On the adhesion dynamics of Plasmodium falciparum infected red blood cells

2017-03-20T00:48:32Z (GMT) by Emma Hodges
Adhesion of malaria parasite-infected red blood cells (iRBCs) to microvascular endothelium is a central event in the pathogenesis of severe falciparum malaria. A biophysical approach was taken to study the dynamics of this adhesive process using two different experimental techniques: dual micropipette aspiration and optical tweezers.
   A dual micropipette adhesion assay was used to investigate the probability and strength of adhesion between an iRBC and an endothelial cell expressed receptor (CD36) that is important in the pathogenesis of malaria. CD36 is an important receptor to study, not only is it one of the best characterized receptor-parasite interactions but it also has an important role in malaria pathology with respect to its role in severe malaria which is still not well understood. In fact CD36 blocking intervene has been looked at as treatment strategy for severe malaria, however little improvement is seen making it vital that we understand more about the adhesion that occurs between CD36 and iRBCs.
   A novel method was employed to assess the adhesion of individual iRBCs for which the shear elastic modulus was also obtained. This allowed for the first time the determination of the influence of membrane rigidity on adhesion. At a constant compression force, an increased membrane stiffness resulted in a decreased contact area. However, an increased membrane stiffness resulted in an increased adhesion force, at a constant contact area. An optical tweezer assay was also employed to measure adhesion. The advantage of this method is that it provides a means of noninvasive manipulation of objects in solution, however there are limitations with respect to the maximum force that can be applied to break the adhesion bonds (~100pN).
   The two experimental methods gave comparable results. The adhesion probability increased with increasing contact time, until approximately 10s where it remained stable at ~ 40%. A model for 2D kinetic adhesion was fitted to obtain a kinetic rate of dissociation, kd = 0.11+/-0.02s-1 and kd = 0.089+/-0.025s-1 for the micropipette and optical tweezer method respectively. The grouped adhesion constant (mrmlKoA) was found to be 0.086 +/- 0.014 using optical tweezers. Increased contact area resulted in an increase in the adhesion strength for both methods. The optical tweezermethod further showed an increased contact time correlated to an increased adhesion strength.
   A limitation with these experimental techniques is that they are inherently non-equilibrium in nature. Consequently, a Langevin simulation was developed to model the detachment of a bead held in an optical trap, from a membrane to which it is initially bound to explore the use of fluctuation theorems to obtain equilibrium values from non-equilibrium work trajectories. The equilibrium free energy of binding was obtained for various tweezer pulling rates using fluctuation theorems. Further, umbrella sampling was used to obtain the equilibrium probability of detachment for a variety of trap potentials.