Fig. 3.
In vitro targeted EC cell imaging and enhanced internalization. a Flow cytometry of HET-1A, OE33, and KYSE-30 cells indicates that the αvβ3 integrins are overexpressed in EC cells. The fluorescence signals from OE33 and KYSE-30 cells shifted clearly to right compared to that from HET-1A cells, indicating that EAC and ESCC cell lines contain higher expression of plasma membrane integrin αvβ3 than HET-1A cells. b Confocal fluorescence images of the KYSE-30 cells after incubation with RGD-f-PNPs/EPI conjugates for 30 min and 3 h. The blue and red color represents the fluorescence signal of the RGD-f-PNPs and EPI, respectively. After 30 min of treatment, a significant blue fluorescence was observed from the KYSE-30 plasma membranes, which can be attributed to the binding between the RGD peptide moieties and overexpressed integrins at the plasma membrane. The data suggest that RGD peptide moieties could enhance the specific binding between RGD-f-PNPs and plasma membrane integrins. After 3 h of the treatment, clear red fluorescence was observed at the cell nucleus, indicating that RGD-f-PNPs/EPI has internalized into the cell nucleus. Scale bar: 10 μm. c Confocal fluorescence images of KYSE-30 and HET1-A cells incubated with RGD-f-PNPs after 0 and 20 min. A significant blue fluorescence of the RGD-f-PNPs was only observed from the KYSE-30 cell membrane after 20 min of treatment, which can be attributed to the specific binding between RGD moiety and integrin on the cell membrane. The data suggest that RGD-f-PNPs can selectively target EC cells vs. normal epithelial cells. Scale bar: 20 μm