Green Tea Epigallocatechin Gallate Research Articles

Green Tea Catechins Attenuate Human Primary Pterygium Cell Survival and Migration Via Modulation of ERK p42/p44 and p38 Pathways.

Pterygium belongs to an ocular surface disease with triangular-shaped hyperplastic growth, characterized by conjunctivalization, inflammation, and connective tissue remodeling. We previously demonstrated neoplastic-like properties of pterygium cells. Green tea catechin, (-)-epigallocatechin gallate (EGCG), has been shown to possess antitumorigenic properties; herein, we aimed to determine the effects of green tea catechins on human primary pterygium cell survival and migration and compared to that on patients' conjunctival cells. Both human primary pterygium and conjunctival cells expressed EGCG receptor, the 67 kDa laminin receptor. Seven-day treatment of green tea extract (Theaphenon E; 16.25 μg/mL) and EGCG (25 μM) attenuated pterygium cell proliferation by 16.78% (p < 0.001) and 24.09% (p < 0.001) respectively, without significantly influencing conjunctival cells. Moreover, green tea extract (16.25 μg/mL) and EGCG (25 μM) treatments also hindered pterygium cell migration by 35.22% (p < 0.001) and 25.20% (p = 0.019), respectively, but not conjunctival cells. Yet, green tea extract and EGCG treatments did not significantly induce pterygium cell apoptosis. Furthermore, green tea extract and EGCG treatments significantly increased the phosphorylation of p38 protein but reduced the phosphorylation of p42/p44 protein in pterygium cells. In summary, this study revealed that green tea extract and EGCG attenuated human primary pterygium cell survival and migration in vitro without damaging conjunctival cells, suggesting a novel potential therapeutic approach for primary pterygium treatment.

Potential role of green tea extract and epigallocatechin gallate in preventing bisphenol A-induced metabolic disorders in rats: Biochemical and molecular evidence

BackgroundBisphenol A (BPA) is an artificial chemical widely used in the production of polycarbonate plastics and epoxy resins. Accumulating evidence indicates that BPA exposure is associated with metabolic disorders. The beneficial effects of green tea and epigallocatechin gallate (EGCG), major catechin present in green tea, on alleviating BPA-induced metabolic disorders have been shown in various studies. PurposeProtective effects of green tea extract and EGCG on BPA-induced metabolic disorders and possible underlying mechanisms were investigated. MethodsRats were randomly divided into control, green tea extract (50 and 100 mg/kg, IP), EGCG (20 and 40 mg/kg, IP), BPA (10 mg/kg, gavage), BPA plus green tea extract (25, 50, and 100 mg/kg, IP), BPA plus EGCG (10, 20, and 40 mg/kg, IP), and BPA plus vitamin E (200 IU/kg, IP). After two months, body weight, blood pressure, biochemical blood tests, hepatic malondialdehyde (MDA), and glutathione (GSH) were assessed. By enzyme-linked immunosorbent assay, serum levels of insulin, leptin, adiponectin, TNFα, and IL-6, and by western blotting, hepatic insulin signaling (IRS-1, PI3K, Akt) were measured. ResultsBPA increased body weight, blood pressure, and MDA, decreased GSH, elevated serum levels of low-density lipoprotein cholesterol, total cholesterol, triglyceride, glucose, insulin, leptin, TNFα, IL-6, and liver enzymes including alanine aminotransferase and alkaline phosphatase, and lowered high-density lipoprotein cholesterol and adiponectin levels. In western blot, decreased phosphorylation of IRS-1, PI3K, and Akt was obtained. Administration of green tea extract, EGCG, or vitamin E with BPA reduced the detrimental effects of BPA. ConclusionThese findings indicate that green tea extract and EGCG can be effective in preventing or reducing metabolic disorders induced by BPA linked to their antioxidant and anti-inflammatory activity, regulating the metabolism of lipids, and improving insulin signaling pathways.

Suppressing effects of green tea extract and Epigallocatechin-3-gallate (EGCG) on TGF-β- induced Epithelial-to-mesenchymal transition via ROS/Smad signaling in human cervical cancer cells.

