Non-Immune Haemolytic

Anaemias II

Red cell Fragmentation Syndromes

KMensah
MLSD 500
Causes of Acquired Non-Immune Haemolytic Anaemias

• Mechanical trauma to red cells

 Abnormalities in the heart and large blood vessels
 Aortic valve prostheses , severe aortic valve disease
 Microangiopathic haemolytic anaemia
 Haemolytic uraemic syndrome, thrombotic thrombocytopenic purpura, metastatic malignancy,
 ***Malignant hypertension, disseminated intravascular coagulation
 March haemoglobinuria
• Burns
• Infections
 Clostridium perfringens (welchii) , malaria , bartonellosis
• Drugs,* chemicals and venoms
 Oxidant drugs and chemicals, arsine, acute lead
 Poisoning, copper toxicity, venoms of certain spiders and snakes
• Hypersplenism

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ERYTHROCYTE DISORDERS
AS A RESULT OF CHEMICAL
AND PHYSICAL AGENTS
 MECHANISMS OF RED CELL DESTRUCTION
1) ERYTHROCYTE VOLUME EXPANSION AND HYPOTONIC LYSIS
• When large amounts of distilled water gain access to the systemic circulation,
either by intravenous injection or when used as an irrigating solution during
surgery, haemolysis will occur.

• Severe haemolysis may also result from water inhalation in near-drowning.

• Occasionally self-induced hypotonic lysis secondary to water intoxication from

polydipsia in the setting of psychiatric illness or hazing rituals occurs.

• In all cases, haemolysis follows expansion of the erythrocyte volume, transition

to a spherical shape and ultimately cell rupture.
2) DAMAGE TO THE RED BLOOD CELL MEMBRANE
• Bee and wasp stings, as well as contact with caterpillar bristle from
Lonomia obliqua are associated with severe haemolysis.
• In addition, spider and scorpion bites occasionally are followed by
haemolytic anaemia and haemoglobinuria.
• The spiders usually responsible are Loxosceles laeta and Loxosceles
recluse.
• In such cases, sphingomyelinase D is one of the causative toxins. The venom
preferentially hydrolyzes band 3 of the red cell membrane protein.
• Band 3 has dual functions of ion exchange and anchoring of the cell
membrane to the underlying cellular skeleton.
• It appears disruption of the structural role is responsible for cell lysis.
2) DAMAGE TO THE RED BLOOD CELL MEMBRANE
• One of the most intriguing mechanisms of membrane damage is that
induced by a class of pore-forming cytotoxins, usually from Bacillus cereus.

• Toxins using similar mechanisms of haemolysis are found in marine

organisms including jelly fish (Chironex fleckeri), sea cucumbers
(Cucumaria echinata), and sea anemones (Stichodactyla helianthus).

• X-ray crystallography reveals these toxins to be composed of proteins

that associate to span the erythrocyte membrane forming an ion
permeable pore.

• Aged cells appear preferentially damaged.

3) DAMAGE TO SKELETAL OR STRUCTURAL PROTEINS
• Gross haemoglobinaemia was observed in 11 of 40 patients with second and third-
degree burns involving 15 to 65 percent of body surface area.

• Within the first 24 hours following a burn, haemolytic anaemia results from the direct
effect of heat on circulating erythrocytes.

• Blood heated to temperatures above 49°C demonstrates morphologically similar damage

consistent with increased osmotic and mechanical fragility.

• In addition to acute damage, heat decreases erythrocyte resilience.

• A normal erythrocyte in liquid behaves physically as a drop of fluid because the flexible
membrane allows the surface of the cell to rotate around the intracellular contents.
3) DAMAGE TO SKELETAL OR STRUCTURAL
PROTEINS
• These fluid-like properties couple collisional energy between the erythrocyte
membrane and the viscous haemoglobin solution within the cell, allowing
dissipation of collisional energy through the entire cell and ultimately
protecting the cell membrane.

• When heated, the spectrin comprising the erythrocyte skeleton melts, and
upon cooling becomes rigid.

• This rigidity prevents collisional energy dissipation, making such cells

particularly susceptible to membrane damage.

