Novel Mechanisms Related to DHA’s Atheroprotective Effects in Endothelial Cells: Modulation of Histone H3 Modification via Activation of p38MAPK Signaling

Abstract

Abstract Objectives Docosahexaenoic acid (DHA) is known for its protective effects against cardiovascular disease, which has been the leading cause of death worldwide for 2 decades. The molecular mechanisms responsible for DHA's atheroprotective effects, however, remain largely unknown. DHA has been found to activate p38 mitogen-activated protein kinase (MAPK) differently in growing and quiescent human endothelial cells, which represent dysfunctional and healthy states in vivo, respectively. This study was designed to characterize the activation pattern of p38MAPK in endothelial cells in response to DHA treatment and to identify possible downstream targets by which DHA exerts its protective effects. Methods EA.hy926 cells were cultured on Matrigel-coated plates to sub-confluent, confluent, and quiescent states. The cells were treated with DHA to establish concentration (10 μM to 150 μM) and time course (10 min to 24 h) curves, with or without SB202190, a p38MAPK-specific inhibitor. The activation of p38MAPK and its downstream targets was quantified by Western blotting. Histone H3 modifications upon DHA treatment were tested with an ELISA-based kit. Results The activation of p38MAPK by DHA in EA.hy926 cells was concentration-, time-, and growth state-dependent. Upon p38MAPK inhibition, activation of mitogen and stress activated kinase 1 (MSK1), a downstream target of p38MAPK, declined. This reduction was attenuated by low concentrations of DHA in quiescent endothelial cells but not confluent or sub-confluent cells. Since MSK1 can act on histone H3 and other chromatin binding proteins like cAMP response element-binding protein (CREB), H3 modifications were monitored. In the confluent state, DHA caused a 6-fold increase in total H3, with a concomitant decrease in most methylation, acetylation, and phosphorylation marks, including H3K9me1/3, H3K27me2, H3K36me1/3, H3K9ac, H3K18ac, H3S10ph, and H3S28ph. Conclusions This study showed that DHA may exert its effects in endothelial cells via the p38MAPK signalling pathway, and MSK1 may be a downstream effector possibly leading to epigenetic changes. The results provide novel insights regarding DHA's atheroprotective actions and identify new therapeutic targets with potential for treating atherosclerosis. Funding Sources Research Manitoba, St Boniface Hospital Foundation-Research Without Borders.