Transforming growth factor-β (TGF-β)-induced Epithelial-to-mesenchymal transition (EMT) process is a fundamental target for preventing cervical cancer cells' progression and invasion. Green tea and its principal active substance, Epigallocatechin-3-gallate (EGCG), demonstrate anti-tumor activities in various tumor cells. The cell viability of two cervical cancer cell lines, Hela and SiHa, in the experimental groups was examined employing the MTT method, and ROS generation was probed applying 2',7'-dichlorofluorescein diacetate-based assay. The Smad signaling and EMT process was evaluated utilizing western blot analysis and quantitative real-time polymerase chain reaction (qRT-PCR). Chromatin immunoprecipitation (ChIP) and Smad binding element (SBE)-luciferase assays were employed to measure Smad-DNA interaction and Smad transcriptional activity, respectively. EGCG (0-100μmol/L) and green tea extract (0-250μg/ml) suppressed the viability of cancer cells in a dose-dependent manner (p<0.01). Our conclusions affirmed that pre-incubation with green tea extract (80μg/ml) and EGCG (60μmol/L) significantly reversed the impacts of TGF-β in Hela and SiHa cells by decreasing Vimentin, ZEB, Slug, Snail, and Twist and increasing E-cadherin expression. The molecular mechanism of green tea extract and EGCG for TGF-β-induced EMT inhibition interfered with ROS generation and Smad signaling. Green tea extract and EGCG could significantly decrease ROS levels, the phosphorylation of Smad2/3, the translocation, DNA binding, and activity of Smads in cervical cancer cell lines treated with TGF-β1 (p<0.01). EGCG and green tea extract suppressed TGF-β-induced EMT in Hela and SiHa cells, and the underlying molecular mechanism may be related to the ROS generation and Smad signaling pathway.

A green tea extract and epigallocatechin-3-gallate attenuate the deleterious effects of irinotecan in an oral epithelial cell model

ObjectiveTo investigate the ability of a green tea extract and epigallocatechin-3-gallate (EGCG) to protect oral epithelial cells against the deleterious effects of the chemotherapeutic agent irinotecan, with respect to cytotoxicity; reactive oxygen species (ROS) generation; cytokine and matrix metalloproteinase (MMP) production; and cell proliferation and migration. MethodsThe B11 oral keratinocyte and GMSM-K oral epithelial cell lines were used in this study. Cell viability was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay. A fluorometric assay was used to quantify ROS production. Cell proliferation was assessed using a fluorescent cell tracker dye, while a migration assay kit was used to monitor cell migration. Cytokine and MMP secretion was quantified by an enzyme-linked immunosorbent assay. ResultsThe green tea extract and EGCG reduced the cytotoxicity of irinotecan toward oral keratinocyte and epithelial cell lines. Irinotecan-induced intracellular ROS generation by oral keratinocytes was reduced by the green tea extract and EGCG. Irinotecan negatively affected the proliferation and migration of oral keratinocytes in a dose-dependent manner. However, these effects were not neutralized by the green tea extract, while EGCG showed a trend to attenuate the irinotecan-induced decrease in cell migration. The green tea extract and EGCG also had a dose-dependent inhibitory effect on irinotecan-induced secretion of interleukin-6 and interleukin-8 by oral epithelial cells. Lastly, the irinotecan-induced decrease in the secretion of MMP-2 and MMP-9 by oral epithelial cells was partially restored by the green tea extract and EGCG. ConclusionsThe green tea extract and EGCG, through anti-cytotoxic, anti-oxidative, and anti-inflammatory properties, may protect the oral mucosa against the deleterious effects of the chemotherapeutic agent irinotecan and may be of interest for treating oral mucositis.

Combination of resveratrol and green tea epigallocatechin gallate induces synergistic apoptosis and inhibits tumor growth invivo in head and neck cancer models.

