Research Article

Mitogenic Effects of Phosphatidylcholine Nanoparticles on MCF-7 Breast Cancer Cells

Figure 5

EGFR protein content and immunocytochemistry. MCF-7 breast cancer cells were incubated for 24 hours with PC nanoparticles dispersed at 0.1 and 0.01% (w/v) in buffer pH 5.0 and buffer pH 7.0 or vehicle (Ct) in absence (a) or presence (b) of serum for immunoblot studies, but only with PC dispersions at 0.1%, pH 7.0, or vehicle (control) in absence of serum for immunocytochemistry (c). Western blotting was performed as described in M & M. Membranes were reprobed to asses actin content and demonstrate equal protein loading in all lanes. Representative immunoblots are shown ((a) and (b)). EGFR quantification was performed by scanning densitometry and expressed as percent of values measured for control, nonstimulated breast cancer cells. Data are expressed as the mean ± S.E.M. of the indicated number ( ) of independent experiments. Statistical analysis was performed by ANOVA. Different letters denote significant difference at , whereas results with the same letter are not statistically different from each other. For EGFR immunocytochemistry (c), cells were washed, fixed, permeabilized, blocked, and incubated with the anti-EGFR antibody, the Cy3-conjugated secondary antibody, and Hoechst. Finally covers were mounted and examined by epifluorescence microscopy. Representative merged images of control and PC-nanoparticles-treated cells are shown (c).