EGCG Group Research Articles

The effects of EGCG supplementation on pancreatic islet α and β cells distribution in adult male mice

Tea and tea products are widely used as the most popular beverage in the world. EGCG is the most abundant bioactive tea polyphenol in green tea, which has positive effects on the prevention and treatment of diabetes. However, the impact of EGCG exposure on glucose homeostasis and islets in adult mice have not been reported. In this study, we studied glucose homeostasis and the morphological and molecular changes of pancreatic islet α and β cells in adult male mice after 60 d of exposure to 1 and 10 mg/kg/day EGCG by drinking water. Glucose homeostasis was not affected in both EGCG groups. The expression of pancreatic duodenal homebox1 (Pdx1) in β cells was upregulated, which might be related to increased insulin level, β cell mass and β cell proliferation in 10 mg/kg/day EGCG group. The expression of aristaless-related homeobox (Arx) in α cells did not change significantly, which corresponded with the unchanged α-cell mass. The significant reduction of musculoaponeurotic fibrosarcoma oncogene homolog B (MafB) positive α-cells might be associated with decreased glucagon level in both EGCG groups. These results suggest that EGCG supplementation dose-dependent increases β cell mass of adult mice and affects the levels of serum insulin and glucagon. Our results show that regular tea drinking in healthy people may have the possibility of preventing diabetes.

Does Epigallocatechin Gallate as a Radiation Protective Agent Reduce the Anti-Tumor Effect of Radiotherapy in Postoperative Breast Cancer Radiotherapy?

Based on the previous encouraging results, we further explored whether EGCG would have a protective effect on potential tumor lesions, that is, reduce the efficacy of radiotherapy. We selected patients with stage III breast cancer with or without EGCG. The local control rate, distant metastasis rate, DFS and OS were compared between the two groups. Patients with stage III breast cancer who were treated with EGCG and radiotherapy was selected from a phase II clinical study (ClinicalTrials.gov, No. NCT02580279). Each patient was matched with one control patient without EGCG From the medical database of our hospital matching for age and stage. The control group of stage-and age-matched patients was selected at random from the medical database of our hospital RESULTS: We identified 43 EGCG patients and 43 matched controls. The median age was 45 years (range: 26-67). Between the two groups, there were no obvious differences in the baseline demographic or clinical features. When compared to the placebo group, the mean radiation-induced dermatitis index (RIDI) in the EGCG group was substantially lower (2.56±1.14 vs 3.36±1.16 T = -3.232, P = 0.002). Repeated measures ANOVA indicated the significant differences in the RTOG score during the course of radiotherapy between the two groups (F = 9.611 P = 0.003). The patients mostly experienced RID two or three weeks after starting radiotherapy, although in the EGCG group, it appeared later (3.19±0.91 weeks) than it did in the placebo group (2.67±0.84 weeks), P = 0.008. The median follow-up for patients in the EGCG group at the time of data collection was 50.6 months with 95% confidence interval (CI) from 43.9 to 57.3. While it was 48.6 months (95% CI: 43.4-53.9) for patients in the control group. There was no significant difference in overall survival (OS), disease free-survival (DFS) and freedom from locoregional (LRF) and distant failure (DMF) (P > 0.05). At the data cut-off (December 2021), the 4-year DFS with EGCG was 71.4% compared to 65.4% with conventional therapy, and the 4-year OS was 77.0% compared to 80.3%. The prophylactic use of EGCG solution reduced the RID score of stage III breast cancer patients without negatively impacting the therapeutic effect of radiotherapy on the tumor. EGCG is safe and feasible choice for RID for breast cancer during radiotherapy.

Effects of humic substances with urea on protozoal population and fermentation in the rumen of sheep

The aim was to investigate the effects of humic substances (HS) combined with urea, on pH, ammonia, total volatile fatty acids (∑VFA), protozoal population, amylolytic and cellulolytic activities in rumen fluid (RF) of 12 female crossbred merino sheep (n = 6 in experimental EG and control CG groups). The basic daily ration consisted of 1.25 kg grass hay, 0.25 kg cereal grains – equally wheat and barley (daily intake of crude protein 113.45 g, crude fibre 394.19 g, crude fat 17.27 g and ash 71.85 g) and 10 g urea. HS were applied at 20 g.day-1 per animal in EG on day 2–18. Increased protozoal cells (P<0.05) of Entodinium spp. by 243.19, of Diplodinium spp. by 5.11 and of Ophryoscolex spp. by 2.06 103.mL-1 were observed in EG on day 3. The effects on the amylolytic and cellulolytic activities as well as pH values were not significant. Ammonia was higher (P<0.05) in EG by 5.78 at 3 h and by 12.75 at 6 h on day 2, by 10.62 at 9 h on day 3, and by 7.84 mg.100 mL-1 at 6 h on day 4 after feeding. ∑VFA were increased (P<0.05) by 3.89 mmol.L-1 at 3 h on day 18 after feeding. The feed intake of HS combined with urea had significantly positive effects on the course of ammonia and protozoal population in RF.

