Non-Immune Hemolytic Disorders (NIHDs)

Non-Immune Hemolytic Disorders (NIHDs)

• NIHDs are characterized by positive routine hemolytic tests but a negative anti-human immunoglobulin (Coombs) test.
• Direct and Indirect Antiglobulin Tests are negative for NIHDs.
• NIHDs are classified as hereditary or acquired.

Hereditary Non-Immune Hemolytic Disorders

• Hereditary NIHDs are mainly intra-corpuscular.
• They include membranopathies, haemoglobinopathies, and enzymopathies.
• A rare hereditary form of Thrombotic Thrombocytopenic Purpura (TTP) is also included.
• These disorders present with intravascular or extravascular haemolysis.

Acquired Non-Immune Hemolytic Disorders

• Acquired NIHDs are mainly extra-corpuscular.
• They include red cell fragmentation, march haemoglobinuria, infections, chemical and physical agents, paroxysmal nocturnal harmoglobinuria (PNH), and acquired disorders of the red cell membrane.
• Acquired NIHDs present with intravascular haemolysis.

Red Cell Fragmentation Syndrome/Mechanical Hemolytic Anemias

• This syndrome results from physical damage to red cells in circulation.
• Presence of Schistocytes, which are diagnostic red cell fragments, characterize the syndrome
• Schistocytes, or Schizocytes, are fragmented red cells that can take on different shapes, such as triangular, helmet, or comma-shaped with pointed edges
• Schistocytes are usually microcytic without a central pallor area, deformability is the same as normal red cells, but lifespan is reduced.
• Normally, everyone has Schistocytes, ranging from 0.2-0.5%
• Higher counts are found in Red Cell Fragmentation Syndromes and are counted from a peripheral blood film.

Red Cell Fragmentation Syndrome

• The cellular components of blood, coagulation system, fibrinolytic system and the endothelial lining of blood vessels have an intimate and complex relationship
• Red cell integrity is easily destroyed by contact with abnormal surfaces.
• The most common mechanism appears to be contact between red cells and abnormal surfaces with some degree of adherence.
• Red Cell Fragmentation syndrome is divided into Macroangiopathic and Microangiopathic Hemolytic Anemias.

Macroangiopathic Hemolytic Anemias/Cardiac Hemolytic Anemias

• Cardiac Haemolysis occurs after cardiac surgery involving prosthetic valves, patches, or vessels.
• Cardiac Haemolysis is rare, occurring in 3% of patients with prosthetic heart valves, and has become less common with newer materials like homografts and xenografts.
• Haemolysis is associated with artificial materials that are present together with turbulent blood flow.
• Damage to red cells may come from abnormal surfaces, calcified or stenotic heart valves, artificial heart valves, and arterial grafts.
• Damage may come from arteriovenous malformations such as Cavernous haemangioma (Kasabach-Merit Syndrome).
• Arteriovenous malformations (AVMs) happen when a group of blood vessels forms incorrectly
• Arteries and veins are unusually tangled and form direct connections, bypassing normal tissues.
• AVMs usually happen during development before birth or shortly after.
• Blood flows directly from arteries to veins usually at high velocity, there are no capillaries.
• AVMs can occur anywhere in the body.

Microangiopathic Hemolytic Anemias (MAHA)

• MAHA is intravascular haemolysis with red cell fragmentation caused by destruction of red cells in an abnormal micro-circulation.
• Three major pathological lesions can give rise to MAHA: presence of fibrin strands in circulationplatelet aggregation, and vasculitis.
• Fibrin strands in circulation cause red cells to pass through the strands and become fragmented and is common in Disseminated Intravascular Coagulopathy (DIC), Meningococcal sepsis and Carcinomatosis
• Platelet aggregation results when platelets adhere to small vessels forming aggregates seen in Thrombotic Thrombocytopenic Purpura (TTP) and Haemolytic Uraemic Syndrome and Homograft rejection (micothrombi in transplanted organ)
• Vasculitis results in roughened endothelial surfaces as a result of inflammatory reactions, and cells become fragment when they collide with the roughened endothelial cells, this is seen in Polyarteritis nodosa, Systemic Lupus, Wegener’s granulomatosis Renal cortical necrosis, and Acute glomerulonephritis

Haemolytic Uraemic Syndrome (HUS)