Despite widespread interest in chemoprevention and therapy due to the high margin of safety of dietary natural compounds, clinical intervention with single agents has failed to yield the expected outcomes, mostly due to poor bioavailability and low potency. Combinations of natural agents with synergistic effects are gaining increasing attention. In the present study, in vitro and in vivo antitumor effects of a combination of two natural dietary agents, green tea epigallocatechin gallate (EGCG) and resveratrol were investigated. It was revealed that their combination at low doses (at which single agents induce minimal apoptosis) synergistically increased apoptosis (combination index <1) in head and neck cancer cell lines. Synergistic apoptosis was also supported by caspase-3 and PARP cleavage. The combination also significantly inhibited growth of xenografted head and neck tumors in nude mice as supported by significant inhibition of tumor volume, tumor weight and Ki67 expression, and increase in TUNEL-positive cells. Mechanistic studies revealed that the combination inhibited AKT-mTOR signaling both in vitro and in vivo. In addition, overexpression of constitutively active AKT protected cells from apoptosis induced by the combination of EGCG and resveratrol. Collectively, the present results for the first time suggest that the combination of EGCG and resveratrol has synergistic growth inhibitory effects and provide an important rationale for future clinical development for chemoprevention and treatment of head and neck cancer.

Formulation and Optimization of Nanospanlastics for Improving the Bioavailability of Green Tea Epigallocatechin Gallate.

The present study aimed to investigate the potential of nanospanlastics for boosting the bioavailability of epigallocatechin gallate (EGCG). EGCG has valuable effects like anti-inflammation, anti-oxidation, and anti-tumorigenesis. Unfortunately, it has a low oral bioavailability due to its limited permeation and poor stability. To overcome these pitfalls, EGCG was fabricated as a nanospanlastic. Nanospanlastics are flexible nanovesicles that are composed of surfactants and edge activators (EAs). EAs improve the deformability of spanlastics by acting as a destabilizing factor of their vesicular membranes. EGCG-loaded spanlastics were prepared by an ethanol injection method, according to 23 factorial design, to explore the impact of different independent variables on entrapment efficiency (EE%), % drug released after 12 h (Q12h), and particle size (PS). In vitro characterization, ex vivo intestinal permeation test, and pharmacokinetic study of the optimized formula were performed. A newly developed RP-HPLC technique was adopted for the estimation of EGCG. The optimized formula (F4) demonstrated more prolonged drug release and a significant improvement in the EE%, permeability, deformability and stability than the corresponding niosomes. The pharmacokinetic study investigated that F4 had a more sustained drug release and a higher bioavailability than the conventional niosomes and free drugs. Nanospanlastics could be a promising approach for improving the bioavailability of EGCG.

Polyphenols in foods: Classification, methods of identification, and nutritional aspects in human health.

Polyphenols widely exists in various foods, including main crops, fruits, beverages and some wines. Famous representatives of polyphenols, such as resveratrol in red wine, (-)-epigallocatechin gallate in green tea, chlorogenic acid in coffee, anthocyanins in colored fruits, procyanidins in grape seed have become hot research topics in food science and nutrition. There have been thousands of papers on the biochemistry, chemistry, nutritional values and population-based investigations of dietary polyphenols. In this chapter, we reviewed the published articles and database of dietary polyphenols to draw a profile for the classification, structural identification, and biological activities mainly based on enzymes, cell bioassay and animal models, as well as the population-based investigation results. The typical compound and its health benefits for each category of polyphenols was also introduced. The identification of dietary polyphenols could be solved by combined spectroscopy methods, of which the liquid chromatography tandem mass spectrometry is highlighted to greatly increase the efficiency on structural identification. Although the population-based investigation showed some controversial results for health benefits, the multi-functions of dietary polyphenols on preventing metabolic syndromes, various cancers and neurodegenerative disease have attracted much attention.

저온 플라즈마 처리된 녹차 유래의 최종당화산물 생성 억제물질의 동정

Green tea (Camellia sinensis) with abundant phenolic compounds is considered to a major dietary source of flavan 3-ols such as (−)-epigallocatechin gallate (EGCG). Here, we assessed the effects of non-thermal dielectric barrier discharge (DBD) plasma on green tea extract using spectroscopy and bioassays. Plasma-induced structural changes in EGCG in green tea resulted in the isolation of two methylene-bridge dimers, oolonghomobisflavan A (1) and B (2). The structures of these compounds were characterized using nuclear magnetic resonance and mass spectroscopy data. The symmetric dimer 1 connected by a methylene linkage was the most potent inhibitor of advanced glycation end products (AGEs) formation (IC50, 8.2±0.5 μM). The radical scavenging capacity of against hydroxyl, and stereoisomers 1 and 2 was more potent than that of the positive control ((+)-catechin). These major products purified by treating green tea with DBD plasma for 60 min were quantified by HPLC. Our results provide evidence that structural changes of EGCG in green tea extract induced by plasma might enhance biological efficacy.

