Endocytosis of PEGylated polymeric mesoscale nanoparticles is dynamin- and macropinocytosis-dependent.

Nanotechnology is rapidly transforming medicine by enabling versatile platforms for targeted delivery, controlled release, and intracellular transport of therapeutic payloads. Polymeric mesoscale nanoparticles (MNPs) are 300 to 500 nm in diameter with a PEGylated surface that exhibit unique renal tropism, specifically toward renal tubular epithelial cells. Despite their well-described therapeutic applications and route of localization to the tubules, we do not yet understand their physicochemical stability and cellular internalization mechanisms. In this study, we investigated the stability of MNPs under stress conditions by subjecting them to repeated freeze-thaw cycles and varying storage conditions to evaluate the effects on particle size and polydispersity index. MNPs demonstrated negligible changes in size and PDI up to 4 freeze-thaw cycles. We found that both empty and dye-loaded MNPs demonstrated negligible change in size under standard -20°C storage conditions. While empty MNPs were only stable at room temperature for one day, and not at 37°C, dye-loaded nanoparticles were stable for at least eight days under both storage conditions. We then performed