Fetal Hemoglobin, The Neonatal Red Cell, and 2,3-Diphosphoglycerate

Symposium on Pediatric Hematology

Fetal Hemoglobin, The Neonatal Red Cell, and 2,3-Diphosphoglycerate Frank A. Oski, M.D. *

Five years have now elapsed since the original reports 7 • 15 concerning the modulating effects of organic phosphates on hemoglobin's affinity for oxygen appeared. This important observation rekindled interest in the role of the red cell in oxygen transport. In this short space of time a voluminous literature dealing with both the biochemical and clinical ramifications of this subject has accumulated. Since several comprehensive reviews of this topic are now available,1114.31 this paper will focus on only those aspects of the problem that are of special interest to the pediatrician, after a preliminary discussion of oxygen transport.

OXYGEN TRANSPORT AND RELEASE Human tissue metabolism is critically dependent on an adequate supply of oxygen. The oxygen transport system in man is the erythrocyte. The erythrocyte serves this role because it contains the iron-protein conjugate, hemoglobin. The red cell's primary function is to bring oxygen to the tissues in adequate quantities, at a sufficient partial pressure, to permit its rapid diffusion from the blood. The ultimate supply of oxygen to the cell is determined by a number of factors which include: the content of oxygen in the inspired air; the partial pressure of oxygen in the inspired air; the pulmonary and alveolar ventilation; the diffusion of oxygen from the alveolar air to the capillary bed; the cardiac output; the blood volume; the hemoglobin concentration; and the passive diffusion of oxygen from the capillaries to the cells. The initial passive diffusion of oxygen from the lungs and its final release to the tissues is determined in large part by the affinity of hemoglobin for oxygen. The oxygen-hemoglobin equilibrium curve reflects the affinity of hemoglobin for oxygen (Fig. 1). As blood circulates in the normal lung, arterial oxygen tension rises from 40 mm. Hg and reaches approximately *Professor of Pediatrics, State University of New York, Upstate Medical Center, Syracuse, New York Supported in part by grants from The John A. Hartford Foundation, Inc., and U.S. Public Health Service (HD-06219).

Pediatric Clinics of North America- Vol. 19, No.4, November 1972

907

908

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Oxygen Dissociation

c:::

80

.~

15 ~ 60

~

~ 40

cl' ~

20 • "p 50" Values

40

60

Figure 1. The oxygen dissoeiation curve of normal adult blood. The Po", the oxygen tension at 50 per cent oxygen saturation, is approximately 27 mm. Hg. As the curve shifts to the right, the oxygen affinity of hemoglobin decreases and more oxygen is released at a given oxygen tension. With a shift to the left, the opposite effects are observed. A decrease in pH or an increase in temperature decreases the affinity of hemoglobin for oxygen.

80

110 mm. Hg, sufficient to ensure at least 95 per cent saturation of the arterial blood. The shape of the curve is such that a further increase in the oxygen tension in the lung results in only a very small increase in the degree of saturation of the blood. As blood travels from the lung, the oxygen tension falls, as oxygen is released to the tissues from hemoglobin. In the normal adult when the oxygen tension has fallen to approximately 27 mm. Hg, at a pH of 7.4 and a temperature of 37° C., 50 per cent of the oxygen bound to hemoglobin has been released. The P 50 , the whole blood oxygen tension at 50 per cent oxygen saturation, is thus stated to be 27 mm. Hg. When the affinity of hemoglobin for oxygen is reduced, more oxygen is released to the tissues at a given oxygen tension. In such situations the oxygen-hemoglobin equilibrium curve is shifted to the right of normal. It has long been recognized that increases in blood acidity, carbon dioxide content, ionic concentration, or temperature are capable of decreasing the affinity of hemoglobin for oxygen and thus shifting the curve to the right. When the affinity of hemoglobin for oxygen is increased, such as occurs with alkalosis or a decrease in temperature, the equilibrium curve appears shifted to the left and the tension must drop lower than normal before the hemoglobin releases an equivalent amount of oxygen. A certain partial pressure gradient is needed for the transfer of oxygen from capillary to tissue. Below this critical capillary oxygen tension, diffusion is impaired and tissue hypoxia results. Kety23 and Landis and Pappenheimer25 have stated that the end-capillary oxygen tension necessary for adequate tissue oxygenation is at least 20 mm. Hg. In support of this figure is the observation of Opitz and Schneider29 who demonstrated that brain oxygen uptake decreased when the venous oxygen tension fell into the 20 to 25 mm. Hg range. Similarly, Berne and associates 9 found that coronary vasodilatation occurred when the arterial oxygen pressure

