Effects of the vascular endothelial growth factor receptor-2 (VEGFR-2) inhibitor SU5416 on in vitro cultures of Plasmodium falciparum.

Casper Hempel,Nils Hoyer,Jørgen A Kurtzhals,Trine Staalsø

doi:10.1186/1475-2875-13-201

Casper Hempel, Nils Hoyer + Show 2 more

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https://doi.org/10.1186/1475-2875-13-201

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Abstract

BackgroundVascular endothelial growth factor (VEGF) is taken up by parasitized red blood cells during malaria and stimulates intra-erythrocytic growth of Plasmodium falciparum in vitro. The cause and consequence of this uptake is not understood.MethodsPlasmodium falciparum was cultured in vitro. Parasite growth and intracellular VEGF levels were assessed using flow cytometry. Intracellular VEGF was visualized by fluorescence immunocytochemistry. Phosphorylated tyrosine was measured by western blotting. In vivo assessment of intra-erythrocytic VEGF was performed in Plasmodium berghei ANKA-infected C57BL/6 mice.ResultsVEGF accumulated intracellularly in infected red blood cells, particularly in schizonts. In vitro growth of P. falciparum was unchanged when co-cultured with the anti-VEGF antibody bevacizumab or with an anti-VEGF receptor-1 peptide. In contrast, the VEGF receptor-2 inhibitor, SU5416, dose-dependently inhibited growth. None of the treatments reduced intracellular VEGF levels. Thus, the anti-parasitic effect of SU5416 seemed independent of VEGF uptake. SU5416 reduced phosphorylated tyrosine in parasitized red blood cells. Similarly, the broad-spectrum tyrosine kinase inhibitor genistein dose-dependently inhibited P. falciparum growth and reduced tyrosine phosphorylation. Neither bevacizumab nor anti-VEGF receptor-1 peptide affected tyrosine kinase activity. Finally, in vivo uptake of VEGF in P. berghei ANKA was demonstrated, analogous to the in vitro uptake in P. falciparum, making it a possible model for the effects of VEGF signalling in vivo during malaria.ConclusionsInhibition of VEGFR-2 signalling reduces intra-erythrocytic growth of P. falciparum, likely due to tyrosine kinase inhibition. Internalisation of VEGF in P. falciparum-infected red blood cells does not rely on VEGF receptors. The function of in vivo uptake of VEGF can be studied in rodent malaria models.

Highlights

Vascular endothelial growth factor (VEGF) is taken up by parasitized red blood cells during malaria and stimulates intra-erythrocytic growth of Plasmodium falciparum in vitro
Sequestration of parasitized red blood cells (PRBCs) followed by local cerebral hypoxia and accumulation of hypoxia inducible factor (HIF)-1α is a possible cause of VEGF production during cerebral malaria (CM)
VEGF uptake in PRBCs Culture of P. falciparum in serum-enriched growth medium resulted in uptake of VEGF into PRBCs, determined by immunocytochemistry on day 6 after the first addition of serum (Figure 1)

Summary

Introduction

Vascular endothelial growth factor (VEGF) is taken up by parasitized red blood cells during malaria and stimulates intra-erythrocytic growth of Plasmodium falciparum in vitro. Sequestration of parasitized red blood cells (PRBCs) in cerebral blood vessels, resulting in local hypoxia and neuronal damage, is a key event in the pathogenesis of CM [2]. In nonimmune travellers and Kenyan children with malaria, VEGF is increased in both brain tissue and blood [4,5]. HIF-1α, which has a short half-life, was undetectable in human brain tissue post mortem, but a HIF-1α associated protein, DEC-1 was upregulated in neurons [9]. Sequestration of PRBCs followed by local cerebral hypoxia and accumulation of HIF-1α is a possible cause of VEGF production during CM

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