Timing and Mechanisms of Nanodiamond Uptake in Colon Cancer Cells

Introduction: As nanodiamonds become more and more widely used for intracellular labelling and measurements, the task of delivering these nanoparticles inside cells becomes more and more important. Certain cell types easily take up nanodiamonds, while others require special procedures. Methods: In previous research, we found that HT-29 cells (a colon cancer cell line), which are notoriously difficult in the context of nanodiamond internalization, show increased uptake rates, when pre-treated with trypsin-ethylenediaminetetraacetic acid (trypsin-EDTA). However, the uptake mechanism has not been studied before. This article focuses on a more detailed investigation of the reasons underlying this phenomenon. We start by identifying the timing of fluorescent nanodiamond (FND) uptake in trypsin-EDTA pre-treated cells. We then use a combination of chemical inhibitors and Immunocytochemistry to identify the main pathways employed by HT-29 cells in the internalization process. Results and Discussion: We investigate how these pathways are affected by the trypsin-EDTA pre-treatment and conclude by offering possible explanations for this phenomenon. We found that nanodiamonds are internalized via different pathways. Clathrin-mediated endocytosis proves to be the dominating mechanism. Trypsin-EDTA treatment increases particle uptake and affects the uptake mechanism.