Synthetic Red Blood Cell-Specific Glycolytic Intermediate 2,3-Diphosphoglycerate (2,3-DPG) Inhibits Plasmodium falciparum Development In Vitro

Abstract

Mechanisms of malaria parasite interaction with its host red blood cell may provide potential targets for new antimalarial approaches. Pyruvate kinase deficiency has been associated with resistance to malaria in both experimental models and population studies. Two of the major pyruvate kinase deficient-cell disorders are the decrease in ATP and the increase in 2,3-biphosphoglycerate (2,3-BPG) concentration. High levels of this metabolite, only present in mammalian red blood cell, has an inhibitory effect on glycolysis and we hypothesized that its accumulation may also be harmful to the parasite and be involved in the mechanism of protection provided by that enzymopathy. We examined the effect of a synthetic form, 2,3-DPG, on the Plasmodium falciparum intraerythrocytic developmental cycle in vitro. Results showed an impairment of parasite growth with a direct effect on parasite maturation as significant lower progeny emerged from parasites that were submitted to 2,3-DPG. Further, adding the compound to the culture medium did not result in any effect on the host cell, but instead the metabolic profile of an infected cell became closer to that of a non-infected cell.

2,3-Diphosphoglycerate and the Protective Effect of Pyruvate Kinase Deficiency against Malaria Infection—Exploring the Role of the Red Blood Cell Membrane

“…These effects on the deformability of noninfected cells and on the RBC zeta potential were not sufficient to impair the efficient reinvasion of new RBCs upon exposure to 2,3-DPG. Thus, it cannot significantly contribute to the substantial decrease in the number of parasites that start the following cycle, as previously observed [ 15 ]. However, in the reinvasion assay, there was never a mixture with 2,3-DPG between the initial culture conditions before the magnetic selection of mature parasites, and the second condition in which the enriched parasites were re-cultured in previously treated RBC, by then washed and cultured in a medium without 2,3-DPG addition.…”

“…We have been investigating how the glycolytic metabolite 2,3-DPG may be involved in the protective effect against malaria caused by PKD. Morais et al [ 15 ] showed that, when 2,3-DPG 8 mM was added daily into a culture medium, there was a sharp reduction in parasite densities and a reduction in parasite progeny at the end of an intraerythrocytic developmental cycle. Furthermore, a shift of the metabolic profile of iRBC treated with 2,3-DPG, making it closer to that of niRBC, was observed.…”

“…Previous results hinted the involvement of the RBC-specific metabolite 2,3-DPG in the protective mechanism afforded by PKD. Morais et al [ 15 ] showed that mature schizonts divide into a lower number of merozoites when cultured in the presence of the compound than those cultured at standard conditions, indicating that the parasites are not developing as efficiently. They also observed that there are no changes in the ATP levels inside the host cell and that the metabolomic profile of noninfected cells either treated or untreated did not differ significantly, while the profile of iRBCs treated with 2,3-DPG became more similar to the profile of niRBCs than to that of untreated iRBCs, which suggests a main effect of the addition of the compound on the parasite rather than on the host cell.…”

“…Healthy-type 0 RBCs were collected from adult volunteer donors ( n = 5). To rule out blood variants that could bias the effect on parasite growth, a sample of whole blood from each donor was used for DNA extraction and subsequent molecular diagnosis of the polymorphisms most common in Portugal, linked to hemoglobinopathies and RBC enzymopathies associated with malaria protection (HBB—hemoglobin subunit beta, pklr —pyruvate kinase, liver and red blood cell and g6pd genes), as described by Morais et al [ 15 ]. All donors were wild types for all studied genes.…”

“…and may be involved in the mechanism of protection against infection provided by PKD. In fact, Morais et al [ 15 ] observed an impairment of P. falciparum growth in vitro under the effect of 2,3-DPG added to the culture medium. As 2,3-DPG affects membrane proteins, altering the host cell’s membrane, it may contribute to an increased RBC clearance, as well as also interfere with the parasite ability to infect new RBCs.…”