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© 2006 American Society for Nutrition J. Nutr. 136:2512-2518, October 2006

Nutrition and Disease

Epigallocatechin Gallate Supplementation Alleviates Diabetes in Rodents1

Swen Wolfram*, Daniel Raederstorff, Mareike Preller, Ying Wang, Sandra R. Teixeira2, Christoph Riegger and Peter Weber

DSM Nutritional Products Ltd, Department of Human Nutrition and Health, CH-4002 Basel, Switzerland

* To whom correspondence should be addressed. E-mail: swen.wolfram{at}dsm.com.


   ABSTRACT
TOP
 ABSTRACT
Introduction
 Materials and Methods
 Results
 Discussion
 LITERATURE CITED
 
As the prevalence of type 2 diabetes mellitus is increasing at an alarming rate, effective nutritional and exercise strategies for the prevention of this disease are required. Specific dietary components with antidiabetic efficacy could be one aspect of these strategies. This study investigated the antidiabetic effects of the most abundant green tea catechin, epigallocatechin gallate (EGCG, TEAVIGO), in rodent models of type 2 diabetes mellitus and H4IIE rat hepatoma cells. We assessed glucose and insulin tolerance in db/db mice and ZDF rats after they ingested EGCG. Using gene microarray and real-time quantitative RT-PCR we investigated the effect of EGCG on gene expression in H4IIE rat hepatoma cells as well as in liver and adipose tissue of db/db mice. EGCG improved oral glucose tolerance and blood glucose in food-deprived rats in a dose-dependent manner. Plasma concentrations of triacylglycerol were reduced and glucose-stimulated insulin secretion was enhanced. In H4IIE cells, EGCG downregulated genes involved in gluconeogenesis and the synthesis of fatty acids, triacylgycerol, and cholesterol. EGCG decreased the mRNA expression of phosphoenolpyruvate carboxykinase in H4IIE cells as well as in liver and adipose tissue of db/db mice. Glucokinase mRNA expression was upregulated in the liver of db/db mice in a dose-dependent manner. This study shows that EGCG beneficially modifies glucose and lipid metabolism in H4IIE cells and markedly enhances glucose tolerance in diabetic rodents. Dietary supplementation with EGCG could potentially contribute to nutritional strategies for the prevention and treatment of type 2 diabetes mellitus.


   Introduction
TOP
 ABSTRACT
 Introduction
Materials and Methods
 Results
 Discussion
 LITERATURE CITED
 
In Asia, green tea is a widely consumed beverage and, for centuries, has been regarded to possess considerable health-promoting effects (1). In recent years, research has mainly focused on the effects of green tea in relation to the prevention of cancer (2) and cardiovascular disease (3). Furthermore, the anti-inflammatory (4), antiarthritic (5), antibacterial (6), antiangiogenic (7), antioxidative (8), antiviral (9), neuroprotective (10) and cholesterol-lowering effects (11) of green tea and isolated green tea constituents are under investigation.

The health-promoting effects of green tea are mainly attributed to its polyphenol content. Green tea is a rich source of polyphenols, especially of flavanols and flavonols, which represent ~30% of fresh leaf dry weight (1). Catechins are the predominant form of the flavanols and are mainly composed of epigallocatechin gallate (EGCG)3, epigallocatechin (EGC), epicatechin gallate (ECG), and epicatechin (EC) (12). Recently, many of the aforementioned beneficial effects of green tea were attributed to its most abundant catechin, EGCG (13–15). It was previously shown that EGCG exerts potent antiobesity effects in mouse and rat models of diet-induced obesity, which are at least partly mediated via a direct impact of EGCG on adipose tissue (16,17). EGCG causes a dose-dependent decrease of in vitro adipocyte differentiation and downregulates the mRNA expression of several lipogenic genes in adipose tissue.

