Skip to main content
. 2023 Jun 8;14:3372. doi: 10.1038/s41467-023-39046-8

Fig. 5. Astrocytes export cholesterol to oligodendrocytes to regulate their survival and remyelination.

Fig. 5

a Primary astrocytes treatment plan and collection of astrocyte-conditioned media (ACM). b Nrf2+ (yellow) astrocytes (Aldh1l1+; magenta), with Hoechst in blue, following treatment with CDDOTFEA or CDDOTFEA, then Luteolin, or CDDOTFEA then CS-6253. Scale bar, 50 µm. c Mean percentage of astrocytes expressing nuclear Nrf2 ± s.e.m. One-way ANOVA and Tukey’s multiple comparisons test; aP = 0.0286, bP = 0.0085, cP = 0.0178. ANOVA summary P = 0.0023, F = 6.994. n = 6 independent litters (no treatment, CDDO, CDDO/Luteolin, CDDO/CS-6253). d Astrocyte expression of genes involved in cholesterol and Nrf2 signalling following CDDOTFEA treatment, represented as Log2FC over no treatment condition ± s.e.m. One-tailed Wilcoxon test for 2−ΔΔCt, P value = 0.0156. n = 4 independent litters (Nfe2l2, Gclc, Nqo1, Hmgcs1, Fdps) and n = 3 independent litters (Mvk, Fdft1). e Astrocyte expression of genes after CDDOTFEA treatment followed by Luteolin treatment, represented as Log2FC over CDDOTFEA condition ± s.e.m. Kolmogorov–Smirnov tests on 2−ΔΔCt, Hmgcs1 P = 0.0286. n = 4 independent litters (Hmgcs1, Fdps) and n = 3 independent litters (Mvk, Fdft1). f Astrocyte expression of genes after CDDOTFEA treatment followed by CS-6253 treatment, represented as Log2FC over CDDOTFEA condition ± s.e.m. Kolmogorov–Smirnov tests on 2−ΔΔCt, Hmgcs1 P = 0.0476, Fdps P = 0.0476, Mvk P = 0.0286. n = 4 independent litters (Hmgcs1, Fdps) and n = 3 independent litters (Mvk, Fdft1). g ACM applied to oligodendrocytes to track the uptake of Bodipy-FL-C12 exported from astrocytes. h Percentage of TPPP/p25+ cells cholesterol which are Bodipy+ ± s.e.m. Kruskal–Wallis test and Dunn’s multiple comparisons test, aP = 0.0561, bP = 0.0360. n = 4 independent litters. i Oligodendrocytes (TPPP/p25+ Sox10+; magenta/cyan) in unconditioned astrocyte media (AST) or following exposure to ACM, and uptake of Bodipy-FL-C12 (yellow). Hoechst in blue. Scale bar, 50 µm. j Immature oligodendrocyte lineage cells (Sox10+ TPPP/p25-) were Bodipy negative (arrows). Scale bar, 50 µm. k Mean percentage of TPPP/p25+ cells which are Tunel+ ± s.e.m., normalised to oligodendrocyte (OL) media control. Kruskal–Wallis test and Dunn’s multiple comparisons test, P value = 0.0349, aP = 0.0083, bP = 0.0265, cP = 0.0265. n = 4 independent litters. l Apoptotic (Tunel+; yellow) TPPP/p25+ Sox10+ oligodendrocytes (magenta/cyan), in OL media or AST control, or following exposure to ACM. Scale bar, 50 µm. m Brain explants myelinated for 14 days in vitro (DIV), were demyelinated with LPC, then fixed at 5 days post-LPC (dpl) when remyelination is initiated. n Remyelination index ± s.e.m. for AST: slice culture media (SC) control, or following exposure to ACM. Two-tailed unpaired Student’s t-test with Welch’s correction, aP = 0.0048, t = 4.842; bP = 0.0125, t = 3.723. n = 3 mice/condition (AST:SC media, SC media: ACM), n = 4 mice/condition (CDDO, CDDO+Luteolin, CDDO + CS-6253), n = 3 mice/condition (AST:SC media, ACM no treatment). o Explants exposed to AST or ACM from astrocytes following no treatment or exposure to CDDOTFEA, CDDOTFEA then Luteolin, and CDDOTFEA then CS-6253 stained for myelin basic protein (MBP; magenta) and neurofilament (NF; green). Scale bar, 50 µm. p Explants myelinated for 14 DIV, were demyelinated with LPC, then treated with the ABCA1 inhibitor PSC833 or vehicle control from 2 to 7 dpl when remyelination is underway. q Explants treated with vehicle control or PSC833 (5 µM) and stained for MBP (magenta) and neurofilament (NF; green). Scale bar, 50 µm. r Remyelination index ± s.e.m. for vehicle or PSC833-treated brain explants. One-way ANOVA with Tukey´s multiple comparisons test, aP = 0.0052, bP = 0.0026, cP = 0.0007. ANOVA summary (F = 17.11 and P value = 0.0008). n = 3 mice/condition. Source data is provided with this paper. The images in 5a, g, m, p were created with Biorender.com.