1 research outputs found
Mechanisms of Nanoparticle Internalization and Transport Across an Intestinal Epithelial Cell Model: Effect of Size and Surface Charge
This
study investigated the effect of nanoparticle size (50 and
100 nm) and surface charge on their interaction with Caco-2 monolayers
as a model of the intestinal epithelium, including cell internalization
pathways and the level of transepithelial transport. Initially, toxicity
assays showed that cell viability and cell membrane integrity were
dependent on the surface charge and applied mass, number, and total
surface area of nanoparticles, as tested in two epithelial cell lines,
colon carcinoma Caco-2 and airway Calu-3. This also identified suitable
nanoparticle concentrations for subsequent cell uptake experiments.
Nanoparticle application at doses below half maximal effective concentration
(EC<sub>50</sub>) revealed that the transport efficiency (ratio of
transport to cell uptake) across Caco-2 cell monolayers is significantly
higher for negatively charged nanoparticles compared to their positively
charged counterparts (of similar size), despite the higher level of
internalization of positively charged systems. Cell internalization
pathways were hence probed using a panel of pharmacological inhibitors
aiming to establish whether the discrepancy in transport efficiency
is due to different uptake and transport pathways. Vesicular trans-monolayer
transport for both positively and negatively charged nanoparticles
was confirmed via inhibition of dynamin (by dynasore) and microtubule
network (via nocodazole), which significantly reduced the transport
of both nanoparticle systems. For positively charged nanoparticles
a significant decrease in internalization and transport (46% and 37%,
respectively) occurred in the presence of a clathrin pathway inhibitor
(chlorpromazine), macropinocytosis inhibition (42%; achieved by 5-(<i>N</i>-ethyl-<i>N</i>-isopropyi)-amiloride), and under
cholesterol depletion (38%; via methyl-β-cyclodextrin), but
remained unaffected by the inhibition of lipid raft associated uptake
(caveolae) by genistein. On the contrary, the most prominent reduction
in internalization and transport of negatively charged nanoparticles
(51% and 48%, respectively) followed the inhibition of lipid raft-associated
pathway (caveolae inhibition by genistein) but was not significantly
affected by the inhibition of clathrin pathway