The immunomodulatory effect of docosahexaenoic acid (DHA) on the RAW264.7 cells by modification of the membrane structure and function

Abstract

Background: DHA can regulate various physiological functions of cells. Our group has clarified the immunomodulatory activity and molecular mechanism of DHA on RAW264.7 cells. However, the effect of DHA on the membrane fatty acid environment and the activation of signaling pathways on the cell membrane is still not clear. Methods: In this study, we evaluated the fluidity, the potential and the fatty acid, phospholipid and protein composition of the RAW264.7 cell membrane by DHA treatment. Results: The fluidity of the RAW264.7 cell membrane was increased by DHA treatment. The results of membrane potential analysis suggested that DHA (2.4 μM) significantly reduced the surface potential of the cell membrane, which might influence the fluidity of cell membranes. In addition, the fatty acids and phospholipids were measured and the results indicated that DHA treatment (2.4 μM) altered the lipid environment and the composition of phospholipids on the RAW264.7 cell membrane. Then the LC-MS/MS-based label free quantitative proteomics approach was applied to identify a total of 86 differential proteins in the 2.4 μM DHA and control groups (>2.0-fold change or <0.5-fold change in protein expression); these proteins are most frequently related to the cell response to stimuli and the response to stress. These results suggested that DHA could alter the fluidity, the potential, the fatty acid and phospholipid composition of the RAW264.7 cell membrane, eventually affecting the proteins of the cell membrane, especially the changes in Siglec 1, iNOS, GPR120, Ras and MEK expressions (validated by western blot analysis), which are likely associated with the activation of the intracellular signaling pathway in RAW264.7 cells by DHA treatment.