Evaluation of green tea extract and epigallocatechin gallate effects on bisphenol A-induced vascular toxicity in isolated rat aorta and cytotoxicity in human umbilical vein endothelial cells.

This study was designed to assess bisphenol A (BPA)-induced vascular toxicity, the effectiveness of green tea extract and epigallocatechin gallate (EGCG) against BPA toxicity, and possible underlying mechanisms. In isolated rat aorta, contractile and relaxant responses as well as malondialdehyde levels were evaluated. Cell viability and effects on the protein levels of apoptotic (bax, bcl2, and caspase-3), autophagic (LC3), and cell adhesion molecules were calculated using the MTT method and western blotting in human umbilical vein endothelial cells (HUVECs). BPA increased aorta MDA levels (p < .0001) and decreased vascular responses to KCl [20 and 40 mM (p < .0001), 80 mM (p < .001)], phenylephrine [10-8 , 10-6 , and 10-5 M (p < .001), 10-7 and 10-4 M (p < .0001)], and acetylcholine [10-6 M (p < .01), 10-5 and 10-4 M (p < .0001)]. In HUVECs, BPA enhanced the levels of LC3A/B, bax/bcl2 ratio, cleaved caspase-3, and vascular cell adhesion molecule-1. Green tea extract, EGCG, and vitamin E co-treatment with BPA diminished the toxic effects of BPA. These findings provide evidence that green tea extract and EGCG possess beneficial effects in preventing BPA-induced vascular toxicity through increasing the antioxidant activities and the regulation of signaling pathways.

EGCG Mediated Targeting of Deregulated Signaling Pathways and Non-Coding RNAs in Different Cancers: Focus on JAK/STAT, Wnt/β-Catenin, TGF/SMAD, NOTCH, SHH/GLI, and TRAIL Mediated Signaling Pathways.

Decades of research have enabled us to develop a better and sharper understanding of multifaceted nature of cancer. Next-generation sequencing technologies have leveraged our existing knowledge related to intra- and inter-tumor heterogeneity to the next level. Functional genomics have opened new horizons to explore deregulated signaling pathways in different cancers. Therapeutic targeting of deregulated oncogenic signaling cascades by products obtained from natural sources has shown promising results. Epigallocatechin-3-gallate (EGCG) has emerged as a distinguished chemopreventive product because of its ability to regulate a myriad of oncogenic signaling pathways. Based on its scientifically approved anticancer activity and encouraging results obtained from preclinical trials, it is also being tested in various phases of clinical trials. A series of clinical trials associated with green tea extracts and EGCG are providing clues about significant potential of EGCG to mechanistically modulate wide ranging signal transduction cascades. In this review, we comprehensively analyzed regulation of JAK/STAT, Wnt/β-catenin, TGF/SMAD, SHH/GLI, NOTCH pathways by EGCG. We also discussed most recent evidence related to the ability of EGCG to modulate non-coding RNAs in different cancers. Methylation of the genome is also a widely studied mechanism and EGCG has been shown to modulate DNA methyltransferases (DNMTs) and protein enhancer of zeste-2 (EZH2) in multiple cancers. Moreover, the use of nanoformulations to increase the bioavailability and thus efficacy of EGCG will be also addressed. Better understanding of the pleiotropic abilities of EGCG to modulate intracellular pathways along with the development of effective EGCG delivery vehicles will be helpful in getting a step closer to individualized medicines.