• The ensuing damage to the erythrocyte membrane structure results in splenic

sequestration and cell removal.
 4) OXIDANT DAMAGE
• Although oxygen is a powerful oxidizing agent, quantum mechanical properties of
the oxygen molecule prevent spontaneous oxidation of biologic membranes.

• When bound to haemoglobin, oxygen has significantly different quantum mechanical

properties and occasionally, an exceptionally reactive superoxide molecule escapes.

• It is estimated that 2 to 3 percent of total haemoglobin would be oxidized daily in

the absence of enzyme systems to protect against escaped superoxide.

• Haemolysis following oxidation is thought to occur via eryptosis.

• In addition to oxidative stress, osmotic shock and certain toxic ions, including gold
and aluminum, may act through eryptosis.
OXIDANT DAMAGE
a) Oxygen Gas and Ozone
• Haemolytic anaemia can occur when ambient oxygen (O2) concentration is
increased markedly.

• Hyperbaric oxygenation has been associated with acute haemolysis.

• Ozone (O3), which has been widely used in some countries for a variety of
therapeutic purposes.

• Had no apparent effect in vivo on red cell enzymes and intermediates at

the 30 mcg/mL concentration commonly infused, but did produce some in
vitro haemolysis at that concentration.
 OXIDANT DAMAGE
b) Arsenic Hydride
• Arsenic exposure is a major cause of anaemia in regions with high environmental
concentration. Egs include Bangladesh’s tainted water supply, and areas of China
where arsenic laden coal is used.
• Arsine gas (arsenic hydride, AsH3) is the most erythrotoxic form of arsenic
and inhalation of arsine gas is a well-recognized cause of haemolytic anaemia.
• Arsine is formed during many industrial processes, including the reaction of
hydrogen with available arsenic compounds in metallurgic processes.
• The arsenic is usually a contaminant, so contact with arsenic compounds may not
be apparent from the patient’s history.
• The mechanism of erythrocyte damage occurs via oxidation of sulfhydryl
groups in the erythrocyte membrane and associated cytoskeleton and
decreased levels of reduced glutathione in erythrocytes exposed to AsH3 are
observed.
OXIDANT DAMAGE
c) Chlorates and Chloramines
• Sodium and potassium chlorate are oxidative drugs that produce methaemoglobinaemia,
Heinz bodies, and haemolytic anaemia.

• Although it has been presumed that the mechanism of haemolysis is similar to that
resulting from other oxidative drugs, enigmatically, no cases have been observed in
patients deficient in glucose-6-phosphate dehydrogenase (G6PD).

• Haemolytic anaemia with Heinz body formation has also occurred in patients undergoing
dialysis when the water contained a substantial amount of chloramines.

• Oxidative damage to the red cells of these patients was demonstrated by the presence
of Heinz bodies, a positive ascorbate-cyanide test, and methaemoglobinaemia
 OXIDANT DAMAGE
d) Formaldehyde
• Leaching of formaldehyde from plastic used in water filters employed
for haemodialysis is also a cause of haemolytic anaemia.

• The low level of formaldehyde in the contaminated water does not result
in a fixative effect, but instead induces metabolic changes within the
red cells.
OTHER IDIOPATHIC CAUSES OF HAEMOLYSIS
THROUGH PHYSICAL & CHEMICAL AGENTS
a) NEOCYTOLYSIS
• Astronauts experience significant anaemia after space flight even in the
presence of normal or elevated ambient oxygen concentration.

• The mechanism of this anaemia remains elusive with intriguing observations

of altered erythropoietin levels and radiolabeling studies of astronaut
erythrocytes suggesting selective haemolysis of young erythrocytes less
than 12 days old.

• In addition to space flight, neocytolysis has been invoked to explain the

anaemia resulting when high-altitude dwellers relocate to sea level.
 LEAD
• Lead intoxication in children generally results from ingestion of flaking lead paint
or chewing lead-painted articles.

• Lead poisoning tends to be more severe in iron-deficient children, as a relatively

close relationship exists between blood lead levels and haematocrit.

• In adults, lead poisoning primarily occurs as the result of inhalation of lead

compounds from industrial processes such as battery manufacture, or ingestion
of food having leached lead from pottery or dishes.

