Type 2 Diabetes Mellitus: Role of Inflammation and Innate Immunity in The Pathogenesis of Endothelial Dysfunction and Atherosclerosis

Study design: prospective, controlled, randomized study. All eligible patients will be randomly assigned in a 1:1 manner to receive aspirin or placebo.

In each recruited subjects we will do:

1. Anamnestic clinical information and Anthropometric measurements
2. Electrocardiogram, echocardiogram and ultrasound assessment of carotid intima-media thickness (IMT) and flow-mediated dilatation (FMD)
3. Ankle-Brachial Index measurement (ABI)
4. blood samples from an antecubital vein after an overnight fast and urine samples at baseline, after 3 days and after 30 days of aspirin (100 mg/day) administration.

Laboratory Methods: Blood samples will be immediately centrifuged at 2,000 rpm for 20 min at 4°C, and the supernatant was collected and stored at -80°C until measurement. All measurements will be done blinded. Samples will be tested in duplicate, and those showing values above the standard curve will be re-tested with appropriate dilutions.Analysis of urinary and platelet isoprostane:Urinary PGF2α-III was measured by a previously described and validated EIA assay method. Ten millilitre urine were extracted on a C-18 SPE column; the purification was tested for recovery by adding a radioactive tracer (tritiated PGF2α-III) (Cayman chemical). The eluates were dried under nitrogen, recovered with 1ml of buffer, and assayed in a PGF2α-III specific EIA kit (Cayman chemical). Urinary PGF2α-III concentration was corrected for recovery and creatinine excretion and expressed as pg/mg of creatinine. PRP was then centrifuged 20 min at 800 g to concentrate platelets and the pellet was suspended in Tyrode buffer to obtain a final platelet concentration of 5x108/mL. PGF2α-III content was measured by a validated EIA assay method as previously described and expressed as pg/mg platelet protein.