Proteomic Insights Into Carotid Atherosclerotic Plaque Progression: Unraveling Molecular Mechanisms From American Heart Association Classifications IV to VI

Atherosclerosis (AS) is a progressive chronic disease, where plaque rupture can lead to acute clinical events. Recognizing the transition from stable to unstable plaques, as defined by the American Heart Association (AHA) classification, is essential, with the AHA's pathological assessment serving as the gold standard for evaluating plaque conditions. Utilizing proteomics and integrative omics techniques, this study aimed to uncover proteomic profiles across different AHA classifications and pinpoint potential therapeutic targets. We utilized Data-Independent Acquisition (DIA) to analyze 87 human carotid plaques, ensuring stringent quality control, resulting in a comprehensive dataset of 6143 proteins. Differential proteomics and protein-protein interaction network analyses were employed to discern expression patterns in AHA IV, V, and VI plaques. Integration with single-cell RNA sequencing pinpointed key proteins driving plaque progression. Mendelian randomization (MR) assessed causal links between identified proteins and atherosclerosis- related diseases. Key proteins—CD44, GAL-1, TREM2, SMAD3, and IL-6R—emerged as pivotal in transitioning carotid plaques from AHA type IV to VI, signifying increased instability. These proteins exhibited differential expression patterns: CD44 and GAL-1 predominated in the fibrous cap, while TREM2, SMAD3, and IL-6R localized to the lipid core. Validation across independent samples and single-cell datasets reinforced their importance. MR analyses affirmed CD44, SMAD3, and IL-6R's causative roles in atherosclerosis-related diseases. This study provides unprecedented insights into the proteomic landscape of atherosclerosis, highlighting specific proteins crucial for the progression of carotid plaques. The identified proteins, particularly CD44, SMAD3, and IL-6R, not only serve as potential biomarkers but also offer promising avenues for targeted therapeutic interventions, emphasizing their significance in understanding and combating atherosclerosis-related disorders.