(-)-Epigallocatechin Gallate Inhibits the Pacemaker Activity of Interstitial Cells of Cajal of Mouse Small Intestine

The effects of (-)-epigallocatechin gallate (EGCG) on pacemaker activities of cultured interstitial cells of Cajal (ICC) from murine small intestine were investigated using whole-cell patch-clamp technique at 30℃ and Ca²⁺ image analysis. ICC generated spontaneous pacemaker currents at a holding potential of -70 mV. The treatment of ICC with EGCG resulted in a dose-dependent decrease in the frequency and amplitude of pacemaker currents. SQ-22536, an adenylate cyclase inhibitor, and ODQ, a guanylate cyclase inhibitor, did not inhibit the effects of EGCG. EGCG-induced effects on pacemaker currents were not inhibited by glibenclamide, an ATP-sensitive K⁺ channel blocker and TEA, a Ca²⁺-activated K⁺ channel blocker. Also, we found that EGCG inhibited the spontaneous [Ca²⁺]_i oscillations in cultured ICC. In conclusion, EGCG inhibited the pacemaker activity of ICC and reduced [Ca²⁺]_i oscillations by cAMP-, cGMP-, ATP-sensitive K+ channel-independent manner.     

Exposure and toxicity of green tea polyphenols in fasted and non-fasted dogs

Standardized green tea extract was evaluated for exposure and toxicity in Beagle dogs following oral dosing by capsules. The main component (−)-epigallocatechin gallate (EGCG) accounted for 56–72% of the material. A 9-month chronic study (0, 200, 500, and 1000 mg/kg/day) was done in fasted dogs to take advantage of the reported improved catechin bioavailability with fasting. Extensive morbidity, mortality, and pathology of many major organs led to its early termination at 6.5 months and prevented identification of the toxicity mechanisms. A follow-up 13-week study examined the exposure to and toxicity of the extract. In general, toxicities were less severe than in the chronic study during the same interval. Dosing in a fed state resulted in considerably lower and less variable exposure than found under fasted conditions. Toxicity was less frequent and of lesser severity with lower exposure but limited sample size and large variability prevented reaching that definitive conclusion. Differences in mortality and morbidity between the preliminary terminated chronic and follow-up subchronic studies with the same dose of the same drug lot and similar exposure were not fully resolved as there may be other as yet unclear confounding factors.     

棓丙酯对大鼠 小鼠肠平滑肌运动的影响

目的：观察不同浓度棓丙酯对大鼠离体小肠收缩活动及小鼠在体小肠推进运动的影响。方法：采用大鼠离体肠管和小鼠小肠推进运动实验,考察棓丙酯对肠平滑肌运动的影响。结果：棓丙酯在10~80μmol/L浓度范围内剂量依赖性抑制大鼠离体小肠平滑肌收缩（P〈0.05,P〈0.01）,拮抗乙酰胆碱及组胺引起的促进收缩作用（P〈0.01）,且相加阿托品、槲皮素引起的抑制收缩作用（P〈0.01）。棓丙酯高剂量组（200mg/kg）、中剂量组（100mg/kg）对小鼠在体小肠推进运动有明显的抑制作用（P〈0.05,P〈0.01）。结论：棓丙酯对大鼠、小鼠肠平滑肌运动具有明显抑制作用。     

HPLC法同时测定绿茶中表没食子儿茶素没食子酸酯和咖啡因的含量

目的:建立同时测定绿茶中表没食子儿茶素没食子酸酯(EGCG)和咖啡因含量的方法。方法:采用高效液相色谱法。色谱柱为DiamonsilTMC18(200mm×4.6mm,5μm),流动相为甲醇-0.1%冰醋酸水溶液(25:75),检测波长为278nm,流速为1.0mL.min-1,柱温为30℃。结果:EGCG和咖啡因的进样量分别在1.25～10.00μg(r=0.9994)和0.25～2.00μg(r=0.9996)范围内与各自峰面积积分值呈良好的线性关系;平均加样回收率分别为98.06%和99.37%,RSD分别为1.05%和1.44%(n均为6)。结论:该方法简便、准确、重复性好,可用于绿茶的质量控制。     

表没食子儿茶素没食子酸酯对高糖环境下体外小鼠足细胞活性氧的影响

目的　观察不同浓度表没食子儿茶素没食子酸酯（EGCG）对高糖造成氧化应激状态下体外小鼠足细胞损害的作用并探讨其机制。 方法　以高糖（25 mmol/L）培养的小鼠足细胞为研究对象,维生素E培养为对照。首先以MTT法检测细胞活力,再在激光共聚焦显微镜下以CM-H2DCFDA荧光探针观察不同浓度EGCG（0.2、10、100 μmol/L）刺激足细胞6、12、24 h后活性氧（ROS）生成,并以流式细胞仪定量分析ROS平均荧光强度。RT-PCR法检测足细胞内ROS产生的主要通路NADPH氧化酶各亚基mRNA（ph22phox、p47phox、p67phox）的表达。 结果　高糖刺激下6 h,足细胞ROS生成显著增加（P < 0.01）。正常糖组和甘露醇组培养12 h ROS生成无显著增加（P > 0.05）。EGCG 0.2 μmol/L作用6 h可显著降低高糖环境下体外小鼠足细胞ROS水平（P < 0.01）。与高糖组比较,EGCG（100 μmol/L）显著减少NADPH氧化酶亚基p22phox和p67phox mRNA表达（均P < 0.05）。与维生素Ｅ组比较,EGCG（0.2 μmol/L）和维生素E（0.2 mmol/L）协同作用组显著减少p47phox mRNA表达（P < 0.05）。 结论　EGCG能缓解高糖环境下体外足细胞氧化应激状态,对高糖培养下足细胞有保护作用。     