The prevalence of type 2 diabetes mellitus (T2DM) is estimated to increase dramatically during the next few years, reaching 300 million by 2025 (18). The increasing prevalence of diabetes is largely due to the rapid spread of obesity, which is considered the most important risk factor for T2DM (19). T2DM is characterized by an imbalance between insulin secretion of the pancreatic beta cells and insulin action on skeletal muscle, adipose tissue, and liver (20). It is generally accepted that loss of the early insulin secretory response initially leads to postprandial hyperglycemia (21). As the disease progresses, the increase in endogenous glucose production (EGP) by the liver contributes to the development of clinical fasting hyperglycemia (22). Therefore, enhancing early insulin secretion and reducing endogenous glucose production have become important goals for the treatment of T2DM.

A study by Waltner-Law et al. (23) provided compelling in vitro evidence that EGCG decreases glucose production of H4IIE rat hepatoma cells. The investigators showed, furthermore, that EGCG mimics insulin, increases tyrosine phosphorylation of the insulin receptor and the insulin receptor substrate, and reduces gene expression of the gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK). If these effects are relevant for the in vivo situation, then EGCG possesses the potential to exert strong antidiabetic effects. Recently, green tea and green tea extracts were demonstrated to beneficially modify glucose metabolism in experimental models of T2DM (24,25). On the other hand, there is only one in vivo study suggesting a glucose-lowering effect of EGCG (26). In that study, EGCG was injected into lean and obese Zucker rats. This route of administration ensured suprapharmacologic plasma concentrations of EGCG and resulted in markedly decreased blood glucose and insulin levels. However, it is unclear whether these observations were due to a direct glucose lowering effect of EGCG or an anorectic effect caused by suprapharmacologic plasma concentrations of EGCG.

It was shown that EGCG ameliorates cytokine-induced beta-cell damage in vitro (27) and prevents the decrease of islet mass induced by treatment with multiple low doses of streptozotocin in vivo (28). However, in the latter study, streptozotocin was coinjected with EGCG, which possesses strong antioxidative activity (29). It is unclear whether the protective effects observed in this study were due to direct inactivation of the injected streptozotocin.

Thus, the antidiabetic effects of EGCG are still not entirely clarified. The in vivo relevance of potentially antidiabetic green tea catechins remains to be demonstrated. Therefore, we conducted an in vivo study to explore the antidiabetic effects of dietary supplementation with the most abundant green tea catechin, EGCG. Additionally, we assessed the effects of EGCG on the expression of genes involved in lipid and glucose metabolism in H4IIE rat hepatoma cells as well as in liver and adipose tissue of db/db mice.


   Materials and Methods
TOP
 ABSTRACT
 Introduction
 Materials and Methods
Results
 Discussion
 LITERATURE CITED
 
    Animal care. The present study was performed with permission of the "Kantonale Veterinäramt Basel" according to Swiss law. The investigation conformed to the National Institutes of Health (NIH) Guide for the Care and Use of Laboratory Animals (30). Mice and rats were individually housed in standard macrolon type 3 and 4 cages, respectively. All animals were maintained on a 12-h light (300 Lux) and 12-h dark cycle at a humidity of 55–60% and a temperature of 23 ± 1°C. All animals consumed modified AIN-93 diets (Provimi Kliba AG) (31) and water ad libitum.

    Diabetes alleviation in db/db mice. The effect of dietary EGCG (TEAVIGO, DSM Nutritional Products) supplementation on T2DM was investigated by utilizing the db/db mouse model (BKS.Cg-m+/+ Leprdb). TEAVIGO is a highly purified extract from green tea leaves (Camellia sinensis) containing >94% EGCG, <5% other catechins (<3% epicatechin gallate). Male db/db mice were purchased from Jackson Laboratories at an age of 5 wk. After an acclimation period of 2 wk, mice consumed a modified AIN-93 diet containing EGCG at concentrations of 2.5, 5.0, or 10.0 g/kg of diet (EGCG 0.25%, 0.5%, or 1% w:w, n = 9/group) or placebo (control, n = 9) for 7 wk. Five mice receiving the thiazolidinedione rosiglitazone (Avandia, GlaxoSmithKline) at a concentration of 72.0 mg/kg of diet (Rosi 0.0072%) were used as a positive control group. Food intake and body weight were determined weekly. Blood glucose levels in fed mice were measured weekly (Glucotrend, Roche Diagnostics). After 5 wk of dietary EGCG supplementation, an oral glucose tolerance test was performed. Before application of an oral glucose load (1 g/kg, Sigma), blood glucose levels were determined in food-deprived mice. Blood glucose levels were measured 15, 30, 45, 60, 90, 120, 150, and 180 min after glucose application (Glucotrend, Roche Diagnostics). After 6 wk of dietary EGCG supplementation, a modified intraperitoneal insulin tolerance test was performed. Immediately prior to the test, all mice were deprived of food. Short-acting insulin (1 IU/kg, Actrapid, Novo Nordisk Pharma GmbH) was administered and blood glucose levels were measured 30, 60, 90, 120, 180, 240, 300, and 360 min after administration. At the end of the study (7 wk), blood samples were collected in the morning from fed mice and analyzed for glucose, free fatty acids, and triacylglycerol concentrations (Hitachi 912 Automatic Analyser). Plasma insulin was determined by use of an enzyme immumometric assay kit (Mercodia AB) in fed mice.