Influence of green tea extract in the color of composite resin restorations

The aim of this study was to evaluate the color change of composite resin restorations in Class I cavity preparation with different depths, submitted to challenge of thermocycling in coffee, after the use of green tea extract (EGCG) as treatment on the dentin surface. Forty-eight human molars were divided into 6 groups according to dentin treatment and depth of restoration (n = 8): Group C3- Control/3 mm; Group C4- Control/4 mm; Group C5- Control/5 mm; Group EGCG3- EGCG/3 mm; Group EGCG4- EGCG/4 mm; and Group EGCG5- EGCG/5 mm. The teeth of the control groups were restored by the bulk fill technique (Filtek Bulk Fill), conditioning the dentin surface only with universal bonding system (Single Bond Universal). The teeth of the EGCG groups were also restored by the bulk filling technique, but conditioning the dentin surface with 0.5% EGCG for 30 s prior to the application of the adhesive system. Initial and final color readings were performed according to the CIE L*a*b* scale in UV-2450 spectroscope, before and after challenge of thermal cycling in coffee. The color change (ΔE) was then calculated based on the formula ΔE = [(ΔL*)2+(Δa*)2+(Δb*)2]½. The ΔE data were submitted to statistical tests of normality, two-way ANOVA and Tukey test to compare the means (p < 0.05). There was no statistically difference for both study factors analyzed (EGCG application and restoration depth), as well as the interaction between both, after aging in coffee (p > 0.05). It was concluded that the previous application of EGCG did not cause a significant color change at the dentin-resin interface.

Fructus mume Extracts Alleviate Diarrhea in Breast Cancer Patients Receiving the Combination Therapy of Lapatinib and Capecitabine.

Lapatinib and capecitabine have been widely used in the therapy of breast cancer. However, long-term use of lapatinib and capecitabine often causes the most common side effect diarrhea, which limit the medicine use. Fructus mume (F. mume) has been proved to be effective to treat chronic diarrhea with few side effects. The compounds from F. mume were extracted by using an ethanol method. Extracts of F. mume (EFM) were analyzed by HPLC. We investigated the protective effects of EFM on the diarrhea caused by lapatinib and capecitabine. From March 1st, 2016 to June 1st, 2017, 208 breast cancer patients with diarrhea caused by lapatinib and capecitabine were recruited. The patients were evenly assigned into two groups: EG group (the patients took 100 mg EFM daily) and CG group (the patients took placebo daily). The effects of EFM on diarrhea and gastrointestinal symptoms were measured by a semiquantitative method seven-point Likert scale. Overall quality of life was measured by SF-36 questionnaire and Hospital Anxiety and Depression Scale (HADS). The HPLC analysis showed that there were three components in EFM, including citric acid, 5-hydroxymethylfurfural (5-HMF), and chlorogenic acid. Breast cancer types were observed by using Hematoxylin and eosin (H&E) stain. The breast cancer can be divided into leaflet, gland and fibroblast types. Patient age, skin metastases, treatment, and grade 1 diarrhea were significant risk factors associated with for grade 2 diarrhea. EFM reduced diarrhea and gastrointestinal symptoms by reducing the average scores of the diarrhea symptom and seven-point Likert scale, and improved life quality of patients significantly by improving SF-36 scores and reducing HADS scores when compared to that in the CG group after 6-week therapy and further 4-week follow-up (P < 0.05). EFM may be a potential choice for the diarrhea therapy in breast cancer patients.

Combined effects of resveratrol and epigallocatechin-3-gallate on post thaw boar sperm and IVF parameters

Frozen-thawed boar semen suffer a fertility decrease that negatively affects its widespread use. In recent years supplementing frozen-thawed boar sperm with different antioxidants gave interesting and promising results; the aim of the present work was to study the effect of supplementing boar sperm thawing medium for 1 h with combination of epigallocatechin-3-gallate (EGCG, 50 μM) and Resveratrol (R, 2 mM), on boar sperm motility (assessed by CASA), viability, acrosome integrity, mitochondrial function, lipid peroxidation and DNA integrity (assessed by flow cytometry), protein tyrosine phosphorylation (assessed by immunofluorescence) and on in vitro fertilization (IVF).Our results demonstrate that sperm motility is negatively affected by R (alone or associated with EGCG, p < 0.05) in comparison to control and EGCG groups both at 1 h and 4 h; this effect is evident both in average motility parameters and in single cells kinematics, studied by cluster analysis, that showed the presence of a specific cell population with simil-hyperactivated features in R group (p < 0.01).Viability, acrosome integrity, mitochondrial functionality and lipid peroxidation are not influenced by the addition of the antioxidants; finally, DNA integrity is negatively influenced by R (both alone or associated with EGCG) both at 1 h and 4 h incubation (p < 0.05). Finally, tyrosine phosphorylated protein immunolocalization, used as capacitation parameter, is not affected by the different treatments.Penetration rate is strongly enhanced by R, both alone or associated with EGCG (p < 0.05); EGCG increases penetration rate as well but to a lower extent.Our findings demonstrate that the combination of R and EGCG could positively affect frozen-thawed boar sperm fertility in vitro; the effect is evident also in R groups, thus demonstrating that this antioxidant is predominant, and no synergic effect is present. Some insights are needed to understand if, in particular R (that showed the strongest effect) could be profitably used for artificial insemination in vivo, given the detrimental effect of this molecule on both sperm motility and DNA integrity.