THE NEONATAL RED CELL AND 2,3-DIPHOSPHOGLYCERATE

909

was 22 to 24 mm. Hg and that loss of myocardial function was severe at an oxygen tension of 10 to 12 mm. Hg. A critical capillary oxygen tension cannot be precisely defined for all tissues under all metabolic conditions because of the inherent variability in requirements from tissue to tissue. Benedixen and Laver6 suggest that the term "average critical range" be employed and place this value between 20 and 30 mm. Hg. At a normal pH and temperature this represents an oxygen saturation in the range of 35 to 55 per cent. Thus a normal individual with a hemoglobin concentration of 16 gm. per 100 mi. having an arterial oxygen content of 22.2 volumes per cent (16.0 x 1.39 mi., the oxygen-carrying capacity of 1 gm. of hemoglobin) would have between 7.7 and 12.1 volumes per cent of oxygen unavailable to the tissues. The average oxygen uptake is approximately 5 volumes per cent. Contributing to this average is the very small arteriovenous oxygen difference in the kidneys, 1.5 volumes per cent, and the large arteriovenous oxygen difference of 11.5 volumes per cent in working muscle and the heart. These latter organs have no oxygen reserve because they are extracting as much oxygen as possible from the blood at the optimal partial pressure for diffusion. Under conditions of increased oxygen need or decreased blood oxygen-carrying capacity, such as occurs with anemia, the body may improve oxygen delivery to the tissues by either improving oxygen transport or changing the affinity of hemoglobin for oxygen. Improved oxygen transport is achieved by increasing the cardiac output or by increasing the oxygen-carrying capacity of the blood by a rise in the hemoglobin concentration. An increase in cardiac output is a rapid compensation but is limited in its magnitude. Increasing cardiac output requires an increase in oxygen consumption which tends to offset its compensatory advantage. The second mechanism of improving oxygen transport, that of decreasing the affinity of hemoglobin for oxygen, allows more oxygen to be delivered to the tissues at an equivalent or even higher partial pressure of oxygen. As a result the end capillary tissue oxygen gradient is maintained above the critical range despite a decrease in the degree of saturation of the blood. It is this latter mechanism which results from the interaction of red cell organic phosphate compounds with hemoglobin that has most recently been recognized as a common, rapid, and efficient means of increasing tissue oxygen delivery. The basis for this interaction stems from the properties of both the hemoglobin and the glycolytic metabolism of the human red cell.

Fetal Hemoglobin and the Organic Phosphates Benesch and Benesch7 and Chanutin and Curnish 15 demonstrated that a variety of organic phosphates, when added to adult hemoglobin in solution, had the ability to reduce the affinity of hemoglobin for oxygen. Of the organic phosphates tested, the compound 2,3-diphosphoglycerate (2,3-DPG) appeared to be the most potent modifier of hemoglobin function. Of the organic phosphates normally found in human erythrocytes, 2,3-DPG is present in highest concentration, averaging approximately 5

910

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Table 1.

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Effect of Added Diphosphoglycerate on Oxygen Affinity of Adult and Fetal Hemoglobin':' MATERNAL

SAMPLE

(n = 8) Mean Standard Value Deviation

FETAL

(n = 8) Mean Standard Value Deviation

---------

p:')()

Whole Blood

31.0

±1.4

24.3

±2.4

19.7

±1.3

19.7

±0.9

29.4

±1.5

22.4

±1.4

p.'lU

Dialyzed hemoglobin P so Dialyzed hemoglobin + DPG

Half saturation pressures (P oo ) in mm. Hg at pH 7.20 (in blood pHc = 7.20) and 37°C., in maternal and fetal blood and hemoglobin solutions without and with DPG (30 millimoles per gm. of hemoglobin). n

=

number of samples

':'From Bauer, C., Ludwig, I., and Ludwig, M.: Life Sciences, 7:1339, 1968.

