CD31 Positive-Extracellular Vesicles from Patients with Type 2 Diabetes Shuttle a miRNA Signature Associated with Cardiovascular Complications

Innovative biomarkers are needed to improve the management of patients with type 2 diabetes mellitus (T2DM). Blood circulating miRNAs have been proposed as a potential tool to detect T2DM complications but the lack of tissue specificity, among other reasons, has hampered their translation to clinical settings. Extracellular vesicle (EV)-shuttled miRNAs have been proposed as an alternative approach. Here, we adapted an immunomagnetic bead-based method to isolate plasma CD31 positive (⁺) EVs to harvest vesicles deriving from tissues relevant for T2DM complications. Surface marker characterization showed that CD31⁺ EVs were also positive for a range of markers typical of both platelets and activated endothelial cells. After characterization, we quantified 11 candidate miRNAs associated with vascular performance and shuttled by CD31⁺EVs in a large (n=218), cross-sectional cohort of patients categorized as T2DM without complications, T2DM with complications, and controls. We found that 10 of the tested miRNAs are affected by T2DM, while the signature composed by miR-146a, -320a, -422a, -451a efficiently identified T2DM patients with complications. Furthermore, another CD31⁺EV-shuttled miRNA signature, i.e. miR-155, -320a, -342-3p, -376, and -422a, detected T2DM patients with a previous major adverse cardiovascular event. Many of these miRNAs significantly correlate with clinical variables held to play a key role in the development of complications. In addition, we show that CD31⁺ EVs from patients with T2DM are able to promote the expression of selected inflammatory mRNAs, i.e. CCL2, IL-1α, and TNFα, when administered to endothelial cells in vitro. Overall, these data suggest that the miRNA cargo of plasma CD31⁺ EVs is largely affected by T2DM and related complications, encouraging further research to explore the diagnostic potential and the functional role of these alterations.