1156
DOI 10.1002/mnfr.201300621
Mol. Nutr. Food Res. 2014, 58, 1156–1159
FOOD & FUNCTION
Voluntary exercise and green tea enhance the expression of genes related to energy utilization and attenuate metabolic syndrome in high fat fed mice Sudathip Sae-tan1 , Connie J. Rogers2 and Joshua D. Lambert1 1 2
Department of Food Science, The Pennsylvania State University, University Park, PA, USA Department of Nutritional Sciences, The Pennsylvania State University, University Park, PA, USA
Obesity and metabolic syndrome are growing public health problems. We investigated the effects of decaffeinated green tea extract (GTE) and voluntary running exercise (Ex) alone or in combination against obesity and metabolic syndrome in high fat (HF) fed C57BL/6J mice. After 16 wk, GTE + Ex treatment reduced final body mass (27.1% decrease) and total visceral fat mass (36.6% decrease) compared to HF-fed mice. GTE + Ex reduced fasting blood glucose (17% decrease), plasma insulin (65% decrease), and insulin resistance (65% decrease) compared to HF-fed mice. GTE or Ex alone had less significant effects. In the skeletal muscle, the combination of Ex and GTE increased the expression of peroxisome proliferator-activated receptor-␥ coactivator-1␣ (Ppargc1a), mitochondrial NADH dehydrogenase 5 (mt-Nd5), mitochondrial cytochrome b (mt-Cytb), and mitochondrial cytochrome c oxidase III (mt-Co3). An increase in hepatic expression of peroxisome proliferator-activated receptor-␣ (Ppara) and liver carnitine palmitoyl transferase-1␣ (Cpt1a) and a decrease in hepatic expression of stearoyl-CoA desaturase 1 (Scd1) mRNA was observed in GTE + Ex mice. GTE + Ex was more effective than either treatment alone in reducing diet-induced obesity. These effects are due in part to modulation of genes related to energy metabolism and de novo lipogenesis.
Received: August 23, 2013 Revised: October 17, 2013 Accepted: October 18, 2013
Keywords: Catechins / Fatty acid oxidation / Green tea / Mitochondrial biogenesis / Voluntary exercise
Additional supporting information may be found in the online version of this article at the publisher’s web-site
Approximately 34% of adults in the United States are classified as obese (i.e. BMI ≥ 30) [1]. Obesity, in addition to having normal BMI but a high percentage of body fat, represents a risk factor for metabolic syndrome [2,3]. A traditional strategy Correspondence: Dr. Joshua D. Lambert, Department of Food Science, The Pennsylvania State University, 332 Food Science Building, University Park, PA 16802, USA E-mail: [email protected] Fax: +1-814-863-6132 Abbreviations: Acox1, acyl-CoA oxidase; Cpt1a, carnitine palmitoyl transferase-1␣; Ex, voluntary exercise; GTE, decaffeinated green tea extract; HF, high fat; HOMA-IR, homeostasis model assessment of insulin resistance; LF, low fat; mt-Co3, cytochrome c oxidase subunit III; mt-Cytb, cytochrome b; mtNd5, NADH dehydrogenase 5; Ppara, peroxisome proliferatoractivated receptor-␣; Ppargc1a, proliferator-activated receptor-␥ coactivator-1␣; Scd1, stearoyl-CoA desaturase 1; Srebf1, sterol regulatory element-binding protein-1c C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
for the treatment of obesity and metabolic syndrome is life style intervention including diet and exercise. Regular physical activity is useful in weight loss due to increased energy expenditure [4]. Animal studies have examined the effects of forced exercise on reduction of body weight and improvement of symptoms related to metabolic syndrome [5]. This use of forced exercise can have significant impacts on physiological outcome but in its laboratory permutations (i.e. forced running or exhaustive swimming) is less translatable to humans. For example, Sprague-Dawley rats with voluntary access to a running wheel had an 18% greater reduction in body weight than rats treated with forced treadmill [6]. Involuntary (forced) exercise has been shown to elicit greater increase in glucocorticoids than voluntary running [7–9]. Although fewer studies have examined the impact of voluntary exercise (Ex) on weight loss, these have shown that voluntary running reduced fat mass, insulin resistance, and inflammation [10, 11]. www.mnf-journal.com
Mol. Nutr. Food Res. 2014, 58, 1156–1159
1157
Figure 1. The effect of GTE, Ex, or GTE + Ex on body mass (A), fat mass (B), and glycemic markers (C) and (D) in high fat fed male C57BL/6J mice. Body weight and fasting blood glucose were determined weekly and biweekly throughout the experiment, respectively. Visceral fat mass and fasting insulin was determined in the plasma at the end of the experiment. HOMA-IR was determined using the final fasting blood glucose and fasting plasma insulin levels measured in the experiment. N = 12 for LF-mice, N = 22 for other groups, and error bars represent the SD. Body weight and blood glucose data were analyzed by two-way ANOVA with Bonferroni posttest. Different symbols indicate pairwise comparisons between each treatment to HF-fed mice (a, HF versus LF; b, HF versus GTE + Ex; c, HF versus GTE; d, HF versus Ex). Error bars were removed from fasting blood glucose to improve clarity. For fasting blood glucose *P < 0.05 compared to HF. For the other parameters, values with different letters are statistically significantly different by one-way ANOVA with Tukey’s posttest, P < 0.05.