millimoles per mI. of red cells, and thus is quantitatively the most important with respect to modulating hemoglobin's affinity for oxygen. After the role of 2,3-DPG in altering the affinity of adult hemoglobin for oxygen was recognized, numerous investigators examined its interaction with fetal hemoglobin in an attempt to determine why the fetal erythrocyte demonstrates a higher affinity for oxygen than the adult erythrocyte. In 1930, Anselmino and Hoffman 2 first observed that the oxygen affinity of human fetal blood was greater than that of maternal blood. Fetal blood had a P 50 value some 6 to 8 mm. Hg lower than that of the normal adult. In 1953, Allen, Wyman, and Smith1 demonstrated that although intact fetal cells possess a higher affinity for oxygen than do the red cells of adults, when adult and fetal hemoglobin solutions were dialyzed against the same buffer, the resulting oxygen affinities were identical. This puzzling observation was resolved by the finding that the affinity of fetal hemoglobin for 2,3-DPG is far less than that of adult hemoglobin. 5.19,27,38 Table 1, taken from the work of Bauer and associates,5 illustrates that when 2,3-DPG is added to solutions of fetal hemoglobin the decrease in oxygen affinity produced by this compound is much less than that observed with adult hemoglobin. The fetal hemoglobin obtained from the red cells of patients with beta-thalassemia behaves in a manner identical to that of fetal hemoglobin obtained from the erythrocytes of the newborn infant.27 From these studies it appears that the major reason that the blood of the newborn infant possesses an oxygen-hemoglobin equilibrium curve that is shifted to the left of that of the normal adult is due to the failure of fetal hemoglobin to bind 2,3-DPG to the same degree as does adult hemoglobin. The position of the oxygen-hemoglobin equilibrium curve in the neonate is determined by the relative proportions of adult and fetal hemo-

I

I

THE NEONATAL RED CELL AND 2,3-DIPHOSPHOGLYCERATE

911

globin present and the red ceIl2,3-DPG concentration. Infants with similar fetal hemoglobin concentrations can have different P 50 values if they differ significantly in their red ce1l2,3-DPG concentrations; alternatively, infants with similar 2,3-DPG concentrations may have dissimilar P 50 values if they differ in their per cent fetal hemoglobin. The need to consider both the proportion of adult and fetal hemoglobin and the 2,3-DPG content of the cells explains why previous investigators failed to find a direct relationship between fetal hemoglobin values alone and the position of the curve. Employing all three variables, Orzalesi and Hay30 developed the term, "effective DPG fraction." The effective DPG fraction bears a precise relationship to the P 50 value and is similar to the "functioning DPG fraction" of Delivoria-Papadopoulos and associates. IH The observed difference in interaction between adult and fetal hemoglobin and 2,3-DPG helped to facilitate provisional determination of the site on the hemoglobin molecule at which 2,3-DPG is bound. Benesch and co-workers originally proposed that 2,3-DPG bound to adult hemoglobin somewhere on the beta chains. 8 This hypothesis was based on their finding that 2,3-DPG bound to deoxyhemoglobin in a 1: 1 molar ratio, and bound to beta4 hemoglobin (hemoglobin H) both in the oxy and deoxy states, but did not bind to isolated alpha chains. It was reasoned that since the hemoglobin tetramer is a symmetric molecule composed of dimer half molecules separated by a diad axis of symmetry that a single binding site common to both half molecules and more accessible in the deoxy confirmation would be necessary to explain the observed experimental data. It was also reasoned that since 2,3-DPG has five titratable protons per molecule and at a physiologic pH possesses four negative charges, the binding site would presumably be positively charged. DeVerdier and Garby l9 suggested the beta-143 histidine as a likely binding site. This positively charged amino acid residue is located at the entrance to the central cavity and could form electrostatic bonds with anions such as 2,3-DPG. In fetal hemoglobin, this position on the gamma chain is occupied by the neutral amino acid serine, which could not bind negatively charged compounds. Bunn and Briehp2 have provided convincing evidence to indicate that the beta-143 histidine cannot be the sole site of binding in view of the fact that fetal hemoglobin does show some interaction with 2,3-DPG. In their study they noted the changes in oxygen affinity produced by the addition of 2,3-DPG to a number of purified, naturally occurring, and naturally modified hemoglobins. Hemoglobin A was found to demonstrate a nearly twofold decrease in oxygen affinity following the addition of 2,3-DPG, while the change observed with hemoglobin F was only 20 per cent. A 23 per cent decrease in oxygen affinity was noted with hemoglobin All'. This minor hemoglobin component accounts for about £3 per cent of the hemoglobin prepared from adult hemolysates and differs from hemoglobin A only in having a hexose residue bound to the N-terminal groups of the beta chains by a Schiff base linkage. No change in oxygen affinity was noted when 2,3-DPG was added to hemoglobin Fl. This minor component of hemoglobin F comprises about 15 per cent of the total, and differs from hemoglobin F in that each N-terminus of the gamma chain is covalently linked to acetyl residues. Thus, when both the beta-143 histidine and the beta chain N-

