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Chapter 6
6.2.2 Uptake experiments
The ability of the different compounds to be internalized in the cell nuclei was then assessed
in vivo: RH30 cells (a rabdomiosarcoma line) were incubated with 10 µmol/L of peptide 1,
standard PNA 2 or modified PNAs 3 and 4 and analyzed after 3 hours. Fluorescence
microscopy images showed that after 3 hours in the RH30 cell line, high intracellular levels
with main nuclear localization could be observed for the NLS peptide 1 (Figure 6-5), as
expected. The unmodified PNA 2, on the other hand, does not show any nuclear
internalization, and a very low cellular uptake, confirming the lack of recognition from cargo
proteins, as reported in previous data 24 . Incubation of cells with PNA 3, instead, showed a
very intense red fluorescence signal in the nucleus, with a behavior comparable to that
displayed by the NLS peptide.
Figure 6-5: Fluorescence microscopy analysis (left) of uptake of RH30 cells treated for 3 hours with
NLS peptide 1, unmodified PNA 2, modified PNA 3, and modified PNA 4 without proline. Optical
images are also reported (right)
The uptake in the nuclear region and the homogeneous diffusion is consistent with a receptormediated
internalization process, showing no difference in behavior between the NLS peptide
1, for which the importin-mediated nuclear transport mechanism has been demonstrated, and
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