Abstract
Cell-penetrating peptides (CPPs) are known to interact with cell membranes and by doing so enhance cellular interaction and subsequent cellular internalization of nanoparticles. Yet, the early events of membrane interactions are still not elucidated, which is the aim of the present work. Surface conjugation of polymeric nanoparticles with cationic CPPs of different architecture (short, long linear, and branched) influences the surface properties, especially the charge of the nanoparticles, and therefore provides the possibility of increased electrostatic interactions between nanoparticles with the cell membrane. In this study, the physicochemical properties of CPP-tagged poly(lactic-co-glycolic acid) (PLGA) nanoparticles were characterized, and nanoparticle-cell interactions were investigated in HeLa cells. With the commonly applied methods of flow cytometry as well as confocal laser scanning microscopy, low and similar levels of nanoparticle association were detected for the PLGA and CPP-tagged PLGA nanoparticles with the cell membrane. However, single particle tracking of CPP-tagged PLGA nanoparticles allowed direct observation of the interactions of individual nanoparticles with cells and consequently elucidated the impact that the CPP architecture on the nanoparticle surface can have. Interestingly, the results revealed that nanoparticles with the branched CPP architecture on the surface displayed decreased diffusion modes likely due to increased interactions with the cell membrane when compared to the other nanoparticles investigated. It is anticipated that single particle approaches like the one used here can be widely employed to reveal currently unresolved characteristics of nanoparticle-cell interaction and aid in the design of improved surface-modified nanoparticles for efficient delivery of therapeutics.