Complement in hemolytic anemia

Abstract

Complement is increasingly being recognized as an important driver of human disease, including many hemolytic anemias. Paroxysmal nocturnal hemoglobinuria (PNH) cells are susceptible to hemolysis because of a loss of the complement regulatory proteins CD59 and CD55. Patients with atypical hemolytic uremic syndrome (aHUS) develop a thrombotic microangiopathy (TMA) that in most cases is attributable to mutations that lead to activation of the alternative pathway of complement. For optimal therapy, it is critical, but often difficult, to distinguish aHUS from other TMAs, such as thrombotic thrombocytopenic purpura; however, novel bioassays are being developed. In cold agglutinin disease (CAD), immunoglobulin M autoantibodies fix complement on the surface of red cells, resulting in extravascular hemolysis by the reticuloendothelial system. Drugs that inhibit complement activation are increasingly being used to treat these diseases. This article discusses the pathophysiology, diagnosis, and therapy for PNH, aHUS, and CAD.

Figure 1

Model of complement dysregulation in aHUS (A) and PNH (B). Initiators of the complement cascade (classical, alternative, and lectin pathway, as well as thrombin and plasmin/kallikrein) lead to C3 activation and C3 convertase formation, which is augmented the amplification loop (proximal complement). Consequently, membrane attack complexes (MAC) are formed on the membrane of target cells (terminal complement), leading to complement-mediated death. Production of C5a results in tissue factor and thrombin activation in a positive feedback loop. Complement dysregulation results from loss-of-function mutations in regulatory factors (Factor H, I, and THBD/thrombomodulin in aHUS and CD55, CD59 in PNH) shown in red, gain-of-function mutations (C3 and Factor B in aHUS) shown in green, and DGKE mutations in aHUS shown in gray, indicating the unknown effect on complement cascade. Eculizumab effectively inhibits terminal complement activation in both disorders by blocking the cleavage of C5 to C5a and C5b.

Figure 2

Frequency of genetic lesions in aHUS. CFH indicates complement factor H; CFI, complement factor I; THBD, thrombomodulin.

Figure 3

A) Model of Ham test for PNH (top) and indirect Ham test for aHUS diagnosis (bottom). The Ham test evaluates the effect of acidified serum on patient's cells, whereas the modified test evaluates the effect of patient's serum on GPI-anchored protein–deficient reagent cells. Both tests use absorbance changes as readouts of cell viability. GPI-AP indicates glycosylphosphatidylinositol-anchored proteins. B) WST-1 cell viability assay in patients with aHUS. Percentage of nonviable reagent cells in aHUS after exposure to acute-phase serum, remission serum on eculizumab (Ecu), and remission serum off eculizumab compared with acute or remission serum from TTP patients.