Autoimmune haemolytic anaemia

Introduction

Immune haemolytic anaemia (IHA) is a clinical condition in which igG and/or igM antibodies bind to red blood cells surface antigen and initiate red blood cell destruction via the complement system and reaction of the endothelia system. Immune haemolytic anaemia is classified as either auto immune, allo immune, or drug induce  base on the antigenic stimulus responsible for the immune response.

Auto immune haemolytic anaemia is characterized by the production of antibodies directed against self red blood cells. Since  the antibodies are usually directed against high-incidence antigens, they often exhibit reactivity against allogenic red blood cell as well .Auto immune haemolytic is fairly uncommon    disorder, with estimation of the incidence at 1-3 cases per 100,000 per year (Pirofsky and Williams, 2004). In contrast allo immune haemolytic anaemia requires exposure to allogenic red blood cells and the resulting allo antibody shows no reactivity towards autologous red blood cells. Sources of allogenic red blood cell include pregnancy, blood product transfusion and transplantation.

Drug induce immune haemolytic anaemia is the final classification of immune haemolytic anaemia. Drug induced antibodies can recognize either intrinsic red blood cell antigen or red blood cell-bound drug. Antibodies that react with intrinsic red blood cell are serologically indistinguishable from auto antibodies. In contrast, antibodies that react against red blood cell- bound drug require the drug for haemolysis.

Pathogeneses of auto immune haemolytic anaemia

The degree of haemolysis depends on the characteristics of the bound antibody (e.g. quantity, specificity, thermal amplitude, ability to fix complement, ability to bind macrophages) as well as the target antigen (density, expression, patient age etc.). IgG antibodies are relatively poor activator of classical complement pathway, but they (in particular igG1G3 antibodies) are recognized by the fc receptor on various phagocytic cells (Abraham, Gelfand,and Jandi, 2008).

Therefore igG red blood cell generally are eliminated  by phagocyte of the reticuloendothelia system, since the reticuloendothelia cell also have receptor for complement factors C3b and I C3b, this complement component, if present can potentiate the extravascular  haemolysis  (Ehlenberger and Nussenzweig, 2008). On the other hand, igM sensitized red blood cell are generally associated with a combination of intravascular and extra vascularhaemolysis. Intra vascular haemolysis occur because igM antibodies unlike igG antibodies readily activate the classical complement path way and produce cytolysis.

However, due to  the presence of regulatory red blood cell protein such as decay accelerating factor (DAF, CD55) and membrane inhibitor of reactive lyses (MIRLCD59), overwhelming complement activation usually is required to  produce clinically evident  intravascular haemolysis, e.g. as seen  ABO- incompatible blood transfusion. More commonly, igM sensitized red cell undergo extra vascularhaemolysis.  While reticuloendothelia cell do not have receptor for the FC fragment of igM antibodies with comparable activity to the receptors directed against the FC fragment igG, they do not have receptor for abundant red blood  cell-bound C3b and IC3b  resulting from the complement activation where as  the spleen is the principal site of igG-associated extra vascular, while the kupffer cells in the liver are the principal effectors of igM –associated extra vascularhaemolysis

Classification of autoimmune haemolytic anaemia

Cases of the autoimmune haemolyticanaemia are generally classified according to the characteristics temperature reactivity of the red blood cell autoantibody. Warm autoantibody reacts most strongly near 370C and exhibit decreased affinity at lower temperatures. Cold autoantibodies on the other hand, bind to red cells most strongly near 0-40C  and typically shows little affinity at physiologic temperature.

Occasionally, patients have a combination of warm and cold antibodies. Cases of auto immune haemolyticanaemia are subdivided further on the basis of aetiology. By definition   idiopathic or primary autoimmune haemolyticanaemia shows no apparent association with an underlying disorder. Depending on the patient population studied, a suspected secondary cause of autoimmune haemolyticanaemia has been determine in 20-80% of reported series; these include lymphoproliferitive disorders autoimmune disorder, immunodeficiency disorder and tumours (Sokol, Hewitt and Stamps, 2002). Lymphoproliferative warm  and cold autoimmune   haemolyticanaemia.  Idiopathic disease is more common in women and has peak incidence in the fourth and fifth decades of  life. The demographic of secondary disease correspond  with those  of the various underlying illnesses

Autoantibodies aetiology

The aetiology of most red blood cell autoantibodies is not well understood. Given the association between AIHA and other autoimmune disorders, generalized immune system dysfunction likely plays a role.

