Effects of sevuparin on rosette formation and cytoadherence of Plasmodium falciparum infected erythrocytes

Abstract

In severe falciparum malaria cytoadherence of parasitised red blood cells (PRBCs) to vascular endothelium (causing sequestration) and to uninfected red cells (causing rosette formation) contribute to microcirculatory flow obstruction in vital organs. Heparin can reverse the underlying ligand-receptor interactions, but may increase the bleeding risks. As a heparin-derived polysaccharide, sevuparin has been designed to retain anti-adhesive properties, while the antithrombin-binding domains have been eliminated, substantially diminishing its anticoagulant activity. Sevuparin has been evaluated recently in patients with uncomplicated falciparum malaria, and is currently investigated in a clinical trial for sickle cell disease. The effects of sevuparin on rosette formation and cytoadherence of Plasmodium falciparum isolates from Thailand were investigated. Trophozoite stages of P. falciparum-infected RBCs (Pf-iRBCs) were cultured from 49 patients with malaria. Pf-iRBCs were treated with sevuparin at 37°C and assessed in rosetting and in cytoadhesion assays with human dermal microvascular endothelial cells (HDMECs) under static and flow conditions. The proportion of Pf-iRBCs forming rosettes ranged from 6.5% to 26.0% (median = 12.2%). Rosetting was dose dependently disrupted by sevuparin (50% disruption by 250 μg/mL). Overall 57% of P. falciparum isolates bound to HDMECs under static conditions; median (interquartile range) Pf-iRBC binding was 8.5 (3.0-38.0) Pf-iRBCs/1000 HDMECs. Sevuparin in concentrations ≥ 100 μg/mL inhibited cytoadherence. Sevuparin disrupts P. falciparum rosette formation in a dose dependent manner and inhibits cytoadherence to endothelial cells. The data support assessment of sevuparin as an adjunctive treatment to the standard therapy in severe falciparum malaria.

Fig 1

(A) Effect of sevuparin on rosetting (%) of P. falciparum (n = 47) is dose dependent. The data show the median with interquartile range of rosettes formed at each concentration of sevuparin. (B) Median (interquartile range) % disruption of rosetting at each concentration of sevuparin. Sevuparin significantly disrupted rosette formation, p < 0.001.

Fig 2

(A) The effects of sevuparin on Pf-iRBC adherence (n = 28). Median (interquartile range) adherence number of Pf-iRBCs binding per 1000 HDMECs at each concentration of sevuparin. (B) The inhibition of sevuparin on cytoadherence of P. falciparum (n = 28). Median (interquartile range) inhibition effect on cytoadhesion. The cytoadherence of patient isolates was significantly inhibited by sevuparin at concentrations ≥ 100 μg/mL (all p < 0.05).

Fig 3. The correlation between the adherent number of Pf-iRBCs binding per 1000 HDMECs and rosetting number (%) of P. falciparum (n = 28), (r_s = 0.152, p = 0.439).

Fig 4. Median (interquartile range) Pf-iRBCs adhering to HDMECs under shear stress at 0.05 Pa, 0.5% parasitaemia and 1% haematocrit at different concentrations of sevuparin (n = 10).