Fig. 3
From: Neuronal SphK1 acetylates COX2 and contributes to pathogenesis in a model of Alzheimer’s Disease
Neuronal SphK1 induces SPMs secretion by COX2 acetylation. a Protein levels of LxA4, RvE1, and RvD1 were detected by using ELISA in CM of neurons derived from WT, APP/PS1, APP/PS1/SphK1 tg, and SphK1 tg mice (n = 6–8 per group). b mRNA levels of COX2 and LOX-15 in neurons derived from cortex of WT, APP/PS1, APP/PS1/SphK1 tg, and SphK1 tg mice (n = 4–6 per group). c Quantification of neuronal COX2 (n = 6 per group) and neuronal LOX-15 (n = 4–6 per group). d Acetylation assay of COX2 protein in neurons derived from WT, APP/PS1, APP/PS1/SphK1 tg, and SphK1 tg mice. [14C] aspirin-treated neuron was positive control. Sonicated neurons incubated in the presence of [14C] acetyl-CoA for 2 h at 37 °C and then COX2 was purified and analyzed on scintillation counter (n = 3–6 per group). e Representative chromatograms of blank, 15-R-LxA4 standard, and 15-R-LxA4 in WT samples (left panel). Molecular MS scanning from the peak at retention time 6.8 min (right upper panel) and MS/MS fragmentation pattern of 15-R-LxA4 from the peak at retention time 6.8 min (right lower panel). f Representative chromatograms (left panel) and quantification of 15-R-LxA4 in neurons derived from WT, APP/PS1, APP/PS1/SphK1 tg, and SphK1 tg mice with acetyl-coA treatment (24 h after 2.5 mM acetyl-CoA treatment) (right panel, n = 6 per group). All data analysis was done on 9-month-old mice. a−d, f One-way analysis of variance, Tukey’s post hoc test. *P < 0.05, ***P < 0.001. All error bars indicate s.e.m.