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terminals are fully reactive, as in hemoglobin A, maximal modification of oxygen affinity occurs following the addition of 2,3-DPG. When the nonalpha chain N-terminal is blocked (hemoglobin Ale) or the beta-143 histidine is absent owing to its replacement with an uncharged serine in the gamma chain (hemoglobin F), reactivity to 2,3-DPG is reduced in nature. When the N-terminal is blocked and the beta-143 histidine is absent (hemoglobin F I), the hemoglobin molecule is completely inert to the effects of 2,3-DPG. Greer and Perutz,21 utilizing a model of hemoglobin built to the 3.5 angstrom coordinates, were able to fit a single molecule of 2,3DPG in the entrance of the central cavity of hemoglobin when it was in the deoxy confirmation. The stereochemistry of the oxygenated molecule did not allow for 2,3-DPG binding at this site.

Fructose ~ SOH

NADH NAD

J'I

Sorbitol NADP~ NADPH AR Glucose

..JI

Glycogen

t

Hexose Monophosphate Shunt

I HK

ATP ..... ADP ~

, PGM Glucose-1-P .. • Glucose-6-P

G-6-PO

r,

6-Phosphogluconate

NADP NADPH ~ NADP 6-PGO Fructose-6-P..... __ NADPH

+GPI

ATP)

- - - - __

-----Ribulose-5-P

PFK

ADP

Fructose-1 ,6-di P L -_ _ _ _--:'..L-_ _ _ _--...J I

~/

J"Aldolase......

"

ii

Dihydroxyacetone-P

.. Glyceraldehyde-3-P NAD Pi PGD DPG,!--- NADH ____ 1,3-Diphosphoglycerate

TPI

2,3-Diphosphoglycerate

~

:~:j

PGK

3-Phosphoglycerate

!

PGM

2-Phosphoglycerate

l

Enolase

2 -Phosphoenol pyruvate ADP

,I

ATP..I+ PK

Pyruvate NADH)j LD NAD1 H Lactate

Figure 2. The metabolism of the human erythrocyte. HK, hexokinase; GPI, phosphoglucose isomerase; PFK, phosphofructokinase; TPI, triosephosphate isomerase; PGD, phosphoglyceraldehyde dehydrogenase (glyceraldehyde phosphate dehydrogenase); PGK, phosphoglyceric acid kinase; PGM, phosphoglyceromutase; DPGM, diphosphoglyceratemutase; DPGP, diphosphoglycerate phosphatase; PK, pyruvate kinase; LDH, lactic dehydrogenase; G-6-PD, glucose-6-phosphate dehydrogenase; 6-PGD, phosphogluconic dehydrogenase; PGM, phosphoglucomutase; SDH, sorbitol dehydrogenase; AR, aldose reductase.