The Green Tea Catechin, EGCg, Preserves Both Muscle and Bone in Aging Sarcopenic Rats

ObjectivesSarcopenia is the progressive decline in muscle mass, strength and function that is exacerbated with age. It is well known that there is an overlap between sarcopenia and osteopenia/osteoporosis in the aging population. Thus, addressing both muscle and bone loss during aging is important. Epigallocatechin gallate (EGCg) is the most abundant green tea catechin and has well established anti‐inflammatory and antioxidant benefits. Green tea consumption has also been associated with bone health. In this study we examined the impact of chronic EGCg treatment on muscle mass and bone structure in aging rats.MethodsAged male Sprague‐Dawley rats (21 months) were fed the following diets ad libitum: Control group (n=9) fed AIN93M diet, and a treatment group (n=10) fed AIN93M diet +EGCg (200 mg/kg) for 8 weeks. At the end of study, gastrocnemius muscles were weighed, and histological analysis was performed to measure muscle fiber cross sectional area (CSA) using hematoxylin &amp; eosin staining. Bones were analyzed by MicroCT imaging (Burker) of the tibia at ~22 μm resolution and images were processed using Inveon Research Workplace software. All bone samples were analyzed for trabecular bone volume, trabecular number, trabecular spacing, trabecular thickness, trabecular pattern factor and bone density.ResultsGastrocnemius muscle wet weights were significantly higher (p&lt;0.05) and fiber CSA tended to be higher (p&lt;0.06) in the EGCg group compared to controls. Tibia bone volume/total volume density was significantly greater (p=0.03) in animals treated with EGCg compared to control. Significant differences were found in trabecular microstructure, with a significant increase in trabecular number per given volume (p=0.01) and a corresponding significant decrease in trabecular spacing (p=0.02) in EGCg‐treated rats compared to control. No significant group differences were found in overall bone density or trabecular thickness. Differences in mean values between control and EGCg groups were determined by student’s t‐test.ConclusionsThese data present novel findings on the osteoprotective mechanism of EGCg on the aging bone through an increase in trabecular bone volume, trabecular number and trabecular spacing. Furthermore, the preservation of muscle mass and muscle fiber CSA suggests that the use of green tea catechin, EGCg, could be a potential nutritional therapy to prevent the progression of muscle and bone loss during sarcopenia.Support or Funding InformationAbbott Nutrition

Solid-Phase Extraction of Catechins from Green Tea with Deep Eutectic Solvent Immobilized Magnetic Molybdenum Disulfide Molecularly Imprinted Polymer.

A type of molecular-imprinted polymer with magnetic molybdenum disulfide as a base and deep eutectic solvent as a functional monomer (Fe3O4@MoS2@DES-MIP) was prepared with surface molecular imprinting method. It was applied as the adsorbent for the selective recognition and separation of (+)-catechin, (−)-epicatechin, (−)-epigallocatechin, (−)-epicatechin gallate, and (−)-epigallocatechin gallate in green tea in the process of magnetic solid-phase extraction (MSPE) combined with high-performance liquid chromatography (HPLC). The structure of Fe3O4@MoS2@DES-MIP was characterized by Fourier transform infrared spectroscopy and field emission scanning electron microscopy. The adsorption properties and selective recognition ability on (−)-epigallocatechin gallate and the other four structural analogues were examined and compared. The results show that the polymer has excellent selective recognition ability for (−)-epigallocatechin gallate, and its adsorption capacity was much higher than that of structural analogues. The Fe3O4@MoS2@DES-MIP not only has the special recognition ability to template a molecule, but also can be separated by magnets with high separation efficiency and can be used in MSPE.

Induction of Endoplasmic Reticulum Stress Pathway by Green Tea Epigallocatechin-3-Gallate (EGCG) in Colorectal Cancer Cells: Activation of PERK/p-eIF2α/ATF4 and IRE1α.

Epigallocatechin-3-gallate (EGCG) is the most abundant bioactive polyphenolic compound among the green tea constituents and has been identified as a potential anticancer agent in colorectal cancer (CRC) studies. This study was aimed to determine the mechanism of actions of EGCG when targeting the endoplasmic reticulum (ER) stress pathway in CRC. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay was performed on HT-29 cell line and normal cell line (3T3) to determine the EGCG toxicity. Next, western blot was done to observe the expression of the related proteins for the ER stress pathway. The Caspase 3/7 assay was performed to determine the apoptosis induced by EGCG. The results demonstrated that EGCG treatment was toxic to the HT-29 cell line. EGCG induced ER stress in HT-29 by upregulating immunoglobulin-binding (BiP), PKR-like endoplasmic reticulum kinase (PERK), phosphorylation of eukaryotic initiation factor 2 alpha subunit (eIF2α), activating transcription 4 (ATF4), and inositol-requiring kinase 1 alpha (IRE1α). Apoptosis was induced in HT-29 cells after the EGCG treatment, as shown by the Caspase 3/7 activity. This study indicates that green tea EGCG has the potential to inhibit colorectal cancer cells through the induction of ER stress.

