Relaxometric studies of erythrocyte suspensions infected by Plasmodium falciparum: a tool for staging infection and testing anti-malarial drugs

Abstract

Purpose: Malaria is a global health problem with the most malignant form caused by Plasmodium falciparum (P. falciparum). Parasite maturation in red blood cells (RBCs) is accompanied by changes including the formation of paramagnetic hemozoin (HZ) nanocrystals, and increased metabolism and variation in membrane lipid composition. Herein, MR relaxometry (MRR) was applied to investigate water exchange across RBCs' membrane and HZ formation in parasitized RBCs.

Methods: Transverse water protons relaxation rate constants (R₂ = 1/T₂ ) were measured for assessing HZ formation in P. falciparum-parasitized human RBCs. Moreover, water exchange lifetimes across the RBC membrane (τ_i ) were assessed by measuring longitudinal relaxation rate constants (R₁ = 1/T₁ ) at 21.5 MHz in the presence of a gadolinium complex dissolved in the suspension medium.

Results: τ_i increased after invasion of parasites (ring stage, mean τ_i / ${\tau }_i^0$ = 1.234 ± 0.022) and decreased during maturation to late trophozoite (mean τ_i / ${\tau }_i^0$ = 0.960 ± 0.075) and schizont stages (mean τ_i / ${\tau }_i^0$ = 1.019 ± 0.065). The HZ accumulation in advanced stages was revealed by T₂ -shortening. The curves reporting R₂ (1/T₂ ) vs. magnetic field showed different slopes for non-parasitized RBCs (npRBCs) and parasitized RBCs (pRBCs), namely 0.003 ± 0.001 for npRBCs, 0.009 ± 0.002, 0.028 ± 0.004 and 0.055 ± 0.002 for pRBCs at ring-, early trophozoite-, and late trophozoite stage, respectively. Antimalarial molecules dihydroartemisinin and chloroquine elicited measurable changes in parasitized RBCs, namely dihydroartemisinin modified τ_i , whereas the interference of chloroquine with HZ formation was detectable by a significant T₂ increase.

Conclusions: MRR can be considered a useful tool for reporting on P. falciparum blood stages and for screening potential antimalarial molecules.

Keywords: Gd-complex; Magnetic Resonance Relaxometry; Malaria; Plasmodium falciparum; Red blood cells; chloroquine; dihydroartemisinin; hemozoin; water exchange rate; water permeability.