Intracellular Fate Investigation of Bio-Eliminable Polymeric Nanoparticles by Confocal Laser Scanning Microscopy

Abstract

This is a study of the in vitro cytotoxicity and intracellular fate of poly[(glycylglycinemethacrylamide)-co-N-(2-hydroxypropylmethacrylamide _y)] bio-eliminable polymer samples and relative nanoparticles in Balb/c 3T3 cloned A31 mouse embryo fibroblasts cell line by using Confocal Laser Scanning Microscopy (CLSM). Nanoparticles were prepared by co-precipitating the polymers with fluorescein labeled human serum albumin (HSA-FITC) as the fluorescent probe and as the model protein drug. The toxicity of the polymers containing 25, 50 and 100%, respectively, of (glycylglycinemethacrylamide) _x (GGMA) was investigated in terms of cytoskeleton morphology by exposing cells to various concentrations of polymers for 24 h. Under normal culture conditions, fibroblast cells exhibit characteristic spreading and shape, however, when the cell cultures were subjected to chemical, metabolic or physical stress, their morphology changed reducing their visibility. The polymers with the lower glycine content exert a lower toxicity even at high concentrations [8.5mg/mL]. The cellular uptake of nanoparticles was determined by incubating fibroblasts with HSA-FITC loaded particles and HSA-FITC alone at three different time points. The results indicate that nanoparticles were up-taken by the cells in a time dependent fashion. Preliminary evaluation of the intracellular fate of the prepared nanoparticles indicate their possible lysosomal escape.