To determine the efficacy of low-dosage treatment, EGCG was administered orally by gavage at dosages of 30 mg · kg–1 · d–1 (EGCG 30, n = 10) and 100 mg · kg–1 · d–1 (EGCG 100, n = 10) for 2 wk in a separate set of mice (10 wk of age). EGCG was dissolved in distilled water and a total volume of 0.1 mL was administered daily. The control group (control, n = 10) was administered 0.1 mL distilled water. The thiazolidinedione rosiglitazone (Avandia, GlaxoSmithKline) was suspended in distilled water and administered at a dosage of 10 mg · kg–1 · d–1 in a positive control group (Rosi 10, n = 5). After 2 wk of application, an oral glucose tolerance test was carried out as described above. Plasma concentrations of insulin in fed mice were also determined.

    Diabetes alleviation in ZDF rats. The effect of dietary EGCG supplementation on T2DM was further investigated in ZDF rats (ZDF/GmiCrl-Leprfa). Male ZDF rats were purchased from Charles River at an age of 5 wk. After an acclimation period of 2 wk, rats consumed a modified AIN-93 diet containing EGCG at a concentration of 5 g/kg of diet (EGCG 0.5% w:w, n = 10) or placebo (control, n = 10) for 10 wk. Ten rats receiving the thiazolidinedione rosiglitazone (Avandia, GlaxoSmithKline) at a concentration of 36.0 mg/kg of diet (Rosi 0.0036%) were used as a positive control group. Food intake and body weight were determined twice per wk. After 10 wk of treatment, an oral glucose tolerance test was carried out as described above. The plasma concentration of insulin was determined prior and 15, 30, and 45 min after glucose administration, as described above. At the end of the study, blood of all rats was collected, the plasma prepared and stored at –80°C until analysis of total EGCG plasma concentration was performed, as previously described (32).

    Real-time quantitative TaqMan RT-PCR. Real-time quantitative TaqMan RT-PCR was used to quantify gene expression as previously described (16). Details are provided in the online supplemental material.

    Gene regulation of glucose and lipid metabolism in rat H4IIE hepatoma cells. The influence of EGCG on glucose and lipid metabolism-related genes were studied in rat hepatoma cell line H4IIE cells (ATCC Global Bioresource Center). Cells were cultured and gluconeogenesis was induced as described by Waltner-Law et al. (23). Cells were starved for 4 or 24 h before being treated with either insulin 10 nmol/L (positive control), or EGCG at 50 or 100 µmol/L. Final DMSO concentration was standardized to 0.5%. Details are provided in the online supplemental material. Glucose-6-phosphotase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK) expression levels were checked by real-time PCR as previously described (16). DNA microarray analysis was performed to investigate the effect of EGCG on genome-wide expression as previously described (33). Details are provided in the online supplemental material.

