Purpose: Hypoxia and reoxygenation (H/R) occur in a wide variety of important clinical conditions such as myocardial infarction. H/R injury is a complex phenomenon involving not only intracellular damage processes but also an injurious inflammatory response. Docosahexaenoic acid (DHA), an n-3 polyunsaturated fatty acid, has long been proved to be protective against several types of cardiovascular disease. However, its beneficial effect during H/R is inconclusive. In this study, we employed an in vitro model to examine whether DHA is protective against H/R-induced cell damage in human coronary artery smooth muscle cells (HCASMCs).
Methods: HCASMCs in the absence or presence of DHA (1, 3, 10, and 30 μM) were subjected to control or H/R treatment using a modular incubator chamber to create hypoxic condition. Cell viability was evaluated by MTT assay. Spectrophotometric and spectrofluorometric assays were used to measure the generation of nitric oxide (NO) and reactive oxygen species (ROS), respectively. Inflammatory cytokines were determined by enzyme-linked immunosorbent assay. Intracellular calcium mobilization was estimated microfluorimetrically using calcium indicator dye, fura 2-acetomethyl ester.
Results: Hypoxia/reoxygenation caused significant injury in cultured HCASMCs. DHA at low concentrations (1, 3, and 10 μM) did not afford protection, whereas at 30 μM, it caused deleterious effects, presumably by enhancing the production of NO, ROS, IL-1β, and IL-6 and altering the intracellular calcium dynamics.
Conclusions: Our results do not support the protective function of DHA in H/R-injured coronary arterial smooth muscle cells.