SEQUESTRATION, VIRULENCE AND FUTURE INTERVENTIONS IN PLASMODIUM FALCIPARUM MALARIA

The Plasmodium falciparum infected erythrocyte (iRBC) withdraws from the peripheral circulation to hide in the deep microvasculature during maturation. This phenomenon, referred to as sequestration, is believed to cause extensive clogging of the vessels and to contribute to the development of severe malaria. Adhesion of the iRBC to the endothelium (cytoadhesion) as well as to uninfected and infected erythrocytes (rosetting and autoagglutnation respectively) is thought essential in this process. This interaction with host cell receptors is mediated by parasite derived proteins of which P. falciparum erythrocyte membrane protein 1 (PfEMP1) is the most studied. The investigations performed within this thesis aim to further elucidate the correlation between PfEMP1-mediated adhesive events and clinical manifestations of severe malaria. This is accomplished by analysis of fresh clinical parasite isolates and in vivo studies in a new animal model. Furthermore, immunization and adjuvant drug treatment, both targeting the adhesive properties here shown associated with severe disease, have been evaluated in our new model. Examination of the binding properties of iRBC from children with different forms of severeor mild malaria, revealed the total adhesive capacity and the capability to simultaneously adhere to several receptors to be associated with severe disease. It also demonstrated that the levels of binding to heparin and blood group A as well as the rosetting rate are significantly higher with iRBC from children with severe malaria as compared to mild disease. These findings implicate the adhesive features of a specific binding domain, the DBL-1α domain of the PfEMP1 to be important in bringing about of severe malaria. By the injection of human iRBC into non-manipulated rats it was shown that parasites sequester in the lungs of the animals and induce pathological changes in a strain specific manner. Additional in vitro experiments confirmed the adhesion of iRBC to the rat endothelium and demonstrated the involved receptors to be the same as those of the human endothelium. The approach of injecting human iRBC was further explored in non-human primates. As adhesion of iRBC to heparin was found associated with severe disease, we generated a new heparin-like glycan devoid of anticoagulant activity to study anti sequestering effects. A substantial decrease in sequestration in vivo was achieved with this substance in both rats and primates, suggesting drugs based on glycans as promising candidates for adjuvant treatment of severe malaria. The FCR3S1.2 parasite is well recognized by sera from children in a malaria endemic region and exhibit all the adhesive features here associated with severe disease. It further sequesters and induces specific pathology in our rat model and is therefore suggested as a severe malaria model parasite. Immunization of animals with the DBL-1α domain of the PfEMP1 of this parasite is here demonstrated to generate antibodies recognizing iRBC and capable of inhibiting sequestration in vivo in both rats and primates. This indicates this domain to be a promising component in a future vaccine. In conclusion we have established a role for the DBL-1α domain of PfEMP1 in bringing about severe malaria and demonstrated how immunization with this domain, or targeting of its adhesive properties with a new glycan, can inhibit sequestration in vivo. We have also established a new animal model in which sequestration and possibly additional pathological events can be studied.