• HUS is an acquired disorder mainly affecting infants and children under 4 years old, and is rare after puberty.
• HUS cases can be sporadic or epidemic (occurring in clusters).
• HUS presents with a triad of intravascular haemolysis, renal failure, and thrombocytopenia.
• Gasser (Europe) and Brain et al. emphasized the presence of red cell fragments, diffuse intravascular coagulation, and microangiopathic haemolytic anaemia.
• The cause of HUS is unknown but host and environmental factors are implicated.
• A host factor includes an abnormal reduced production of prostacyclin, which limits adherence of platelets to endothelium, is a powerful vasodilator and anti-platelet aggregation.
• Another Host factor is Thromboxane A2, a prostaglandin, is produced by platelets only after platelets adhere to the endothelium and has physiological properties opposite to prostacyclin.
• Occurrence in different members of a family suggests constitutional and genetic factors with HUS development.
• Environmental factors associates with various infective agents, such as viruses, rickettsia, and bacteria.
• Epidemic forms commonly involve infection with Eschericia coli 0157 (verotoxin producing).
• Diarrhoea is the commonest presenting symptom, and is usually bloody, and associated with vomiting, and usually last for a day or weeks
• Clinical features include heamoglobinuria, proteinuria from acute renal failure, uraemiaanaemia, thrombocytopenia, jaundice, and CNS signs, such as convulsion or coma.
• Laboratory features include:
  • Lots of red cell fragments
  • Anisocytosis, Poikilocytosis
  • Polychromatic cells, Nucleated red cells
  • Severe thrombocytopenia
  • PT, PTTK, and TT are usually normal
• HUS management is mainly supportive before renal failure ensues.
• Other forms of management:
  • red cell transfusionplatelet transfusion rarely needed except there is bleeding.
  • coagulation factors replacement with Fresh Frozen Plasma (FFP) if there is evidence of DIC.
  • Corticosteroids and use agents that inhibits platelet aggregation.

Thrombotic Thrombocytopenic Purpura (TTP)

• TTP is a clinical syndrome with fever, anaemia, thrombocytopenia, neurological abnormalities, and renal Failure.
• Platelet aggregation is seen in small blood vessels.
• TTP's aetiology is unknown or not fully understood.
• Possible etiology
  • The disorder could be congenital or acquired.
  • In both types is persistent presence of unusually large Von Willebrand Factor (vWF) and due to abnormal platelet aggregation
  • The metaloprotease called ADAMTS13 normally degrades WVF or VWF cleaving protein
  • The ADAMTS13 gene is located on chromosome 9q34.
  • ADAMTS13 is predominantly produced by the hepatocytes by vascular endothelial cells, but in limited quantity.
  • ADAMTS 13 level is significantly reduced, often to levels lower than 10% or is completely absent in patients with TTP.
  • In the congenital form, there are somatic mutations at the ADAMTS13 gene, and in the acquired form, autoantibodies develop against ADAMTS13.
• Another possible aetiology is a deficiency of serum factors that promote prostacyclin production by endothelial cells, because Prostacyclin inhibits platelets adhesion and aggregation to endothelial cells.
• TTP is more common amongst teenagers and young adults, and has a 50% mortality rate with conservative management
• Both sexes affected equally.
• Clinical features inlcude sudden onset, fever, neurological abnormalities predominate, purpura may or may not be early
• Clinical and laboratory feature include:
  • anaemia
  • marked haemoglobinuria
  • jaundice
  • renal failure is not common.
  • Laboratory features:
  • Fragmented and contracted red cells
  • polychromasia
  • thrombocytopenia
  • proteinuria is universal though it may be intermitent.
• Management of TTP includes
  • Plasmapharesis (plasma exchange by plasmapharesis with replacement using Fresh Frozen Plasma + Plasma Proteins + Platelets)
  • Prostacyclin analogues (infusion of prostacyclin analogues may be of benefit especially if there responds to FFP)
  • Splenectomy
  • Removal of stimuli (e.g. using broadspectrum antibiotics for infective agents and/or termination of pregnancy)

March Haemoglobinuria

• Haemoglobinuria following running is known for 100+ years.
• Caused by damage to red cells between small bones of feet during prolonged marching or running.
• Seen in joggers and is a benign condition.
• No red cell fragments on blood film.

Infections

• Infections can cause haemolytic anaemia in many ways.
• This includes acute hemolytic anaemia of G6PD deficiency and microangiopathic haemolytic anaemia (e.g., pneumococcal and meningococcal septicaemia).
• Malaria may cause haemolysis by extra vascular destruction of parasitised red cells or direct intravascular destruction of red cells when the sporozoites break out of red cells.
• Black water fever is an acute intravascular haemolysis associated with acute renal failure in Falciparum malariae infestation, and is a rare, devastating complication.
• The onset is acute, with marked haemoglobinaemia, haemoglobinuria, and rapid progression to oliguric renal failure.
• Black Water fever was first described among whites treated with quinine, but now reported among blacks in areas that were previously free from malaria, hence it is associated with high degree of parasitaemia and lack of immunity.
• Babesiosis is a rare disorder caused by Babesia, an intraerythrocytic, thick borne protozoan.
  • Ixodes vicinus associated with cattle in Europe
  • Ixodes dammini associated with rodents and deer in America.
• Commonly seen in splenectomized patients where it is fatal with acute onset.
• Other infective agents include Toxoplasmosis (caused by Toxoplasma gondii) in adults with severe immunosuppression, Oroya fever (caused by Bartonella bacilliformis) Clostridium perfringens Psittacosis and Yersinia enterocolitica, and Viruses like Dengue, Yellow fever and West African haemorrhagic fever.

