Abstract 254: 14, 15-EET Protects Heart Pericytes by Delaying I/R Injury-induced Calcium Overloading
Background - Pericytes are an important cellular component of the blood vessel wall of the arteries, arterioles, and microvessels of the heart; they provide structural integrity and regulate vessel diameter by contracting and relaxing dynamically in response to vasoactive stimuli. It has been suggested that pericytes contribute to coronary no-flow due to pericyte constriction following myocardial infarction, thus worsening outcome. It has also been demonstrated that intracellular calcium is involved in perciyte constriction. Our previous findings indicate that cardiomyocytes are protected following ischemia/ reperfusion injury (I/R) by the eicosanoid 14,15-EET. Since 14,15-EET is protective following I/R and a vasodilator, we tested the hypotheses that I/R injury induces calcium overloading, which injures peciytes, and that 14, 15-EET is able to block this process.
Methods and Results - We isolated and cultured pericytes from the mouse heart ventricle by 3G5 antibody Dynabead sorting. Pericytes were characterized by multiple immunocytochemical markers for contractile proteins, cytoskeletal protein, and cell surface receptors (alpha-smooth muscle actin, calponin-1, NG2, vimentin, CD31,smoothlin, and fibroblast protein-1). Cultured pericytes were subjected to 5 hours of oxygen and glucose deprivation, with or without 14,15-EET, followed by 15 hours of re-oxygenation in the absence of 14,15-EET. Calcium imaging and cell death during re-oxygenation were assessed by Fluo-4 and propidium iodide respectively. Digital images were taken with confocal microscope (Nikon Eclipse Tie-A1RSi). The brightness of the green fluorescent signals represents the relative level of intracellular calcium and the red fluorescent signals represent the cell death. We found that calcium signal peak (overloading) occurred during re-oxygenation, immediately followed by cell death. This process was delayed by 14,15-EET treatment during oxygen and glucose deprivation. The cell death at 5h, 10h, and 15h of re-oxygenation was 57.2%, 71.1%, and 85.3% in control group, and 19.9%, 35.3%, and 58.3% in 14,15-EET treated group.
Conclusions - Our data suggests that 14,15-EET-induced protection in pericytes is mediated through the calcium signaling pathway.
Author Disclosures: Z. Cao: None. C.M. Davis: None. S. Kaul: None. N.J. Alkayed: None.
- © 2015 by American Heart Association, Inc.