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Comparative Study
, 163 (3), 1147-56

Foreign Body-Type Multinucleated Giant Cell Formation Is Potently Induced by Alpha-Tocopherol and Prevented by the Diacylglycerol Kinase Inhibitor R59022

Affiliations
Comparative Study

Foreign Body-Type Multinucleated Giant Cell Formation Is Potently Induced by Alpha-Tocopherol and Prevented by the Diacylglycerol Kinase Inhibitor R59022

Amy K McNally et al. Am J Pathol.

Abstract

Multinucleated foreign body giant cells (FBGCs) form by monocyte-derived macrophage fusion on implanted biomedical devices and are believed to mediate oxidative damage to biomaterial surfaces. Our in vitro system of human macrophage culture and interleukin (IL)-4-induced FBGC formation was developed to study the macrophage fusion mechanism and the physiological significance of FBGCs on implanted biomaterials and at other sites of chronic inflammation. Here, we demonstrate that the antioxidant vitamin E (90% alpha-tocopherol) moderately induces macrophage fusion and increases IL-4-induced FBGC formation. Moreover, purified alpha-tocopherol, but not beta-, gamma-, or delta-tocopherol, most remarkably induces macrophage fusion, leading to cultures of confluent FBGCs below normal plasma concentrations. This is not observed with the similar antioxidants probucol or Trolox, suggesting that the alpha-tocopherol effects on FBGC formation are independent of its antioxidant activity. Consistent with the reported activation of diacylglycerol kinase by alpha-tocopherol, the diacylglycerol kinase inhibitor R59022 completely abrogates FBGC formation. R59022 inhibition of IL-4-induced FBGC formation is reversed by alpha-tocopherol, suggesting that FBGC formation involves diacylglycerol kinase activation. This study suggests a novel role for diacylglycerol kinase in the mechanism of macrophage fusion/FBGC formation at sites of chronic inflammation and reveals that the pleiotropic lipophilic compound, alpha-tocopherol, is a highly potent macrophage fusion factor.

Figures

Figure 1.
Figure 1.
Effects of purified tocopherols on FBGC formation. a: Dose dependency of α-tocopherol. b: Comparison of tocopherols. The indicated concentrations of purified α-tocopherol (circles), β-tocopherol (squares), γ-tocopherol (triangles), or δ-tocopherol (diamonds) were added to day 3 macrophages, and the cultures were fixed and stained with May-Grünwald/Giemsa on day 7. Results are expressed as percent FBGC formation ± SEM, n = 3 monocyte donors that are different for a and b.
Figure 2.
Figure 2.
Vitamin E- or α-tocopherol-induced FBGC formation, and reversal of R59022 inhibition of IL-4-induced FBGC formation by α-tocopherol. Day 3 macrophages were otherwise untreated (a) or treated with 50 μmol/L vitamin E (b), 50 μmol/L α-tocopherol (c), 10 ng/ml IL-4 (d), 10 ng/ml IL-4 plus 10 μmol/L R59022 (e), or 10 ng/ml IL-4 plus 10 μmol/L R59022 plus 100 μmol/L α-tocopherol (f), and fixed and stained with May-Grünwald/Giemsa on day 7. Scale bar, 100 μm.
Figure 3.
Figure 3.
Effects of purified tocopherols on IL-4-induced FBGC formation. The indicated concentrations of purified α-tocopherol (circles), β-tocopherol (squares), γ-tocopherol (triangles), or δ-tocopherol (diamonds) were added to day 3 macrophages together with 10 ng/ml of IL-4, and the cultures were fixed and stained with May-Grünwald/Giemsa on day 7. Results are expressed as percent FBGC formation ± SEM, n = 3 monocyte donors.
Figure 4.
Figure 4.
Effect of R59022 on α-tocopherol-induced FBGC formation. The indicated concentrations of purified α-tocopherol were added to day 3 macrophages in the absence (circles) or the presence of 5 μmol/L (squares) or 15 μmol/L (triangles) of R59022, and the cultures were fixed and stained with May-Grünwald/Giemsa on day 7. Results are expressed as percent FBGC formation ± SEM, n = 3 monocyte donors.
Figure 5.
Figure 5.
Effect of R59022 on macrophage/FBGC adhesion and FBGC formation in IL-4-treated cultures. The indicated concentrations of R59022 were added to day 3 macrophages, and the cultures were fixed and stained with May-Grünwald/Giemsa on day 7. Results are expressed as percent adhesion (circles) or percent FBGC formation (squares) ± SEM, n = 3 monocyte donors.
Figure 6.
Figure 6.
Effect of R59022 on IL-4-induced FBGC formation, and reversal of R59022 inhibition of IL-4-induced FBGC formation by α-tocopherol. The indicated concentrations of purified α-tocopherol were added to day 3 macrophages together with 10 ng/ml of IL-4 in the absence (circles) or the presence of 5 μmol/L (squares), 10 μmol/L (triangles), or 15 μmol/L (diamonds) of R59022, and the cultures were fixed and stained with May-Grünwald/Giemsa on day 7. Results are expressed as percent FBGC formation ± SEM, n = 3 monocyte donors.

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