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Chapter 33 Pathobiology of the Human Erythrocyte and Its Hemoglobins 449

humans appears to be remarkably uniform under normal circum- span. Increasing phosphatidyl serine exposure and reduced amino-
stances, spanning approximately 120 days from release of the reticu- phospholipid translocase activity during aging might induce oxidative
locyte into circulation to sequestration of the senescent RBC in the damage to the cell. It is probable that these mechanisms leading to
reticuloendothelial cells of the liver and spleen. The precise signal, or cell destruction are not mutually exclusive, that no single effect
signals, marking RBCs for destruction remain unknown, as does the predominates, and that these events occur at different times at dif-
underlying pathophysiology within the RBC or on its surface. ferent sites of RBC damage.
However, several interrelated theories have emerged; these are dis- Regardless of the mechanism(s) fostering eventual senescence and
cussed only briefly because they are mentioned in other chapters. destruction of RBCs, the process itself involves components clinically
RBCs accumulate surface blemishes during their lives in the cir- useful for assessment of anemias associated with accelerated destruc-
culation. These appear to result in part from the accumulation of tion. Chief among these is the generation of indirect or unconjugated
small amounts of oxygen damage to membrane structures. The bilirubin, the byproduct of heme catabolism occurring within the
altered regions are sensed by the reticuloendothelial cells during reticuloendothelial cells. In markedly accelerated states of RBC
passage of the erythrocytes through the liver and spleen. Removal or destruction, hypertrophy of the liver and spleen can also occur,
pitting of these damaged regions from RBC membranes can be docu- providing a useful physical indicator of hemolytic anemia. These
mented microscopically; small amounts of normal membrane are also indirect clinical features, coupled with the reticulocyte count, remain
lost during the process. more useful for detecting clinical hemolysis than complicated studies
The biconcave disk shape of the RBC, so important to its disten- of RBC kinetics.
sibility, depends on a high ratio of surface area to volume. This
requires redundant membrane surface area. The membrane surface
2
area of the normal biconcave disk is approximately 140 µm . To HEMOGLOBIN SYNTHESIS, STRUCTURE, AND FUNCTION
enclose a sphere containing a normal RBC volume (≈90 fL), only
2
approximately 95 µm would be needed. Progressive loss of mem- Basic Features
brane surface by means of the pitting phenomenon should ultimately
cause the aging erythrocyte to assume a more rigid spherical shape. Hbs are the major oxygen-carrying pigments of the body. They are
A sphere is inevitably far less distensible and far less capable of passing packaged into RBCs in quantities sufficient to carry enough oxygen
through small apertures than a disk, especially in the sluggish and from the lungs to the tissues to meet the needs of those cells for oxida-
tortuous circulation of the spleen. This geometric mechanism can tive metabolism. These quantities are enormous—almost 2 pounds
lead to the eventual destruction of the RBC. of Hb are present in the body of a reasonably sized human at any
RBCs progressively lose some of the critical enzymes needed for given time. Because free Hb in the bloodstream is catabolized and
intermediary metabolism and antioxidant capacity. G6PD levels, for excreted renally in a matter of minutes, packaging in erythrocytes is
example, progressively decline during the circulating life span, as do essential to preserve the newly synthesized molecules for the entire
levels of several other enzymes. The decline of certain enzymes can 4-month life span of the RBC. Otherwise, the caloric and biosyn-
be used as a crude means of estimating the relative age of different thetic resources needed to replace daily losses of Hb would be pro-
RBC populations. The biochemical or oxidative mechanism of hibitive. The RBC’s major function is to encase Hb and protect it so
destruction postulates that aged RBCs are eventually depleted of it can function as an oxygen transporter for a prolonged period. An
critical enzymes needed for maintenance of redox status. Oxidation additional function of Hb is to modulate vascular tone by its transport
of critical membrane proteins, lipids, and Hb would then ensue, of nitric oxide (NO) and possibly nitrous oxide.
causing distortion and rigidity of the RBC membrane, with acceler- The cellular content of blood influences its viscosity; in particular,
ated loss as previously described. The end product would be sphero- the hemodynamics are adversely compromised by the presence of too
cytes incapable of traversing the splenic vascular bed and escaping many circulating erythrocytes because blood viscosity correlates
engulfment by the reticuloendothelial cell. especially with hematocrit. To provide for adequate oxygen transport
It has been proposed that an immune-type mechanism can con- (i.e., enough Hb molecules) in a number of RBCs compatible with
tribute to normal and pathologic RBC senescence. This hypothesis tolerable viscosity, each cell must enclose a high concentration of Hb
is based on the observation that oxidative damage, regardless of cause, (32–35 g per 100 mL of cytoplasm). This concentration is close to
promotes a clustering, or capping, of oligomers of band 3 on the RBC the solubility limit of Hb in physiologic solutions. It follows that even
surface. Under normal circumstances, band 3 molecules form mono- minor perturbations within these molecules (e.g., oxidation) or in the
mers, dimers, or tetramers. Higher order aggregates appear to be milieu (e.g., changes in pH or ionic strength) can have potentially
recognized by an endogenous isoantibody possessed by all people. devastating effects on the solubility of Hb. Because polymerized or
Any RBC accumulating oxidative damage from wear and tear in the precipitated Hbs derange intracellular viscosity, trigger proteolytic
circulation, from depletion of enzymes, or from internal pathologic reactions that lead to oxidative damage of erythrocytes, and compro-
processes such as denaturation of Hb in certain hemoglobinopathies mise oxygen transport, it is not surprising that the fate of the RBC
can accumulate these aggregates. The aggregates would then be is inextricably interwoven with the state of its enormous complement
bound by antibody and be removed by the reticuloendothelial cells of Hb molecules.
as antigen–antibody complexes, using the same means used by reticu-
loendothelial cells to recognize any immune complex. This mecha-
nism could also provide for the pitting or polishing of damaged RBC Hemoglobin Structure
membranes. All three of the proposed mechanisms are interrelated
by their inception with oxidative damage. The Hb tetramer consists of two pairs of unlike globin polypeptide
Other membrane-related changes might influence RBC destruc- chains, each associated with a heme group. Normal Hb has two
tion. Bcl-X L , a suppressor of apoptosis, is present in erythrocyte α-globin and two non–α-globin chains; the interaction of these
membranes, and its antagonization may promote cell death. This may chains is responsible for the quaternary structure of the Hb molecule
be mediated by calcium accumulation and phosphatidylserine expo- and normal oxygen transport. Functionally, the second exon of each
sure. Cholesterol and fatty acids accumulate on the aging RBC globin gene encodes the major component of the heme-binding
membrane and might be targets for oxidation induced by reactive pocket, and the α and non-α contacts are regulated by the third exon.
oxygen species. RBCs are removed from the circulation by splenic The behavior of Hb is determined by its primary structure, the
macrophages, probably by several mechanisms. SHPS-1, a surface covalent linking of amino acids to form the polypeptide globin. The
glycoprotein and a member of the immunoglobulin superfamily that higher order structures of Hb depend on the sequence of amino acid
interacts with RBC membrane CD47, is abundant in macrophages. residues that make up the globin chain. The α-globin chains contain
Studies using mice expressing a mutant SHPS-1 suggested that this 141 residues, and the β-globin–like chains are 146 amino acids long
molecule might negatively regulate phagocytosis, influencing cell life (Fig. 33.1). There is considerable homology among these globins,