(P23-006-25) EPA and Gut Metabolite Urolithin A Mitigate Extreme Heatstroke-Induced NLRP3 Inflammasome Activation in Microglial Cells

Objectives: Global warming and extreme heat exposure pose significant health threats, particularly for vulnerable populations such as older adults. Heatstroke (HS) is an emerging risk factor to exacerbate neuroinflammation and its associated neurodegenerative diseases. However, the mechanisms underlying HS-induced neuronal damage and the efficacy of dietary interventions in mitigating these effects remain poorly understood. This study hypothesized that HS activates the NLRP3 inflammasome, an innate immune pathway leading to cytotoxic IL-1β release, which could be mitigated by anti-inflammatory dietary compounds. The objectives were to elucidate the mechanisms by which HS promoted neuroinflammation and to assess the preventive potential of eicosapentaenoic acid (EPA) and urolithin A (UroA), a gut-derived metabolite of ellagic acid.

Methods: To simulate cerebral HS, characterized by elevated core body temperature and reduced oxygenation, murine BV2 microglial cells were exposed to hyperthermic conditions (41°C) with hypoxia (~1% O₂) for 1 hour. To model metabolic susceptibility or pre-existing low-grade inflammatory conditions, BV2 cells were primed with LPS prior to HS exposure. BV2 cells were treated with EPA (50 µM), UroA (10 µM), or their combination for 48 hours. Cell viability, IL-1β release, and pro-inflammatory gene expression were measured. NLRP3 inflammasome activation by HS was further confirmed using J774 macrophages, stably expressing pro-IL-1β-Gaussia luciferase.

Results: HS induced cell death, IL-1β release, and increased IL-1β reporter activity, with these effects being further exacerbated by pre-existing inflammation. Notably, EPA and UroA effectively attenuated HS-induced IL-1β release, indicating suppression of NLRP3 inflammasome activation. Moreover, combined EPA and UroA treatment exhibited synergistic effects in mitigating NLRP3 inflammasome activation, even under pre-existing inflammatory conditions.

Conclusions: Dietary intake of EPA or UroA-producing foods could provide protective effects against HS-induced neuroinflammation by suppressing NLRP3 inflammasome activation. These findings highlight novel dietary intervention strategies targeting HS-induced neuroinflammation, particularly for vulnerable populations.

Funding Sources: University of Massachusetts Faculty Research Grant, USDA-HATCH