Catechin conjugated with fatty acid inhibits DNA polymerase and angiogenesis

Catechins in green tea have anticancer and antiangiogenesis activities, with epigallocatechin-3-gallate (EGCG) being the most potent antiangiogenic tea catechin. This study examined whether chemical modification of catechin enhanced anticancer and antiangiogenic effects. Catechin, conjugated with fatty acid (acyl-catechin), strongly inhibited DNA polymerase, HL-60 cancer cell growth, and angiogenesis. Catechin conjugated with stearic acid [(2R,3S)-3',4',5,7-tetrahydroxyflavan-3-yl octadecanoate; catechin-C18] was the strongest inhibitor in DNA polymerase alpha and beta and angiogenesis assays. Catechin-C18 also suppressed human endothelial cell (HUVEC) tube formation on the reconstituted basement membrane, suggesting that it affected not only DNA polymerases but also signal transduction pathways in HUVECs. These data indicate that acyl-catechins target both DNA polymerases and angiogenesis as anticancer agents. These results suggest that acylation of catechin is an effective chemical modification to improve the anticancer activity of catechin.     

Epigallocatechin-3-O-gallate inhibits TNFalpha-induced monocyte chemotactic protein-1 production from vascular endothelial cells

Monocyte chemotactic protein-1 (MCP-1) plays a pivotal role in the recruitment of monocytes and thus in the development of inflammatory cardiovascular diseases. Epigallocatechin-3-O-gallate (EGCG), the major catechin derived from green tea, has multiple beneficial effects to reduce cardiovascular disease but the effects of EGCG on vascular endothelial MCP-1 production is not known. In this study, we investigated the mechanisms by which EGCG may inhibit tumor necrosis factor-alpha (TNFalpha)-induced MCP-1 production in bovine coronary artery endothelial cells. TNFalpha increased MCP-1 production in both a concentration and time-dependent manner. Inhibitors of phosphatidylinositol-3-OH kinase (PI-3 kinase), LY294002 and wortmannin, decreased TNFalpha-induced MCP-1 production. EGCG prevented TNFalpha-mediated MCP-1 production and reduced phosphorylation of Akt (Ser473). In addition, EGCG attenuated TNFalpha mediated down-regulation of TNFalpha receptor 1 (TNFR1), but not TNFR2. In conclusion, EGCG inhibited TNFalpha-induced MCP-1 production. Moreover, EGCG inhibited Akt phosphorylation as well as TNF activation of TNFR1, which subsequently resulted in reduced MCP-1 production. These data provide a novel mechanism where the green tea flavonoid, EGCG, could provide direct vascular benefits in inflammatory cardiovascular diseases.     

Green tea constituent epigallocatechin-3-gallate inhibits angiogenic differentiation of human endothelial cells

Several independent research studies have shown that consumption of green tea reduces the development of cancer in many animal models. Epidemiological observations, though inconclusive, are suggesting that green tea consumption may also reduce the risk of some cancers in humans. These anti-carcinogenic effects of green tea have been attributed to its constituent polyphenols. Angiogenesis is a crucial step in the growth and metastasis of cancers. We have investigated the effect of the major polyphenolic constituent of green tea, epigallocatechin-3-gallate (EGCG), on the tube formation of human umbilical vein endothelial cells (HUVEC) on matrigel. Tube formation was inhibited by treatment both prior to plating and after plating endothelial cells on matrigel. EGCG treatment also was found to reduce the migration of endothelial cells in matrigel plug model. The role of matrix metalloproteinases (MMP) has been shown to play an important role during angiogenesis. Zymography was performed to determine if EGCG had any effect on MMPs. Zymographs of EGCG-treated culture supernatants modulated the gelatinolytic activities of secreted proteinases indicating that EGCG may be exerting its inhibitory effect by regulating proteinases. These findings suggest that EGCG acts as an angiogenesis inhibitor by modulating protease activity during endothelial morphogenesis.     