THE NEONATAL RED CELL AND 2,3-DIPHOSPHOGLYCERATE

913

2,3-Diphosphoglycerate Metabolism in the Erythrocytes of the Newborn The level of red cell 2,3-DPG gradually increases with gestation and at term its concentration within the infant's erythrocytes is similar to that of the normal adult. IS. 30 The 2,3-DPG levels fall transiently during the first several days of life and then rise. 22 By the end of the first week of life, in the term infant, the 2,3-DPG levels are considerably higher than they are at birth. IS . The red cell synthesizes 2,3-DPG from 1,3-DPGin the presence of the enzyme diphosphoglycerate mutase (Fig. 2). The 2,3-DPG formed is then eventually hydrolyzed to 3-phosphoglycerate and inorganic phosphate by the enzyme 2,3-diphosphoglycerate phosphatase. The conversion of glyceraldehyde-3-phosphate to 1,3-DPG is regulated by the NAD/NADH ratio and the concentration of inorganic phosphate, both co-factors for the glyceraldehyde-3-phosphate dehydrogenase reaction. The conversion of 1,3-DPG to either 2,3-DPG or 3-PGA is governed in part by the concentration of free, or unbound, 2,3-DPG within the cell, the level of 3phosphoglycerate, and the ADP/ATP ratio. High concentrations of free 2,3-DPG inhibit 2,3-diphosphoglycerate mutase, the enzyme responsible for its synthesis. Increased concentrations of ADP facilitate conversion of 1,3-DPG to 3-PGA. High levels of 3-PGA appear to inhibit the phosphoglycerate kinase reaction and direct synthesis of 1,3-DPG to 2,3-DPG. To add to the complexity of this process of regulation, it has now been demonstrated that 2,3-DPG itself may inhibit several important glycolytic steps.IO, :34 These include hexokinase, phosphofructokinase, phosphoglycerate kinase, and pyruvate kinase. When normal adult cells are incubated under nitrogen the level of 2,3-DPG rises rapidly.32 This response has been explained in two ways. The increase in deoxygenated hemoglobin results in increased binding of 2,3-DPG, thus relieving the product inhibition of the 2,3diphosphoglycerate mutase, and facilitates further 2,3-DPG synthesis. With deoxygenation, the intracellular pH also rises as a consequence of absorption of protons by the deoxygenated hemoglobin molecule. This increase in intracellular pH results in an augmentation of glycolysis with increased 2,3-DPG synthesis. 3 Regardless of the precise reason for this rise in 2,3-DPG, it does not occur in the cells of the neonate under similar experimental conditions. The red cells from the newborn infant also differ from those of the adult with respect to their 2,3-DPG stability. When these cells are incubated in room air in appropriate buffers, the decline in 2,3-DPG levels is six times as rapid as that observed in cells from adults. 35 , 39 The activity of 2,3-DPG mutase and of 2,3-DPG phosphatase, the enzymes responsible for 2,3-DPG synthesis and hydrolysis, are the same or greater in the erythrocytes of the newborn infant. 35 ,37 This fall in 2,3-DPG has been variously attributed to a block in glycolysis proximal to the point of 2,3DPG synthesis,39 an increased flow of metabolites through the phosphoglycerate kinase step with a by-pass of the 2,3-DPG cycle,35 and to an increase in the rate of 2,3-DPG breakdown in the fetal erythrocyte,37 in the presence of normal production. At present, the precise reason for this

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alteration in 2,3-DPG metabolism in the red cells of the neonate remains unknown.

CHANGES IN HEMOGLOBIN-OXYGEN AFFINITY IN THE FETAL AND NEONATAL PERIOD In the term infant the oxygen-hemoglobin equilibrium curve gradually shifts to the right and the P 50 value approximates that of the normal adult by approximately 4 to 6 months of age 18 (Table 2). A significant increase in P 50 can be observed during the first week of life. 18 This increase is a result of the rise in the red ce1l2,3-DPG level. It is tempting to speculate that it is caused by the transient rise in the serum inorganic phosphate level which so commonly occurs during this period. In the premature infant, born with lower 2,3-DPG levels and higher fetal hemoglobin values, the shift in the position of the curve is far more gradual. 18 In all infants the position of the curve appears to be directly correlated with the "effective"30 or "functioning" DPG fraction. 18 Red cell 2,3-DPG levels are profoundly decreased in infants with severe respiratory distress. 18.20 This decrease, with its associated lowering of the P oo value, is most marked in infants with profound acidosis. It would appear that the infant, both in utero and ex utero, is far less capable of responding to hypoxia by increasing his red cell level of 2,3-DPG. This lack of response to hypoxia is in contrast to the rapid response observed in adults in a variety of situations characterized by an increase in the level of deoxygenated hemoglobin. Such conditions include anemia, ascent to high altitude, cyanotic heart disease, and chronic lung disease. When adult erythrocytes are transfused into the fetus or newborn infant they retain their characteristic oxygen-hemoglobin equilibrium curve. Novy and associates 28 and Mathers et al.2 6 have shown that following intrauterine transfusions the adult cells retain their normal properties for periods as long as 8 weeks. In these instances where the fetal oxygenhemoglobin equilibrium approximates that of the maternal blood, no deleterious effects have been observed. 28 This observation has resulted in a reexamination of the belief that a difference in hemoglobin-oxygen affinity between mother and infant is required for adequate oxygen delivery to the fetus in utero. These findings, in association with observations. Table 2.