Epigallocatechin-3-gallate prevents Non-alcoholic fatty liver disease by modulating liver function, lipid profiles and M1/M2 macrophage polarization

Abstract Background Here, we examine the therapeutic efficacy of Epigallocatechin-3-gallate, the most potent catechin in green tea, on nonalcoholic fatty liver disease (NAFLD), a growing epidemic in humans. Methods NAFLD was induced by high-fat diet (HFD) with 30% fructose in drinking water for 8 weeks. Mice were 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). Liver histology, serum lipid prolife, liver function as well as liver M1/M2 polarization were evaluated. Results At the end of the experiment, the mean body weight of LE and HE mice were 39.7g and 45.7g, while it was 51.9g in the MD group (P&amp;lt; 0.05). Similarly, liver weights in the LE and HE groups (means = 1.44g, and 1.68g, respectively) were lower than that in MD (2.94g, P &amp;lt; 0.01). Liver dysfunction observed in the MD group (mean AST=133.2 U/L and ALT=183.4 U/L), was significantly improved in both LE (44.8 U/L, 86.8 U/L, respectively, P &amp;lt; 0.001) and HE groups (83.63 U/L, 117.6 U/L, respectively P &amp;lt; 0.05). Serum cholesterol, HDL, triglyceride levels were increased in the MD group, but lowered in both LE and HE mice (p&amp;lt;0.01). Hepatic FACS, RT-PCR and IHC studies demonstrated that the HE group had lower F4/80+ CD86+ (0.3 ×103 vs 1.5 × 104), F4/80+ MHC II+ (0.7 ×104 vs 1.3 × 104), and higher F4/80+ CD206+(2.96 ×104 vs 0.3 × 104), F4/80+ CD23+ (1.27 ×104 vs 0.65 × 104) hepatic infiltration, accompanied by down-regulated CD68 and up-regulated CD163 mRNA expression. Conclusion EGCG subdued NAFLD despite consumption of HDF and high fructose water, marked by improved liver function and serum lipid profile, as well as M1/M2 polarization. EGCG should be investigated as a therapeutic modality in human NAFLD.

Inflammation and oxidative stress in corneal tissue in experimental keratitis due to Fusarium solani: Amelioration following topical therapy with voriconazole and epigallocatechin gallate.

Combined antifungal and antioxidant therapy may help to reduce oxidative stress in fungal keratitis. Experimental Fusarium solani keratitis was induced by application of F.solani conidia to scarified cornea (right eye) of 16 rabbits (another four rabbits were negative controls [Group I]). Five days later, F.solani-infected animals began receiving hourly topical saline alone (Group II), voriconazole (10mg/mL) alone (Group III), epigallocatechin gallate (EGCG, 10mg/mL) alone (Group IV) or voriconazole and EGCG (Group V). Twenty days post-inoculation, corneal lesions were graded. After animal sacrifice, excised corneas underwent histopathological and microbiological investigations. Corneal tissue levels/activities of interleukin 1 beta (IL-1β) and tumour necrosis factor alpha (TNF-α) gene mRNA transcripts, matrix metalloproteinase (MMP) 2 and 9 proteins, malondialdehyde (MDA) and reduced glutathione (GSH), and superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), were also measured. Clinical and histopathological scores (severity of corneal lesions; [P<.05]) and mean levels (P<.05) of IL-1β and TNF-α mRNA transcripts, MMP 2, MMP 9 and MDA were Group II>Groups IV and III>Groups V and I. Mean SOD, CAT, GPx and GSH levels (P<.05) were Group II<Groups IV and III<Groups V and I. Topical voriconazole with EGCG apparently reduces inflammation in experimental F.solani keratitis, as manifested by improved clinical, histological, microbiological and molecular parameters.