麦没食子儿茶素没食子酸酯对TNF-α诱导的内皮细胞中PAI-1表达的影响及机制研究

目的探讨绿茶的主要成分麦没食子儿茶素没食子酸酯(EGCG)在血管内皮细胞中对肿瘤坏死因子-α(TNF-α)所诱导的纤溶酶原激活物抑制物-1(PAI-1)表达的影响及机制。方法利用人脐静脉内皮细胞体外培养方法,分别与TNF-α和/或EGCG孵育,运用蛋白免疫印迹方法检测内皮细胞中磷酸化细胞外调节蛋白激酶1/2(p-ERK1/2)和肿瘤坏死因子受体1(TNFR1)蛋白表达,应用酶联免疫吸附法方法检测细胞液中PAI-1水平。结果 TNF-α以浓度依赖和时间依赖方式增加内皮细胞中PAI-1的表达。EGCG可抑制TNF-α所诱导的PAI-1的表达,并可抑制ERK1/2磷酸化。TNF-α的刺激可使TN-FR1表达明显减少,而EGCG可抑制这一作用。结论 EGCG可能通过抑制ERK1/2的磷酸化,从而抑制TNF-α所诱导的PAI-1的表达,同时可减少TNF-α对TNFR1的抑制作用,在改善肥胖、胰岛素抵抗及相关心血管疾病中起着重要作用。     

Complex effects of different green tea catechins on human platelets

Epigallocatechin gallate (EGCG), a major component of green tea, has been previously shown to inhibit platelet aggregation. The effects of other green tea catechins on platelet function are not known. Pre-incubation with EGCG concentration-dependently inhibited thrombin-induced aggregation and phosphorylation of p38 mitogen-activated protein kinase and extracellular signal-regulated kinases-1/2. In contrast EGCG stimulated tyrosine phosphorylation of platelet proteins, including Syk and SLP-76 but inhibited phosphorylation of focal adhesion kinase. Other catechins did not inhibit platelet aggregation. Interestingly, when EGCG was added to stirred platelets, a tyrosine kinase-dependent stimulation of platelet aggregation was observed. The two other catechins containing a galloyl group in the 3' position (catechin gallate, epicatechin gallate) also stimulated platelet aggregation, while catechins without a galloyl group (catechin, epicatechin) or the catechin with a galloyl group in the 2' position (epigallocatechin) did not.     

Green tea catechins reduce invasive potential of human melanoma cells by targeting COX-2, PGE2 receptors and epithelial-to-mesenchymal transition

Melanoma is the most serious type of skin disease and a leading cause of death from skin disease due to its highly metastatic ability. To develop more effective chemopreventive agents for the prevention of melanoma, we have determined the effect of green tea catechins on the invasive potential of human melanoma cells and the molecular mechanisms underlying these effects using A375 (BRAF-mutated) and Hs294t (Non-BRAF-mutated) melanoma cell lines as an in vitro model. Employing cell invasion assays, we found that the inhibitory effects of green tea catechins on the cell migration were in the order of (-)-epigallocatechin-3-gallate (EGCG)>(-)-epigallocatechin>(-)-epicatechin-3-gallate>(-)-gallocatechin>(-)-epicatechin. Treatment of A375 and Hs294t cells with EGCG resulted in a dose-dependent inhibition of cell migration or invasion of these cells, which was associated with a reduction in the levels of cyclooxygenase (COX)-2, prostaglandin (PG) E(2) and PGE(2) receptors (EP2 and EP4). Treatment of cells with celecoxib, a COX-2 inhibitor, also inhibited melanoma cell migration. EGCG inhibits 12-O-tetradecanoylphorbol-13-acetate-, an inducer of COX-2, and PGE(2)-induced cell migration of cells. EGCG decreased EP2 agonist (butaprost)- and EP4 agonist (Cay10580)-induced cell migration ability. Moreover, EGCG inhibited the activation of NF-κB/p65, an upstream regulator of COX-2, in A375 melanoma cells, and treatment of cells with caffeic acid phenethyl ester, an inhibitor of NF-κB, also inhibited cell migration. Inhibition of melanoma cell migration by EGCG was associated with transition of mesenchymal stage to epithelial stage, which resulted in an increase in the levels of epithelial biomarkers (E-cadherin, cytokeratin and desmoglein 2) and a reduction in the levels of mesenchymal biomarkers (vimentin, fibronectin and N-cadherin) in A375 melanoma cells. Together, these results indicate that EGCG, a major green tea catechin, has the ability to inhibit melanoma cell invasion/migration, an essential step of metastasis, by targeting the endogenous expression of COX-2, PGE(2) receptors and epithelial-to-mesenchymal transition.     