1

Oxygen Transport in Term Infants TOTAL

HEMOGLOBIN

INFANTS

AGE

(g./100 m!. blood)

19 18 14 10 14 8 8

1 day 5 days 3 weeks 6-9 weeks 3-4 months 6 months 8-11 months

17,8 ± 2,0 16,2 1.2 12,0 1.3 10,5 1.2 10,2 0,8 11.3 0,9 11.4 0,6

NO. OF

O 2 CAPACITY (m1.!100 mI. blood) 24,7 22,6 16,7 14,7 14,3 14,7 15,9

2,8 2,2 1.9 1.6 1.2 0,6 0,8

P,,(l

AT pH 7,40 (mm, Hg)

19.4 20,6 22,7 24.4 26,5 27,8 30,3

± ± ± ± ± ± ±

1.8 1.7 1.0 1.4 2,0 1.0 0.7

2.3·DPG

FETAL

m,umolesi

HEMOGLOBIN

m!.RBC

(Sf of total)

5433 6850 5378 5560 5819 5086 7381

± ± ± ± ± ± ±

1041 996 732 747 1240 1570 485

77,0 ± 7,3 76,8 ± 5,8 70,0 ± 7.33 52,1 11.0 23,2 16,0 4,7 2,2 1.6 1.0

j

THE NEONATAL RED CELL AND 2,3-DIPHOSPHOGLYCERATE

915

in situations where mothers with hemoglobin variants with very high affinities for oxygen have given birth to apparently normal infants,33 suggest that a higher fetal than maternal blood affinity for oxygen is not an absolute necessity for intrauterine existence. Before this concept is totally embraced it should be recognized that the intrauterine transfusion of lambs appears to produce subtle evidence of hypoxic stress. 4 Exchange transfusion following birth produces rapid alterations in the infant's oxygen-hemoglobin equilibrium curve. 17 The early changes produced are a function of the storage characteristics of the blood employed. The use of fresh heparinized blood or blood stored in the anticoagulant citrate-phosphate-dextrose, for periods up to 1 week,36 produce a prompt increase in the P 50 value of the infant to that of the normal adult. Blood stored in the conventional anticoagulant, acid-citratedextrose, promptly demonstrates a fall in 2,3-DPG and an associated increase in hemoglobin-oxygen affinityY Alterations in P so produced by exchange transfusions performed with ACD blood are related to its length of storage. Blood stored in ACD for periods of four to five days or longer may actually transiently decrease the P so value in the newborn recipient. 17

IMPLICATIONS OF CURVE SHIFTS FOR THE NEWBORN INFANT Although the oxygen-carrying capacity of the blood decreases during the first 3 to 4 months of life because of the progressive fall in total hemoglobin concentration, this fall is accompanied by an increase in the oxygen-unloading capacity owing to the gradual shift to the right in the position of the oxygen-hemoglobin equilibrium curve. As a result, at a mixed venous oxygen tension of 40 mm. Hg, arbitrarily selected as the normal central venous oxygen tension at rest, the 3 month old infant is delivering more oxygen to his tissues than the newborn infant despite the fact that his hemoglobin has fallen from 17.0 gm. per 100 ml. to approximately 11.0 gm. per 100 ml. remains to be demonstrated whether the newborn infant with red cells that are relatively unresponsive to hypoxia is handicapped in clinical situations accompanied by an increase in the concentration of deoxygenated (hemoglobin such as occurs with respiratory or cardiac disorders. The effect of exchange transfusion with 2,3-DPG rich blood in such situations is currently under investigation. The results of preliminary studies appear encouraging. 16 The fetal red cell appears to be uniquely designed for survival in a hypoxic environment. The decreased interaction between fetal deoxyhemoglobin and 2,3-DPG appears to prevent inordinate shifts to the right in the position of the curve in utero, thus ensuring that the fetal blood will not have a lower affinity for hemoglobin than that of the mother. The lack of binding of deoxygenated fetal hemoglobin to ATP prevents the depletion of red cell membrane ATP with its attendent increase in cellular rigidity. Adult cells at low partial pressures of oxygen become progres-

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sively less deformable. 24 If this were to occur in utero the fetal circulation would be severely compromised. The fetal cell, perhaps as a consequence of its exposure to a low oxygen tension environment, however, lacks the capacity to deal with oxidative stresses mediated via hydrogen peroxide generation. This is reflected in the characteristic neonatal red cell relative deficiencies of both catalase and glutathione peroxidase, the enzymes responsible for the detoxification of hydrogen peroxide. In addition, in the intrauterine milieu, the plasma also lacks the normal antioxidant, vitamin E or D-alpha tocopherol, which makes the cell even more vulnerable to the effects of increased oxygen tension. Thus, although the cell appears admirably suited for intrauterine existence, it may be metabolically handicapped under normal aerobic conditions. Teleologically speaking, the cell appears programmed for premature selfdestruction in order that it may be replaced by a new adult type erythrocyte that is capable of responding to the usual environmental stresses.