Role and mechanism research on reversal of 5-fluorouracil resistance by epigallocatechin gallate in gastric cancer drug-resistance cells lines SGC-7901/5-FU

Drug-resistance gastric cancer cell line (SGC-7901/5-FU) was established by high doses of repeated impact joint drug concentration increment methods. The cell viability of the parent cell line and the drug-resistance cell line were determined by standard MTT assay. Cell survival rate of drug-resistance was calculated by the formula [(A490 of the treatment group / A490 of the control group) × 100%]. Cell half inhibitory concentration (IC50) and resistance index (RI) were calculated by the Graphpad prime 6.0 software(RI=IC50 value of drug-resistance cells / IC50 value of parent cells). The apoptosis rate of SGC-7901/5-FU cells was quantified by flow cytometry after staining with annexin-V and PI. Western blot was used to detect the protein expression of drug-resistance-related proteins (ABCG2, P-gp, MDR-1 and GST-π) and apoptosis-related proteins (PARP, Survivin, Bax and bcl-2). IC50 value was significantly increased in drug-resistant cells compared with parental cells [(64.7±3.9) mg/L and (4.1±0.3) mg/L, respectively, t=26.46, P=0.000], and the RI was 15.6. Proliferation activity in the drug-resistant cells was higher than that in parental cells at different 5-FU concentrations (all P<0.05). In drug-resistant cells, the IC50 value of 5-FU combined with EGCG group obviously decreased compared with 5-FU group [(7.3±0.1) mg/L and (63.1±1.4) mg/L respectively, t=40.84, P=0.000], and the RI was 0.12. Proliferation activity in drug-resistant cells was significantly decreased after EGCG treatment at different 5-FU concentrations (all P<0.05). Cell apoptosis rates in control group, 5-FU group, EGCG group and 5-FU combined with EGCG group were (3.0±1.0)%, (7.0±1.3)%, (6.0±1.2)% and (18.0±1.4)%, while apoptosis rate in 5-FU combined with EGCG group was significantly higher than those of other 3 groups(F=129.5, P=0.000). Western blot revealed that after EGCG treatment, the expression levels of drug-resistance-related proteins (ABCG2, P-gp, MDR-1 and GST-π) in the drug-resistant cell line SGC-7901/5-FU decreased significantly; the expression levels of apoptosis marker protein PARP and pro-apoptotic protein Bax increased significantly; and the expression levels of anti-apoptotic protein Survivin and Bcl-2 decreased significantly (all P<0.05). EGCG can reduce the resistance of gastric cancer resistant cell line SGC-7901/5-FU, whose role may be via the inhibition of the expression of drug-resistance-related proteins, and the elevation of the protein expression ratio of PARP/Survivin and Bax/Bcl-2.

Effect of epigallocatechin gallate, green tea extract and chlorhexidine application on long-term bond strength of self-etch adhesive to dentin

This study evaluated the effect of dentin pretreatment with metalloproteinase inhibitors on long-term microtensile bond strength (μTBS) of self-etch adhesive (SEA) to dentin. Thirty-two dentin fragments received pretreatments (n=8): C: control (not treated); GT: 2% green tea extract; CLX: 2% chlorhexidine; EGCG: 2% epigallocatechin gallate. SEAs (Clearfil SE Bond) and resin composite were applied to dentin surface. After 24h, resin-blocks were cut into sticks submitted to μTBS testing immediately or after 6 months of storage in water. Two-way ANOVA and Tukey test (α=5%) indicated that CLX group showed μTBS values statistically superior to the other groups at immediate testing, whereas those of EGCG group were the lowest (p=0.018). After 6 months, a significant increase in μTBS values was observed for all groups, except for CLX (p=0.018). In conclusion, μTBS increased in dentin pretreatment with EGCG and green tea groups, and in the non-treated group.

Protective effect of acetylated epigallocatechin gallate on melanocytes from oxidative stress-induced damage

