20.5 Cell binding and uptake

Polyplexes enter the cell and interacts with anionic proteoglycans by a transmembrane protein called syndecans. The presence of cationic surface charge is essential for binding and cellular uptake. The type of cell and polymer used, determines the internalization of the complex and thus its transfection efficiency. Linear PEI-polyplexes uses a clathrin-coated pit pathway for transfection in African green monkey kidney (COS-7) and hepatocellular carcinoma cells (HUH-7). Both lipid raft and clathrin dependent pathway are used by branched polyplexes such as in HeLa cells. But the lipid based pathway is much more efficient in transfection.

20.6 Escape from endosome

PEI based polyplexes and PMAM have high transfection efficiency as they behave as proton sponge. Proton sponge hypothesis states that only few nitrogen atoms are protonated at normal physiological pH. When the pH gets lowered such as in endosomes, more number of nitrogen atoms get protonated and a gradient is created that causes influx of chloride ion. The increase chloride ion influx causes water influx and thus endosome swells and gets ruptured. Proton sponge activity is absent in polymers with buffer capacity at pH 5. Polymers based on PEI and PMAM buffers the interior of endosomes leading to decreased acidification, high chloride content and increased volume. Protonable amine groups if removed decreases transfection efficiency.

20.7 Dissociation of polyplexes

After the endosomal escape polyplex must reach nucleus and also DNA need to dissociate from carrier. Thus polyplexes having low molecular weight and tending to dissociate rapidly have better transfection ability as compared to high molecular weight polyplexes. PEI based polyplexes shows slow dissociation from DNA when analyzed using fluorescence microscopy. Reducible PLL polymers have been made that can be degraded by intracellular environment causing fast releases of DNA and its enhanced expression.