Nanoparticle agglomeration restricts uptake into living cells
Ultrafine particles form clumps in biological fluids; this curtails their diffusion and limits their uptake.
Size matters when it comes to nanoparticles entering lung cells. New research posted this week to ES&T’s Research ASAP website (es051043o) shows that the finer the particles, the faster they agglomerate in biological fluids, such as the lung’s airway surface liquid. This aggregation alters the motility of the nanoparticles, with the smaller ones—which are considered the most toxic—diffusing the most slowly. As a consequence, cells accumulate less particles. These results suggest that the smallest particles might be less dangerous than previously thought, because their uptake can be reduced for purely physical reasons.
“We were really surprised to find that the uptake rate at low, physiologically relevant concentrations was limited by physical transport properties in the solution rather than by the cells’ ability to engulf cerium oxide nanoparticles,” says Wendelin Stark, head of the functional materials laboratory at the Swiss Federal Institute of Technology Zurich and the paper’s corresponding author.
Researchers in Stark’s group exposed human lung fibroblasts—cells that form connecting tissue and are involved in the formation of asbestosis—to suspensions of different-sized cerium oxide nanoparticles, ranging in diameter from 20 to 150 nanometers (nm). They observed that the smallest particles immediately massed together. The scientists believe that this agglomeration takes place because adsorption of the proteins present in the cell culture medium decreases the particles’ surface charge, and this offsets the electrostatic repulsion that usually keeps them apart. Interestingly, protein adsorption led to very similar changes in surface charge in a whole series of industrially important oxide nanoparticles. This suggests that agglomeration is the common fate of many nanoparticles in biological fluids, unless the particles are surface-modified.
Stark says he chose to concentrate on cerium oxide powder, which is also known as ceria, because these nanoparticles are increasingly important in automotive catalystic converters, which are used to control pollution, as well as for industrial applications such as the polishing of computer chips. Moreover, ceria particles are rather persistent in living cells.
In addition, “cells [normally] do not contain cerium; that’s why whole-cell elemental analysis makes a lot of sense,” says Silvia Diabaté, a toxicologist with the Institute of Toxicology and Genetics of the Forschungszentrum Karlsruhe (Germany). In fact, this method’s high sensitivity offers for the first time a way to explore what happens when living cells are exposed to nanoparticles at concentrations as low as 0.1 parts per million.
When they examined the cells, Stark’s group found only agglomerates of ceria nanoparticles, which were all contained in cellular containers known as vesicles. Peter Gehr, an expert on pulmonary particle uptake at the University of Bern (Switzerland), says that in his uptake studies he has never found such vesicles enclosing the silica or polystyrene model nanoparticles that he uses. Instead, he has found particles smaller than 200 nm in diameter to be highly mobile and to cross biological membranes.
Stark says that his group’s findings are important for in vitro studies but that significant steps are still needed before the data can be applied to in vivo systems. Gehr agrees, explaining that isolated cells bathed in an artificial cell-culture medium, as in the ES&T study, cannot be directly compared with the situation in the lung, where various cell types interact to form closed cellular assemblies confining the lung’s enormous surface area. “But even then, with a tightly closed cellular layer beneath the airway surface liquid, particulate matter enters the organism in several distinct ways,” Gehr adds.
Anyway, “measuring the amount of ceria taken up still doesn’t really tell you much about the toxicity,” Diabaté says, adding that studies examining the effects of titanium dioxide nanoparticles in rats have established that smaller particles are more toxic, even if their uptake rate varies in different cell types. “We didn’t see any violent responses; the exposed cells didn’t die. But we are [continuing] on this subject,” says Stark. “We definitely have a lot of work in front of us.” —ORI SCHIPPER