Tea (Camellia sinensis) is a beverage with worldwide popularity. It is rich in catechins with (−)-epigallocatechin-3-gallate is the most abundant [12]. A number of animal studies have shown the preventive effects of green tea polyphenols against obesity [3]. A recent meta-analysis of 11 human trials with green tea preparations reported a 1.31 kg average body weight loss in intervention groups compared to control groups [13]. Although previous studies have shown that forced running or exhaustive swimming in combination with tea catechins can attenuate obesity in mice, to our knowledge there have been no reports on the effects of green tea in combination with Ex on metabolic syndrome in high fat (HF) fed mice [14, 15]. We report the results of our study on the preventive effects of combination treatment with Ex and decaffeinated green tea extract (GTE) in a HF-fed mouse model of metabolic syndrome (see Supporting Information for detailed methodology). Ex or GTE (Supporting Information Table 1) alone resulted in decreased final body weight compared to HF-fed mice (Supporting Information Table 2; 12.0 and 9.4%, respectively, Fig. 1A). Treatment with the combination of GTE and Ex yielded the lowest final body weight (27%, Fig. 1A) and for most of the experiment, the body weight of the combination treated mice overlapped with the low fat (LF) fed controls. There were minor differences (>10%) in average weekly energy intake per mouse in GTE + Ex-treated mice compared to HF-fed controls (Supporting Information Fig. 1A). Both Ex-treated mice and GTE + Ex-treated mice had the similar average daily running activity (Supporting Information C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Fig. 2A). Previously, it has been reported that cotreatment with exhaustive swimming and 0.5% catechins reduced rate of body weight gain by 33% compared to HF controls [14]. GTE and Ex treatments decreased retroperitoneal fat mass (31 and 33%, respectively): Ex treatment also decreased intestinal fat mass (26.2%) compared to HF mice (Fig. 1B). The combination treatment with GTE and Ex reduced retroperitoneal fat mass and intestinal fat mass back to those found in LF-fed control mice. Both GTE and Ex, as single treatments, increased epididymal fat mass (42 and 31% increase). More study is needed to understand this phenomenon. Fasting blood glucose levels in GTE + Ex-treated mice were significantly lower than the other groups, whereas GTEor Ex-treatment alone had no significant effect (Fig. 1C). Fasting plasma insulin levels and homeostasis model assessment of insulin resistance (HOMA-IR) were significantly lower in the GTE + Ex-treated mice (Fig. 1D) compared to HF mice. HOMA-IR and insulin values in combination- treated mice were not significantly different from LF-fed controls. GTE or Ex alone had less pronounced effects on insulin levels and HOMA-IR (Fig. 1D). Although previous studies have reported that treatment with (−)-epigallocatechin-3-gallate and GTE had a beneficial effect on both fasting blood glucose and fasting plasma insulin, results of exercise have been mixed [11, 16, 17]. The effect of GTE and Ex treatment on the expression of genes related to energy utilization in the skeletal muscle was analyzed by real-time PCR (Supporting Information Table 3, Fig. 2A). GTE, Ex, and GTE + Ex-treatment significantly decreased the expression of acyl-CoA oxidase (Acox1) www.mnf-journal.com
1158
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Mol. Nutr. Food Res. 2014, 58, 1156–1159
Figure 2. The effect of GTE, Ex, or GTE + Ex treatment on genes related to lipid metabolism and energy utilization in skeletal muscle and liver tissue from high fat fed, male C57BL6/J mice. (A) Skeletal muscle mRNA expression of Acox1, Ppargc1a, mt-Nd5, mt-Cytb, mt-Co3 was examined by RT-PCR. (B) In the liver, mRNA expression of Ppara, Srebf1, Cpt1a, Scd1 was measured by real-time PCR. Expression for all genes was normalized to Gapdh and calculated fold-change compared to HF-fed mice. N = 12 for LFmice, N = 22 for other groups, and error bars represent the SD. Bars with different superscript letters indicate that the values are statistically significantly different by one-way ANOVA with Tukey’s posttest (P < 0.05).
