Quantitative bioimage analytics enables measurement of targeted cellular stress response induced by celastrol-loaded nanoparticles.

The cellular stress response, which provides protection against proteotoxic stresses, is characterized by the activation of heat shock factor 1 and the formation of nuclear stress bodies (nSBs). In this study, we developed a computerized method to quantify the formation and size distribution of nSBs, as stress response induction is of interest in cancer research, neurodegenerative diseases, and in other pathophysiological processes. We employed an advanced bioimaging and analytics workflow to enable quantitative detailed subcellular analysis of cell populations even down to single-cell level. This type of detailed analysis requires automated single cell analysis to allow for detection of both size and distribution of nSBs. For specific induction of nSB we used mesoporous silica nanoparticles (MSNs) loaded with celastrol, a plant-derived triterpene with the ability to activate the stress response. To enable specific targeting, we employed folic acid functionalized nanoparticles, which yields targeting to folate receptor expressing cancer cells. In this way, we could assess the ability to quantitatively detect directed and spatio-temporal nSB induction using 2D and 3D confocal imaging. Our results demonstrate successful implementation of an imaging and analytics workflow based on a freely available, general-purpose software platform, BioImageXD, also compatible with other imaging modalities due to full 3D/4D and high-throughput batch processing support. The developed quantitative imaging analytics workflow opens possibilities for detailed stress response examination in cell populations, with significant potential in the analysis of targeted drug delivery systems related to cell stress and other cytoprotective cellular processes.