Regulation of intracellular glutathione levels in erythrocytes infected with chloroquine-sensitive and chloroquine-resistant Plasmodium falciparum

Abstract

Malaria is one of the most devastating tropical diseases despite the availability of numerous drugs acting against the protozoan parasite Plasmodium in its human host. However, the development of drug resistance renders most of the existing drugs useless. In the malaria parasite the tripeptide glutathione is not only involved in maintaining an adequate intracellular redox environment and protecting the cell against oxidative stress, but it has also been shown that it degrades non-polymerized ferriprotoporphyrin IX (FP IX) and is thus implicated in the development of chloroquine resistance. Glutathione levels in Plasmodium -infected red blood cells are regulated by glutathione synthesis, glutathione reduction and glutathione efflux. Therefore the effects of drugs that interfere with these metabolic processes were studied to establish possible differences in the regulation of the glutathione metabolism of a chloroquine-sensitive and a chloroquine-resistant strain of Plasmodium falciparum. Growth inhibition of P. falciparum 3D7 by D,L-buthionine-( S, R )sulphoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase (gamma-GCS), and by Methylene Blue (MB), an inhibitor of gluta thione reductase (GR), was significantly more pronounced than inhibition of P. falciparum Dd2 growth by these drugs. These results correlate with the higher levels of total glutathione in P. falciparum Dd2. Short-term incubations of Percoll-enriched trophozoite-infected red blood cells in the presence of BSO, MB and N, N (1)-bis(2-chloroethyl)- N -nitrosourea and subsequent determinations of gamma-GCS activities, GR activities and glutathione disulphide efflux revealed that maintenance of intracellular glutathione in P. falciparum Dd2 is mainly dependent on glutathione synthesis whereas in P. falciparum 3D7 it is regulated via GR. Generally, P. falciparum Dd2 appears to be able to sustain its intracellular glutathione more efficiently than P. falciparum 3D7. In agreement with these findings is the differential susceptibility to oxidative stress of both parasite strains elicited by the glucose/glucose oxidase system.

Stage independent chloroquine resistance and chloroquine toxicity revealed via spinning disk confocal microscopy

“…That is, the heme-based toxicity of CQ can only be observed while Hz is being synthesized. Since glutathione levels have been suggested to contribute to CQR and since Dd2 parasites apparently contain elevated total glutathione [42] perhaps other reactive species generated by CQ in the S DV are the basis of CQ toxicity at S. Alternatively, although much work remains to be done, our data (Fig. 10) also support minor, but continued, Hb metabolism in S. Hz microcrystals made in S that are unresolvable by optical microscopy (Fig.…”

Plant homologs of the Plasmodium falciparum chloroquine-resistance transporter, Pf CRT, are required for glutathione homeostasis and stress responses

“…The identification of the malaria parasite PfCRT as a CLT homolog supports this theory, because it contains a relic plastid derived from an early endosymbiotic event with a cyanobacterium (31,32). Interestingly, both clt mutants and chloroquine-resistant plasmodia carrying PfCRT mutations have altered GSH homeostasis (33). This observation, combined with the shared evolution of these proteins, suggests a level of functional conservation between CLTs and PfCRT.…”

Blood schizontocidal activity of methylene blue in combination with antimalarials againstPlasmodium falciparum

“…Antagonism with AMO and CQ is surprising as MB (like 4-aminoquinolines) concentrated in Plasmodium food vacuole where it inhibits the formation of hemozoin (Atamna et al, 1996). It is also known to inhibit parasite glutathione reductase (Färber et al, 1998) enhancing chloroquine activity (Meierjohann et al, 2002). We thus decided to study the interaction between MB and known antimalarials with a slightly different model from that of Akoachere et al (2005).…”

“…Moreover, MB inhibits parasite glutathione reductase thus jeopardizes glutathione functionality (Färber et al, 1998). Instead of antagonise, this phenomenon should enhance chloroquine activity, which is closely related to glutathione levels (Meierjohann et al, 2002). We thus re-examined herein the activity of MB on African and South-American P. falciparum strains, determined the most sensitive parasitic stage and studied the association of MB with classical antimalarial drugs, as the spread of multidrug-resistant Plasmodium strains favours the use of combinations to protect the individual compounds from emergence of resistance [WHO-TDR, 2006].…”