The relationship between AIHA and lymphoproliferative disorder and other neoplasm likewise suggest generalized dysfunction of immune surveillance. The immune system has many control point that keep a balance between the need to tolerate self –antibodies and the need to respond appropriately to foreign antigen. Immune self tolerance occurs centrally with developing lymphocyte precursors via clonal deletion or clonal anergy, and peripherally with mature  T and B cells via down regulation of the immune response.

Disruption of any of these processes may be a potential cause of autoimmune disease. Less generalized autoimmune process also exist for example autoimmune disease can arise from the response to a foreign antigen if the foreign antigen shows sufficient homology with self –antigen. Another theoretical source of the auto-antibodies is a malignant B cells clone, but red blood cell auto-antibodies generally are polyclonal. On the bases of the studies with other autoimmune  disorder such as ankylosing  spondylitis and multiple  sclerosis,  both genetic and environmental factor likely play a role in the production of red blood cell auto-antibody

Signs and symptoms

  • Paleness of the skin
  • Fatigue
  • Fever
  • Confusion
  • Light headiness
  • Dark urine

Laboratory diagnosis (serology)

Two criteria must be met to diagnose autoimmune haemolytic anaemia: serological evidence of autoantibody and clinical or laboratory evidence of haemolysis. serological evidence an auto autoantibody is provided by a positive auto control and direct antiglobulin test  (DAT, direct comb’s test) result and subsequent identification of an autoantibody  in the red  blood cell elute and possibly the serum. Serum reactivity with autologous red blood cells generally indicates the presence of an autoantibody but, it does not exclude the presence of an alloantibody.

In addition if the patient have been transfuse recently,alloantibodies  bound to the transfuse cells  can produce a mixed field  positive auto control. While the autocontrol measures invitro binding of red blood cells, the DAT indicate whether there is in vivo red cell binding of either igG or C3d. the DAT involves mixing the individual’s anti-coagulated red blood cell with polyspecific anti-human globulin (AHG) with anti- igG and anti C3d activities; if positive, the sample can be tested separately (“split”) with reagent specific for anti –igG or anti-C3d. A positive DAT is non-specific for a red cell autoantibody. Additional primary cause of a positive DAT includes an acute or  delayed haemolytic transfusion reaction, haemolytic disease of the new born, transplantation, drug induce antibodies, and administration of various therapies including intravenous immunoglobulins  (IVIG), Rh immune globulins (RIG), anti-lymphocyte globulins. A positive DAT test may arise from secondary to sickle cell disease,  thalasamia, renal disease, multiple myeloma, auto immune diseases, AIDS and other disease with elevated serum globulins (Clark, Tanley and Wallas, 2000).

Given the broad differential for a positive DAT, supplemental serological testing is necessary to ascertain the cause. The antibody screen detects both unbound autoantibodies and alloantibodies. Antibody bound to red blood cell can be eluted off the cell surface and subsequently identified in theelute. Both the red cell elute and the serum sample can be examine for soluble auto-antibodies and/or allo-antibodies using diagnostic red blood cell panels and indirect anti- globulin testing. Drug dependant antibodies, On the other hand, may not be detected unless the drug is added for invitro testing. Generally autoantibodies react with all panel red blood cells (i.e. pancreative), whereas  allo-antibodies exhibit antigen specificity, only reacting with specific antigen  positive cells.

Other technique may be necessary to distinguish between the two types. For example an alloantigen to a high incidence antigen in a post transfusion setting can mimic an auto- antibodies with a positive DAT (mixed field) and reaction with all panel cells. In addition, auto-antibodies may exhibit apparent specificity. Auto-adsorption uses autologous red cell to adsorb auto-antibodies prior to repeating serum testing for auto-antibodies. If an antibody exhibit specificity, then demonstration that autologous red blood cell are negative for the corresponding antigen confirms that it is an allo- antibody ; in the absent of a recent transfusion, a positive result that it is an autoantibody. Auto-adsorption can also be used to cross check donor’s red blood cell units for patient with warm auto-antibodies (only for patient who have not been transfuse recently).

