What happens to negatively charged lipid vesicles upon interacting with polycation species?

1. What happens to negatively charged lipid vesicles upon interacting with polycation species? Ali DURAN POLYMER TECHNOLOGY

2. INTRODUCTION • Synthetic polyelectrolytes are widely used now in medicine and biology, in particular for stimulation of immune response, • This requires to study the behaviour of polyelectrolytes in biological environment and especially, their interaction with cells, • It is known that a cell membrane usually carries a net negative charge, • Therefore, this study focused on synthetic polycations interacting with mixed vesicles composed of neutral phosphatidylcholine and diphosphatidylglycerol (cardilipin, CL2-) carrying two negative headgroups,

3. RESULTS AND DISCUSSION • It was found that binding of both CP(2) and CP(2,16) to solid and liquid DPPC-CL2-vesicles accompanied by neutralization of the liposome surface charge, registered by measuring EPM of polycation-vesicle complexes, • As follows from the figures, the largest particle size was observed at EPM=0, • Further increase in polycation concentration resulted in overcharging of the complex species and a decrease in particle size, • In this polycation concentration range, the complex species were stabilized against aggregation by the abundant positive charge of the adsorbed polycation,

4. RESULTS AND DISCUSSION • The cardinal difference in the behavior of solid and liquid vesicles revealed in the amount of negative CL2- molecules formed salt bonds with the polycation units. • As expected, in the case of the solid vesicles, the neutralizing amount of the polycation, measured at EPM=0, was equal to a half amount of CL2- involved in the vesicular membranes, • However, in the case of liquid liposomes, the EPM=0 point corresponded to the total amount of the membrane CL2- ,

5. RESULTS AND DISCUSSION • In addition, polycation adsorption induced lateral lipid segregation in the DPPC-CL membrane revealed in increase in the bilayer melting point and sharpening of DSC curves, • As a result, the initial membrane divided into two two-dimensional microphases: one composed of neutral DPPC molecules, and other of CL molecules kept together due to ionic contacts of their negative headgroups with positive polycation units,

6. RESULTS AND DISCUSSION • In other words, polycation adsorption on negatively-charged liquid vesicles led to a sharp charge asymmetry in the vesicular membrane,

7. RESULTS AND DISCUSSION • However, the liquid vesicles retained their integrity when adsorbed CP(2) unless Vexceeded a certain critical value, • At higherV values, polycation adsorption resulted in irreversible vesicle disruption indicated by a sharp conductivity increase in the systems containing the vesicles filled with NaCl,

8. Adsorption of polycations on the surface of the labeled EL-CL2- vesicles resulted in quenching of FITC fluorosence. It was found that CP(2) could be completely removed from the surface of liquid EL-CL2- vesicles either by recomplexation with an excess of PAA or by addition of an excess of a simple salt.

9. CONCLUSIONS • It was found that the negative 3:1 PAA-CP(2,16) complex bound to negative vesicles as a whole, • Therefore, complexation with hydrophobized polycation can be used to enhance the affinity of negative biologically active macromolecules to biomembranes, • Interaction of polycations with vesicles depending on their composition, phase state of the lipid bilayer and chemical structure of a polycation can be accompanied by lateral lipid segregation, highly accelerated transmembrane migration of lipid molecules, incorporation of adsorbed polycations into vesicular membrane as well as aggregation and disruption of vesicles,

10. CONCLUSIONS • The electrostatically adsorbed polycation, CP(2), can be completely removed from the liquid vesicle membrane by an increase in simple salt concentration or by recomplexation with polyanions, • This results in resumption of the initial lateral and transmembrane lipid distribution in vesicle bilayer, • In contrast, complexation of polycation carrying hydrophobes, CP(2,16), with negative vesicular membranes is controlled by both electrostatic and hydrophobic interactions, • Such polycations retain contact to the vesicles even in concentrated salt solutions and in the presence of competing polyions apparently due to incorporation of hydrophobic polycation fragments into the hydrophobic part of the membrane.

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