Inhibitory Effects of Green Tea and (–)-Epigallocatechin Gallate on Transport by OATP1B1, OATP1B3, OCT1, OCT2, MATE1, MATE2-K and P-Glycoprotein

Green tea catechins inhibit the function of organic anion transporting polypeptides (OATPs) that mediate the uptake of a diverse group of drugs and endogenous compounds into cells. The present study was aimed at investigating the effect of green tea and its most abundant catechin epigallocatechin gallate (EGCG) on the transport activity of several drug transporters expressed in enterocytes, hepatocytes and renal proximal tubular cells such as OATPs, organic cation transporters (OCTs), multidrug and toxin extrusion proteins (MATEs), and P-glycoprotein (P-gp). Uptake of the typical substrates metformin for OCTs and MATEs and bromosulphophthalein (BSP) and atorvastatin for OATPs was measured in the absence and presence of a commercially available green tea and EGCG. Transcellular transport of digoxin, a typical substrate of P-gp, was measured over 4 hours in the absence and presence of green tea or EGCG in Caco-2 cell monolayers. OCT1-, OCT2-, MATE1- and MATE2-K-mediated metformin uptake was significantly reduced in the presence of green tea and EGCG (P < 0.05). BSP net uptake by OATP1B1 and OATP1B3 was inhibited by green tea [IC₅₀ 2.6% (v/v) and 0.39% (v/v), respectively]. Green tea also inhibited OATP1B1- and OATP1B3-mediated atorvastatin net uptake with IC₅₀ values of 1.9% (v/v) and 1.0% (v/v), respectively. Basolateral to apical transport of digoxin was significantly decreased in the presence of green tea and EGCG. These findings indicate that green tea and EGCG inhibit multiple drug transporters in vitro. Further studies are necessary to investigate the effects of green tea on prototoypical substrates of these transporters in humans, in particular on substrates of hepatic uptake transporters (e.g. statins) as well as on P-glycoprotein substrates.     

Proposed mechanisms of (-)-epigallocatechin-3-gallate for anti-obesity

Green tea catechins (GTCs) are polyphenolic flavonoids formerly called vitamin P. GTCs, especially (-)-epigallocatechin-3-gallate (EGCG), lower the incidence of cancers, collagen-induced arthritis, oxidative stress-induced neurodegenerative diseases, and streptozotocin-induced diabetes. Also, inhibition of adipogenesis by green tea and green tea extract has been demonstrated in cell lines, animal models, and humans. The obesity-preventive effects of green tea and its main constituent EGCG are widely supported by results from epidemiological, cell culture, animal, and clinical studies in the last decade. Studies with adipocyte cell lines and animal models have demonstrated that EGCG inhibits extracellular signal-related kinases (ERK), activates AMP-activated protein kinase (AMPK), modulates adipocyte marker proteins, and down-regulates lipogenic enzymes as well as other potential targets. Also, the catechin components of green tea have been shown to possess anti-carcinogenic properties possibly related to their anti-oxidant activity. In addition, it was shown that dietary supplementation with EGCG could potentially contribute to nutritional strategies for the prevention and treatment of type 2 diabetes mellitus. In this review, the biological activities and multiple mechanisms of EGCG in cell lines, animal models, and clinical observations are explained.     

