BIOMEDICAL APPLICATIONS IV: CARBON NANOTUBE-NUCLEIC ACID COMPLEXES FOR BIOSENSORS, GENE DELIVERY AND SELECTIVE CANCER THERAPY - Carbon Nanotubes: From Bench Chemistry to Promising Biomedical Applications - page 120

CNTs consisted of pristine, un-functionalised SWCNTs that they were non-
covalently conjugated with DNA. The authors suggested an endocytotic
pathway for cell internalisation on the basis of absent uptake at 4 ° C or in
presence of NaN 3 (two known conditions that inhibit endocytosis). Moreover,
after pre-treating the cells with either sucrose (hypertonic treatment) or a
K + -depleted medium prior to exposure to SWCNT conjugates, cell cytometry
showed a remarkable reduction of internalised cells, thus indicating the
particular clathrin pathway for an endocytotic cellular uptake of SWCNTs.
It is important to note that although these results were conirmed by
additional assays and controls, they were speciic for the samples used in
the experiments: since only pristine and oxidised SWCNTs were employed
in this investigation, the results might not correspond to the cellular uptake
of MWCNTs or other functionalised nanotubes, especially in view of the
contradictory results obtained by Pantarotto et al. 61 In the latter case, the
researchers used CNTs bearing a TEG chain and a peptide, and their indings
did not conirm those obtained by the group of Dai, since endocytosis was
not involved. Therefore, further investigations are needed to disclose the real
internalisation mechanism of CNTs, which do differ remarkably on the basis
of several unique characteristics and preparative protocols. To support this
statement, Becker et al. 62 demonstrated that there is a length dependence in
the uptake of DNA-wrapped SWCNTs, so the eventual internalisation process
is inluenced by this aspect more than other factors, including nanotubes'
functionalisation or chirality. In fact the results showed a selective rejection of
certain SWCNTs by the incubated cells on the basis of their length. Interestingly,
the longer fractions of (335 ± 27) nm and (253 ± 26) nm, respectively, collected
through SEC, were internalised without affecting the viability of the cells.
Conversely, shorter SWCNT fractions (below 253 nm) exhibited decreased
metabolic activity and cell survival, suggesting that shorter tubes could be
more toxic to cells than longer SWCNTs. This was established for several
cell lines, including A549 (human alveolar basal epithelial cells), MC3T3-E1
(clonal murine calvarial) and A10 (embryonic rat thoracic aorta medial layer
myoblasts) cells. However, despite this apparently general phenomenon of
length-dependent uptake in these in vitro experiments, the exact threshold
process is not understood yet and it could vary with cell type and nanotube
samples.
5.2.2 Carbon nanotubes and RNA
The use of ribonucleic acid (RNA) has shown several advantages for potential
therapeutic applications: contrary to DNA, RNA cannot integrate directly
into the host chromosome, and hence it is less prone to become mutagenic. 63