(0.7-, 0.6-, and 0.6-fold, respectively) compared to HFfed mice, whereas the expression of proliferator-activated receptor-␥ coactivator-1␣ (Ppargc1a) (1.5-fold), NADH dehydrogenase 5 (mt-Nd5) (1.5-fold), cytochrome b (mt-Cytb) (1.5fold), and cytochrome c oxidase subunit III (mt-Co3) (1.3-fold) were significantly increased in GTE + Ex-treated mice compared to HF-fed mice. Ex-treatment tended to increase the expression of Ppargc1a, mt-Nd5, mt-Cytb, and mt-Co3, but the effect was not statistically significant. Changes in the expression of these genes have been associated with increased mitochondrial fatty acid oxidation capacity in skeletal muscle [18]. To our knowledge this is the first report to examine the effects of GTE in combination with Ex [19]. Acox1 expression in the skeletal muscle was decreased in all treatment groups. Acox1 is involved in peroxisomal -oxidation, and previous studies have demonstrated that the expression of Acox1 is inversely associated with gene expression related to mitochondrial -oxidation [20]. The effect of GTE and Ex on the expression of genes related to fat metabolism in the liver was also analyzed by RT-PCR. Gene expression analysis showed that mice treated with GTE, Ex or GTE + Ex had 1.5-, 1.7-, and 2.2-fold increased expression of peroxisome proliferator-activated receptor-␣ (Ppara), respectively, compared to HF-fed mice (Fig. 2B). The expression of carnitine palmitoyl transferase-1␣ (Cpt1a) was significantly increased in the GTE-treated mice and GTE + Ex-treated mice (1.2- or 1.3-fold) compared to HF-fed mice (Fig. 2B). Cpt1a is regulated by Ppara [21]. The increase of Ppara and Cpt1a may indicate enhanced hepatic fatty acid C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
oxidation. Mice treated with Ex or GTE + Ex had significantly reduced expression of stearoyl-CoA desaturase 1 (Scd1) (0.5and 0.5-fold, respectively) compared to HF-fed mice (Fig. 2B). This decrease in hepatic expression of Scd1 suggests a decrease in hepatic de novo lipogenesis, but further studies at the enzyme level are required. The expression of sterol regulatory element binding transcription factor 1 (Srebf1) gene was significantly increased in the GTE + Ex-treated mice (1.4-fold) compared to HF-fed mice but not in mice treated with GTE or Ex (Fig. 2B). Although hepatic expression of Srebf1 increased, the ratio of Ppara to Srebf1 mRNA expression was increased by 139.2, 143.5, and 158.6% in mice treated with GTE, Ex or GTE + Ex, respectively, compared to HF-fed mice. This increased ratio suggests that all treatments modulate the fatty acid metabolism in the liver by favoring fatty acid oxidation rather than de novo lipogenesis. A previous study found that obese patients had higher ratio of Srebf1/Ppara [22]. In summary, we report for the first time that Ex in combination with GTE reduced symptoms of metabolic syndrome related to HF-fed mice more significantly than either treatment alone. These phenotypic changes may result in part from enhanced energy metabolism and decreased de novo lipogenesis, and involve peroxisome proliferator-activated receptor signaling. The authors thank KAG, DD, ALB, TX, and SCF for technical assistance. This study was supported by the National Institutes of Health (AT004678) to JDL. The authors have declared no conflict of interest. www.mnf-journal.com
Mol. Nutr. Food Res. 2014, 58, 1156–1159
References
1159 [12] Yang, C. S., Maliakal, P., Meng, X., Inhibition of carcinogenesis by tea. Ann. Rev. Pharmacol. Toxicol. 2002, 42, 25–54.
