96 Cell culture composition influences cellular responses to particulate matter-associated metals

Abstract Background In vitro cell culture models provide a reproducible, cost-effective assessment of the effects of exposure to particulate matter (PM) and its components. While much consideration is paid to the cells, how composition and complexity of culture media may influence PM studies is poorly understood. Objective To determine the effects of culture media composition on immortalised alveolar epithelial cell (AEC) responses to vanadium pentoxide (V2O5), a tracer of heavy fuel oil combustion associated with respiratory and cardiovascular effects. Methods AECs were exposed to 0 to 100 µM V2O5 for 24 h in five different media (DCCM-1, DMEM:F12 (1:1), Hybridoma (Sartorius), Hybridoma (ThermoFisher) or Plasmax (CancerTools.org)). Expression of metal-, oxidative stress-, and viral response-genes was measured by RT-qPCR, and inflammatory mediator release by ELISA. Composition of media and post-exposure AECs was measured by inductively coupled plasma-mass spectrometry. Results In DCCM-1, V2O5 elicited upregulation of metallothionein and antioxidant gene expression (MT1G: 120-fold, HMOX1: 13-fold, both P < 0.001), which was muted in other media. Media-dependent differences in IL-6 release were also observed. Conversely, V2O5-mediated inhibition of antiviral genes (IFIT1, ISG15: 0.07 to 0.10-fold, P < 0.05) was robust across media. V2O5 elicited a concentration-dependent increase in intracellular vanadium across all media, but in DCCM-1, which contained higher copper concentrations, this was accompanied by increased intracellular copper. Supplementation of DMEM:F12 with copper replicated the effects of V2O5 in DCCM-1. Conclusions V2O5 induces metallothionein and antioxidant responses in a copper-dependent manner. The mechanism of copper entry remains under investigation. These findings highlight the need for careful consideration and reporting of cell culture conditions.