Skip to main content
[Preprint]. 2024 Jul 3:2024.07.01.601474. [Version 1] doi: 10.1101/2024.07.01.601474

Figure 6. Cholesterol regulates activation of human astrocytes.

Figure 6.

A) Schematic representation of the experimental design. B) Lipid droplet staining in iAstrocytes following 24h treatment with cholesterol. C) Normalized membrane MHC-I levels (stained for anti-HLA Class I Heavy Chain) in vehicle versus cholesterol treated ApoE3 iAstrocytes determined by flow cytometry. n=6 (K2.1J) and n=8 (B037) from 4 independent experiments. ****P<0.0001 Unpaired t-test. D) Normalized Il-6 secretion in vehicle versus cholesterol treated ApoE3 iAstrocytes n=6 (K2.1J) and n=12 (B037) from 4 independent experiments. ****P<0.0001 Unpaired t-test. E) Fold change of phospholipid species with indicated number of double bonds (unsaturation) in cholesterol treated vs control iAstrocytes (BIONi037 ApoE3). n=6 wells from 3 independent experiments. F-G) Representative histogram and quantification (G) of normalized MHC-I membrane levels determined by flow cytometry (stained for anti-HLA Class I Heavy Chain) in response to indicated treatment conditions in iAstrocytes. n=6 (K2.1J) and n=6 (B037) wells from 3 independent experiments per line. *P<0.05 One-way ANOVA with Dunnett’s multiple comparison correction. H) Secreted Il-6 levels in medium of ApoE3 iAstrocytes that were pre-treated with vehicle, exogenous cholesterol (10mM) or atorvastatin (0.5mM for one hour and then treated for 24 hours with increasing doses of TNF/Il-1α/C1q (in presence of vehicle, atorvastatin or exogenous cholesterol). n=5 biological replicates (n=2 Kolf2.1J and n=3 BIONi037). **p<0.01 intercept difference by linear regression model. Relative Il-6 levels with vehicle 0.25 times cocktail dose set at 1. I) Relative changes in membrane MHC-I levels determined by flow cytometry (stained for anti-HLA Class I Heavy Chain) in ApoE3 or ApoE4 iAstrocytes treated with cholesterol. n=6 (K2.1J) and n=6 (B037) wells from 3 independent experiments. BIONi037 (B037) and Kolf2.1J (K2.1J). J) ApoE4 decreases HLA expression and immune function in human glia by increased cholesterol storage in cholesteryl esters.