[1] Flegal, K. M., Carroll, M. D., Ogden, C. L., Curtin, L. R., Prevalence and trends in obesity among US adults, 1999–2008. J. Am. Med. Assoc. 2010, 303, 235–241.
[13] Hursel, R., Viechtbauer, W., Westerterp-Plantenga, M. S., The effects of green tea on weight loss and weight maintenance: a meta-analysis. Int. J. Obesity 2009, 33, 956–961.
[2] Main, M. L., Rao, S. C., O’Keefe, J. H., Trends in obesity and extreme obesity among US adults. J. Am. Med. Assoc. 2010, 303, 1695–1696.
[14] Murase, T., Haramizu, S., Shimotoyodome, A., Nagasawa, A. et al., Green tea extract improves endurance capacity and increases muscle lipid oxidation in mice. Am. J. Physiol. 2005, 288, R708–R715.
[3] Sae-tan, S., Grove, K. A., Lambert, J. D., Weight control and prevention of metabolic syndrome by green tea. Pharmacol. Res. 2011, 64, 146–154. ´ J. P., St-Jean, L. et al., [4] Tremblay, A., Nadeau, A., Despres, Long-term exercise training with constant energy intake. 2. Effect of glucose metabolism and resting energy expenditure. Int. J. Obesity 1990, 14, 75–84. [5] Gauthier, M. S., Couturier, K., Charbonneau, A., Lavoie, J. M., Effects of introducing physical training in the course of a 16-week high-fat diet regimen on hepatic steatosis, adipose tissue fat accumulation, and plasma lipid profile. Int. J. Obesity Rel. Metabol. Disord. 2004, 28, 1064–1071. [6] Narath, E., Skalicky, M., Viidik, A., Voluntary and forced exercise influence the survival and body composition of ageing male rats differently. Exp. Gerontol. 2001, 36, 1699–1711. [7] Kawashima, H., Saito, T., Yoshizato, H., Fujikawa, T. et al., Endurance treadmill training in rats alters CRH activity in the hypothalamic paraventricular nucleus at rest and during acute running according to its period. Life Sci. 2004, 76, 763– 774. [8] Hayes, K., Sprague, S., Guo, M., Davis, W. et al., Forced, not voluntary, exercise effectively induces neuroprotection in stroke. Acta Neuropathol. 2008, 115, 289–296. [9] Liu, Y. F., Chen, H. I., Wu, C. L., Kuo, Y. M. et al., Differential effects of treadmill running and wheel running on spatial or aversive learning and memory: roles of amygdalar brainderived neurotrophic factor and synaptotagmin I. J. Physiol. 2009, 587, 3221–3231. [10] Rogers, C. J., Zaharoff, D. A., Hance, K. W., Perkins, S. N. et al., Exercise enhances vaccine-induced antigen-specific T cell responses. Vaccine 2008, 26, 5407–5415. [11] Bradley, R. L., Jeon, J. Y., Liu, F.-F., Maratos-Flier, E., Voluntary exercise improves insulin sensitivity and adipose tissue inflammation in diet-induced obese mice. Am. J. Physiol. 2008, 295, E586–E594.
