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. 2021 Aug 23;6(16):e150099. doi: 10.1172/jci.insight.150099

Figure 4. Pharmacological PPARα inhibition with GW6471 prevents APP knockdown–induced increases in the expression of PPAR target genes and synaptic activity in cortical cultures.

Figure 4

Primary cultures of rat cortical cells expressing a shRNA targeting endogenous APP (shAPP) or a scrambled shRNA encoding GFP (shScra-GFP). At 13–14 DIV, cells were treated (+) or not (-) with PPARα antagonist GW6471 for 24 hours. (A) Representative immunoblot of cell lysates, 4 independent experiments (the lanes were run on the same gel but were noncontiguous). (B) APP expression/α-tubulin ratios (n = 7 of each analyzed in 4 independent experiments), mean ± SEM; Student’s t test (APP, P < 0.001). (C and D) Real time PCR analyses for Ppara, Acox1, Cpt1a, and Pdk4 mRNA levels (n = 6–8 for each condition analyzed in 6 independent experiments). Results were normalized to Rpl32 mRNA and compared with respective untreated (-) shScra-GFP control cells. Results are shown as mean ± SEM; Brown-Forsythe and Welch 1-way ANOVA tests followed by Dunnett’s T3 multiple comparisons test: (-) shAPP vs. (-) shScra-GFP: Ppara mRNA, P = 0.006, Acox1 mRNA, P = 0.008, Cpt1a mRNA, P = 0.043; Pdk4 mRNA, P = 0.0009; (+) shScra-GFP vs. (-) shScra-GFP: Cpt1a mRNA, P = 0.026; (+) shAPP vs. (-) shAPP: Cpt1a mRNA, P = 0.010. (E) RMP (n = 8–10 cells per group analyzed in 3 independent experiments). (F) Representative traces of total synaptic activity and (G) mean values of synaptic events’ frequency (n = 12–17 cells per group analyzed in 6 independent experiments). (H) Cumulative probability plot of the amplitude distribution (n = 15–27 cells per group in 7 independent experiments). (E–H) Brown-Forsythe and Welch 1-way ANOVA tests followed by Dunnett’s T3 multiple comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001, #P < 0.05.