Unraveling the C-reactive protein complement-cascade in destruction of red blood cells: potential pathological implications in Plasmodium falciparum malaria

Abstract

Background: Deficiencies of the complement-regulatory proteins on RBC (RBC_Mal) of patients with Plasmodium falciparum were reported. Here, we sought to determine the role of affinity-purified C-reactive protein from patients (CRP_Mal), in modulating the complement-regulatory proteins and downstream effect on complement-cascade. Methods: CRP_Mal was characterized by analytical ultracentrifuge and electrophoretic analysis. Surface plasmon resonance, Western blotting, co-immuno-precipitation, flow-cytometry and ELISA determined the binding of CRP_Mal with RBC_Mal. Modifications of membranes for RBC_Mal-CRP_Mal binding were explored by scanning electron microscopy, osmotic and turbulence fragility, hydrophobicity and oxyhemoglobin release. Flow-cytometry, ELISA, Western blotting and Scatchard analysis monitored the status of complement-regulatory proteins on RBC_Mal. Complement-activation via CRP_Mal was quantified by C3-deposition and hemolysis. Results: CRP_Mal binds specifically to RBC_Mal through distinct molecules. Such binding altered the normal discoid-shape of RBC_Mal with increased membrane fluidity and hydrophobicity. In the presence of CRP, RBCMal showed reduced complement-regulatory proteins (CR1 or CD35, CD55 and CD59) with decreased affinity. These changes caused enhanced C3-deposition and complement-mediated hemolysis. Conclusion: Taken together, we have established the contributory effect of CRP_Mal causing decreased complement-regulatory proteins, possibly providing a new mechanism of complement-fueled RBC_Mal destruction refractory to erythrophagocytosis and may account for pathogenesis of anemia.