Objective To investigate the protective effect of acetylated epigallocatechin gallate (AcEGCG) against H2O2 - induced oxidative damage to human epidermal melanocytes, and to explore its possible mechanism. Methods Human epidermal melanocytes were isolated and cultured in vitro. Some melanocytes were classified into a H2O2 group induced by H2O2 only, EGCG groups and AcEGCG groups induced by H2O2 after pretreatment with different concentrations of EGCG and AcEGCG, respectively. Three concentrations (10, 20 and 40 μmol/L) of EGCG or AcEGCG were used to treat melanocytes for 1 hour in MTS assay and lactate dehydrogenase (LDH) leakage assay and for 2 hours in Western blot assay, while only one concentration (40 μmol/L) was used to treat melanocytes for 0.5, 1, 2 and 4 hours respectively in flow cytometry assay. Some melanocytes treated with only culture medium and 0.1% dimethyl sulphoxide (DMSO) served as the control group. After additional culture, MTS assay was performed to determine cell survival rate, flow cytometry to detect the level of reactive oxygen species (ROS) in melanocytes, Western blot to measure the expressions of caspase-9 and caspase-3 proteins. Lactate dehydrogenase (LDH) kit was used to detect the leakage of LDH to culture medium. Statistical analysis was carried out by using one-way analysis of variance for comparisons of multiple group means followed by Student-Newman-Keuls-q (SNK-q) test for multiple comparisons. Results Compared with the control group, the H2O2 group showed significantly decreased cell survival rate (22.99% ± 0.53%, P < 0.01), but increased LDH leakage level (36.58% ± 0.73%, P < 0.01), intracellular ROS level (19.08 ± 0.57, P< 0.01), as well as caspase-9 (2.65 ± 0.079, P< 0.01) and caspase-3 (2.36 ± 0.057, P< 0.01) expressions. In comparison with the H2O2 group, the cell survival rate was significantly higher in the 10-, 20- and 40-μmol/L AcEGCG groups (79.50% ± 3.62%, 86.52% ± 5.13%, 97.81% ± 5.21%, respectively, all P < 0.01) and EGCG groups (43.19% ± 1.68%, 63.34% ± 3.60%, 70.82% ± 2.1%, respectively, all P< 0.01). However, the 10-, 20- and 40-μmol/L AcEGCG groups and EGCG groups all showed a significant decrease in the expression levels of caspase-9 (AcEGCG groups: 1.44 ± 0.067, 1.26 ± 0.059 and 1.10 ± 0.072 respectively; EGCG groups: 2.31 ± 0.085, 2.13 ± 0.091 and 1.35 ± 0.064 respectively, all P < 0.05) and caspase-3 (AcEGCG groups: 1.70 ± 0.053, 1.57 ± 0.057 and 1.24 ± 0.068 respectively, all P < 0.05; EGCG groups: 2.09 ± 0.076, 1.98 ± 0.093 and 1.79 ± 0.056 respectively, all P< 0.05) compared with the H2O2 group. Similarly, a significant reduction was observed in the leakage level of LDH in these AcEGCG and EGCG groups (all P < 0.01) and in ROS levels in the 40-μmol/L AcEGCG and EGCG groups when compared with the H2O2 group. Conclusions AcEGCG has a stronger protective effect against H2O2-induced oxidative damage to human epidermal melanocytes compared with EGCG, which may be realized through clearance of free radicals, antioxidant effects, and decrease of caspase-9 and caspase-3 expressions. Key words: Melanocytes; Oxidative stress; Vitiligo; Caspase 3; Caspase 9; Epigallocatechin gallate

Oral administration of naturally occurring chitosan-based nanoformulated green tea polyphenol EGCG effectively inhibits prostate cancer cell growth in a xenograft model

In preclinical animal models, several phytochemicals have shown excellent potential to be used as effective agents in preventing and treating many cancers. However, the limited bioavailability of active agents could be one reason for their restricted usefulness for human consumption. To overcome this limitation, we recently introduced the concept of nanochemoprevention by encapsulating useful bioactive food components for their slow and sustained release. Here, we report the synthesis, characterization and efficacy assessment of a nanotechnology-based oral formulation of chitosan nanoparticles encapsulating epigallocatechin-3-gallate (Chit-nanoEGCG) for the treatment of prostate cancer (PCa) in a preclinical setting. Chit-nanoEGCG with a size of <200nm diameter and encapsulating EGCG as determined by dynamic light scattering and transmission electron microscope showed slow release of EGCG in simulated gastric juice acidic pH and faster release in simulated intestinal fluid. The antitumor efficacy of Chit-nanoEGCG was assessed in subcutaneously implanted 22Rν1 tumor xenografts in athymic nude mice. Treatment with Chit-nanoEGCG resulted in significant inhibition of tumor growth and secreted prostate-specific antigen levels compared with EGCG and control groups. In tumor tissues of mice treated with Chit-nanoEGCG, compared with groups treated with EGCG and controls, there was significant (i) induction of poly (ADP-ribose) polymerases cleavage, (ii) increase in the protein expression of Bax with concomitant decrease in Bcl-2, (iii) activation of caspases and (iv) reduction in Ki-67 and proliferating cell nuclear antigen. Through this study, we propose a novel preventive and therapeutic modality for PCa using EGCG that addresses issues related to bioavailability.

