Aspartame exacerbates cerebral ischemia-reperfusion injury via mitochondrial dysfunction and ERK/CREB1 pathway...

Aspartame, a widely used non-nutritive sweetener, has been associated with potential neurotoxicity. However, it remains unclear whether aspartame consumption can exacerbate cerebral ischemia-reperfusion injury (CIRI), a major contributor to stroke outcomes, and the underlying mechanisms are poorly understood.

In vivo, male C57BL/6 J mice were given free access to 0.1% or 0.2% (w/v, equivalent to human acceptable daily intake level) aspartame, for 7 days, followed by bilateral common carotid artery occlusion (BCCAO) to induce CIRI. In vitro, hippocampal neural stem cells (NSCs) were subjected to oxygen-glucose deprivation (OGD) with aspartame. Cell injury, general and mitochondrial reactive oxygen species (ROS) burden, and mitochondrial function were assessed. Gene Ontology (GO), KEGG enrichment, and protein-protein interaction (PPI) network analyses were performed to identify potential targets. The ERK/CREB1 signaling pathway was evaluated by western blotting and pharmacological modulation.

Aspartame significantly increased the infarct volume and aggravated neuronal damage in BCCAO-treated mice. In NSCs, aspartame, but not acesulfame or sucralose, selectively enhanced OGD-induced apoptosis, accompanied by mitochondrial depolarization and excessive ROS accumulation, while showing minimal effects under normoxia conditions. GO/KEGG and PPI analyses highlighted ERK/CREB1 as an important node in aspartame-induced neurotoxicity. Consistently, aspartame suppressed the phosphorylation of ERK1/2 and CREB1. The ERK activator LM22B-10 and mitochondria-targeted antioxidant Mito-TEMPO partially reversed mitochondrial dysfunction, apoptosis and ERK/CREB1 suppression. Additionally, aspartame increased the mRNA expression of pro-inflammatory cytokines (TNF-α, IL-1β and IL-6) and increased NF-κB p65 phosphorylation and reduced the proportion of Tuj1-positive cells, which were mitigated by ERK activation.

Aspartame exacerbates CIRI-associated injury in a stress-dependent manner, involving mitochondrial dysfunction, ROS accumulation, and ERK/CREB1 suppression. Future studies are warranted to explore the long-term neurobehavioral outcomes and validate these mechanisms in clinical scenarios.