    Statistical analysis. Data from db/db mice and ZDF rats are expressed as means ± SEM for animals in each group. Statistical significance of the mean differences between dietary groups was tested by 1-way ANOVA or repeated-measures ANOVA. Homogenity of variance was tested using Levene's test. When ANOVA revealed significant differences, means were compared using LSD post-hoc analysis (SPSS 13.0). Differences were considered significant at P < 0.05. RT-PCR data were analyzed by 1-way ANOVA. If significant differences were found, Dunnett's test for multiple comparison (Statistica, version 5.5A, StatSoft) was used to compare each group to the control group. Significance was determined at P < 0.05. The gene chip data analysis was carried out using RACE-A (Roche Affymetrix Chip Experiment Analysis), a Roche proprietary software package for differential expression analysis, as described (33). Gene expression in EGCG treated cells was compared with that of solvent-treated cells. EGCG-induced changes in gene expression levels are reported as percentage of control relative to solvent-treated cells, which were set to 100%. Changes were considered significant at P < 0.05 for a single gene regulation by EGCG.


   Results
TOP
 ABSTRACT
 Introduction
 Materials and Methods
 Results
Discussion
 LITERATURE CITED
 
    Diabetes alleviation in db/db mice. Dietary EGCG supplementation for 5 wk improved oral glucose tolerance in db/db mice in a dose-dependent manner (Fig. 1A, B). When EGCG was applied orally by gavage for 2 wk, a dosage of 100 mg · kg–1 · d–1 improved oral glucose tolerance (Fig. 1D, E).


Figure 1
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  		Figure 1  Blood glucose concentrations during an oral glucose tolerance test (oGTT) (A, D), calculated area under the curve (AUC) of the oGTT (B, E) and blood glucose concentrations of food-deprived db/db mice (C, F). Mice consumed diets supplemented with 0.25%, 0.5%, or 1% EGCG (n = 9) or 0.0072% rosiglitazone (n = 5) for 5 wk (A, B, C), or were administered orally by gavage 30 or 100 mg · kg–1 · d–1 EGCG (n = 10) or 10 mg · kg–1 · d–1 rosiglitazone (n = 5) for 2 wk (D, E, F). Values are means ± SEM. Means without a common letter differ, P < 0.05.

 
A pronounced decrease of glucose levels was observed in food-deprived db/db mice treated with EGCG (Fig. 1C, F). Dietary supplementation with 0.25%, 0.5%, and 1% EGCG resulted in reductions of glucose levels in food-deprived mice by 23.0%, 35.2%, and 47.6%, respectively. Compared with control mice, dietary supplementation with 0.25%, 0.5%, and 1% EGCG decreased glucose levels in fed mice by 2.4%, 11.6%, and 36.9%, respectively (Table 1).


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  		TABLE 1 Plasma concentrations of glucose, insulin, free fatty acids, and triacylglycerol in fed db/db mice and ZDF rats after supplementation with EGCG or rosiglitazone1

 
In mice that were deprived of food immediately before the test, the glucose-lowering effects of insulin were not significantly altered by EGCG treatment during the early time points of the insulin tolerance test. In control mice, blood glucose concentrations rebounded between 1 and 3 h after insulin administration to 69.9% of baseline. However, in mice treated with 1% EGCG, this rebound was not observed (Fig. 2A). The plasma concentrations of triacylglycerol were reduced by 0.25%, 0.5%, and 1% EGCG (Table 1). The plasma concentrations of insulin (plasma collected in the morning at 0800 from fed mice) were increased by EGCG supplementation in a dose-dependent manner (Table 1). In a previous study (16), we observed that EGCG supplementation tended to reduce plasma insulin concentrations in fed animals in 2 models of diet-induced obesity (C57BL/6J mice and Sprague Dawley rats) (for comparison, see Fig. 2B).


Figure 2
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  		Figure 2  Blood glucose concentrations during an intraperitoneal insulin tolerance test of control db/db mice (n = 9) and db/db mice fed a diet containing 1% EGCG (n = 9) for 6 wk (A). There were effects of time (P < 0.001), group (P = 0.011), and time x group (P < 0.001). Plasma insulin concentrations of fed C57BL/6J mice that consumed a high fat diet (control, n = 8) or a high fat diet supplemented with 1% EGCG (n = 8) for 5 mo, Sprague Dawley rats (SD) fed a high fat diet for 5 mo followed by either a high fat diet alone (control, n = 10) or a high fat diet supplemented with 1% EGCG for 4 wk (n = 10), and db/db mice fed an AIN-93 diet (n = 9) or an AIN-93 diet supplemented with 1% EGCG (n = 9) (B). Insulin concentrations of Sprague Dawley rats and C57BL/6J mice were obtained from a previous study (16). Values are means ± SEM. Means without a common letter differ, P < 0.05.