Inhibition of prostate tumor angiogenesis by the tumor suppressor CEACAM1

We have previously shown that CEACAM1, a cell-adhesion molecule, acts as a tumor suppressor in prostate carcinoma. Expression of CEACAM1 in prostate cancer cells suppresses their growth in vivo. However, CEACAM1 has no effect on the growth of prostate cancer cells in vitro. This difference suggests that the antitumor effect of CEACAM1 may be due to inhibition of tumor angiogenesis, perhaps by increased secretion of antiangiogenic molecules from the cells. In this study, we have demonstrated that expression of CEACAM1 in DU145 prostate cancer cells induced the production of a factor or factors that specifically blocked the growth of endothelial but not epithelial cells. Conditioned medium from the CEACAM1-expressing cells but not control luciferase-expressing cells inhibited endothelial cell migration up a gradient of stimulatory vascular endothelial growth factor in vitro and inhibited corneal neovascularization induced by basic fibroblast growth factor in vivo. Moreover, conditioned medium from CEACAM1-expressing cells induced endothelial cell apoptosis in vitro. Only medium conditioned by CEACAM1 mutants that were able to suppress tumor growth in vivo could cause endothelial cell apoptosis. These observations suggest that CEACAM1-mediated tumor suppression in vivo is, at least in part, due to the ability of CEACAM1 to inhibit tumor angiogenesis.     

EGCG inhibited bladder cancer SW780 cell proliferation and migration both in vitro and in vivo via down-regulation of NF-κB and MMP-9

Epigallocatechin-3-gallate (EGCG), the bioactive polyphenol in green tea, has been demonstrated to have various biological activities. Our study aims to investigate the antiproliferation and antimigration effects of EGCG against bladder cancer SW780 cells both in vitro and in vivo. Our results showed that treatment of EGCG resulted in significant inhibition of cell proliferation by induction of apoptosis, without obvious toxicity to normal bladder epithelium SV-HUC-1 cells. EGCG also inhibited SW780 cell migration and invasion at 25–100 μM. Western blot confirmed that EGCG induced apoptosis in SW780 cells by activation of caspases-8, -9 and -3, Bax, Bcl-2 and PARP. Besides, animal study demonstrated that EGCG [100 mg/kg, intraperitoneal (i.p.) injection daily for 3 weeks] decreased the tumor volume significantly in mice bearing SW780 tumors, as well as the tumor weight (decreased by 68.4%). In addition, EGCG down-regulated the expression of nuclear factor-kappa B (NF-κB) and matrix metalloproteinase (MMP)-9 in both protein and mRNA level in tumor and SW780 cells. When NF-κB was inhibited, EGCG showed no obvious effect in cell proliferation and migration. In conclusion, our study demonstrated that EGCG was effective in inhibition SW780 cell proliferation and migration, and presented first evidence that EGCG inhibited SW780 tumor growth by down-regulation of NF-κB and MMP-9.     

Epigallocatechin-3-gallate (EGCG), a green tea polyphenol, suppresses hepatic gluconeogenesis through 5'-AMP-activated protein kinase

Epigallocatechin-3-gallate (EGCG), a main catechin of green tea, has been suggested to inhibit hepatic gluconeogenesis. However, the exact role and related mechanism have not been established. In this study, we examined the role of EGCG in hepatic gluconeogenesis at concentrations that are reachable by ingestion of pure EGCG or green tea, and are not toxic to hepatocytes. Our results show in isolated hepatocytes that EGCG at relatively low concentrations (相似文献   

Neutrophil restraint by green tea: inhibition of inflammation,associated angiogenesis,and pulmonary fibrosis

Neutrophils play an essential role in host defense and inflammation, but the latter may trigger and sustain the pathogenesis of a range of acute and chronic diseases. Green tea has been claimed to exert anti-inflammatory properties through unknown molecular mechanisms. We have previously shown that the most abundant catechin of green tea, (-)epigallocatechin-3-gallate (EGCG), strongly inhibits neutrophil elastase. Here we show that 1) micromolar EGCG represses reactive oxygen species activity and inhibits apoptosis of activated neutrophils, and 2) dramatically inhibits chemokine-induced neutrophil chemotaxis in vitro; 3) both oral EGCG and green tea extract block neutrophil-mediated angiogenesis in vivo in an inflammatory angiogenesis model, and 4) oral administration of green tea extract enhances resolution in a pulmonary inflammation model, significantly reducing consequent fibrosis. These results provide molecular and cellular insights into the claimed beneficial properties of green tea and indicate that EGCG is a potent anti-inflammatory compound with therapeutic potential.     