Catechin modulates inflammation in TNBS induced colitis. (P6238)

Abstract This study assessed the effect of catechin in modulating the inflammation of colitis. Thirty rats were randomized into three groups: Group I: chemically induced with colitis and received no treatment, Group II: chemically induced with colitis and received daily injection of EGCG, Group III: control group provided with water only. In groups I and II, colitis was induced by TNBS solution. Rats were sacrificed on days 3, 10 and 17 and biopsies of distal colons were assessed by histology and molecular techniques for ROS and changes in cytokines like IL-6 and IL1-β. Colitis induced by TNBS and non-treated with EGCG group, showed high inflammatory scores resulting in colonic mucosa and submucosa damage, necrosis, absence of goblet cells, infiltration of inflammatory cells, crypt abscesses and granulomas. Infiltration of inflammatory cells and tissue damage peaked at day 17 after the induction. At day 3 and day 10, the treated rats with EGCG showed amelioration in the severity of inflammation, but at day 17, the state of the colonic crypts and goblet cells was close to normal. The number of mast cells in the colon of treated rats decreased by more than 30% at day 17 compared to the non- treated rats. In addition, the expression of ROS decreased significantly. IL-1B and IL-6 expressions decrease with time by 50% in the treated group. Our data confirmed the anti-inflammatory effect of EGCG in TNBS-induced colitis and provided a clue to a potential therapeutic agent.

Anti-Edema Effect of Epigallocatechin against Acute Spinal Cord Injury in Rats and its Correlation with the P38MAPK/NF-κb Signaling Pathway

Study Design: This study investigated the outcome of epigallocatechin (EGCG) against edema after spinal cord injury (SCI). Objective: To perform the evidence correlation of EGCG with p38MAPK\NF-κB\AQP4 signaling pathway, also explore the effect of EGCG on edema after SCI, and clarify the possible mechanism of the EGCG reducing the edema after SCI. Summary of background data: Although some studies show that EGCG have a protective effect on nerve cells, there are few documents covering the relation between EGCG and SCI. On the basis of confirming significant inhibition of EGCG against edema after SCI, our research group intended to take the lead in further demonstrating the mechanism of anti-edema of EGCG against SCI. Methods: 160 rats were randomly divided into sham - surgery, SCI, EGCG and specific p38MAPK inhibitor SB203580 treatment groups, every group contained 40 rats. Acute SCI models were established in rats by vascular clip method. We used sensitive ELISA to examine the level of TNF-α and IL-1β, Western blot to examine the expression of p38MAPK/NF- κB/AQP4 signaling pathway related protein, the change of water content in spinal cordto evaluate the effect of EGCG against edema after SCI therapy. Results: Both EGCG and SB203580 groups could significantly reduce the releasing of TNF-α and IL-1β (P<0.05), while EGCG had a better effect (P<0.05), thus EGCG could significantly inhibit the phosphorylation status of p38MAPK reduce the expression of NF-κB p65 and AQP4 (P<0.05), and decrease the water content after SCI (P<0.05), similar to SB203580. Conclusion: The experimental findings indicate that EGCG can protect secondary SCI by potential mechanism of regulating p38MAPK\NF -κB\AQP4 signaling pathwayand thus reduce edema after SCI.

Effects of cocktail therapy on cell proliferation and Collagen I and TIMP-1 mRNA expression in rat hepatic stellate cells

AIM: To investigate the effects of cocktail therapy on cell proliferation and collagen I (Col I) and tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNA expression in rat hepatic stellate cells. METHODS: Hepatic stellate cells (HSC-T6) cultured in vitro were exposed to taurine (taurine group), epigallocatechin gallate (EGCG; EGCG group), genistein (genistein group), and a combination of these three drugs (cocktail-treated group) for 24 h, respectively. Cell proliferation was determined by methyl thiazol tetrazolium (MTT) assay. The expression of Col Ⅰ and TIMP-1 mRNAs was measured by semi-quantitative reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: Cell proliferation was significantly inhibited in the taurine group, EGCG group, genistein group and cocktail-treated group compared with the control group (0.237 ± 0.007, 0.216 ± 0.009, 0.242 ± 0.008 and 0.130 ± 0.004 vs 0.452 ± 0.011, respectively; all P＜0.05). The reduced rate of cell proliferation was more significant in the cocktail-treated group than in other treatment groups (all P＜0.05). The expression levels of Col I and TIMP-1 mRNAs markedly decreased in all treatment groups when compared with the control group (all P＜0.05). The expression levels of Col Ⅰ and TIMP-1 mRNAs were significantly lower in the cocktail-treated group than in other treatment groups (P＜0.05). CONCLUSION: Cocktail therapy significantly inhibits cell proliferation and down-regulates Col Ⅰ and TIMP-1 mRNA expression in HSCs. Cocktail therapy may be more effective than monotherapy in the treatment of liver fibrosis.