 
The cumulative food intake between control mice and mice supplemented with 0.25%, 0.5% or 1% EGCG did not differ (Table 2). However, the positive control rosiglitazone caused a significant reduction in cumulative food intake. Conversely, body weight increased in mice treated with rosiglitazone compared with control mice. EGCG supplementation did not exert a significant influence on final body weight. When EGCG was applied orally by gavage, cumulative food intake and body weight did not change compared with control mice (Table 2). Rosiglitazone applied orally by gavage for 2 wk increased body weight (Table 2).


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  		TABLE 2 Body weight, food intake and adipose tissue mass of db/db mice and ZDF rats after supplementation with EGCG or rosiglitazone1

 
    Diabetes alleviation in ZDF rats. Dietary EGCG supplementation for 10 wk improved oral glucose tolerance in ZDF rats (Fig. 3A, B). Plasma insulin concentrations of EGCG-treated rats were increased at 15 min after glucose application compared with control rats (Fig. 3C). Plasma concentrations of free fatty acids were lower in EGCG supplemented rats (Table 1). Supplementation of 0.5% EGCG resulted in total EGCG plasma concentrations of 97.9 ± 17.7 µg/L. EGCG was not detectable in the plasma of control ZDF rats nor in ZDF rats treated with rosiglitazone.


Figure 3
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  		Figure 3  Blood glucose concentrations during an oral glucose tolerance test (oGTT) of untreated ZDF rats, ZDF rats supplemented with 0.5% EGCG, and ZDF rats treated with 0.0036% rosiglitazone (5 mg · kg–1 · d–1) for 10 wk (A). Calculated area under the curve (AUC) of the oGTT (B). Plasma insulin concentrations before and 15, 30, and 45 min after glucose administration (C). Values are means ± SEM, n = 10/group. Means without a common letter differ, P < 0.05.

 
    Gene expression in liver and adipose tissue of db/db mice. The mRNA expression of glucokinase (GK) was increased in the livers of mice supplemented with EGCG in a dose-dependent manner (Table 3). Rosiglitazone treatment increased GK mRNA levels to a similar extent. A decrease of PEPCK mRNA expression was observed in adipose tissue of EGCG supplemented mice (Table 3). High-dose EGCG supplementation increased acyl-CoA oxidase-1 (ACO-1) and carnitine palmitoyl transferase-1ß (CPT-1) mRNA expression in liver and adipose tissue (Table 3). In adipose tissue, rosiglitazone treatment caused a pronounced upregulation of CPT-1 mRNA expression (830% in untreated control mice; Table 3).


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  		TABLE 3 Gene expression in liver and adipose tissue of db/db mice supplemented with EGCG or rosiglitazone for 7 wk12

 
    Gene regulation of glucose and lipid metabolism in rat H4IIE hepatoma cells. Real-time quantitative TaqMan RT-PCR analysis showed that EGCG decreased the Dex/cAMP-induced gene expression of the gluconeogenic enzymes PEPCK and G6Pase in a time- and dose-dependent manner. Expression of PEPCK was reduced by 11.2 or 32.2% after 4 h and 46.6 or 69.2% after 24 h at 50 or 100 µmol/L EGCG, respectively (Table 4). Expression of G6Pase was decreased by 16.1 or 58.1% after 4 h and 47.2 or 87.4% after 24 h at 50 or 100 µmol/L EGCG, respectively (Table 4).


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  		TABLE 4 Gene expression in H4IIE rat hepatoma cells after treatment with EGCG for 4 or 24 h12

 
DNA microarray analysis revealed that EGCG significantly modulates expression of genes involved in lipid and glucose metabolism. Overall lipid metabolism was downregulated, as seen by reduced expression of genes involved in fatty acid synthesis (fatty acid synthase and stearoyl-CoA desaturase 1 and 2), oxidation (short-chain acyl-CoA dehydrogenase) and activation (acyl-CoA synthetase 2), as well as triacylglycerol synthesis (glycerol-3-phosphate acyltransferase) and cholesterol synthesis (3-hydroxy-3-methylglutaryl-CoA synthase 1, mevalonate kinase). Genes involved in gluconeogenesis (glucose-6-phosphatase, fructose-1,6-bisphosphatase 1) were downregulated, whereas genes involved in glycolysis (phosphofructokinase) and glucose transport (glucose transporter 1) were increased (Table 5).


