Blocking Glycolysis in Human Astrocytes Dials Down Brain Inflammation

Astrocytes are the brain's most abundant glial cells and play a central role in maintaining neural homeostasis. When chronically activated, they drive neuroinflammation implicated in Alzheimer's disease, HIV-associated neurocognitive disorders, Parkinson's disease, and traumatic brain injury. A key but underexplored factor is how the metabolic environment shapes astrocyte inflammatory responses—a gap this study directly addresses.

Using primary human fetal-derived astrocytes, the team tested 2-deoxyglucose (2-DG), a glucose analogue that competitively inhibits glycolysis and serves as a caloric restriction mimetic. Cells were pre-treated with 2-DG at 10, 20, or 50 mM before stimulation with IL-1β (20 ng/mL for 24 hours). Seahorse extracellular flux analysis showed that IL-1β alone modestly increased both oxygen consumption rate (OCR) and extracellular acidification rate (ECAR), reflecting a shift toward glycolytic metabolism. Adding 20 or 50 mM 2-DG significantly reversed both parameters, confirming effective glycolytic inhibition even in an inflammatory context.

RT-qPCR and RNA-sequencing confirmed that 2-DG substantially reduced IL-1β-induced upregulation of TNF, IL-6, and complement component C3—key mediators of neuroinflammatory cascades. Transcriptomic analysis revealed broad gene expression changes across immune signaling, metabolic, and structural pathways. Critically, ATAC-sequencing demonstrated that these transcriptional shifts coincided with measurable changes in chromatin accessibility, suggesting that metabolic reprogramming rewires the epigenetic landscape of astrocytes. Motif analysis of differentially accessible regions pointed to transcription factors involved in NF-κB and inflammatory signaling as likely mediators.

Parallel experiments explored whether alternative metabolic substrates could replicate these anti-inflammatory effects. Glucose deprivation alone reduced cytokine expression, and supplementation with the ketone body β-hydroxybutyrate (BHB, 30 mM)—elevated during ketogenic diets—produced similar anti-inflammatory effects. Importantly, combining BHB or glucose deprivation with 2-DG produced additive reductions in cytokine expression, suggesting complementary or synergistic mechanisms.

These results provide mechanistic support for the hypothesis that caloric restriction and ketogenic diets exert neuroprotective effects partly by reprogramming astrocyte immunometabolism at both the transcriptional and epigenetic levels. The finding that chromatin accessibility changes accompany transcriptomic shifts implies durable, potentially heritable alterations in astrocyte inflammatory tone. While the study is limited by its in vitro, single cell-line design, it lays important groundwork for investigating glycolysis-targeting strategies as adjunct therapies for neuroinflammatory diseases.