Figure 2.

Activatable cell penetrating peptides (ACPPs) can report gelatinase activation by cellular uptake of fluorescence. (A) Structure of ACPPs: a cell-penetrating polycationic part (green) is attached to a polyanionic sequence (orange) via a cleavable linker (blue). In the presence of active gelatinases (MMP-2/-9), the ACPP is cleaved. The cell-penetrating polycationic part coupled to a TAMRA fluorophore is released and can then enter into surrounding cells. (B) Cellular uptake of TAMRA fluorophore induced by NMDA or glutamate transient perfusion. Rat hippocampal neurons (DIV14) were stimulated for 10 min with NMDA (100 μM) or glutamate (50 μM), with or without the broad-spectrum Matrix Metalloproteinase (MMP) inhibitor Ca-DTPA (5 mM), then incubated for 2 h 30 in presence of 1 μM gelatinase ACPPs or an ACCP containing an uncleavable scrambled sequence as a linker (scrambled-ACPP). Hoechst labeled nuclei (blue). Scale bar 20 μm. (C) Quantification of the number of TAMRA fluorescent cells in gelatinase-ACPP treated cultures in control, NMDA or glutamate stimulated conditions. The graph represents mean values ± SEM (n > 30 fields for each condition, in three independent experiments). One-way ANOVA test (p-value = 0.0264) was performed followed by Tukey’s multiple comparison test where *p < 0.05 (control vs. stimulations).