Unveiling the Molecular Mechanism of Intestinal Metabolite para ‐Cresol in Modulating Neuroinflammation and Synaptic Dysfunction: Implications for Autism Spectrum Disorder

ABSTRACT Autism spectrum disorder (ASD) is a diverse group of neurodevelopmental disorders that share similar behavioral patterns. Many individuals with ASD also exhibit gastrointestinal disturbances, likely linked to alterations in the composition and activity of intestinal bacteria, resulting in the overproduction and release of toxic metabolites into the systemic circulation. These toxins may reach the central nervous system (CNS) and activate microglia, which release inflammatory cytokines, ultimately impairing neuronal function. This microbiota‐gut‐brain axis has been suggested to play a crucial role in the pathogenesis of ASD. Para ‐cresol ( p ‐Cresol) is one of these intestinal metabolites and could potentially contribute to ASD, as its urinary levels are elevated in autistic children under the age of 8 and correlate with symptom severity. Here, we aim to investigate the effect of p ‐Cresol on various brain‐cell types to speculate on their specific contributions to ASD‐related synaptic dysfunctions. Immunocytochemistry assays revealed a significant decrease in excitatory (vesicular glutamate transporter VGLUT, postsynaptic density protein 95 PSD95) and inhibitory (vesicular GABA transporter VGAT) synaptic markers in p ‐Cresol‐treated neurons during synaptogenesis. These effects were exacerbated in SH3 and multiple ankyrin repeat domains 3 (Shank3) knockdown neurons, which exhibit increased susceptibility of their synapses. p ‐Cresol also induced a dose‐dependent inflammatory response in both astrocytes and microglia, characterized by the overexpression and release of inflammatory cytokines and chemokines such as interleukin 6 (IL6), interleukin 1β (IL1β), and C‐C‐ motif chemokine ligand 3 (CCL3). Our results suggest that p ‐Cresol exerts cell‐specific effects on astrocytes and microglia, and the release of inflammatory cytokines and chemokines in response to p ‐Cresol treatment may contribute to synaptic dysfunction, in addition to its direct effect on neurons. image