Mechanistic studies of in vitro cytotoxicity of poly(amidoamine) dendrimers in mammalian cells

Abstract

Poly(amidoamine) (PAMAM) dendrimer nanoparticles have been demonstrated to elicit a well defined cytotoxicological response from mammalian cell lines, the response increasing systematically with dendrimer generation and number of surface amino groups. In this work, using generation G4, G5, and G6 dendrimers, this systematic response is furthermore demonstrated for the generation of reactive oxygen species, lysosomal activity, and the onset of apoptosis and levels of DNA damage. The results are consistent with a pathway of localisation of PAMAM dendrimers in the mitochondria leading to ROS production causing oxidative stress, apoptosis and DNA damage. ROS production is co-located in the mitochondria, and both generated levels and timescales are systematically generation dependent (G4<G5<G6). Flow cytometry confirms that with increasing dose, the percentage of healthy and early apoptotic cells decreases, whereas the late apoptotic and necrotic cell populations increase. This process is again systematically generation dependent. DNA damage as measured using the TUNEL assay further demonstrates a systematic trend, G4, G5 and G6 showing 4.69%, 25.87% and 89.63% DNA breakage respectively. Increases in lysosomal activity at timescales of ~24h are observed in HaCaT but not SW480 cells upon low concentration PAMAM exposure. Overall, significant differences are observed between the responses of the dermal cell line, HaCaT, and the colon cell line, SW480, and it is suggested that these can be understood in terms of differing intrinsic antioxidant levels.