1

REFERENCES 1. Allen, D. W., Wyman, T., and Smith, C. A.: The oxygen equilibrium of fetal and adult human hemoglobin. J. BioI. Chern., 203:84,1953. 2. Anselmino, K. T., and Hoffman, F.: Die Ursachen des Icterus Neonatorum. Arch. Gynak., 143:477,1930. 3. Astrup, P.: Red cell pH and oxygen affinity of hemoglobin. New Eng. J. Med., 283:202, 1970. 4. Battaglia, F. C., Bowes, W., McGaughey, H. R., Makowski, E. L., and Meschia, G.: The effect of fetal exchange transfusion with adult blood upon fetal oxygenation. Pediat. Res., 3:60, 1969. 5. Bauer, C., Ludwig, I., and Ludwig, M.: Different effects of 2,3-diphosphoglycerate and adenosine triphosphate on the oxygen affinity of adult and foetal human hemoglobin. Life Sciences, 7:1339, 1968. 6. Benedixen, H. H., and Laver, M. B.: Hypoxia in anesthesia: A Review. Clin. Pharmacol. Ther., 6:510, 1965. 7. Benesch, R, and Benesch, R. E.: The effect of organic phosphates from the human erythrocyte on the allosteric properties of hemoglobin. Bichem. Biophys. Res. Commun., 26:162,1967. 8. Benesch, R, Benesch, R E., and Enoki, Y.: The interaction of hemoglobin and its subunits with 2,3-diphosphoglycerate. Proc. Nat. Acad. Sci. U.S.A., 61 :1102,1968. 9. Berne, R. M., Blackman, J. R, and Gardner, T. H.: Hypoxemia and coronary blood flow. J. Clin. Invest., 36:1101, 1957. 10. Beutler, E.: 2,3-Diphosphoglycerate affects enzymes of glucose metabolism in red blood cells. Nature New Biology, 232:20, 1971. 11. Brewer, G. J., and Eaton, J. W.: Erythrocyte metabolism: interaction with oxygen transport. Science, 171 :1205, 1971. 12. Bunn, H. F., and Briehl, R W.: The interaction of 2,3-diphosphoglycerate with various human hemoglobins. J. Clin. Invest., 49:1088,1970. 13. Bunn, H. F., May, M. H., Kocholaty, W. F., and Shields, C. E.: Hemoglobin function in stored blood. J. Clin. Invest., 48:311, 1969. 14. Chanutin, A.: Red Cell2,3-Diphosphoglycerate (DPG) Metabolism and Function in Health and Disease. Publication of the United States Army Medical Research and Development Command,1971. 15. Chanutin, A., and Curnish, R. R: Effect of organic and inorganic phosphates on the oxygen equilibrium of human erythrocytes. Arch. Biochem., 121 :96, 1967. 16. Delivoria-Papadopoulos, M., Miller, L. D., Tunnessen, W. W., Jr., and Oski, F. A.: The effect of exchange transfusion on altering mortality in infants weighing less than 1300 gm. at birth and its role in the management of severe respiratory distress syndrome. Prog. Soc. Pediat. Res., 6:82, 1972. 17. Delivoria-Papadopoulos, M., Morrow, G., III, and Oski, F. A.: Exchange transfusion in the newborn infant with fresh and "old" blood: The role of storage on 2,3diphosphoglycerate hemoglobin-oxygen affinity, and oxygen release. J. Pediat., 79:898, 1971.