C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
[15] Shimotoyadome, A., Haramizu, S., Inaba, M., Murase, T. et al., Exercise and green tea extract stimulate fat oxidation and prevent obesity in mice. Med. Sci. Sports Exerc. 2005, 37, 1884–1892. [16] Bose, M., Lambert, J. D., Ju, J., Reuhl, K. R. et al., The major green tea polyphenol, (-)-epigallocatechin-3-gallate, inhibits obesity, metabolic syndrome, and fatty liver disease in highfat-fed mice. J. Nutr. 2008, 138, 1677–1683. [17] Huang, P., Li, S., Shao, M., Qi, Q. et al., Calorie restriction and endurance exercise share potent anti-inflammatory function in adipose tissues in ameliorating diet-induced obesity and insulin resistance in mice. Nutr. Metab. 2010, 7, 59–67. [18] Gao, C. L., Liu, G. L., Liu, S., Chen, X. H. et al., Overexpression of PGC-1beta improves insulin sensitivity and mitochondrial function in 3T3-L1 adipocytes. Mol. Cell Biochem. 2011, 353, 215–223. [19] Li, L., Pan, R., Li, R., Niemann, B. et al., Mitochondrial biogenesis and peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) deacetylation by physical activity: intact adipocytokine signaling is required. Diabetes 2011, 60, 157–167. [20] Chance, D. S., Wu, S. M., McIntosh, M. K., Inverse relationship between peroxisomal and mitochondrial beta-oxidation in HepG2 cells treated with dehydroepiandrosterone and clofibric acid. Proc. Soc. Exp. Biol. Med. 1995, 208, 378–384. [21] van Raalte, D. H., Li, M., Pritchard, P. H., Wasan, K. M., Peroxisome proliferator-activated receptor (PPAR)-alpha: a pharmacological target with a promising future. Pharm. Res. 2004, 21, 1531–1538. [22] Pettinelli, P., Del Pozo, T., Araya, J., Rodrigo, R. et al., Enhancement in liver SREBP-1c/PPAR-alpha ratio and steatosis in obese patients: correlations with insulin resistance and n3 long-chain polyunsaturated fatty acid depletion. Biochim. Biophys. Acta 2009, 1792, 1080–1086.
www.mnf-journal.com
DOI 10.1002/mnfr.201300621
Mol. Nutr. Food Res. 2014, 58, 1156–1159
FOOD & FUNCTION
Voluntary exercise and green tea enhance the expression of genes related to energy utilization and attenuate metabolic syndrome in high fat fed mice Sudathip Sae-tan1 , Connie J. Rogers2 and Joshua D. Lambert1 1 2
Department of Food Science, The Pennsylvania State University, University Park, PA, USA Department of Nutritional Sciences, The Pennsylvania State University, University Park, PA, USA
Obesity and metabolic syndrome are growing public health problems. We investigated the effects of decaffeinated green tea extract (GTE) and voluntary running exercise (Ex) alone or in combination against obesity and metabolic syndrome in high fat (HF) fed C57BL/6J mice. After 16 wk, GTE + Ex treatment reduced final body mass (27.1% decrease) and total visceral fat mass (36.6% decrease) compared to HF-fed mice. GTE + Ex reduced fasting blood glucose (17% decrease), plasma insulin (65% decrease), and insulin resistance (65% decrease) compared to HF-fed mice. GTE or Ex alone had less significant effects. In the skeletal muscle, the combination of Ex and GTE increased the expression of peroxisome proliferator-activated receptor-␥ coactivator-1␣ (Ppargc1a), mitochondrial NADH dehydrogenase 5 (mt-Nd5), mitochondrial cytochrome b (mt-Cytb), and mitochondrial cytochrome c oxidase III (mt-Co3). An increase in hepatic expression of peroxisome proliferator-activated receptor-␣ (Ppara) and liver carnitine palmitoyl transferase-1␣ (Cpt1a) and a decrease in hepatic expression of stearoyl-CoA desaturase 1 (Scd1) mRNA was observed in GTE + Ex mice. GTE + Ex was more effective than either treatment alone in reducing diet-induced obesity. These effects are due in part to modulation of genes related to energy metabolism and de novo lipogenesis.