Development of a Whole-organism Model to Screen New Compounds for Sun Protection

We used zebrafish as a whole-organism model to screen new compounds for sun protection activity. First of all, we designed a series of UVB exposure experiments and recorded the phenotypic changes of zebrafish embryos. Results showed that 100 mJ/cm(2) of UVB given six times separated by 30 min intervals is the best condition. Fin malformation (reduced and/or absent fin) phenotypes are the most evident consequences after exposure to UVB. Each fin was affected by UVB, including pelvic, ventral, caudal, and dorsal fin, but pelvic fin seemed to be the most sensitive target after UVB exposure. We furthermore carried out "prevention" and "treatment" experiments using green tea extract and/or (-)-epigallocatechin (EGCG) to test this whole-organism model by observing the morphological changes of all fins (especially pelvic fin) after UVB exposure. Effects of UVB, green tea extract and EGCG on fin development were assessed using the Kaplan-Meier analysis, log-rank test and Cox proportional hazards regression. Results showed that a zebrafish pelvic fin in the UVB + green tea (treatment) group is 5.51 (range from 2.39 to 14.90) times, one in the UVB + green tea (prevention) group is 7.04 (range from 3.11 to 18.92) times, and one in the 25 ppm of EGCG (prevention) group is 22.19 (range from 9.40 to 61.50) times more likely to return to normal fin than one in the UVB only group. On the basis of these observations, we believe this model is effective for screening the higher stability and lower toxicity of new compounds, such as small chemicals which are derivative from EGCG or other dietary agents for sun protection.

차 폴리페놀화합물의 사이토카인 생성 및 항암능에 대한 영향

국산 미생물 발효차의 폴리페놀 색소성분들인 데아플라빈(TF)과 데아루비긴(TR) 및 EGCG를 macrophage cell line (RAW264.7에 적용하여 nitric oxide 합성 및 사이토카인 생성을 평가하였다. 사이토카인 생성은 TF, TR 및 EGCG를 RAW264.5 cell에 적용하였을 때, <TEX>$80\;{\mu}g/ml$</TEX> 농도에서 대조군과 LPS 촉진 처리에 비하여 nitric oxide 생성은 약 1.5배 증가하였다. IL-6, <TEX>$TNF-{\alpha}$</TEX> 및 GM-CSF는 TF, TR 및 EGCG 농도에 의존적으로 증가하였다. <TEX>$TNF-{\alpha}$</TEX> 생성은 크게 증가하였으며, 이는 TF, TR 및 EGCG가 사이토카인 생성을 통하여 면역증강 효과를 가질 것으로 나타났다 TF, TR 및 EGCG는 총 페놀 함량에 비례하여 항산화능을 나타내었으며, 암세포 증식을 유의적으로 억제하였다. 이들 폴리페놀물질의 억제효과는 그 성분들의 항암촉진작용 및 항산화활성에 의한 것으로 판단된다. Theaflavins (TF) and thearubigins (TR) are constituents of tea pigments which are polyphenols derived from Korean fermentation tea. After TF, TR and [(-) epigallocatechin-3-gallate](EGCG) have been applied to macrophage cell line (RAW264.7) nitric oxide (NO) synthesis and cytokines production were estimated. Cytokines production by enzyme linked immune-sorbent assay (ELISA) determined. NO production was increased by about 1.5-folds at the dose of <TEX>$80\;{\mu}g/ml$</TEX> compared to control and lipopolysaccharide (LPS) stimulation when TF, TR and EGCG were applied to a RAW264.7 cell. Interleukin-6 (IL-6), Tumor necrosis factor (<TEX>$TNF-{\alpha}$</TEX>) and granulocyte-macrophage colony stimulating factor (GM-CSF) increased depended on concentrations of TF, TR and EGCG. The production of tumor necrosis <TEX>$factor-{\alpha}$</TEX> increased highly in TR, TF and EGCG group with LPS. These results suggest that TF, TR and EGCG have immune-enhancement effect through the cytokine production. TF, TR and EGCG inhibited cancer cell viability, the anticancer effect of these polyphenols may explain the anti-tumor promotion action and antioxidant activity of these tea constituents.

Radiomodifying responses of vitamin E and epigallocatechin gallate supplementation during irradiation of implanted tumors