In addition to the customary serum antibody identification, a red cell elute should be tested for antibodies if the positive DAT SHOWS igG reactivity. IgM auto-antibodies generally detach from the red cell surface in vivo, so they are not detected directly in the DAT and are not present in the elute. Elution may also concentrate any existing bound antibodies and thus improve sensitivity. Auto-antibodies are approximately 80% of the cases of AIHA (Issit,Pavone, and Goldfinger, 2002). Together with the DAT, result from serum and red blood cell elute antibody studies help distinguish the forms of AIHA

Laboratory diagnosis (haemolysis)

The diagnosis of AIHA also requires manifestations of haemolytic anaemia. From 0.007% to 0.1% of healthy blood donors and from 0.3% to 8% of hospital patient have positive DAT, without clinical evidence of immune haemolytic anaemia (IHA) (Judd, Barns, andStaner, 2008). Haemolysis in AIHA can either be extra vascular or intravascular. Typically intravascular haemolysis has a rapid and aggressive presentation, whereas extra vascularhaemolysis is milder. A CBC with peripheral smear, bilirubin, LDH (In particular iso-enzyme1), haptoglobin, and urine haemoglobin are the basic test use to evaluate and differentiate intravascular and extra vascularhaemolysis

Procedure for direct anti-globulin test

Specimen: Anticoagulated whole blood

Technique

  1. Wash patient’s red cells three times with saline and prepare a 5% suspension.
  2. Add 0.02ml of washed red cells to 0.4ml of anti-human globulin serum (AHGS) in a test tube.
  3. Allow it to stand for five minutes and centrifuge at 100g for 15 seconds
  4. Examine the tube for agglutination macroscopically and microscopically.

Result:Agglutination indicates the presence in vivo sensitization of red blood cells by an antibody showing that, the direct comb’s test is positive

Treatment

Much literature exist regarding the treatment of AIHA. Efficiency of treatment depends on the correct diagnosis of either warm or cold type AIHA.

Warm type AIHA is usually a more insidious, not treatable by simply removing the underlying cause. First line therapy for this is usually the use of steroids medication such as prednisone. If steroids medication did not improve the condition, removal of the spleen (splenectomy) may be considered.

Therapy to suppress the immune system is usually given if the person does not respond to steroids splenectomy. Medication such as rituximab (Retuxan), danazol, cyclophosphamide (Cytoxan), azothioprine (Imuran) or cyclosporine have been used. Blood transfusion are given with caution because of the potential that blood may not be compatible and may cause further haemolysis.

Cold agglutinin disease is treated by avoiding cold or sometimes with rituximab. Removing the underlying cause is also important.

Conclusion

Autoimmune haemolytic anaemia causes the destruction of an individual’s red blood cells by either or both warm and cold autoantibody from the person’s own immune system. Prevention of this disease especially the warm AIHA is unknown since the causes are not well understood. It is therefore advisable to carried out a screen test for AIHA for every anaemic patient that want to undergo blood transfusion or organ transplantation in order to treat the disease before transfusion to prevent further haemolysis of the red blood cells that will be transfuse In the other hands the cold AIHA can be prevented by avoiding the cold.

References

Abrahamson, N. Gelfand, E.W. Jandi,J.H. andRosen,F.S. (2008).The interaction between human monocyte and red cells.specificity      for igG subclasses and igG fragments. J. Exp Med 132: 1207-1215

Clark,J.A., Tanley, P.C. andWallas,C.H. (2000). Evaluation of patient with positive direct tests and more reactive elutes discovered during pretransfusion testing. Immunohematology8:9-12

Ehlenberger, A.G. andNussezweig, V. (1997).The role of membrane receptor for C3b and   C3d in phagocytosis.J Exp Med 145:357-371.

Issit,P.D., Pavone,B.G., Goldfinger, D., Zwikder, H., Issit,C.H., Tessel,J.A., Krovand, S.W. and Bell, C.A. (2002).Anti Wrb and other autoantibodies responsible for positive direct antiglobulin test in 150 individuals.Br J Haematol 36:5 -18

Judd,W.J., Bernes, B.A., Staner,E.A., Oberman,H.A., Averill,D.B. and Butch,S.H. (2008). The evaluation of a positive direct antiglobulin test (autocontrol) in pretransfusion testing revisited. Transfusion 26:220-224

Pirofsky, B. (2004).Autoimmunization and autoimmune haemolyticanaemia. Baltimore: Williams and Wingins.

Sakol, R.J., Hewitt, S. and Stamps,B.K. (2002). Autoimmune haemolysis: An 18-year study of 865 cases referred to a regional transfusion centre. Br Med J 282:2023-2027.

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