Sublytic Terminal Complement Components Induce Eryptosis in Autoimmune Haemolytic Anaemia Related to IgM Autoantibodies

Abstract

BACKGROUND/AIMS: Eryptosis, the suicidal death of red blood cells (RBCs), is characterized by phosphatidylserine (PS) exposure at the cell surface. It can be catalysed by a variety of abnormal conditions and diseases. Until now, the many questions surrounding the physiology and pathophysiology of eryptosis have not been sufficiently answered. Recently, we demonstrated IgM and IgA autoantibodies (aab) to induce PS exposure on circulating RBCs of patients with autoimmune haemolytic anaemia (AIHA). However, it remained unclear how these aab lead to eryptosis.

METHODS: Serum and plasma samples from patients with clinically relevant AIHA of cold type were used to induce eryptosis in O RBCs. Serum containing fresh complement from healthy donors, antibodies to complement component, and complement factor depleted sera were added to examine the influence of the complement on PS-exposure. RBC bound annexin V PE were analysed by flow cytometry.

RESULTS: Eryptosis related to IgM aab was found to be dependent on complement activation and could be effectively inhibited by EDTA, serum heat inactivation and anti-C5. PS exposure increased with sequential activation of the sublytic terminal complement components C5b6, C5b-7 and was most significant at the C5b-8 stage. A decrease was observed following the formation of the lytic membrane attack complex C5b-9, either because of lysis of eryptotic RBCs or because of inhibition of eryptosis by C9.

CONCLUSION: Our findings reflect new aspects on RBC destruction in AIHA as well the impact of the terminal complement complexes on the RBC membrane. The striking differences to nucleated cell apoptosis may even have physiological meaning of RBC acting as a buffer of the complement system.