Introduction Increasing attention has focused on the therapeutic potential of green tea polyphenols owing to the anti-inflammatory, proapoptotic, and multiple other effects they have produced in a variety of mammalian cell lines and animal models of disease. (-)-Epigallocatechin-3-gallate (EGCG), a potent anti-inflammtory compound, makes up 40% of the polyphenolic fraction of green tea. ECGC has also proven to be a potent inhibitor of nuclear factor κB (NF-κB), a key regulatory factor in the control of intestinal inflammation. We investigated whether the anti-inflammatory effects of EGCG also apply to human peripheral blood monocytes.
Aim To evaluate the ability of EGCG to inhibit the production of proinflammatory cytokines and to induce apoptosis in human PBMCs in vitro in the presence or absence of lipopolysaccharide (LPS).
Methods PBMCs were preincubated for 1 hour in the presence of increasing concentrations of EGCG (0-125 ng/μL) prior to incubation for 16 hours with 10 ng/mL LPS. The anti-inflammatory effect of EGCG was measured by the reduction of TNF-α production, as well as other inflammation-related cytokines, in LPS-stimulated cells. The effect of ECGC on apoptosis of LPS-activated PBMCs was measured by annexin V-FITC/PI (propidium iodide) staining. UV light exposure was used as a positive control for apoptosis studies.
Results In the absence of EGCG, LPS increased TNF-α production from a mean of 186 pg/mL to 503 pg/mL. Pretreatment of cells with increasing concentrations of EGCG (5-125 ng/μL) proportionally reduced TNF-α production by 20%, 33%, 38%, and 45% at 5, 25, 50, and 125 ng/μL EGCG, respectively. Reductions in TNF-α production were not caused by loss of viability of cultured cells, as assessed by mitochondrial reduction of MTT. An additional indicator of viability, interferon-γ production, was unaffected by the administration of EGCG until the highest concentration was reached. EGCG did increase apoptosis in a concentration-dependent fashion from 1.5% in untreated cells to 55% at the highest tested concentration.
Discussion This study demonstrates the ability of EGCG to induce anti-inflammatory effects on human PBMCs and the ability of EGCG to induce apoptosis in LPS-stimulated human PBMCs. In light of the fact that tissue macrophages constitute a significant source of proinflammatory cytokines in inflammatory bowel disease (IBD), we will proceed to investigate the effects of EGCG in vitro and in vivo on peripheral blood monocytes and mucosal tissue macrophages from human subjects with IBD.
Work supported by K23 DK073750-02 (G.W.D.) and K23 AA014235 (M.W.L.).
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