(A) Brainstem neurons from hA53T transgenic mice and substantia nigra neurons from PFF-injected mice accumulate iron measured using iCP-MS (see Figure S1 for sorting paradigm). n= 10 biological replicates per group.
(B) Neuronal iron accumulation secondary to PFFs occurs via a non-cell-autonomous mechanism measured using iCP-MS. n= 10 biological replicates per group.
(C) PFF-microglia conditioned media (MCM) is sufficient for neuronal iron accumulation measured using iCP-MS. n = 10 biological replicates per group.
(D) 14 secreted factors are significantly elevated in PFF MCM compared with in monomer MCM measured using Procartaplex Cytokine Multiplex assay. n = 10 biological replicates per group.
(E) IL-6 is necessary for PFF-MCM-mediated neuronal iron accumulation measured using iCP-MS. n = 5 biological replicates per group.
(F) Trans-IL-6 signaling is sufficient for neuronal iron accumulation from ferrous, but not ferric, iron sources. Iron accumulation measured using iCP-MS. n = 5 biological replicates per group.
(G) IL-6 overexpression is sufficient for neuronal iron accumulation in vivo. Iron measured in isolated neurons (see Figure S1) using iCP-MS. n = 5 biological replicates per group.
(H) Seven iron transport genes are differentially expressed in neurons secondary to PFF MCM treatment in an IL-6-dependent manner. Gene expression measured using a custom Luminex Quantigene assay. n = 6 biological replicates per group.
(I) IL-6 overexpression is sufficient for increased expression of Zip14, Tfrc, Dmt1, Ftl, Fth, and Hampin vivo. Gene expression measured by qPCR. n = 5 biological replicates per group.
(J) Schematic showing gene expression changes in (H)–(I) (genes of interest in red) overlayed on a diagram of cellular iron transport highlighting upregulation of ferric and ferrous iron import and downregulation of iron export.
(A–C, E–G, and I) Data indicate mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, by unpaired student’s two-tailed t test (A–D, G, and I) and two-way ANOVA (E and F) with Tukey’s honestly significant difference (HSD) post hoc test.