Epigallocatechin-3-gallate (EGCG) inhibits the migratory behavior of tumor bronchial epithelial cells

Background

Many studies associated the main polyphenolic constituent of green tea, (-)-Epigallocatechin-3-gallate (EGCG), with inhibition of cancers, invasion and metastasis. To date, most of the studies have focused on the effect of EGCG on cell proliferation or death. Since cell migration is an important mechanism involved in tumor invasion, the aim of the present work was to target another approach of the therapeutic effect of EGCG, by investigating its effect on the cell migratory behavior.

Methods

The effect of EGCG (at concentrations lower than 10 μg/ml) on the migration speed of invasive cells was assessed by using 2D and 3D models of cell culture. We also studied the effects of EGCG on proteinases expression by RT-PCR analysis. By immunocytochemistry, we analyzed alterations of vimentin organization in presence of different concentrations of EGCG.

Results

We observed that EGCG had an inhibitory effect of cell migration in 2D and 3D cell culture models. EGCG also inhibited MMP-2 mRNA and protein expression and altered the intermediate filaments of vimentin.

Conclusion

Taken together, our results demonstrate that EGCG is able to inhibit the migration of bronchial tumor cells and could therefore be an attractive candidate to treat tumor invasion and cell migration.     

Green tea polyphenol (−)‐epigallocatechin gallate suppressed the differentiation of murine osteoblastic MC3T3‐E1 cells

Recently, various physiological effects of the tea polyphenol catechin for alleviating diseases such as cancer, arteriosclerosis, hyperlipidaemia and osteoporosis have been reported. However, the physiological effect of catechin on bone metabolism remains unclear. We examined the physiological effect of EGCG [(?)‐epigallocatechin‐3‐gallate], which is the main component of green tea catechin, on osteoblast development using the precursor cell line of osteoblasts, MC3T3‐E1, and co‐culture of the osteoblasts from mouse newborn calvaria and mouse bone marrow cells. Although EGCG did not affect the viability and proliferation of MC3T3‐E1 cells, EGCG inhibited the osteoblast differentiation. Furthermore, EGCG did not affect the mineralization of differentiated MC3T3‐E1 cells, and reduced osteoclast formation in co‐culture. These results suggest that EGCG can effectively suppress bone resorption, and can be used as an effective medicine in the treatment of the symptoms of osteoporosis.     

低分子量透明质酸寡糖片段介导内皮细胞增殖的信号通路

为研究低分子量透明质酸寡糖片段(hyaluronan oligosaccharides, o-HA)对血管内皮细胞生长与迁移的影响,及透明质酸（hyaluronan,HA）受体（CD44与RHAMM）在此过程中的作用,首先通过细胞计数、MTT实验、细胞周期分布及单层细胞损伤模型修复实验,观察o-HA对血管内皮细胞（猪髂总动脉内皮细胞,porcine vascular endothelial cell line,PIEC）增殖及创伤愈合的影响．结果显示,o-HA明显促进血管内皮细胞生长,并且能够促进内皮细胞向创伤区迁移.蛋白质免疫印迹分析证明,o-HA作用于PIECs后,细胞Src激酶、ERK-1/2的磷酸化程度增强,c-Myc蛋白、周期蛋白D1表达水平增高.Src 激酶特异性的化学抑制剂PP2可轻度抑制ERK-1/2磷酸化;进而通过抗-CD44与抗-RHAMM抗体分别预先封闭细胞表面相应的特异性受体位点后,再用o-HA刺激细胞,探讨HA受体在o-HA介导PIECs信号传导过程中的作用．结果显示,抗CD44抗体不能抑制o-HA介导的ERK-1/2磷酸化;而抗RHAMM抗体可轻度抑制o-HA介导的ERK-1/2磷酸化．结果提示,o-HA具有促进血管内皮细胞增殖及创伤愈合的作用,其机制可能是通过血管内皮细胞表面受体RHAMM实现的.该作用可能通过激活Src激酶及细胞内MAPK（ERK-1/2）信号通路,启动早期反应基因转录,诱使c-Myc蛋白高表达,从而促进血管内皮细胞生长．该作用也可能与上调细胞周期蛋白 D1的表达有关．     

Green tea extract inhibits angiogenesis of human umbilical vein endothelial cells through reduction of expression of VEGF receptors

