The Effect of Architecture and Molecular Weight on Cell Uptake and Intracellular Distribution of Poly (ethylene Glycol)

Abstract

The aim of this thesis was to investigate interactions of comb-type poly(polyethylene glycol) methyl ether methacrylate (p(PEG-MA)) with in vitro cultured cells and compare to linear PEG counterparts. For this purpose, p(PEG-MA) polymers were at different molecular weights (10,000 g/mol (10K) and 20,000 g/mol (20K)) were synthesized via reversible addition fragmentation chain transfer (RAFT) polymerization. Characterizations of polymers were performed via NMR, GPC and DLS. The effects of polymers on in vitro cultured cells were investigated using cancerogenic A549 and healthy BEAS-2B human lung cell lines. Cytotoxicity of polymers was investigated via MTT assay. Comb-type p(PEG-MA) and linear PEG decreased the cell viability in a dose-dependent manner. However, the lowest cell viability was above 50% indicating no significant cytotoxic effect of both types of polymers. LDH assay was performed to determine the effect of polymers on cell membrane integrity. Both polymers enhanced membrane permeability in a dose- dependent manner. The highest LDH release was less than 2% indicating no significant effect on cell membrane integrity. Cellular uptake of both types of polymers completely diminished at 4oC suggesting energy-dependent internalization mechanism of polymers. Comb-type polymers were found to be taken up more by A549 cells. Comb-type polymers were internalized by A549 cells mainly via actin-dependent pathway, while BEAS-2B cells took up the all polymers via microtubule-dependent pathway. Linear 20K PEG was internalized by A549 cells via dominantly actin and also microtubule-dependent pathways while linear 10K PEG was taken up via actin-dependent pathway. Both comb-type and linear polymers were found to be localized in endocytic vesicles and cytosol. Consequently, comb-type PEG may offer an alternative to linear PEG with better cellular uptake properties and possess different uptake mechanisms in normal and cancer cells that may provide a potential for selective targeting strategies.