Received: August 23, 2013 Revised: October 17, 2013 Accepted: October 18, 2013
Keywords: Catechins / Fatty acid oxidation / Green tea / Mitochondrial biogenesis / Voluntary exercise
Additional supporting information may be found in the online version of this article at the publisher’s web-site
Approximately 34% of adults in the United States are classified as obese (i.e. BMI ≥ 30) [1]. Obesity, in addition to having normal BMI but a high percentage of body fat, represents a risk factor for metabolic syndrome [2,3]. A traditional strategy Correspondence: Dr. Joshua D. Lambert, Department of Food Science, The Pennsylvania State University, 332 Food Science Building, University Park, PA 16802, USA E-mail: [email protected] Fax: +1-814-863-6132 Abbreviations: Acox1, acyl-CoA oxidase; Cpt1a, carnitine palmitoyl transferase-1␣; Ex, voluntary exercise; GTE, decaffeinated green tea extract; HF, high fat; HOMA-IR, homeostasis model assessment of insulin resistance; LF, low fat; mt-Co3, cytochrome c oxidase subunit III; mt-Cytb, cytochrome b; mtNd5, NADH dehydrogenase 5; Ppara, peroxisome proliferatoractivated receptor-␣; Ppargc1a, proliferator-activated receptor-␥ coactivator-1␣; Scd1, stearoyl-CoA desaturase 1; Srebf1, sterol regulatory element-binding protein-1c C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
for the treatment of obesity and metabolic syndrome is life style intervention including diet and exercise. Regular physical activity is useful in weight loss due to increased energy expenditure [4]. Animal studies have examined the effects of forced exercise on reduction of body weight and improvement of symptoms related to metabolic syndrome [5]. This use of forced exercise can have significant impacts on physiological outcome but in its laboratory permutations (i.e. forced running or exhaustive swimming) is less translatable to humans. For example, Sprague-Dawley rats with voluntary access to a running wheel had an 18% greater reduction in body weight than rats treated with forced treadmill [6]. Involuntary (forced) exercise has been shown to elicit greater increase in glucocorticoids than voluntary running [7–9]. Although fewer studies have examined the impact of voluntary exercise (Ex) on weight loss, these have shown that voluntary running reduced fat mass, insulin resistance, and inflammation [10, 11]. www.mnf-journal.com
Mol. Nutr. Food Res. 2014, 58, 1156–1159
1157
Figure 1. The effect of GTE, Ex, or GTE + Ex on body mass (A), fat mass (B), and glycemic markers (C) and (D) in high fat fed male C57BL/6J mice. Body weight and fasting blood glucose were determined weekly and biweekly throughout the experiment, respectively. Visceral fat mass and fasting insulin was determined in the plasma at the end of the experiment. HOMA-IR was determined using the final fasting blood glucose and fasting plasma insulin levels measured in the experiment. N = 12 for LF-mice, N = 22 for other groups, and error bars represent the SD. Body weight and blood glucose data were analyzed by two-way ANOVA with Bonferroni posttest. Different symbols indicate pairwise comparisons between each treatment to HF-fed mice (a, HF versus LF; b, HF versus GTE + Ex; c, HF versus GTE; d, HF versus Ex). Error bars were removed from fasting blood glucose to improve clarity. For fasting blood glucose *P < 0.05 compared to HF. For the other parameters, values with different letters are statistically significantly different by one-way ANOVA with Tukey’s posttest, P < 0.05.
Tea (Camellia sinensis) is a beverage with worldwide popularity. It is rich in catechins with (−)-epigallocatechin-3-gallate is the most abundant [12]. A number of animal studies have shown the preventive effects of green tea polyphenols against obesity [3]. A recent meta-analysis of 11 human trials with green tea preparations reported a 1.31 kg average body weight loss in intervention groups compared to control groups [13]. Although previous studies have shown that forced running or exhaustive swimming in combination with tea catechins can attenuate obesity in mice, to our knowledge there have been no reports on the effects of green tea in combination with Ex on metabolic syndrome in high fat (HF) fed mice [14, 15]. We report the results of our study on the preventive effects of combination treatment with Ex and decaffeinated green tea extract (GTE) in a HF-fed mouse model of metabolic syndrome (see Supporting Information for detailed methodology). Ex or GTE (Supporting Information Table 1) alone resulted in decreased final body weight compared to HF-fed mice (Supporting Information Table 2; 12.0 and 9.4%, respectively, Fig. 1A). Treatment with the combination of GTE and Ex yielded the lowest