Abstract Body: Introduction: Type-2 diabetes increases the risk of stroke; however, the mechanisms remain incompletely understood. Using a rat model of type-2 diabetes that expresses amyloid-forming human amylin in the pancreas (the HIP rat), we previously reported that human amylin deposits in the brain microvasculature causing brain microhemorrhages and neurological deficits. Here, we tested the hypotheses: 1, mechanisms of amylin-mediated cerebral microvascular injury involve damage of the endothelial cell tight junctions; 2, an increase in epoxyeicosatrienoic acids (EETs), protective proteins secreted by the endothelial cells, reduces brain microvasculature and microvascular injury in HIP rats.

Methods: To delineate amylin-mediated neurological deficits and brain microvascular injury, we used HIP rats expressing amyloidogenic human amylin in the pancreas and amylin knockout (AKO) rats intravenously infused with aggregated human amylin. To upregulate EETS, HIP rats were treated for 10 weeks with a soluble epoxide inhibitor, APAU (UC1153) which was added in the drinking water (1 ml drug/L). Biochemical analyses were carried out for investigating the formation of adducts of amylin aggreagtes with tight junction proteins and the peroxidative injury of the endothelial cells.

Results: HIP rats and AKO rats infused with aggregated amylin have increased brain microhemorrhages (HIP 14.86 ± 6.619; p=0.04, and AKO-inj 8.500 ± 5.137; p=0.17), white matter injury (HIP 170.5 ± 43.90; p=0.0022), and neurologic deficits (neurological Z-score -0.5337 ± 0.3505; p>0.01). Vascular amylin deposition provokes peroxidative injury (HIP 4HNE 166.4 ± 44.28; p=0.0027, MDA 45.04 ± 13.15; p=0.005, and AKO-inj 4HNE 182.2 ± 56.54; p=0.012, MDA 44.17 ± 21.84; p=0.07)), loss of endothelial cell coverage (HIP p=0.0002, and AKO-inj p=0.42) and tight junctions in HIP and AKO rats infused with aggregated amylin. Treatment with sEHI reduced vascular amylin deposition (198.6 ± 96.75; p=0.11), prevented loss of tight-junction proteins, and improved cognitive and motor function in HIP rats (neurological Z-score for cognitive function 0.2063 ± 0.4635; p=0.71, and motor function 0.8858 ± 0.1931; p<0.01).

Conclusions: Systemic amylin dyshomeostasis is a mediator of microvascular injury to the brain by damaging the endothelial cell tight junctions. Reducing amylin dyshomeostasis by upregulating EETs reduces amylin-mediated cerebrovascular injury and neurological deficits in the setting of diabetes.