Epidemiological and animal studies have indicated that consumption of green tea is associated with a reduced risk of developing certain forms of cancer. However, the inhibitory mechanism of green tea in angiogenesis, an important process in tumor growth, has not been well established. In the present study, green tea extract (GTE) was tested for its ability to inhibit cell viability, cell proliferation, cell cycle dynamics, vascular endothelial growth factor (VEGF) and expression of VEGF receptors fms-like tyrosine kinase (Flt-1) and fetal liver kinase-1/Kinase insert domain containing receptor (Flk-1/KDR) in vitro using human umbilical vein endothelial cells (HUVECs). GTE in culture media did not affect cell viability but significantly reduced cell proliferation dose-dependently and caused a dose-dependent accumulation of cells in the G1 phase. The decrease of the expression of Flt-1 and KDR/Flk-1 in HUVEC by GTE was detected with immunohistochemical and Western blotting methods. These results suggest that GTE may have preventive effects on tumor angiogenesis and metastasis through reduction of expression of VEGF receptors.     

Lipid raft-associated catechin suppresses the FcepsilonRI expression by inhibiting phosphorylation of the extracellular signal-regulated kinase1/2

The major green tea catechin, (-)-epigallocatechin-3-O-gallate (EGCG), has a suppressive effect on the expression of the high-affinity IgE receptor FcepsilonRI, which is key molecule in the IgE-mediated allergic reactions. Here we show that EGCG binds to the cell surface and highly associates with plasma membrane microdomains, lipid rafts, on the human basophilic KU812 cells. The disruption of these lipid rafts caused a reduction of the amount of raft-associated EGCG and the FcepsilonRI-suppressive effect of EGCG. We also found that EGCG has an ability to inhibit the phosphorylation of the extracellular signal-regulated kinase1/2 (ERK1/2) and that the ERK1/2 specific inhibitor also reduced FcepsilonRI expression. Moreover, the inhibitory effect elicited by EGCG on ERK1/2 was prevented by disruption of rafts. Thus, these results suggest that the interaction between EGCG and the lipid rafts is important for EGCG's ability to downregulate FcepsilonRI expression, and ERK1/2 may be involved in this suppression signal.     

In vitro and in vivo mechanistic study of a novel proanthocyanidin,GC-(4→8)-GCG from cocoa tea (Camellia ptilophylla) in antiangiogenesis

Angiogenesis, the process of blood vessel formation, is critical to tumor growth. Ant-angiogenic strategies demonstrated importance in cancer therapy. Cocoa tea (Camellia ptilophylla), a naturally decaffeinated tea commonly consumed as a healthy drink in southern China, had recently been found to be a potential candidate for antiangiogenesis. A novel proanthocyanidin, GC-(4→8)-GCG, which consisted of gallocatechin and gallocatechin 3-O gallate moieties, was discovered and thought to be one of the effective candidates for antiangiogenesis. Hence, the present study aimed to evaluate the antiangiogenesis activities of GC-(4→8)-GCG in vitro and in vivo, and SU5416 was applied as a positive control. The inhibitory effects of GC-(4→8)-GCG on three important processes involved in angiogenesis, i.e., proliferation, migration and differentiation, were examined using human microvascular endothelial cell line HMEC-1 by MTT assay, scratch assay and tube formation assay, respectively. Using transgenic zebrafish embryos TG(fli1:EGFP)y1/+(AB) as an animal model of angiogenesis, the antiangiogenic effect of GC-(4→8)-GCG was further verified in vivo. Our results demonstrated that GC-(4→8)-GCG significantly inhibited migration (P<.001) and tubule formation (P<.001–.05) of HMEC-1 in dose-dependent manner. Regarding intracellular signal transduction, GC-(4→8)-GCG attenuated the phosphorylation of ERK, Akt and p38 dose-dependently in HMEC-1. In zebrafish embryo, the formation of new blood vessels was effectively inhibited by GC-(4→8)-GCG in a dose-dependent manner after 3 days of treatment (P<.001–.05). In conclusion, these results revealed that our novel proanthocyanidin, GC-(4→8)-GCG might be a potential and promising agent of natural resource to be further developed as an antiangiogenic agent.