Mercury Activates Phospholipase A₂and Induces Formation of Arachidonic Acid Metabolites in Vascular Endothelial Cells

Abstract

Currently, mercury has been identified as a risk factor in cardiovascular diseases among humans. Here, we tested our hypothesis that mercury modulates the activity of the vascular endothelial cell (EC) lipid signaling enzyme phospholipase A(2) (PLA(2)), which is an important player in the EC barrier functions. Monolayers of bovine pulmonary artery ECs (BPAECs) in culture, following labeling of membrane phospholipids with [(3)H]arachidonic acid (AA), were exposed to the inorganic form of mercury, mercury chloride, and the release of free AA (index of PLA(2) activity) and formation of AA metabolites were determined by liquid scintillation counting and enzyme immunoassay, respectively. Mercury chloride significantly activated PLA(2) in BPAECs in a dose-dependent (0 to 50 muM) and time-dependent (0 to 120 min) fashion. Metal chelators significantly attenuated mercury-induced PLA(2) activation, suggesting that cellular mercury-ligand interaction is required for the enzyme activation and that chelators are suitable blockers for mercury-induced PLA(2) activation in ECs. Sulfhydryl (thiol-protective) agents, calcium chelating agents, and cPLA(2)-specific inhibitor also significantly attenuated the mercury-induced PLA(2), suggesting the role of thiol and calcium in the activation of cPLA(2) in BPAECs. Significant formation of AA metabolites, including the release of total prostaglandins, thromboxane B(2), and 8-isoprostane, were observed in BPAECs following their exposure to mercury chloride. Mercury chloride induced cytotoxicity as observed by the altered cell morphology and enhanced trypan blue uptake, which was attenuated by the cPLA(2) inhibitor AACOCF(3). The results of this study revealed that inorganic mercury-induced PLA(2) activation through the thiol and calcium signaling and the formation of bioactive AA metabolites further demonstrated the association of PLA(2) with the cytotoxicity of mercury in ECs. Overall, the results of the current study underscore the importance of PLA(2) signaling in mercury-induced endothelial dysfunctions.

Role of Mercury Toxicity in Hypertension, Cardiovascular Disease, and Stroke

“…37,38 The clinical consequences of these and other pathophysiologic mechanisms explains the wide variety of cardiovascular diseases caused by mercury including CHD, MI, arrhythmias, abnormal heart rate variability, generalized atherosclerosis, sudden death, CVA, carotid artery stenosis, renal dysfunction, and hypertension. 4,5,6,7,8,9,13,15,19,26,28, Mercury activates phospholipase A2 (PLA-2) and induces formation of arachidonic acid metabolites such as total prostaglandins, thromboxane B2, and 8-isoprostane in vascular endothelial cells and activates vascular endothelial cell phospholipase D. 26,[51][52][53][54] Many of the cardiovascular consequences of mercury are mitigated by concomitant intake of fish containing omega-3 fatty acids and by the intake of selenium. 8,[45][46][47][48][49][50] Even very low levels of chronic mercury exposure promote endothelial dysfunction as a result of increased inflammation, oxidative stress, reduced oxidative defense, reduction in nitric oxide (NO) bioavailability, which increases the risk of CVD and CVA.…”

Redox-active antioxidant modulation of lipid signaling in vascular endothelial cells: vitamin C induces activation of phospholipase D through phospholipase A2, lipoxygenase, and cyclooxygenase

“…Release of AA from the cellular membrane phospholipids is widely assayed as an index of PLA 2 activity [18,37]. BLMVECs in 35-mm dishes (5 × 10 5 cells/dish) were labeled with carrier-free [ 3 H]AA (5 μCi/ml) in complete EC media containing 10% FBS, nonessential amino acids, antibiotic, and growth factor for 12 h at 37°C in 5% CO 2 -95% air.…”

“…The COX-and LOX-catalyzed formation of AA metabolites in BLMVECs cultured in 35mm dishes (5 × 10 5 cells/dish), following their exposure to vitamin C at different concentrations (mM) in MEM for 1 and 2 h, was determined by utilizing the commercially available EIA kits (Cayman Chemical Co., Ann Arbor, MI) according to Mazerik et al [37].…”

Methylmercury Increases and Eicosapentaenoic Acid Decreases the Relative Amounts of Arachidonic Acid‐Containing Phospholipids in Mouse Brain

“…Dietary supplementation with marine fish oil rich in n-3 PUFAs, including EPA and docosahexaenoic acid (DHA), leads to various favorable effects caused by the modification of the lipid composition [20,36,37]. MeHg has a negative effect and may activate PLA 2 and subsequently release ARA, leading to toxic effects [9,10,38,39]. The above evidences suggest that both EPA and MeHg influence the membrane phospholipid composition in biological systems, and through a favorable modification of membrane phospholipid composition and inhibition of ARA metabolism, EPA may alleviate the MeHg-induced toxicity.…”