Endothelial cell (EC) activation and dysfunction have been linked to a variety of chronic vascular inflammatory
disease states including insulin resistance (IR), obesity, and atherosclerosis—the major cause of morbidity and
mortality in Western Societies. Proinflammatory cytokines and proatherogenic risk factors such as
hyperlipidemia activate key signaling pathways such as NF-kB and MEKK/JNK and decrease protective ones
such as AKT/eNOS signaling, an effect that confers pro-adhesive and pro-thrombotic properties to ECs.
Therefore, suppressing the inflammatory response in the vascular endothelium may provide a novel
therapeutic approach to limit IR/obesity, and atherothrombosis.
MicroRNAs (miRNAs) are small, single-stranded, non-coding RNAs capable of repressing gene expression at
the post-transcriptional level and are involved in a variety of pathophysiological processes including the
regulation of immune and inflammatory responses. During this grant period, we identified miR-181b as a nodal
regulator of endothelial cell quiescence through its regulatory effects on two major signaling pathways – NF-kB
and AKT/eNOS, and have uncovered novel targets for therapy. Preliminary and published observations now
identify endothelial miR-181b as a critical determinant of systemic inflammation, obesity/IR, and atherogenesis.
Furthermore, our studies highlight the relevance of microRNA-dependent targets in response to ligand-specific
signaling in ECs–an emerging concept in the microRNA field. Moreover, we demonstrate that methotrexate
(MTX), a clinically relevant therapy that suppresses endothelial activation, can function in a miR-181b-
dependent manner. These observations provide the foundation for the central hypothesis that endothelial miR-
181b, via inhibitory effects on NF-¿B and MEKK/JNK and inductive effects on AKT/eNOS signaling, regulates
systemic vascular inflammation, IR/obesity, and atherosclerosis. To better understand the precise role of miR-
181b in stimuli-specific regulation of EC activation, three aims are proposed. In Aim1, we will delineate the
molecular basis for miR-181b's ability to inhibit endothelial NF-¿B and MEKK/JNK and induce AKT/eNOS
pathways in response to divergent stimuli. In Aim2, we will determine the effect of altered endothelial miR-181b
expression on obesity/insulin resistance and atherothrombosis. In Aim3, we will explore the molecular
mechanisms by which methotrexate (MTX) rescues miR-181b expression in ECs, and we will determine
whether the anti-inflammatory effects of MTX are miR-181b-dependent. The results of these studies will
provide insights regarding miR-181b-mediated control of endothelial activation and the functional
consequences of miR-181b sufficiency and deficiency on insulin resistance and atherogenesis. Furthermore,
studies exploring the link between miR-181b and MTX may provide insights that can be exploited for
therapeutic gain in the treatment of a broad spectrum of chronic inflammatory disorders.