Analyzing the Biological Entity of Nanomaterials - Safety of Nanomaterials along Their Lifecycle - page 60

a mechanistic understanding of nanomaterial behavior in interaction with living
systems. The hypothesis underlying the nanoGEM work package APQ, dealing with
the in situ characterization of nanomaterials, is that only a full characterization of
nanoparticles in the biological system would enable us to derive structure-activity
relationships. Before we go into details of how interactions with nanoparticles and
the biological environment can be studied, and how this challenging task was dealt
with in the project nanoGEM, we would like to spend a few more words on how such
interactions will influence the interactions with cells.
As pointed out by the group of Kenneth Dawson, the entity that cells will finally
interact with will not be the pristine nanomaterial but a nanomaterial covered by sev-
eral layers of biomolecules (Lynch et al. 2007; Lynch, Salvati, and Dawson 2009;
Walczyk et al. 2010). Obviously, the cells do not “see” the original nanomaterial sur-
face; the first contact actually occurs via the biological molecules on the nanopar-
ticle surface, and the nanoparticle-protein complex thus represents a biological entity
(Lynch et al. 2007). This so-called protein corona enables completely different types of
interactions compared to a pristine nanomaterial surface, for example, protein-protein
interactions with different cell surface receptors, which will influence the nanopar-
ticle uptake. Meanwhile there is clear evidence from several groups that nanomaterials
uncoated with any biomolecules (for example, experiments performed in vitro without
any serum addition to the cell culture medium) compared to nanomaterials coated
with biomolecules (i.e., experiments performed in vitro with serum-containing cell
culture medium) have a different uptake rate into cells (Petri-Fink et al. 2008; Bajaj
et al. 2009; Lesniak et al. 2012). For instance, Petri-Fink and coworkers showed that
for iron oxide nanoparticles (A-PVA-SPION) the presence of serum strongly inhibited
cellular uptake due to formation of a protein corona (Petri-Fink et al. 2008). In con-
trast, Lesniak and coworkers showed that silica nanoparticles in the absence of serum
adhered stronger to cell membranes and were internalized with a higher efficiency
(Lesniak et al. 2012). But apart from affecting uptake levels the protein corona also
influenced the intracellular nanoparticle location and the impact on cells.
Interactions of proteins with nanoparticle surfaces may lead to partial unfolding
of the proteins, displaying epitopes that have previously been hidden inside the pro-
teins on the protein surface. These so-called “cryptic epitopes” may be recognized
by specific receptors, and may also lead to adverse effects (Lynch, Dawson, and
Linse 2006), for example, by activation of the immune system. In addition, some
limited data support the hypothesis that some nanoparticles may behave as haptens
and turn antigenic when bound to a protein. Other data suggest that some nanopar-
ticles have very efficient adjuvant properties most probably due to the fact that they
create a local reservoir at the injection site leading to the protection and slow release
of antigens (Zolnik et al. 2010).
Finally, the interaction with biomolecules may also influence agglomeration of
nanoparticles and thus affecting sedimentation under in vitro or biodistribution
under in vivo test conditions. If nanomaterials are ranked according to their toxic
effects in vitro or in vivo , one needs to consider the in situ characterization data
taking into account the dose that will finally reach a cell or a specific tissue, which
is also dependent on agglomeration state, and not rank only according to the dose
applied to the cell culture dish or to the animal.