
group
splitting of pyrimidine nucleosides into pyrimidine base and
ribose-1-phosphate as follows:
Table
protein/mm)UridineDeoxyunidineCytidineDeoxycytidineThymidineHealthy
(@tmoles/gm
persons4.6
(7)Chronic
±0.5
(10)26.1
(10)4.2
and acute myeloid leukemia4.2
(8)Type
±0.5
(8)11.5
(6)3.2
(10)9.8
(10)12.7
±0.5
(9)6.0
±1.2
(9)1.4
±0.1
(4)1.9
±0.1
(4)3.1
±0.5
±0.5
(17)23.1
±2.8
±0.2
±0.6
±1.8
(17)1.5
(9)4.7
(9)10.8
±0.2
(9)19.4
±1.9
(9)0.9
±0.1
(9)1.5
±0.2
(8)15.6
±0.1
(6)2.7
±0.4
(6)1.6
±2.0
±1.3
±0.2
(9)14.5
±0.3
±1.0
±0.6
(9)19.0
±0.8
±0.9
I
Ia2.3
(9)lb5.7
(12)Chronic
lymphoid leukemia0.9
(8)Acute
leukemia, Type II1.0
Activities of pyniinidine nucleoside phosphonylases
parentheses indicate number of persons.
2552
±1.7
±0.2
(11)2.4
±0.5
(11)0.7
in leukocytes
of healthy persons and leukemic patients. Numbers in
±0.3
(8)
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Biochemicalldentification
In blasts of Subgroups Ia and lb (Metabolic Type I), the
activity of glucose-6-phosphate dehydrogenase is lower than
that of normal leukocytes by 3 and 2 times, correspondingly,
and in blasts of Type II, by 10 times. For 6-phosphogluconate
dehydrogenase these ratios are 2.4, 1.5, and 4.8.
Glutathione reductase is, apparently, one of the very few
enzymes whose activity in blasts of metabolic Type II is higher
than in Type I blasts. Moreover, its activity in myeloblasts
(Subgroup Ia) is higher than in undifferentiated monocytic
cells (Subgroup Ib). Evidently, leukemic transformation of
cells is not accompanied by a decrease in the activity of this
enzyme.
Cells
based on the synthesis of new proteins, nucleic acids, and
other important compounds), and ending with the specific
function—retain their effectiveness in cells of all stages of
maturity. In most cases only the relation of oxidative and
glycolytic mechanisms of energy generation changes.
As a result of this work, the problem of aerobic glycolysis is
seen in a new light. Earlier we suggested that aerobic glycolysis
is not a pathologic but a normal physiologic phenomenon con
nected with intense functional activity: mechanical work, in
tensified biosynthesis during proliferation, and so forth (27,
29, 30). Now aerobic glycolysis comes forth as a characteristic
peculiarity of metabolism of cell forms of myeloid nature.
According to this thesis, all elements of bone marrow origin
should have the faculty of aerobic glycolysis. Indeed, it is
known that this ability is inherent in white cells of the bone
marrow (16), as well as in thrombocytes (21 , 37), erythro
cytes, and reticulocytes (24, 27). Evidence is lacking only con
cerning erythroblasts. Investigation of the latter would be very
interesting and important from this point of view. However, at
the present time the impossibility of obtaining the nuclear
precursors of erythrocytes in isolated form makes the deter
mination of their type of metabolism difficult. In agreement
with the conception put forth concerning the division of mye
bid and lymphoid tissue according to the type of metabolism,
erythroblasts should accumulate lactic acid in the presence of
air. This assumption calls for experimental verification.
Opposite to the bone marrow in its metabolism stands
lymphoid tissue, having a purely oxidative metabolism in the
presence of oxygen. As shown in the results set forth, even the
DISCUSSION
On considering the results of these experiments, we are
struck by certain points. One of these is the fact that, with the
exception of the systems of respiration and glycolysis, practi
cally all the enzyme activities in blast cells of acute leukemia
are decreased, sometimes very sharply. This fact may be con
sidered in relation to the observation that proliferative activity
of blast cells in acute leukemia is lowered (3, 9, 13). On the
other hand, we must keep in mind that the enzymes studied
are connected with one or another function in mature dif
ferentiated cells. It is natural that in young undifferentiated
forms, where morphologic processes predominate, they are un
developed. At the same time, the most universal processes
respiration and glycolysis, which give energy to all cell activity
beginning with proliferation and differentiation (which is
4Leukocytes(zmoies/gm
ofBlood
Table
protein/mm)Glucoie-6phosphate
dehydrogenaseGiutathione
dehydrogenase6-Phosphogiuconate
reductaseHealthy
persons196.0
(10)Patients
±12.0
(19)20.7
(5)17.7
with chronic myeloid leukemia164.0
(6)Blasts
±8.5
(15)15.7
(6)11.4
(15)8.6±1.3
(5)13.1±1.4
(17)13.6±2.0
(5)9.1±1.2
±12.8
(4)6.4
±0.4
(2)11.7
±0.8
(2)Lymphocytes
of patients with chronic
lymphoid leukemia30.0
(6)Blasts
±2.3
(13)3.7
±0.3
(6)19.0
±0.2
±2.1
(21)4.3
±0.6
(6)14.9
±0.8
±3.5
±1.6
±2.4
of patients with acute leukemia, Type I
Ia61.4±6.6
(5)lb92.1±5.5
(5)Lymphocytes
ofhealthy
persons60.1
of patients with acute leukemia, Type II18.4
Activities
of glucose-6-phosphate
dehydrogenase,
6-phosphogluconate
dehydrogenase,
and glutathione
±2.5
(9)
reductase
in white
blood cells of healthy persons and leukemic patients. Numbers in parentheses indicate number of persons.