Chemical and Physical Agents

• Dapsone and Salazopyrin cause oxidative damage to red cell membrane, resulting in intravascualar haemolysis where Heinz bodies prominent in red cells.
• High levels of cupper in the blood can cause intravascular haemolysis, as in Wilson's disease.
• Chemical poisoning from lead, chlorate, and arsine can cause severe haemolysis.
• Nitrites present in well water and some vegetable juices can cause methaemoglobinaemia in infants.
• Nitrates cause methaemoglobinaemia.

Paroxysmal Nocturnal Haemoglobinuria (PNH)

• PNH is also called Machiafava-Michaelis syndrome.
• It is a rare, acquired, somatic mutation and clonal intracorpuscular disorder of bone marrow stem cells.
• There is deficient synthesis of glycosylphosphatidylinositol (GPI) anchor protein on the membrane of red cells
• GPI attaches several surface proteins to red cell membrane.
• PNH results from mutation in the X chromosome gene that code for phosphatidylinositol glycan protein A (PIG-A).
• There are no GPI linked proteins because GPI (anchor protein) is absent.
• Because of it's absence CD55 and CD59 are not made, which are Decay Accelerating Factors (DAF) and Membrane inhibitor of reactive lysis (MIRL).
• DAF recognizes C3b and C4b fragment created when C3 and C4 are activated and accelerates the decay of C3 and C5.
• MIRL is a potent inhibitor of MAC (Membrane Attack Complex).
• Red cells become sensitive to lysis by complement resulting into intravascular haemolysis because of absence of DAF and MIRL
• Males and females are equally affected.
• Clinical features includes 2 types
  • classical haemolytic PNH where haemolysis is its main feature
  • Laboratory PNH, detected only by laboratory tests.
• Haemolytic PNH is very rare in infants and young children, but is common in adolescent and young adults up to 40 years.
• Haemoglobinuria is noticed in the first urine after sleep and clears during the day.
• Abdominal pain may be present, often severe, intermittent, and unrelated to meals.
• Thrombotic complications, especially of large veins, are common.
• Haemosiderinuria are constant features, and iron deficiency may develop.
• Splenomely is seen in some patients.
• Laboratory features includes
  • Anaemia
  • Polychromasia
  • Neutropenia
  • Marked thrombocytopenia
  • Tear drop poikilocytes
  • Low Neutrophil Alkaline Phosphatase (NAP) score.
• NAP score measures the proportion of WBCs that contain alkaline phosphatase, with normal levels being between 20 and 150.
• WBCs from the clone are also deficient in GPI.
• Alkaline phosphatase is also an anchor protein like DAF and MIRL, resulting in a low NAP score in PNH.
• PNH diagnosis can be made from the following tests:

1. Ham-Dacie Test or Acidified Serum Lysis Test:

• This test activates the alternate complement pathway and PNH cells will easily be lysed because of sensitivity of complement.

2. Sucrose Lysis Test
3. Thrombin Lysis Test

• Tests 2 and 3 activate the classical complement pathway.
• Diagnosis confirmation is by Flow Cytometry.
• PNH Treatment include:
  • Iron therapy
  • Anti-coagulation with warfarin.
  • Immunosuppression
  • Bone marrow transplantation.
• PNH usually remits with median survival of 10 years.

Acquired Disorders of Red Cell Membrane

• Lipids on the red cell membrane are in equilibrium with lipids of the plasma.
• Changes in the ratio of free cholesterol to phospholipids in plasma may affect red cell and result in haemolysis.
• Mostly pathologies in the liver, such as acute or chronic hepatitis and in Liver cirrhosis.
• Further pathologies include:
  • Gilbert's disease (inherited nonhaemolytic unconjugated hyperbilirubinaemia)
  • Ziever's syndrome: an uncommon disorder seen mainly in alcoholics. Presents with intravascular haemolysis and acute abdominal pain.
  • Wilson's disease: due to high level of copper ion in the blood. Presents with acute intravascular haemolysis, hepatic and neurological features.
  • Vit E deficiency: seen in infants fed with diets rich in polyunsaturated fatty acids. Presents with haemolysis, contracted red cells, and thrombocytosis.