Purpose: Dendrimers are an emerging class of nanoscale drug delivery vehicles, with a potential to delivery therapeutics in a targeted manner. They present a large density of functional groups at the surface that play a key role in the cellular uptake and eventual drug release. The cellular uptake, and the eventual The aim of this study is to understand the influence of surface charge on cellular interactions and intracellular trafficking of polyamidoamine (PAMAM) dendrimers in lung epithelial cells.

Methods PAMAM dendrimers with cationic (G4 amine terminated), anionic (G3.5 carboxyl terminated) and neutral (G4 hydroxyl terminated) surface charge were labeled with fluoroisothiocyanate (FITC). Cell uptake and subsequent intracellular fate was determined by flow cytometry, fluorescence and confocal microscopy using A549 lung epithelial cells. Temperature block and metabolic inhibition (sodium azide and 2-deoxyglucose) were used to investigate if the cellular uptake was an energy dependent mechanism. Various endocytotic blockers including chlorpromazine, sucrose, filipin, nocodazole, ammonium chloride and cytochalasin were used to identify the pathways involved in the transport of different dendrimers.

Results Cellular uptake of all dendrimers increased as a function of incubation time, but the uptake of cationic and neutral dendrimers was much higher than anionic dendrimers. Incubation at 100C and metabolic inhibition significantly reduced the cellular uptake of dendrimers. All dendrimers were found to be localized in lysosomes within 30 minutes. Sucrose reduced the cellular uptake of all the dendrimers by more than 50%. Chlorpromazine, filipin, nocodazole and ammonium chloride did not show any inhibitory effect on endocytotic uptake of cationic and neutral dendrimers, but filipin, noocodazole, and ammonium chloride significantly reduced the cellular uptake of anionic dendrimer.

Conclusions The surface charge and functionality play a significant role on the cellular interactions and subsequent endocytotic uptake into the cell. Anionic dendrimers appear to be mainly taken up by caveolae mediated endocytosis in A549 lung epithelial cells. While cationic and neutral dendrimers are taken in by a non-clathrin and non-caveolae endocytosis mechansim and may be they are transported by electrostatic or non specific fluid phase endocytosis.These findings open up new possibilities of targeting therapeutic agents to specific cell organelles based on surface charge.

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