Fig. 7
From: Astrocytic spermidine insufficiency contributes to enhanced pain sensitivity associated with ApoE4
Spermidine reduced NOS2 expression via the NF-κB pathway. (A) Western blot analysis of the NOS2 expression in the spinal dorsal horn of ApoE3-TR and ApoE4-TR mice in sham or SNI group, with β-actin serving as the loading control. (B) NO levels in the spinal dorsal horn of ApoE3-TR and ApoE4-TR mice in sham or SNI group. (C) Western blot analysis of intracellular NOS2 expressions in ApoE3 and ApoE4 astrocytes with or without CYM stimulation. (D) NO levels in ApoE3 and ApoE4 astrocytes with or without CYM stimulation. (E) Western blot analysis of NOS2 expression in the spinal dorsal horn receiving low-dose (22.35 mM) spermidine or PBS treatment in ApoE3-TR and ApoE4-TR mice following SNI. (F) The content of NO in the spinal dorsal horn of ApoE3-TR and ApoE4-TR mice receiving spermidine gavage or PBS treatment after SNI. (G) Western blot analysis of intracellular NOS2 expression in ApoE3 and ApoE4 astrocytes following CYM stimulation with or without spermidine (5 mM) treatment in mice. (H) NO levels in ApoE3 and ApoE4 astrocytes with or without spermidine treatment after CYM stimulation. (I) The NF-κB pathway was enriched by GSEA in ApoE4-TR mice after SNI injury. (J) Western blot analysis of p65 and p-p65 expression in the spinal dorsal horn of ApoE3-TR and ApoE4-TR mice in sham or SNI group. (K) Western blot analysis of intracellular p65 and p-p65 expression in ApoE3 and ApoE4 astrocytes with or without CYM stimulation. (L) Western blot analysis of p65 and p-p65 expression in the spinal dorsal horn of ApoE3-TR and ApoE4-TR mice after SNI, with or without spermidine (22.35 mM) treatment. (M) Western blot analysis of p65 and p-p65 expression in ApoE3 and ApoE4 astrocytes after CYM stimulation, with or without spermidine (5 mM) treatment. (N) Western blot analysis of NOS2, p65 and p-p65 expression in the spinal dorsal horn of ApoE3-TR and ApoE4-TR mice after SNI and spermidine gavage treatment, with or without intrathecal PMA injection. (O) Western blot analysis of NOS2, p65 and p-p65 expression in ApoE3 and ApoE4 astrocytes after the treatment with CYM and spermidine, with or without PMA addition. n = 3 independent biological replicates per group for all Western blot and NO analysis. All data are expressed as mean ± SEM. Statistical comparisons were conducted with one-way ANOVA followed by Tukey’s post hoc test (B, D, F, H). *P < 0.05, and **P < 0.01