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  		TABLE 5 DNA microarray analysis of H4IIE rat hepatoma cells after treatment with 50 µmol/L EGCG for 24 h12

 

   Discussion
TOP
 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
LITERATURE CITED
 
This study demonstrates that dietary supplementation of EGCG markedly ameliorates T2DM in preclinical models. Thus, to our knowledge, we provide the first evidence that EGCG consumed in the diet exerts potent antidiabetic activity in vivo in a dose-dependent manner.

Our findings clearly show that EGCG enhances oral glucose tolerance in severely diabetic db/db mice and in moderately diabetic ZDF rats. The in vivo and in vitro findings suggest that the reduction of EGP and increase in glucose-induced insulin secretion contribute to the antidiabetic effects of EGCG. EGP is the main determinant of fasting glucose levels. EGCG supplementation causes a pronounced decrease of glucose levels in food-deprived animals, which exceeds the decrease of glucose levels in animals not deprived of food. Additionally, the results of a modified insulin tolerance test performed in db/db mice, which were deprived of food just prior to the test, point in the same direction. No effect of EGCG supplementation during the insulin-mediated decrease in blood glucose was observed. Instead, we observed a significant difference in blood glucose starting 3 h after the injection of rapid-acting insulin. The difference was caused by absence of the rebound in blood glucose in EGCG supplemented db/db mice. This again suggests that EGP is decreased by EGCG. Further euglycemic-hyperinsulinemic clamp studies for the determination of EGP, gluconeogenesis, and glycogenolysis will provide a better mechanistic understanding.

Furthermore, our results imply that EGCG supplementation influences the expression of genes involved in glucose and lipid metabolism in the liver as well as in H4IIE rat hepatoma cells. The same pattern of downregulation of gluconeogenic enzymes and upregulation of glycolytic enzymes was found in vitro and in vivo, suggesting that reduced EGP at least partially contributed to the enhanced glucose tolerance of mice supplemented with EGCG. The observed decrease in PEPCK mRNA expression as a key gluconeogenic enzyme is in line with other published in vitro evidence (23).

To our knowledge, we report for the first time that EGCG supplementation causes a dose-dependent increase in GK mRNA expression in livers of db/db mice. In the liver, an increase in GK activity leads to enhanced glycolysis and hepatic glucose uptake (34). However, in a diet-induced obesity model, EGCG supplementation led to reduced expression of GK mRNA in the liver, suggesting that the upregulation of GK observed in our study may be a secondary effect (17).

The increase in insulin concentrations in fed db/db mice supplemented with EGCG could either be caused by direct stimulation of insulin secretion in response to feeding or by a protective effect of EGCG on the pancreas. The results of our study are in line with recently published data, which suggest that EGCG preserves and protects the pancreas by its strong antioxidative capacity (27,28). This would ultimately lead to enhanced pancreatic function and improved insulin secretion in response to feeding. Our study in ZDF rats showed that EGCG enhances glucose-stimulated insulin secretion. This is especially important in preclinical models such as db/db mice and ZDF rats that are characterized by a progressive decline of pancreatic function due to subsequent beta-cell failure and loss of beta-cell mass.

Interestingly, in 2 models of diet-induced obesity, characterized by moderately increased insulin concentrations in fed animals, EGCG supplementation resulted in decreased insulin levels. There are 2 possible explanations for this finding. First, EGCG supplementation could increase insulin concentrations in fed animals only if glucose levels reach pathophysiologically high concentrations, which is not the case in the utilized diet-induced obesity models. Second, the prevention of diet-induced obesity due to EGCG supplementation could reduce peripheral insulin resistance, thereby reducing levels of circulating insulin.

