Fig. 4

SH-SY5Y cells were exposed to SiPCL-NP at a concentration of (2.6 × 10¹⁰ NPs/ml) for 24 h and cells displaying a cholinergic phenotype were analyzed. NP-exposure was performed in undifferentiated cells (Undiff) or cells differentiated with retinoic acid (RA) or staurosporine (ST) at day in vitro (DIV) 1 (A), DIV4 (B) or DIV6 (C). At the end of the differentiation period (DIV7), high-affinity choline transporter 1-immunoreactive (CHT1-ir) cells were quantitatively assessed. Data represent the mean percentage (%) of CHT1-ir cells of the total number of cells. Error bars correspond to SEM. Significant differences between control- and SiPCL-NP-exposed-groups are labeled with asterisks (*p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001). Representative immunofluorescence microscopy images of CHT1-ir cells (green) are depicted in D–F (D = Undiff NP DIV4, E = RA NP DIV4, F = ST NP DIV4). NPs are shown in red and cell nuclei in blue. Magnification ×630, scale bar = 10 μm