Medin-Induced Pro-inflammatory and Prothrombotic Activation of Coronary Artery Endothelial Cells: A Potential Novel Mediator Linking Aging and Atherosclerosis

Background: Age is the most important risk factor for coronary artery disease (CAD) independent of traditional risk factors. Aging induces classic pro-inflammatory and prothrombotic vascular phenotypic changes whose molecular mediators remain poorly understood. Medin is a common cleavage product protein that accumulates in vasculature with aging and shown to cause endothelial dysfunction. Its role in CAD is unknown. The study aimed to evaluate the effects of medin on human coronary artery endothelial cell (HCAEC) pro-inflammatory and prothrombotic activation and establish the relationship between medin and coronary atherosclerosis in human decedents. Methods: HCAECs were exposed to physiologic dose of medin (5 M) for 20 hours and ribonucleic acid sequencing (RNAseq) with signaling pathway analyses and reverse transcription polymerase chain reaction of select pro-inflammatory and prothrombotic genes performed. Corresponding protein expression was measured by Western blot or enzyme linked immunosorbent assay in HCAECs exposed to medin (5 M) without or with nuclear factor-{kappa}B (NF{kappa}B) inhibitor RO106-9920 (10 M). Coronary arteries from 40 deceased individuals underwent immunohistochemistry and medin and plaque burden were quantified and their relationship evaluated. Results: RNAseq showed predominant pro-inflammatory gene expression changes induced by medin. HCAECs treated with medin showed increased phosphorylated NF{kappa}B, elevated protein expression of interleukin (IL)-6, IL-8, monocyte chemotactic protein (MCP)-1, intercellular adhesion molecule (ICAM)-1, vascular cell adhesion molecule (VCAM)-1 and plasminogen activator inhibitor (PAI)-1 and reduced protein expression of thrombomodulin; these changes were reversed by RO106-9920 co-treatment. In human tissues, coronary artery medin strongly correlated with plaque burden (R=0.76, p<0.0001) and coronary macrophage content (R=0.72, p<0.0001). Coronary arteries from decedents with myocardial infarction had higher medin than those without (5.53{+/-}2.67% versus 0.02{+/-}0.02%, p=0.0005). Conclusions: Medin induced NF{kappa}B-mediated endothelial cell pro-inflammatory and prothrombotic activation and was strongly associated with coronary plaque burden and inflammation. Medin is a novel candidate mediator linking aging and coronary atherosclerosis.