In-Vitro modelling of the lungs response to nanoparticulates
Altin Kocinaj1, Laura Urbano1, Darragh Murnane1
1University of Hertfordshire, College Lane, Campus, Hatfield AL10 9AB
Nanoparticles are abundant in everyday life, with air pollution and consumer products leading to a growing number of concerns for potential health effects of inhaled exposure. There is no standalone alternative for animal toxicity studies, thus we sought to build a predictive cell culture model using high content imaging/analysis (HCA) and traditional viability assays. Human lung epithelial cells (NCI-H441) were seeded for in a 96-well plate and exposed to a panel of nanoparticles for 24 hours, for high content imaging. For the HCA, plated cells were given fluorescent labels which allowed both morphological assessment (such as cell area and diameter, membrane permeability and mitochondrial activity) and cell segmentation for the consequent analysis. Image acquisition was performed using an automated fluorescence imager (EVOS M7000) and was segmented and analysed using the accompanying Celleste program. Morphological differences can be seen and analysed using HCA, with the possible identification of subgroups of cells. There are differences in such parameters as cell area with ~100% of cells exposed to 33ug/ml of copper oxide being below the median cell area of the control or lowest concentration used. Also, a flattening and spread of the 33ug/ml distribution indicating a larger population of cells compared to control with greater membrane permeability. These preliminary results indicate the possible use of HCA to define morphometric cell-by-cell changes and population subgroups that are not able to be elucidated by a single value from standard assays (such as Prestoblue – cell viability).
The use of inexpensive cell culture techniques coupled with the use of HCI/A (high content imaging/analysis) can provide the means of identifying morphometric changes within the cell which would otherwise be ignored. When combined with biochemical assays and expression studies enables a potentially more complete toxicological picture.