Effect of solid lipid nanoparticles system on the stability of Green Tea leaves (Camellia sinensis L. Kuntze) extract as sunscreen

Green tea (Camellia sinensis L.) is known to have activity as antioxidant and sunscreen due to its catechin compound. The major catechin-derived compound is epigallocatechin gallate (EGCG). However, EGCG in green tea leaves is unstable due to its rapid degradation and large molecular mass of 458 daltons, making it difficult to penetrate the skin. The purpose of this research is to formulate solid lipid nanoparticles (SLN) of green tea leaves ethanol extract using high speed homogenization and sonication method, as well as to evaluate SLN ability in improving extract stability as sunscreen using UV-Vis spectrophotometry. SLN formula contained 0,1% green tea leaves ethanol extract, and the most optimal formula was composed of 4% lipid (Precirol ATO 5:Gelucire 44/14) 4:1 and 1% Kolliphor P188 as surfactant. Characterization results showed particle size of 286.7 nm, polydispersity index 0.296, and zeta potential -3.7 mV. Photostability test using 120-minutes irradiation showed decrease in percentage of DPPH free radicals damping at 5.34% extract and 1.77% SLN for UVA, and 7.7% extract and 2.34% SLN for UVB.

Epigallocatechin gallate attenuates arsenic induced genotoxicity via regulation of oxidative stress in balb/C mice

Arsenic is well known genotoxicant which causes the excessive generation of reactive oxygen species (ROS) and inhibition of antioxidant enzyme systems leading to cell damage through the activation of oxidative sensitive signaling pathways. Epigallocatechin gallate (EGCG), the main and active polyphenolic catechin present in green tea, has shown potent antioxidant, free radical scavenging and genoprotective activity in vivo. The present study attempted to investigate antioxidant and geno-protective efficacy of EGCG by regulating arsenic induced oxidative stress in mice. Animals received prophylactic and therapeutic treatments at two different doses (25 and 50mg/kg b.wt.) of EGCG orally for 15days and administered arsenic intraperitoneally at dose of 1.5mg/kg b.wt (1/10th of LD50) for 10days. Arsenic intoxication revealed enhanced ROS production (114%) in lymphocytes; elevated levels of LPO (2-4 fold); reduced levels of hepato-renal antioxidants (approx. 45%) and augmented genomic fragmentation in hepato-renal tissues; increased chromosomal anomalies (78%) and micronucleation (21.93%) in bone marrow cells and comet tailing (25%) in lymphocytes of mice. Both pre and post treatments of EGCG decreased ROS production, restored lipid peroxidation (LPO) and reduced hepato-renal antioxidants levels, reduced the DNA fragmentation, number of chromosomal aberrations (CA), micronucleation (MN), and comet tailing but prophylactic treatment of 50mg/kg b.wt was the most effective treatment in regulating arsenic induced oxidative stress. The effectiveness of this dose was furthermore validated by calculating the inhibitory index. Thus, results of present work empirically demonstrate free radical scavenging, anti-oxidative and genoprotective efficacy of EGCG against arsenic toxicity.

Abstract 209A: Involvement of pRb-E2F pathway in green tea extract-induced growth inhibition of human myeloid leukemia cells

