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. 2023 Sep 21;15(9):2364. doi: 10.3390/pharmaceutics15092364

Figure 4.

Figure 4

D-DPTIP is preferentially taken up by microglia and selectively inhibits microglial nSMase2 activity. (A) Representative hippocampal images showing Cy5–D–DPTIP signal (red) was co-localized with Iba1 (microglia) signal (green), but not with GFAP (astrocyte) or NeuN (neuron) signal (Arrows). (B) Single cells were selected by gating singlets (upper panel) and live cells were selected by gating PI negative signals (lower panel). (C–E) Cy5 positive cells were gated from brain samples of PS19 mice treated with Cy5–D–DPTIP (C), PS19 mice treated with vehicle (D), and WT mice treated with vehicle (E). (F) Cy5 positive cells gated from (C) were used for gating by ACSA2 (astrocyte) and TMEM119 (microglia) and (NeuN) positive signals. Fluorescence Minus One (FMO) are used for negative controls. (G) Quantification of F. Statistics were performed using one-way ANOVA. (H,I) Quantification of nSMase2 activity assay performed on isolated CD11b- cells (H) and CD11b+ cells (I). Statistics were performed using unpaired Student’s t-tests in (H,I) and one-way ANOVA in (G) with Tukey’s multiple comparisons. Bars represent mean ± SEM. * p < 0.05, ** p < 0.01. Scale bar is 30 µm.