Beneficial effect of aurothiomalate on murine malaria

Abstract

Background: Premature death of Plasmodium-infected erythrocytes is considered to favourably influence the clinical course of malaria. Aurothiomalate has previously been shown to trigger erythrocyte death or eryptosis, which is characterized by cell membrane scrambling leading to phosphatidylserine exposure at the cell surface. Phosphatidylserine-exposing cells are rapidly cleared from circulating blood. The present study thus tested whether sodium aurothiomalate influences the intraerythrocytic parasite development in vitro and the clinical course of murine malaria in vivo.

Methods: Human erythrocytes were infected with Plasmodium falciparum BinH in vitro and mice were infected (intraperitoneal injection of 1 x 106 parasitized murine erythrocytes) with Plasmodium berghei ANKA in vivo.

Results: Exposure to aurothiomalate significantly decreased the in vitro parasitemia of P. falciparum-infected human erythrocytes without influencing the intraerythrocytic DNA/RNA content. Administration of sodium aurothiomalate in vivo (daily 10 mg/kg b.w. s.c. from the 8th day of infection) enhanced the percentage of phosphatidylserine-exposing infected and noninfected erythrocytes in blood. All nontreated mice died within 30 days of infection. Aurothiomalate-treatment delayed the lethal course of malaria leading to survival of more than 50% of the mice 30 days after infection.

Conclusions: Sodium aurothiomalate influences the survival of Plasmodium berghei-infected mice, an effect only partially explained by stimulation of eryptosis.

Figure 1

Effects of sodium aurothiomalate on intraerythrocytic amplification and in vitro parasitemia. A. In vitro parasitemia with P. falciparum (left panel) in human erythrocytes as a function of the aurothiomalate concentration (arithmetic means ± SEM, n = 16). *, *** indicate significant difference (p < 0.05, p < 0.001) from the absence of aurothiomalate. Intraerythrocytic DNA amplification (right panel) as a function of the aurothiomalate concentration (arithmetic means ± SEM, n = 12). B. Intraerythrocytic DNA amplification (right panel) as in B for different time periods (arithmetic means ± SEM, n = 8).

Figure 2

Effects of aurothiomalate on phosphatidylserine exposure of infected and noninfected human erythrocytes. Arithmetic means ± SEM (n = 6) of the percentage of annexin V-binding infected (open bars) and non-infected (closed bars) erythrocytes following infection of human erythrocytes with P. falciparum in the presence of 0 - 100 μM aurothiomalate. ##, ### indicate significant difference (p < 0.01, p < 0.001) from non-infected erythrocytes, **, *** indicate significant difference (p < 0.01, p < 0.001) from the absence of aurothiomalate.

Figure 3

Effect of sodium aurothiomalate treatment on phosphatidylserine exposure of infected and non-infected erythrocytes from Plasmodium berghei-infected mice. Arithmetic means ± SEM (n = 6-8) of the percentage of phosphatidylserine-exposing infected (right bars) and non-infected (left bars) erythrocytes taken from animals without (black bars) and with (white bars) sodium aurothiomalate treatment (daily 10 mg/kg b.w. s.c.) on the 22^nd day after infection. ### indicates significant difference (p < 0.001) from absence of sodium aurothiomalate. *** indicates significant difference (p < 0.001) from noninfected erythrocytes.

Figure 4

Effect of sodium aurothiomalate treatment on the parasitemia and survival of Plasmodium berghei-infected mice. A: Original histograms of parasitemia-dependent Syto16 fluorescence in untreated animals (upper panels) and in animals treated daily from day 8 daily with 10 mg/kg b.w. sodium aurothiomalate s.c. (lower panels) 10 (left panels) and 20 (right panels) days after infection with P. berghei. B: Arithmetic means ± SEM of the parasitemia in mice without treatment (open circles, n = 8 mice) or with daily 10 mg/kg b.w. s.c. of sodium aurothiomalate (closed circles, n = 6 mice) as a function of days after infection with P. berghei. Significant difference (* p < 0.05, ** p < 0.01, *** p < 0.001; t-test) from the untreated animals on days 12 - 20. The results presented are one of three independent series. C: Survival of mice without treatment (open circles) or with daily 10 mg/kg b.w. sodium aurothiomalate s.c. (closed circles) as a function of days after infection with Plasmodium berghei. D-E: Arithmetic means ± SEM of the parasitemia in mice without treatment (open circles, n = 4 mice) or with daily 10 mg/kg b.w. s.c. of sodium aurothiomalate (closed circles, n = 4 mice) as a function of days after infection with P. berghei. The parasitemia was determined daily either by staining with Syto16 and subsequent FACS analysis as in B (D) or by daily Giemsa staining of blood smears and light microscopy-dependent analysis (E).

Figure 5

Effect of sodium aurothiomalate treatment on the hematocrit of Plasmodium berghei-infected mice. Arithmetic means ± SEM of packed cell volume (hematocrit) in mice without treatment (open circles, n = 8 mice) or with daily 10 mg/kg b.w. s.c. of sodium aurothiomalate (closed circles, n = 8 mice) as a function of days after infection with P. berghei. * indicates significant difference (p < 0.05; t-test).