9601 Background: Dietary antioxidants may play an antagonistic role during radiation treatment (RT). Antioxidants such as vitamins C and E show radioprotective effects on normal cells in vitro but limited evidence in this regard is available in healthy tissue or tumor cells in vivo. Methods: Syngeneic, murine breast carcinoma (MCa-4) cells were implanted in hind legs of C3H mice. Vitamin E (VE) or epigallocatechin gallate (EGCG, derived from green tea extract) were fed at 500 and 658 mg/kg diet, respectively, for 4 wk until tumors reached 8 mm. Three tumor control dose 50 (TCD50) assays (N=48 mice/assay) were performed to quantify the radiomodifying effect of the antioxidants. Tumor growth data were collected prior to and after RT. Tissue concentrations of VE and EGCG were analyzed by HPLC. Tissue reactions were recorded for 90 d following RT. Angiogenesis specific gene arrays were used to analyze expression in tumor and cultured tumor cells. Results: A 4% increase in TCD50 was observed in mice fed either VE or EGCG supplemented diets relative to the control diet, though these changes were not statistically significant (P=0.30 and 0.20, respectively). The gamma-50 (slope) was 6.2 for control and 3.5 for both experimental arms (P=0.20). Prior to RT, tumor growth was 10% slower in the EGCG group compared to controls (P=0.002). VE concentration in the tumor was twice that of surrounding muscle (382 vs. 189 pmole/mg) but only slightly (4%) slowed tumor growth (P=0.24) prior to RT. There were fewer RT-related soft-tissue reactions (loss of irradiated limb below the tumor) in the EGCG (9.8%) and VE (7.1%) groups compared to control (23.8%) at P=0.09 and 0.04, respectively. EGCG increased VEGF and MMP gene expression in stromal tissue but decreased it in the tumor. Conclusions: TCD50 and gamma-50 were modestly increased in mice fed diets supplemented with VE or EGCG, but these changes were not statistically significant. EGCG significantly inhibited tumor growth rate. Adverse RT-related soft tissue reactions occurred less frequently with antioxidant supplementation, particularly with VE. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Harvard Medical School/Osher Institute; Goldman Philanthropic Partnerships

Effect of tea polyphenols and EGCG on nasopharyngeal carcinoma cell proliferation and the mechanisms involved

Objective: Tea polyphenols present in green tea show cancer chemopreventive effects in many tumor models. Epidemiological studies have also suggested that green tea consumption might be effective in the prevention of certain human cancers. In the present study, we investigated the molecular mechanisms of the inhibition of cell proliferation by tea polyphenols in nasopharyngeal carcinoma (NPC) cell line CNE1-LMP1. Methods: CNE1-LMP1 cells were treated with tea polyphenols at various doses (0, 25, 50, 100, 200 µg/ml) for 24 hours, the morphology of cells was observed by light microscopy, and cell survival rate was determined by MTT assay. At the same time, cell cycle of CNE1-LMP1 was analyzed by flow cytometry. Cyclin D1 transcription was analyzed by cyclin D1 promoterluciferase reporter system and expression of cyclin D1 protein by Western blot analysis. Transactivities of NF-κB and AP-1 was analyzed by Dual-fluorescence reporter gene system. Results: After treatment of CNE1-LMP1 cells with tea polyphenols, the number of proliferating cells was obviously decreased as determined by light microscopy and MTT assay (from 100% to 89.4%, 83.3%, 74.8% and 38.1%). With the increase of tea polyphenols concentrations, the number of cells in S-phase was obviously decreased, and the number of cells in G1-phase from 22.20% to 13.16%, and the number of cells in G0/G1 phase was elevated from 68.5% to 74.08%. It suggests that tea polyphenols could arrest the cell cycle at both of the two checkpoints. Furthermore, transcription and were obviously declined 7–8 folds (100–200 µg/ml tea polyphenols or EGCG group) and expression of cyclin D1 protein also decreased in a dose-dependent manner. Transactivities of NF-κB and AP-1 were obviously down-regulated in CNE1-LMP1 cells. Conclusion: Green tea polyphenols could inhibit cell proliferation, by suppressing the activity of NF-κB and AP-1, and by down-regulation of the transcription of cyclin D1.

A colorimetric study of coffee and tea staining of four composite resins : Gross M. D. and Moser J. B. J. Oral Rehabil.4 (1977) 311–322

The aim of this study was to evaluate the color change of composite resin restorations in Class I cavity preparation with different depths, submitted to challenge of thermocycling in coffee, after the use of green tea extract (EGCG) as treatment on the dentin surface. Forty-eight human molars were divided into 6 groups according to dentin treatment and depth of restoration (n = 8): Group C3- Control/3 mm; Group C4- Control/4 mm; Group C5- Control/5 mm; Group EGCG3- EGCG/3 mm; Group EGCG4- EGCG/4 mm; and Group EGCG5- EGCG/5 mm. The teeth of the control groups were restored by the bulk fill technique (Filtek Bulk Fill), conditioning the dentin surface only with universal bonding system (Single Bond Universal). The teeth of the EGCG groups were also restored by the bulk filling technique, but conditioning the dentin surface with 0.5% EGCG for 30 s prior to the application of the adhesive system. Initial and final color readings were performed according to the CIE L*a*b* scale in UV-2450 spectroscope, before and after challenge of thermal cycling in coffee. The color change (ΔE) was then calculated based on the formula ΔE = [(ΔL*)2+(Δa*)2+(Δb*)2]½. The ΔE data were submitted to statistical tests of normality, two-way ANOVA and Tukey test to compare the means (p < 0.05). There was no statistically difference for both study factors analyzed (EGCG application and restoration depth), as well as the interaction between both, after aging in coffee (p > 0.05). It was concluded that the previous application of EGCG did not cause a significant color change at the dentin-resin interface.