Abstract It has long believed that green tea has many health benefits. However, the clinical research has not provided conclusive evidence on these benefits especially on cancer prevention and treatment. In basic research, there are also many inconsistent reports on the biological activity of green tea extract and its major chemical components (catechins) with both inhibitory and stimulatory effects of EGCG on various cancer cells being reported. The stimulatory or inhibitory activity is often linked to either activation or inactivation of receptor tyrosine kinase. In this study, we present evidence that green tea extract and its major chemical components, Epigallocatechin-3-gallate (EGCG), epigallocatechin (ECG), and epicatechin (EC) inhibit growth of two human myeloid leukemia cells (TF-1a and MV4-11) through the regulation of pRb synthesis and pRb-E2F complexes. Addition of green tea extract to the culture of the myeloid leukemia cells significantly inhibited their proliferation with a substantial portion of cell death and decreased their colony forming cells in soft-agar culture. EGCG, ECG, and EC showed a similar inhibitory effect on these cells with the inhibitory efficiency varied with each catechin and the dose applied. Green tea extract and EGCG had no significant effect on the expression of G1 CDKs and the CDK inhibitors but downregulated the formation of pRb-CDKs examined by Western blot and immunoprecipitation, respectively. Surprisingly, the expression of pRb was markedly upregulated while the phosphorylation of pRb downregulated. The upregulation of pRb was blocked by pre-treatment with cycloheximide, a protein synthesis inhibitor, suggesting a requirement of protein synthesis. In agreement with these results, pRb-E2F complexes were upregulated and E2F DNA binding activity decreased determined by EMSA assay. Since both TF-1a and MV4-11 cell are factor-independent cell lines, the upregulation of pRb-E2F complexes and downregulation of DNA binding activity by green tea extract is most likely through a receptor tyrosine kinase-independent pathway. In summary, this study reveals that green tea extract, EGCG, ECG, and EC inhibited growth in human myeloid leukemia cells via regulating pRb synthesis and the formation of pRb-E2F complexes, leading to the inhibition of the E2F DNA binding activity. In addition, we found that that the stem/progenitor cells derived from the leukemia cell lines tested are more sensitive to the inhibitory effect of green tea extract. We propose that as a beverage one should not consume high amounts of green tea (extract) due to its possible cytotoxic or apoptotic effect, and its significant inhibitory effect on stem/progenitor cells, although the biological effect of green tea extract in vivo could be quite different from in vitro. However, the concentrated green tea extract may have potential for clinical investigation as an inducer of cancer cell death. Citation Format: Darrell Henry, Sebastien Brumaire, Gabriela Alvarez, Beatriz Alvarez, Xiaotang Hu. Involvement of pRb-E2F pathway in green tea extract-induced growth inhibition of human myeloid leukemia cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 209A.

Involvement of pRb-E2F pathway in green tea extract-induced growth inhibition of human myeloid leukemia cells

Both inhibitory and stimulatory effect of EGCG on cancer cells have been reported, which often is linked to receptor tyrosine kinase signaling. In this study, we present evidence that green tea extract and its chemical component, Epigallocatechin-3-gallate (EGCG), inhibit growth of human myeloid leukemia cells through the regulation of pRb synthesis and formation of pRb-E2F complexes. Addition of green tea extract to the culture of TF-1a and MV4-11 myeloid leukemia cells significantly inhibited their proliferation with a substantial portion of cell death being observed. The green tea extract and EGCG had no significant effect on the expression of G1 CDKs and the CDK inhibitors but downregulated the formation of pRb-CDKs. Surprisingly, the expression of pRb was markedly upregulated while the phosphorylation of pRb downregulated. The upregulation of pRb was blocked by pre-treatment with cycloheximide, a protein synthesis inhibitor, suggesting a requirement of protein synthesis. In agreement with these results, pRb-E2F complexes were upregulated and E2F DNA binding activity decreased. Since both TF-1a and MV4-11 are factor-independent cell lines, the upregulation of pRb-E2F complexes and inhibition of DNA binding activity by green tea extract is most likely through a receptor tyrosine kinase-independent pathway. We also found that the stem/progenitor cells derived from these two leukemia cell lines are more sensitive to the inhibitory effect of green tea extract. Our result suggests that concentrated green tea extract and EGCG may have potential for clinical investigation as an inducer of cancer cell death.

Effects of green tea, matcha tea and their components epigallocatechin gallate and quercetin on MCF‑7 and MDA-MB-231 breast carcinoma cells.