DECEMBER 1968
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1968 American Association for Cancer Research.
2553
Joseph F. Seitz and Irma S. Luganova
leukemic transformation of lymphocytes does not produce
aerobic glycolysis. Oxidative metabolism of lymphocytes is
evidently a stable metabolic sign of all lymphoid tissue and its
derivatives.
Thus, we know now of the existence of normal, physiologi
cally sound blood cells having aerobic glycolysis (all myeloid
elements) and of typically neoplastic cells not having aerobic
glycolysis (acute leukemic blasts of Type II, lymphocytes of
patients with chronic lymphoid leukemia). In the light of these
facts, the conception of a so-called cancerous metabolism,
based on the recognition of aerobic glycolysis as a specific sign
of neoplasia, is deprived of experimental basis.
One of the leading phenomena ofleukemic as well as cancer
ous transformation is the conflict between the proliferative
potential of the tissue and its normal functions. The loss of @he
cells' ability to use chemical materials and energy in positive
functional ways in some manner or other opens the pathway
for their utilization for uncontrolled growth. In this light, the
investigation of glycogen in leukemic cells, closely linked with
specific physiologic function, is especially interesting. Detailed
analysis has certainly shown that a marked tendency exists
toward decrease of the glycogen level and activity of glycogen
synthesizing enzymes in leukemic cells. For example, within
the limits of one tissue line—myeloid—the glycogen content
and activity of the enzymes connected with its biosynthesis
progressively falls from the norm to chronic myeloid leukemia
and acute myeloid leukemia. Another regularity stands out no
less sharply.
In relation
to the synthesis
and accumulation
of
glycogen, lymphoid cells are much behind myeloid cells of
approximately the same degree of maturity.
These facts have another aspect. The significant difference in
glycogen content and its turnover rate in leukemic and nonleu
kemic
cells, or myeloid
and lymphoid
cells, reveals an addi
tional possibility of biochemical identification of individual
cell forms. Along with the test for aerobic glycolysis and cer
tain other biochemical parameters, the investigation of glyco
gen in leukocytes in leukemia is widely used in the clinic of
our institute for precision in diagnosis and for obtaining more
profound characteristics of cells of the blood and bone mar
row.
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15. Luck, D. J. Glycogen Synthesis from Uridine Diphosphate Glucose.
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Glycolysis
and Glycogen
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Bio
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17. Luganova, I. S., and Seitz, J. F. Concerning the Qualitative Charac
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Bull. Exptl. Biol. Med., 46: 57—59,1958.
18. Luganova, I. S., and Seitz, J. F. Glycogen Content and Its Metabo
lism in Human Leucocytes in the Normal State and in Leukemia.
Vopr. Med.Khim., 8: 354—361,1962.
19. Luganova, I. S., and Seitz, J. F. The Activity of Enzymes of the
System of Glycogen Synthesis in Human
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ACKNOWLEDGMENTS
21. Luganova,
We wish to express our thanks for the morphologic research to L. M.
Rozanova, V. V. Kuraleva, N. S. Povergo, and V. A. Freydzon and to S.
I. S., Seitz, J. F., and Teodorovich,
V. I. Metabolism
in
Human Thxombocytes. Biokhimiya,23: 405—411,1958.
22. Luganova, I. S., Yegorova, V. A., and Seitz, J. F. Methods of
I. Sherman, for valuable discussionon hematologic questions.
Isolating
Metabolically
Active Leucocytes
and Thrombocytes
from
Human Blood. McI. Vopr. Hematol. and Perelivaniya Krovi, 14:
215—221,1963.
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DECEMBER 1968
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2555
The Biochemical Identification of Blood and Bone Marrow Cells
of Patients with Acute Leukemia
Joseph F. Seitz and Irina S. Luganova
Cancer Res 1968;28:2548-2555.
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