Silica nanoparticle uptake induces survival mechanism in A549 cells by the activation of autophagy but not apoptosis

Abstract

We report here in vitro evaluation of cellular nanoparticle uptake, cytotoxic effect and autophagy as possible survival strategy in lung epithelial cells (A549). The effects of surface charge and presence of serum proteins, as well as the influence of various inhibitors on the uptake of 20 nm monodispersed silica nanoparticles are discussed. Uptake rate of the particles with various functional groups is demonstrated to be similar in the presence of serum proteins, while the uptake rate ranking results in COOH > NH2 > OH under serum free conditions. The uptake process is shown to be actin dependent, and can be partially inhibited by dynasore in the presence of serum proteins. These results suggest a macropinocytotic uptake process that was also confirmed by scanning electron microscopy. In spite of the intensive active uptake, significant cytotoxic effect is detected only at relatively high (above 250 μg/mL) concentrations, while blebbing of the cell surface is observed at already 5 hours of exposure time. By monodansylacadaverin staining and RT-PCR experiments, we show that this blebbing is related to autophagy rather than apoptotic cell death. After the exposure to silica nanoparticles, stress-related and cell-death-related genes are found to be upregulated, but no activation of the apoptotic process was observed. The A549 cells display elevated levels of autophagosomes, however cells do not show typical apoptosis markers such as increased amount of active caspase-3 and release of mitochondrial cytochrome C. Based on these results, we propose here autophagic activity and cross-talk between autophagic and apoptotic pathways as a mechanism that allows the survival of A549 cells under exposure to silica nanoparticles.