Docosahexaenoic Acid Exerts Anti-Inflammatory Action on Human Eosinophils through Peroxisome Proliferator-Activated Receptor-Independent Mechanisms

Abstract

Background: Despite the fact that previous studies have indicated the significant roles of polyunsaturated fatty acids (PUFAs) in the immune system through peroxisome proliferator-activated receptor alpha (PPARα) and PPARγ, the biological functions and the mechanisms of action in eosinophils are poorly understood. Methods: We investigated the functional effects of docosahexaenoic acid (DHA, n-3 PUFA) on human peripheral blood eosinophils, using in vitro systems to test the hypothesis that DHA negatively regulates eosinophil mechanisms through PPARα and PPARγ. Results: Eosinophil apoptosis that spontaneously occurs under normal culture conditions was accelerated in the presence of DHA. In addition, eotaxin-directed eosinophil chemotactic responses were inhibited by pretreatment with DHA, disturbing both the velocity and the directionality of the cell movement. Pharmacological manipulations with specific antagonists indicated that the effects of DHA were not mediated through PPARα and PPARγ, despite the presence of these nuclear receptors. DHA also induced Fas receptor expression and caspase-3 activation that appears to be associated with a proapoptotic effect of DHA. Further, DHA rapidly inhibited the expression of eotaxin receptor C-C chemokine receptor 3 and eotaxin-induced calcium influx and phosphorylation of extracellular signal-regulated kinase. Interestingly, these inhibitory effects were not observed with linoleic acid (n-6 PUFA). Conclusions: The data might explain one of the mechanisms found in previous research showing the favorable effects of n-3 PUFA supplementation on allergic diseases, and provide novel therapeutic strategies to treat eosinophilic disorders.

Adiponectin attenuates human eosinophil adhesion and chemotaxis: implications in allergic inflammation

“…Informed consent was obtained from each subject, and the study protocols were approved by the Ethics Committee of Akita University School of Medicine. Cells were isolated as previously described [21][22][23][24][25][26]. In brief, granulocytes were isolated by sedimentation with 6% dextran followed by centrifugation on 1.088 Percoll (Pharmacia, Uppsala, Sweden) density gradients.…”

“…The stained cells were analyzed using a flow cytometer (FACScan; Becton Dickinson Immunocytometry Systems, San Jose, CA). The expression of CCR3 was assessed as the ratio of the MFI of the sample and the isotype-matched control [25].…”

“…An apoptosis detection kit (Medical Biological Laboratories, Nagoya, Japan) was used to quantitatively determine eosinophils undergoing apoptosis by virtue of their ability to bind to annexin V and propidium iodide (PI) [23,25]. Briefly, eosinophils (1 Â 10 6 cells/mL) were incubated with vehicle or recombinant human adiponectin (R&D Systems, Minneapolis, MN) in RPMI 1640 (Invitrogen, Grand Island, NY) with 10% FBS at 37 C in humidified air with 5% CO 2 for 48 h. The eosinophils were washed and stained with annexin V and PI according to the manufacturer's instructions.…”

“…The cells that migrated and adhered to the lower surface of the membrane were counted for 10 fields by light microscopy. We also investigated chemotaxis using a TAXIScan system (GE, Tokyo, Japan) as previously described [25,26]. In brief, the cells were preincubated with vehicle or adiponectin (1 mg/mL) for 60 min at 37 C, after which aliquots of 1 mL of the cell suspension were injected into one compartment.…”

“…Since transmembrane-migrated adherent cells were measured in our Boyden chamber system, it might have been due to simple inhibition of cell adhesion. To further evaluate the inhibitory effect of adiponectin, we used the TAXIscan system, which is capable of detecting real-time horizontal migration of cells [25,26,29]. The response toward eotaxin was clearly visualized in eosinophils ( Figure 3E).…”