Our results indicate that there is an increase of in vivo fatty acid oxidation due to EGCG supplementation by the upregulation of CPT-1 and ACO-1 mRNA levels in liver and adipose tissue of db/db mice. Furthermore, DNA microarray analysis of H4IIE rat hepatoma cells showed that genes involved in the synthesis of fatty acids, triacylglycerol, and cholesterol are strongly downregulated when treated with EGCG. These results are in line with findings that EGCG prevents diet-induced obesity in rodents (16,17) and that green tea extract promotes fat oxidation in mice and humans (35,36).

Similar to previous findings in diet-induced obesity models (16,17), EGCG supplementation in db/db mice did not influence food intake. Thus, we can exclude the possibility that EGCG enhances glucose tolerance simply by reducing food intake. The results of the present study emphasize a direct effect of EGCG on the improvement of glucose metabolism, which could be mediated through a decrease of hepatic glucose output and an increase of glucose-stimulated insulin secretion.

Applying EGCG orally by gavage resulted in significantly improved glucose tolerance and reduced glucose levels in food-deprived db/db mice at a dosage of 100 mg · kg–1 · d–1. The effects obtained were similar to the effects observed after db/db mice were supplemented with 0.25% EGCG (with respect to food intake and body weight this concentration is equal to 375 mg · kg–1 · d–1) and 0.5% EGCG in ZDF rats (with respect to food intake and body weight this concentration is equal to 437 mg · kg–1 · d–1). This suggests that in rodents the bioavailability of EGCG is reduced by ~4-fold when administered with food. Due to the limited amounts of plasma obtained from db/db mice, we could not determine total EGCG plasma concentrations. Therefore, we analyzed total EGCG plasma concentrations in ZDF rats supplemented with 0.5% EGCG. Total EGCG plasma concentrations of 97.9 µg/L in rats suggest a relatively low bioavailability and a fast metabolism of EGCG. Our findings agree with a recent pharmacokinetics study in mice. Lambert et al. (37) showed that intragastric administration of EGCG at a dosage of 75 mg/kg results in a Cmax of ~128 µg/L total plasma EGCG and a terminal half-life of ~83 min. Furthermore, we showed that in humans an oral intake of EGCG at a dosage of 50 mg (~0.7 mg/kg) results in a Cmax of ~130 µg/L total plasma EGCG and a terminal half-life of ~112 min (32). Those results indicate that rodents must be orally administered 100- to 600-fold more EGCG (depending on whether they are administered by gavage or by feed admixture) to achieve similar plasma concentrations as those found in humans. Total plasma EGCG concentrations shown to be efficacious in mice and rats can be reached by an intake of low-to-moderate dosages of EGCG. Therefore, we expect that our results could have relevance for humans with T2DM.

In conclusion, this study demonstrates that among various green tea catechins, EGCG possesses pronounced antidiabetic efficacy in preclinical models of T2DM. The effects of EGCG are at least partially mediated through reduced hepatic glucose production and enhanced pancreatic function. Our data suggest that supplementation with EGCG could potentially improve glucose tolerance in humans with T2DM. This hypothesis should now be investigated in randomized placebo-controlled trials.


   ACKNOWLEDGMENTS
 
The authors thank Conrad Wyss for skilled animal experimentation, Silvia Manz and Erna Fuhrer for excellent technical assistance, and Arneet Saltzman for a critical reading of the article.


   FOOTNOTES
 
1 Supplemental Materials and Methods are available with the online posting of this paper at jn.nutrition.org. Back

2 Present address: Diabetes and Metabolism, Novartis Institutes for BioMedical Research Inc. (NIBRI), 100 Technology Square, Cambridge, Massachusetts, MA 02139. Back

3 Abbreviations used: EGCG, epigallocatechin gallate; EGP, endogenous glucose production; GK, glucokinase; PEPCK, phosphoenolpyruvate carboxykinase; rosi, rosiglitazone; T2DM, type 2 diabetes mellitus. Back

Manuscript received 15 March 2006. Initial review completed 16 April 2006. Revision accepted 29 June 2006.


   LITERATURE CITED
TOP
 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
 LITERATURE CITED
 

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