Abstract W P201: Angiopoietin-1 Mimetic Peptide Treatment of Stroke Regulates MicroRNA Expression and Promotes Neuroprotection in Type One Diabetic Rats

Abstract

Introduction: Angiopoietin-1 (Ang-1) mediates vascular remodeling. Diabetes decreases Ang1 and disrupts Ang1/Tie2 signaling, impairs vascular maturation and regulates immune response. MicroRNAs have been implicated in vascular diseases and inflammation. Vasculotide (VT) is an Ang1 mimetic peptide that promotes Tie2 activity. We hypothesize that treating type one diabetic rats (T1DM) with VT after stroke regulates miRNA expression decreases pro-inflammatory factors and thereby produces a neuroprotective effect. Methods: T1DM was induced in male Wistar rats with streptozotocin followed by 2h transient middle cerebral artery occlusion (MCAo). These rats were then treated with: 1) PBS control; 2) VT (3 ug/kg, ip injection) just prior to stroke and at 8h and 24h after MCAo. Functional tests were conducted 48h after MCAo. Afterwards, brain hemorrhage, BBB leakage, miRNA expression, RT-PCR, immunostaining and Western blot assays were done. Results: VT treatment did not decrease brain hemorrhage, but significantly decreased Evans Blue leakage (30.6±5.8 ng/mg vs. 17.9±2.0 ng/mg) lesion volume (35.1±3.5% vs 25.8±5.3%) and improved functional outcome after stroke when compared to control (p<0.05). To elucidate the mechanism, miRNA was measured. VT treatment significantly increased miR-451 (81.6 folds), miR-155 (2.4 fold), miR-98 (1.9 fold), and miR-126 (1.5 fold) expression, but decreased miR-200b (3.8 fold). Immunostaining showed that VT significantly decreased the number of apoptotic (45.5±3.7 vs 30.8±2.6) and cleaved-caspase-3 positive cells (45.2±3.5 vs 33.1±4.3), as well as monocyte chemotactic protein-1 (MCP1, 5.02±0.4% vs 1.8±0.2%) and tumor necrosis factor (TNF)-alpha (3.1±0.5% vs 1.5±0.2%). Western blot showed that VT significantly decreased receptor for advanced glycation end products (RAGE), MCP1 and TNFa (3-5 fold) in the ischemic border area compared to T1DM control. Conclusion: VT promotes neuroprotection after stroke in T1DM rats. Regulation of miRNA and neuroinflammatory factor expression and decreased BBB leakage may contribute to the VT induced neuroprotective effects observed after stroke in T1DM rats.