BackgroundPolyphenol catechins from green tea, particularly (−)-epigallocatechin-3-gallate (EGCG), exhibits numerous beneficial health effects, although the mechanisms remain unclear. MethodsIn this study, the mechanism of EGCG-mediated healing in an experimentally injured zebrafish model was examined at the cellular and molecular level using confocal microscopy and gene expression analysis. ResultsThe mechanisms of action of EGCG were shown to involve: (1) reducing neutrophil response (accumulation, travel speed, and distance) and (2) downregulating the expression of IL-1β, TNFα, and related signaling pathways. As determined by dynamic time-lapse tracking studies, the local accumulation of neutrophils with high migration speeds after wounding (n=33 cells, v=0.020 μm/s, d=37.8 μm), underwent significant reduction following treatment with EGCG doses of 300 μM (n=22 cells, v=0.013 μm/s, d=39.5 μm) and 600 μM (n=18 cells, v=0.008 μm/s, d=9.53 μm). Reverse transcription polymerase chain reaction studies revealed that several signature genes in the IL-1β, TNFα, and related signaling pathways were downregulated after EGCG treatment. ConclusionThe convenience, transparency, and simplicity of the zebrafish model facilitate tracking of fluorescent neutrophils in real time, in order to monitor inflammation, and assess the impact of therapeutic agents.

Background Given the potential side effects associated with conventional medical treatment of Inflammatory Bowel Disease (IBD), dietary supplements have been investigated by several groups. We tested if epigallocatechin gallate (EGCG) may be beneficial in IBD. Methods 36 C57BL/6J mice were subjected to dextran sulfate sodium (DSS)-induced IBD and divided into 3 groups: model group (MD; no EGCG), low dose EGCG group (LE, 20 mg/kg/d) and high dose EGCG group (HE, 50 mg /kg/d). Results Both high and low dose EGCG treatment alleviated clinical manifestations of IBD including body weight loss and disease activity index (DAI): HE: 3.3 and LE: 2.8 vs. 8.3 in MD (p < 0.001, n=8–10 mice/group). Macroscopic severity score (MSS) of HE and LE groups were 2.4, and 2.2, respectively, significantly lower than in MD (5.0, P < 0.01). The mean colon length of MD group was 5.8 cm; treatment with EGCG restored colon length (HE: 6.5 cm; LE: 6.4 cm). Intestinal permeability was improved as demonstrated by FITC-Dextran assay (HE: 3.28; LE: 2.85, vs. MD group: 6.69 ng/ml, P < 0.001) .The colitis histology score of HE (1.54) and LE (0.42) groups were significantly lower than that in MD (4.3, P < 0.001). In addition, EGCG treatment reduced colonic levels of pro-inflammatory cytokines: IL-6 (HE: 0.45; LE group: 0.41 vs MD: 0.89 ng/g tissue, P =0.03), MCP-1 (HE: 1.83; LE: 1.87, vs MD: 3.5 ng/g tissue P = 0.01) and TNF-alpha (HE: 8.87; LE: 10 vs MD: 21.7 ng/g tissue, P <0.001). Lower CD3+ T cell and CD68+ macrophage infiltration was also noted in HE and LE mice (P < 0.001), compared to MD controls. Conclusion These results underscore the therapeutic potential of EGCG in the treatment of IBD, which appear to be mediated through multiple inflammatory pathways.

Background The zebrafish represents the simplest and most convenient vertebrate model for undertaking large screens because of its small size, rapid reproduction time and transparency of the body. Here, we adopt a well-established model of inflammation in zebrafish to assess the impact of Epigallocatechin-3-Gallate (EGCG), the main catechin in green tea. Method Tg (MPX:GFP) or Tg (MPX:mCHERRY) transgenic zebrafish were used, bearing GFP- or mCherry-tagged neutrophils, respectively. 4dpf embryos were anesthetized, and inflammation was induced by sterile tailfin-resection. The inflamed groups were separated into those that were non-treated or exposed to EGCG from 0 – 600 uM for 6 hours. 10 fish per group were then mounted alive in methyl cellulose for wide-field fluorescence imaging and analysis was done using ImageJ. Animation of neutrophil migration or accumulation was recorded using an inverted fluorescence confocal microscope till 6-hours after resection. To examine the mechanistic basis of EGCG’s effect, 37 zebrafish inflammatory genes were studied using qPCR. Results The static accumulation of neutrophils at the resection site was highly elevated after wounding (n=32, p<0.0001), and this accumulation was reduced significantly by 31% and 43%, respectively, after exposure to EGCG 300 uM (p<0.05) and 600 uM (p<0.01). Dynamic neutrophil tracking studies are in progress. qPCR studies revealed 6 signature genes in the IL-1 beta pathway to be significantly down-regulated at least 2-fold after EGCG treatment. Conclusion EGCG suppresses inflammation in zebrafish through similar biochemical pathways as in other species. This novel animal model allows facile screening of inflammatory modulators in real-time.