• Lead poisoning from restoring tapestries and producing ceramics also has been
noted.
 LEAD
• Generally, the erythrocyte disorder associated with lead intoxication in
vivo is thought to be a result of interference with normal production of
erythrocytes.

• There is direct evidence that lead inhibits red blood cell (RBC) 5
nucleotidase and results in basophilic stippling and haemolysis.

• The other lead-poisoning–associated morphologic changes are observed

when chronic lead exposure is associated with sideroblastic anaemia, but
are not observed in acute lead poisoning.

• In addition, modest shortening of red cell life span is a relatively constant

feature of lead intoxication.
 LEAD
• Microscopic examination of the blood provides the key diagnostic clue to lead
poisoning.

• Complete observations of the acute haematologic changes, including

erythrocyte distortion, occurring after the intravenous injection of lead in an
attempt to treat malignant disease.

• Lead induces normocytic and slightly hypochromic erythrocytes, with the

hypochromia possibly resulting from coexisting iron deficiency.

• Basophilic stippling of the erythrocytes may be fine or coarse, and the

number of granules seen in each cell may be quite variable.
 COPPER

• Erythrocyte damage has resulted from ingestion of copper sulfate in suicide

attempts and from copper accumulation when hemodialysis fluid is
contaminated by copper pipes.

• Haemolysis in Wilson disease has been attributed to the elevated plasma

copper levels characteristic of that disorder.

• Spherocytic anaemia with a hematocrit below 25 percent may be the

presenting symptom.
 COPPER
• The pathogenesis may be related to oxidation of intracellular
glutathione, haemoglobin, and the reduced nicotinamide adenine
dinucleotide phosphate (NADPH), along with inhibition of G6PD by
copper.

• However, the amount of copper required to inhibit G6PD is large.

• Copper in much lower concentrations inhibits pyruvate kinase, hexokinase,

phosphogluconate dehydrogenase, phosphofructokinase, and
phosphoglycerate kinase, suggesting a global metabolic insult.

• Plasma exchange successfully prevents haemolytic anemia in Wilson

disease.
 RADIATION
• Although decreased red cell survival is part of the complex series of events
occurring after administration of large doses of total body radiation,
erythrocytes appear to be very resistant to the direct effects of radiation.

• Shortened red cell survival after radiation is likely related to red cell loss
through internal bleeding and various secondary events such as infection.
 HAEMOLYTIC ANAEMIA
RESULTING FROM INFECTIONS
WITH MICROORGANISMS
 Mechanism of Red Cell Destruction
• Several distinct mechanisms may lead to haemolysis during infections.

• These include :
 Direct invasion of or injury to the erythrocytes by the infecting
organism, as in malaria, babesiosis, and bartonellosis.

 Elaboration of haemolytic toxins, as by Clostridium perfringens.

 Development of antibodies, either autoantibodies against red cell
antigens or deposition of microbial antigens or immune complexes on
erythrocytes.
 MALARIA
PATHOGENESIS OF THE ANEMIA
• Haemolytic Mechanisms
• Destruction of parasitized red cells appears to occur largely in the spleen,
and splenomegaly typically is present in chronic malarial infection.
• The “pitting” of parasites from infected erythrocytes may also occur in the
spleen.
• The degree of parasitaemia, in part, determines the destruction of infected
erythrocytes.
• Low rates of red cell parasitaemia may have little effect on the development
of anaemia, whereas high rates, for eg,10 percent, may have very significant
effects.
 MALARIA
PATHOGENESIS OF THE ANAEMIA
Haemolytic Mechanisms
• The degree to which anemia develops often seems to be disproportionate to the number
of cells infected with the parasite.

• It is estimated that 10 times the number of uninfected red cells are removed for each
infected red cell, dramatically magnifying the haemolytic rate.

• Osmotic fragility is increased in non-parasitized cells, as well as in cells containing

plasmodia.

• The erythrocyte cation permeability is altered in monkeys with malaria.

• Hemin accumulation facilitates haemolytic cell loss via a process of programmed cell
death, referred to as eryptosis.
 MALARIA
Haemolytic Mechanisms:
• This suicidal death pathway is mediated by increased cell calcium, increased annexin-V
binding, and ceramide formation.

