Modulation of Inflammatory Responses by Green and Black Tea in LPS-induced RAW 264.7 Cells

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Title: Modulation of Inflammatory Responses by Green and Black Tea in LPS-induced RAW 264.7 Cells
Author: Summers, Caroline Rueda
Advisors: Dr. Brenda P. Alston-Mills, Committee Member
Dr. Jonathan C. Allen, Committee Co-Chair
Dr. Gabriel Keith Harris, Committee Chair
Abstract: Recently, the World Health Organization estimated that chronic diseases are responsible for 46% of disease occurrence and 59% of all deaths worldwide. Growing evidence suggests that increasing chronic disease incidence is dependent upon several factors, including inadequate consumption of antioxidant-rich fruits and vegetables and chronic inflammation. Many researchers have previously investigated potential health benefits associated with consuming plant-based foods, and the ability of these foods to prevent or suppress the pathologies linked to chronic diseases. Specifically, plant-based bioactive compounds called polyphenols have been widely studied, and show promise as a therapeutic agent against chronic diseases. Tea is produced from the leaves of the Camellia sinensis plant and, aside from water, is the most popular beverage in the world. Green tea, black tea, and oolong tea are the three major types of commercially produced tea. Black tea comprises over 75% of global tea production and consumption, followed by green tea at approximately 20%, and oolong tea at less than 2%. For centuries, tea has been known for its health effects. It has historically been used as a medicine in China and Japan. Tea polyphenols are particularly thought to have chemopreventive and cardioprotective effects due to their antioxidant and anti-inflammatory properties. Previous in vitro and animal studies have analyzed the anti-inflammatory activity of polyphenols, and the specific mechanisms by which these compounds suppress inflammation. But, to our knowledge, few studies have investigated tea’s anti-inflammatory capacity as a whole food, or compared the potential anti-inflammatory effects of different tea types. Therefore, the objectives of the following study were to evaluate total phenol content in green tea (GT) and black tea (BT); to assess viability of cells exposed to GT, BT, and LPS; and to measure the modulation of inflammatory responses in lipopolysaccharide (LPS)-induced RAW 264.7 mouse macrophages by GT and BT. We predicted that GT and BT would differentially modulate inflammation because of differences in composition. First, the total phenolic content in commercially produced GT and BT was measured using the Folin-Ciocalteu method. Freshly prepared GT had a significantly higher phenolic content than freshly prepared BT (1317.1 + 6.0 GAE, 918.9 + 10.7 GAE). Stored at -80° C, the phenolic content of GT and BT significantly increased at one month (1770.0 + 35.2 GAE, 1124.0 + 19.1 GAE) and two months post-preparation (1587.2 + 21.5 GAE, 1003.2 + 8.6 GAE), then decreased at 3 months post-preparation (1407.8 + 13.4 GAE, 941.8 + 0.5 GAE) (p < 0.05). The second goal of our study was to assess viability in RAW 264.7 cells treated with GT, BT, and LPS. Viability was colorimetrically determined by the Trypan Blue assay and by measured absorbance values from the MTT assay. The GT, BT, and LPS treatments did not produce a cytotoxic effect, and cell viability values for each treatment were not significantly different (p < 0.05). The third objective was to measure prostaglandin E2 (PGE2) and cyclooxygenase-2 (COX-2), two major inflammatory mediators, in tea-treated, LPS-induced RAW 264.7 cells. We analyzed the inflammatory potency of LPS from two E. coli serotypes, 0111:B4 and 055:B5, and found that the latter serotype more significantly stimulated PGE2 production. When applied, GT and BT exhibited anti-inflammatory activity by similarly suppressing PGE2. Western blotting analysis revealed that only GT applied at the highest concentration (5%) significantly inhibited COX-2 expression relative to the positive control (p < 0.05). Further studies are needed to elucidate the exact mechanisms by which tea exerts anti-inflammatory activity on these inflammatory mediators.
Date: 2010-04-16
Degree: MS
Discipline: Nutrition

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