Thesis: Carla Perpiñá Clérigues

Directores/as: Francisco García García y María Pascual Mora

Title: Sex differences in the effects of alcohol consumption on the content of plasma extracellular vesicles: a multi-omics study in humans and mice

Abstract: Alcohol is a widely consumed psychoactive substance whose use leads to severe health and social consequences. Importantly, biological sex plays a critical role in shaping vulnerability to alcohol-related damage. Accumulating evidence indicates that women exhibit a stronger neuroinflammatory response to acute and chronic alcohol exposure, a process in which Toll-like receptor 4 (TLR4) acts as a key molecular mediator. This dissertation investigates the biological mechanisms underlying two patterns of alcohol consumption, focusing on sex-specific responses and the modulatory role of TLR4. The work centers on plasma extracellular vesicles (EVs), small circulating particles involved in intercellular communication that represent a promising source of non-invasive biomarkers. Specifically, the lipidome and miRNome of EVs were analyzed using single-omic and integrative multi-omic bioinformatic approaches across human and animal models. Alcohol exposure induced pronounced sex-dependent alterations in EV lipid composition. Acute alcohol intake in human adolescents revealed predominantly pro-inflammatory lipid profiles in females. In mouse models, lipidomic changes were strongly influenced by both sex and TLR4 expression, highlighting the receptor’s role in shaping alcohol-induced inflammatory responses. In adults with Alcohol Use Disorder (AUD), EV lipid alterations in females were associated with pathways related to neuroinflammation and cancer, whereas in males they were mainly linked to liver toxicity. Chronic alcohol exposure further demonstrated that sex and TLR4 jointly regulate EV lipid remodeling, affecting central processes such as the Lands’ cycle and phospholipid metabolism, and pointing to lipid rafts as relevant hubs for TLR4-related signaling. Integrative multi-omic analyses combining lipidomic and miRNA data from AUD patients identified molecular signatures with diagnostic potential, including sex-specific profiles. Notably, miRNA-related functional alterations showed opposing patterns between males and females in pathways related to inflammation, oxidative stress, and autophagy. Overall, this dissertation demonstrates that plasma EVs carry sex-specific molecular signatures of alcohol-related effects, offering a valuable source of biomarkers. Results and methods are also available through interactive web-based tools.