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THE NEONATAL RED CELL AND 2,3-DIPHOSPHOGLYCERATE

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18. Delivoria-Papadopoulos, M., Roncevic, N. P., and Oski, F. A.: Postnatal changes in oxygen transport of term, premature and sick infants: The role of adult hemoglobin and red cell 2,3-diphosphoglycerate. Pediat. Res., 5:235, 1971. 19. DeVerdier, C.-H., and Garby, L.: Low binding of 2,3-diphosphoglycerate to hemoglobin F. Contribution to the knowledge of the binding site and an explanation for the high-oxygen affinity of fetal blood. Scand. J. Clin. Invest., 23:149,1969. 20. Duc, G., and Engel, K.: Hemoglobin-oxygen affinity and erythrocyte 2,3diphosphoglycerate content in hyaline membrane disease and cardiac malformations. Prog. Soc. Ped. Res., 79, 1970. 21. Greer, J., and Perutz, M.: Cited in Bunn, H. F., and Briehl, R. W.: The interaction of 2,3diphosphoglycerate with various human hemoglobins. J. Clin. Invest., 49:1088, 1970. 22. Hjelm, M.: The content of some organic phosphocompounds in erythrocytes from healthy neonates correlated with the haemoglobin concentration in blood and the age of the subjects. (In press.) 23. Kety, S. S.: Determinants of tissue oxygen tension. Fed. Proc., 16:666, 1957. 24. LaCelle, P. L., and Weed, R I.: Low oxygen pressure: A cause of erythrocyte membrane rigidity. J. Clin. Invest., 49:54a, 1970. 25. Landis, E. M., and Pappenheimer, J. R: Exchange of substances through the capillary walls. In Handbook of Physiology, Circulation II. Washington, D.C., The American Physiological Society, 1963, ch. 29. 26. Mathers, N. P., James, G. B., and Walker, J.: The oxygen affinity of the blood of infants treated by intrauterine transfusion. J. Obstet. Gynec. Brit. Comm., 77:648, 1970. 27. Maurer, H. S., Behrman, R. E., and Honig, G. R: Dependence of the oxygen affinity of blood on the presence of foetal or adult haemoglobin. Nature, 227:388, 1970. 28. Novy, M. J., Frigoletto, F. D., Easterday, C. L., Umansky, I., and Nelson, N. M.: Changes in cord blood oxygen affinity after intrauterine transfusions for erythroblastosis. New Eng. J. Med., 285:589, 1971. 29. Opitz, E., and Schneider, M.: Uber die Sauerstoffversorgung des Gehirns und den mechanismus von Mangelwirkunge. Ergebn. Physiol., 46:126,1950. 30. Orzalesi, M. M., and Hay, W. W.: The regulation of oxygen affinity of fetal blood. I. In vitro experiments and results in normal infants. Pediatrics, 48:857, 1971. 31. Oski, F. A., and Gottlieb, A. J.: The interrelationships between red blood cell metabolites, hemoglobin, and the oxygen equilibrium curve. Prog. Hemat., 7:33, 1971. 32. Oski, F. A., Gottlieb, A. j., Miller, W. W., and Delivoria-Papadopoulos, M.: The effects of deoxygenation of adult and fetal hemoglobin on the synthesis of red cell 2,3diphosphoglycerate and its in vivo consequences. J. Clin. Invest., 49:400, 1970. 33. Parer, J. T.: Reversed relationship of oxygen affinity in maternal and fetal blood. Amer. J. Obstet. Gynec., 108:323, 1970. 34. Ponce, J., Roth, S., and Harkness, D. R.: Kinetic studies on the inhibition of glycolytic kinases of human erythrocytes by 2,3-diphosphoglyceric acid. Biochem. Biophys. Acta, 250:63, 1971. 35. Schriiter, W., and Winter, P.: Der 2,3-diphosphoglyceratsloffwichsel in den erythrocyten neugeborener und erwaschsener. Klin. Wschr., 45:255, 1967. 36. Shafer, A. W., Tague, L. L., Welch, M. H., and Guenter, C. A.: 2,3-Diphosphoglycerate in red cells stored in acid-citrate-dextrose and citrate-phosphate-dextrose: Implications regarding delivery of oxygen. J. Lab. Clin. Med., 77:430, 1971. 37. Trueworthy, R C., and Lowman, J. T.: Intracellular control of the 2,3-diphosphoglycerate concentration in fetal red cells. Proceedings of the Society for Pediatric Research, 1971, p.86. 38. Tyuma, I., and Shimizu, K.: Different response to organic phosphates of human fetal and adult hemoglobins. Arch. Biochem. Biophys., 129:404, 1969. 39. Zipursky, A., LaRue, T., and Israels, L. G.: The in vitro metabolism of erythrocytes from newborn infants. Can. J. Biochem. Physiol., 38:727, 1960. Department of Pediatrics State University of New York Upstate Medical Center 750 East Adams Avenue Syrac\lse, New York 13210