Increased lung cell susceptibility to injury due to autophagic overload after exposure to nanoparticles

Autophagy is an intracellular process that helps maintain cellular homeostasis. In this study, we measured autophagic flux and cellular injury in alveolar epithelial cells (AECs) and lung adenocarcinoma (A549) cells exposed to polystyrene nanoparticles (PNPs: 20 nm, carboxylated, near-infrared dye-labeled) and a secondary insult to explore the effects of limited autophagic capacity. Rat AEC monolayers (RAECMs) and A549 cells were exposed for 24 h to 1) PNP, 2) rotenone (RT: inhibits mitochondrial respiratory chain complex I) or tunicamycin (TN: disrupts protein synthesis and induces the unfolded protein response), or 3) RT or TN for an additional 24 h after 12 h of PNP preincubation. Release of lactate dehydrogenase served as an index of cellular damage. Autophagic flux was assessed using confocal imaging to quantify the rate of autophagosome production. PNP taken up into AEC and A549 cells induced autophagy, eliciting increased autophagic flux and reaching a steady state at ∼10-24 h. When RAECM or A549 cells were exposed to RT or TN alone for up to 24 h, autophagy was also activated, reaching steady-state autophagic flux similar to PNP exposure alone. In the presence of both PNP and RT (or TN), steady-state autophagic flux was similar to that with PNP, RT, or TN alone. Cellular damage due to RT or TN, in both AEC and A549 cells, became more severe after PNP preexposure, indicating that induction of autophagy by PNP makes AEC and A549 cells more susceptible to injury by each of two secondary insults due to finite autophagic capacity.NEW & NOTEWORTHY Autophagy plays a key role in maintaining cellular homeostasis and protecting cells from external stressors. However, our findings show that autophagic flux has a finite capacity, indicating that when autophagic flux is induced by one agent, cells become susceptible to greater injury when exposed to a second injurious agent because autophagic capacity cannot be increased beyond its finite capacity. In this study, we show that in the presence of autophagic flux-inducing nanoparticles, susceptibility to injury by exposure to a second agent (e.g., tunicamycin or rotenone) is increased. These findings provide insight into one mechanism by which exposure to ambient airborne nanoparticles can lead to lung disease.