LDL Cholesterol Modulates Astrocyte Metabolism, Lipid Handling, and Morphology: Evidence From In Vitro and In Vivo...

Astrocytes are the primary antioxidant defense cells of the brain, protecting the central nervous system (CNS) through a controlled inflammatory response and acting as metabolic suppliers to neurons. These cells exhibit morphological, functional, and molecular changes in pathological conditions, such as neurodegenerative diseases. Previous studies have demonstrated a link between hypercholesterolemia, especially elevated levels of low-density lipoprotein (LDL) cholesterol, and brain disorders, including hippocampal astrogliosis. In this context, this study aimed to investigate how LDL cholesterol modulates astrocyte biology. In vitro, high-passage rat C6 astroglial cells were exposed to human LDL cholesterol (50 or 300 μg/mL) for 24 or 48 h. We evaluated lipid accumulation, cholesterol metabolism-related gene expression, astrocyte-related gene expression, reactive species production, antioxidant activity, redox-related gene expression, fatty acid and glucose uptake, cell proliferation, and metabolic activity - 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction. LDL exposure increased intracellular lipid content and downregulated LDL receptor (LDLR), 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), and sterol regulatory element-binding transcription factor 1 (SREBF1) gene expression. LDL exposure altered astrocytic marker expression, as evidenced by increased glial fibrillary acid protein (GFAP) messenger RNA (mRNA) levels at 24 h with 300 μg/mL LDL and at 48 h with 50 μg/mL LDL. LDL cholesterol decreased long-chain fatty acids (LCFA) uptake and superoxide dismutase (SOD) activity at 24 h and increased cluster of differentiation 36 (CD36), also known as fatty acid translocase levels, at 48 h. Nuclear factor erythroid 2-related factor 2 (NRF2) expression was significantly increased after 48 h of incubation with 300 μg/mL LDL. The MTT reduction assay did not indicate decreased cell viability; instead, it revealed increased metabolic activity after 24 h of incubation with 300 μg/mL LDL, with no changes observed in glucose uptake. In vivo, hippocampal astrocytes from young (3-month-old) and middle-aged (14-month-old) LDL receptor knockout (LDLr-/-) and wild-type C57BL/6 mice were analyzed by immunofluorescence and quantitative reverse transcription polymerase chain reaction (RT-qPCR). In the hippocampal Cornu Ammonis 3 (CA3) region, 14-month-old LDLr-/- mice showed an increase in the number of processes compared to 3-month-old wild-type C57BL/6 mice. Aging and genotype influenced astrocyte morphology and expression of genes such as S100 calcium-binding protein B (S100B) and aquaporin-4 (AQP4). Our findings demonstrate that LDL cholesterol induces morphological, metabolic, and molecular changes in astrocytes, both in vitro and in vivo, suggesting that astroglial cells are sensitive to lipid imbalance and may play a role in the brain consequences of hypercholesterolemia.