We investigated the anticarcinogenic potential of green tea and its components epigallocatechin gallate (EGCG) and quercetin, as well as tamoxifen, on MCF-7 and MDA-MB-23 breast cancer cells. Using high-performance liquid chromatography, the quantity of EGCG and quercetin in green tea was analyzed. The receptor status of the cells was confirmed immunohistochemically. Various viability and cytotoxicity tests were later performed to investigate the effects of the substances. After incubating the cells with green tea extract, EGCG, quercetin and tamoxifen, a decrease in viability (MTT test) or proliferation (BrdU assay) was found in all cell tests with varying effects, depending on the assay used. The effects were similar in both cell lines. This work confirmed that EGCG and quercetin are contained in green tea and that both substances in pure form and as green tea have an anticarcinogenic effect on both estrogen receptor-positive and -negative breast cancer cells. This effect could also be demonstrated with tamoxifen in both cell lines (MTT and BrdU assays). These results suggest that the effects observed in these experiments are not generated only via estrogen receptor-mediated pathways.

The importance of inhibition of a catabolic pathway of methotrexate metabolism in its efficacy for rheumatoid arthritis

Methotrexate (MTX), an antifolate, is the anchor drug for the treatment of rheumatoid arthritis (RA). It is inexpensive, effective, and generally safe. When clinical response is inadequate, biological therapies are commonly used in combination with MTX. However, biological agents have safety concerns (i.e. infections, malignancy) and the addition of a biologic agent is expensive, making strategies to improve MTX efficacy important.Inhibition of pathways of folate metabolism involving purine metabolism by MTX, have been traditionally emphasized as important in MTX efficacy. However, inhibition MTX catabolism may also be important. MTX is irreversibly hydroxylated to form 7-hydroxy methotrexate (7-OH-MTX) by aldehyde oxidase (EC 1.2.3.1) (AOX). Catabolism of MTX to 7-OH-MTX is the first metabolic process imposed on an oral dose of MTX and will alter subsequent interactions of MTX with other enzymes. 7-OH-MTX is less potent than MTX in the treatment of rat adjuvant arthritis. RA patients with a low capacity to catabolize MTX to 7-OH-MTX do better clinically than individuals who are rapid formers of 7-OH-MTX. Therefore, altering the catabolism of MTX may be an innovative way to improve MTX efficacy.Raloxifene is a FDA-approved therapy for postmenopausal osteoporosis and for the reduction of invasive breast cancers but has no known activity in RA. Raloxifene is a potent inhibitor of human liver AOX. Postmenopausal women with RA frequently have low bone mineral density and would be candidates for raloxifene and MTX combination therapy. The effect of raloxifene on MTX metabolism has never been studied.Our hypothesis is that in postmenopausal women with RA and osteoporosis treated with MTX and raloxifene, the inhibition of AOX with resultant decreased formation of 7-OH MTX; will increase MTX levels and improve MTX efficacy. This hypothesis could be studied in an open-label, proof of concept clinical study in individuals before and after the addition of raloxifene. Red blood cell MTX and 7-OH-MTX levels and RA disease activity (DAS28) would be measured. In possible future studies, there are dietary substances, as supplements, (e.g. epigallocatechin gallate in green tea and resveratrol) which inhibit human liver AOX which could be evaluated.

Epigallocatechin gallate inhibits hepatitis B virus infection in human liver chimeric mice

BackgroundPersistent hepatitis B virus (HBV) infection causes liver cirrhosis and hepatocellular carcinoma and constitutes a major worldwide health problem. Currently, anti-HBV drugs are limited to peginterferon and nucleos(t)ide analogs, which are costly and have considerable side effects; the development of novel, effective anti-HBV agents is crucial.MethodsCatechins are a major group of compounds found in green tea extract and epigallocatechin gallate (EGCG) has been shown to have antiviral properties, including inhibition of cellular entry by HBV. FRG (Fah−/−/ Rag2−/−/ IL-2Rγ/−) mice were used in this study to generate chimeras carrying human primary hepatocytes, to facilitate investigation of the inhibitory effect of EGCG on HBV infection.ResultsHere, we show the inhibitory effect of EGCG on HBV infection and replication in HuS-E/2 cells. The inhibitory effect of EGCG on HBV infection in vivo was confirmed by monitoring HBV DNA and HBsAg in serum and immunostaining the liver tissues of the human liver chimeric mice.ConclusionsThe effects of EGCG suggest a robust strategy for the treatment of HBV infection and EGCG may have therapeutic potential for the treatment of HBV-associated liver diseases.