• Oxidative damage to red cell lipids occurs and there is an abnormality in the
phosphorylation of membranes of parasitized red cells.

• P. falciparum-infected red cells have a highly irregular surface produced by the

intracellular growth of the plasmodium, but non-parasitized cells often have similar
surface defects.

• Activation of hepatosplenic macrophages enhance red cell clearance supported by red cell
surface changes in both parasitized and unparasitized cells that foster recognition and
erythrophagocytosis by macrophages.
 MALARIA
PATHOGENESIS OF THE ANAEMIA
Haemolytic Mechanisms:
• Both marked loss of red cell deformability and deposition of immunoglobulin G and
complement (C3d), sometimes resulting in a positive direct antiglobulin reaction, may
enhance red cell removal by macrophages.

• Parasite products are part of the immune complexes on the red cell surface.

• The P. falciparum ring surface protein 2 (RSP-2) mediates adhesion of infected red
cells to endothelial cells and is deposited on uninfected cells, undoubtedly providing a
mechanism for removal of these cells by mediating complement-dependent phagocytosis.

• Splenomegaly further enhances red cell removal from the circulation.

 BARTONELLOSIS (OROYA FEVER)

PATHOGENESIS:
• After a sandfly bite, the red blood cells become infected with
Bartonella bacilliformis.
• It is believed that the organism does not grow within the red cell, but
rather adheres to its exterior surface:
• When infected red cells are washed with citrated plasma, free
organisms are found but the red cells are not haemolyzed.
• In hanging-drop cultures, masses of organisms are clearly seen outside
the erythrocytes, while the cells themselves are intact.
• The osmotic fragility of the red cells is normal.
 BARTONELLOSIS (OROYA FEVER)
PATHOGENESIS:
• They are rapidly removed from the circulation, apparently both by liver and
spleen.
• Normal red cells transfused into patients with bartonellosis meet a similar
fate.
• A 130-kDa Bartonella protein that causes erythrocytes to acquire trenches,
indentations, and invaginations has been purified from culture broths and
has been called deformin.
• In addition, two B. bacilliformis genes, designated ialA and ialB, predicted
to encode polypeptides of 170 amino acids (20.1 kDa) and of 186 amino acids
(19.9 kDa), respectively, greatly enhance the ability of Escherichia coli to
invade erythrocytes.
 BABESIOSIS
• Babesiae are intraerythrocytic protozoas known as piroplasms.
• They are transmitted by ticks that may infect many species of wild and
domestic animals.
• Humans occasionally become infected with Babesia microti (North America)
or Babesia divergens (Europe), species that normally parasitize rodents,
and, deer, elk, and cattle, respectively.
• Other Babesia-like piroplasms, such as Babesia WA1, first isolated from a
patient in the state of Washington, and Babesia MO1, isolated in Missouri,
may also produce human disease.
• Once thought to be rare, babesiosis is being recognized with increasing
frequency.
• The disease is usually tickborne in humans, but has also been transmitted
by transfusion.
 CLOSTRIDIUM PERFRINGENS SEPTICEMIA
PATHOGENESIS
• C. perfringens are Gram-positive, encapsulated, spore-forming, anaerobic
bacilli.
• The organism causes gas gangrene in soft tissues.
• The α toxin of C. perfringens is a lecithinase C that reacts with lipoprotein
complexes at cell surfaces, liberating potent haemolytic substances,
lysolecithins.
• This toxin is the agent that causes intravascular haemolysis and its
subsequent effects.
• It has also been suggested that erythrocyte membrane proteolysis plays an
important role in haemolysis
 OTHER INFECTIONS
• Some organisms, among them such common pathogens as Haemophilus influenzae, E. coli,
and Salmonella species, can produce red cell agglutination in vitro, but it is not known
whether this phenomenon is important in initiating in vivo hemolysis.

• Bacteria may also produce destruction of red cells indirectly when bacterial
polysaccharides are adsorbed onto erythrocytes.

• Action of an antibody directed against the antigen-coated cells results in their

agglutination or in complement-mediated lysis.

• The unmasking of T-type antigens by bacteria renders the cell poly agglutinable.

• This may be a rare cause of haemolysis occurring in the course of bacterial infections.
 HAVE A NICE DAY!!!

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