Effects of oral epigallocatechin gallate on the oral pharmacokinetics of verapamil in rats

Verapamil is known to be a P‐glycoprotein (P‐gp) substrate and norverapamil is formed via hepatic cytochrome P450 (CYP 3A) in the rat. Epigallocatechin gallate (EGCG), a flavonoid, was reported to be an inhibitor of both P‐gp and CYP3A. Hence, it could be expected that EGCG could alter the pharmacokinetics of verapamil. In this study, 9 mg/kg verapamil was administered orally to Sprague–Dawley rats 30 min after the oral administration of 2 and 10 mg/kg of oral EGCG. Compared with the controls, the AUC values of both verapamil (74.3% and 111% increase for 2 and 10 mg/kg EGCG, respectively) and norverapamil (51.5% and 87.2% increase for 2 and 10 mg/kg EGCG, respectively) were significantly greater in the presence of EGCG. However, compared with the controls, both the AUC and the relative bioavailability of verapamil were significantly (p<0.01) increased by 74.3–111% in the presence of EGCG. The likely explanation is inhibition of P‐gp. Inhibition of CYP3A would increase the AUC of verapamil but decrease the AUC of norverampil. However, inhibition of P‐gp would lead to an increase of AUC of both verapamil and norverapamil. Copyright © 2009 John Wiley & Sons, Ltd.     

Hepatotoxicity from green tea: a review of the literature and two unpublished cases

Purpose  To review the current literature on suspected green tea-related hepatic reactions and to describe two new cases reported within the framework of the Italian surveillance system of natural health products. Results  A literature search of publication between 1999 and October 2008 retrieved 34 cases of hepatitis. Histological examination of the liver revealed inflammatory reactions, cholestasis, occasional steatosis, and necrosis. A positive dechallenge was reported in 29 cases. There was one reported death. A positive rechallenge occurred in seven cases (20%). In the two new cases, the causality assessment was judged as “possible” according to the RUCAM score. Conclusions  Our analysis of the published case reports suggests a causal association between green tea and liver damage. The hepatotoxicity is probably due to (-)-epigallocatechin gallate or its metabolites which, under particular conditions related to the patient’s metabolism, can induce oxidative stress in the liver. In a few cases, toxicity related to concomitant medications could also be involved.     

(−)-Catechin promotes adipocyte differentiation in human bone marrow mesenchymal stem cells through PPARγ transactivation

Green tea intake has been shown to confer various health benefits to patients suffering from metabolic disorders. Here, we studied the effect of several major green tea polyphenols on adipocyte differentiation in human bone marrow mesenchymal stem cells (hBM-MSCs) and compared it to the effect of representative antidiabetic drugs. (−)-Catechin was the most potent of the eight green tea polyphenols evaluated in promoting adipocyte differentiation in hBM-MSCs, and this effect was dose-dependent. (−)-Catechin increased the mRNA levels of various adipogenic markers, such as adiponectin, peroxisome proliferator-activated receptor gamma (PPARγ), FABP4, and LPL, as measured during adipocyte differentiation in hBM-MSCs. In addition, (−)-catechin upregulated the secretion of adiponectin in hBM-MSC culture. Using a reporter gene assay and a competitive ligand binding study, (−)-catechin also significantly activated PPARγ in a dose-dependent fashion; however, (+)-catechin, the enantiomer of (−)-catechin, was not effective as a PPARγ agonist, which seems to imply that the effect of (−)-catechin on PPARγ is stereospecific. In conclusion, our data suggest that (−)-catechin promotes adipocyte differentiation and increased sensitivity to insulin in part by direct activation of PPARγ, which could be at the basis of the observed pharmacological benefits of green tea intake in reducing the risk of type 2 diabetes.     

Analysis of Flavonoid‐Based Pharmacophores that Inhibit Aggrecanases (ADAMTS‐4 and ADAMTS‐5) and Matrix Metalloproteinases Through the Use of Topologically Constrained Peptide Substrates

Polyphenolic natural products from green tea and red wine have been identified as metalloproteinase inhibitors. Members from the flavonoid and stilbene families found to possess metalloproteinase inhibitory activities include (?)‐epigallocatechin gallate, (?)‐epicatechin gallate and piceatannol, but their minimally active pharmacophores have not been evaluated. The present study has examined compounds that are structural components of or structurally related to (?)‐epigallocatechin gallate, (?)‐epicatechin gallate and piceatannol for inhibition of aggrecanases and four representative matrix metalloproteinases. Piceatannol and pyrogallol were found to inhibit all aggrecanases and matrix metalloproteinases studied, indicating a crucial reliance on multiple hydroxyl groups for (?)‐epigallocatechin gallate, (?)‐epicatechin gallate and piceatannol activity. Differences in K_i values for pyrogallol as determined with two structurally distinct substrates indicated the likelihood that this compound binds in a non‐competitive modality. Further analysis showed that pyrogallol acts as an exosite inhibitor, consistent with the action of (?)‐epigallocatechin gallate. In contrast, piceatannol was shown to be a competitive binding inhibitor and showed no differences in apparent K_i values as determined by distinct substrates, illustrating the benefits of using two structurally distinct substrates to assist the analysis of protease inhibitors. The compounds identified here could be utilized to develop novel metalloproteinase probes or as fragment components of more active inhibitors.