final body weight (27%, Fig. 1A) and for most of the experiment, the body weight of the combination treated mice overlapped with the low fat (LF) fed controls. There were minor differences (>10%) in average weekly energy intake per mouse in GTE + Ex-treated mice compared to HF-fed controls (Supporting Information Fig. 1A). Both Ex-treated mice and GTE + Ex-treated mice had the similar average daily running activity (Supporting Information C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Fig. 2A). Previously, it has been reported that cotreatment with exhaustive swimming and 0.5% catechins reduced rate of body weight gain by 33% compared to HF controls [14]. GTE and Ex treatments decreased retroperitoneal fat mass (31 and 33%, respectively): Ex treatment also decreased intestinal fat mass (26.2%) compared to HF mice (Fig. 1B). The combination treatment with GTE and Ex reduced retroperitoneal fat mass and intestinal fat mass back to those found in LF-fed control mice. Both GTE and Ex, as single treatments, increased epididymal fat mass (42 and 31% increase). More study is needed to understand this phenomenon. Fasting blood glucose levels in GTE + Ex-treated mice were significantly lower than the other groups, whereas GTEor Ex-treatment alone had no significant effect (Fig. 1C). Fasting plasma insulin levels and homeostasis model assessment of insulin resistance (HOMA-IR) were significantly lower in the GTE + Ex-treated mice (Fig. 1D) compared to HF mice. HOMA-IR and insulin values in combination- treated mice were not significantly different from LF-fed controls. GTE or Ex alone had less pronounced effects on insulin levels and HOMA-IR (Fig. 1D). Although previous studies have reported that treatment with (−)-epigallocatechin-3-gallate and GTE had a beneficial effect on both fasting blood glucose and fasting plasma insulin, results of exercise have been mixed [11, 16, 17]. The effect of GTE and Ex treatment on the expression of genes related to energy utilization in the skeletal muscle was analyzed by real-time PCR (Supporting Information Table 3, Fig. 2A). GTE, Ex, and GTE + Ex-treatment significantly decreased the expression of acyl-CoA oxidase (Acox1) www.mnf-journal.com
1158
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Mol. Nutr. Food Res. 2014, 58, 1156–1159
Figure 2. The effect of GTE, Ex, or GTE + Ex treatment on genes related to lipid metabolism and energy utilization in skeletal muscle and liver tissue from high fat fed, male C57BL6/J mice. (A) Skeletal muscle mRNA expression of Acox1, Ppargc1a, mt-Nd5, mt-Cytb, mt-Co3 was examined by RT-PCR. (B) In the liver, mRNA expression of Ppara, Srebf1, Cpt1a, Scd1 was measured by real-time PCR. Expression for all genes was normalized to Gapdh and calculated fold-change compared to HF-fed mice. N = 12 for LFmice, N = 22 for other groups, and error bars represent the SD. Bars with different superscript letters indicate that the values are statistically significantly different by one-way ANOVA with Tukey’s posttest (P < 0.05).
(0.7-, 0.6-, and 0.6-fold, respectively) compared to HFfed mice, whereas the expression of proliferator-activated receptor-␥ coactivator-1␣ (Ppargc1a) (1.5-fold), NADH dehydrogenase 5 (mt-Nd5) (1.5-fold), cytochrome b (mt-Cytb) (1.5fold), and cytochrome c oxidase subunit III (mt-Co3) (1.3-fold) were significantly increased in GTE + Ex-treated mice compared to HF-fed mice. Ex-treatment tended to increase the expression of Ppargc1a, mt-Nd5, mt-Cytb, and mt-Co3, but the effect was not statistically significant. Changes in the expression of these genes have been associated with increased mitochondrial fatty acid oxidation capacity in skeletal muscle [18]. To our knowledge this is the first report to examine the effects of GTE in combination with Ex [19]. Acox1 expression in the skeletal muscle was decreased in all treatment groups. Acox1 is involved in peroxisomal -oxidation, and previous studies have demonstrated that the expression of Acox1 is inversely associated with gene expression related to mitochondrial -oxidation [20]. The effect of GTE and Ex on the expression of genes related to fat metabolism in the liver was also analyzed by RT-PCR. Gene expression analysis showed that mice treated with GTE, Ex or GTE + Ex had 1.5-, 1.7-, and 2.2-fold increased expression of peroxisome proliferator-activated receptor-␣ (Ppara), respectively, compared to HF-fed mice (Fig. 2B). The expression of carnitine palmitoyl transferase-1␣ (Cpt1a) was significantly increased in the GTE-treated mice and GTE + Ex-treated mice (1.2- or 1.3-fold) compared to HF-fed mice (Fig. 2B). Cpt1a is regulated by Ppara [21]. The increase of Ppara and Cpt1a may indicate enhanced hepatic fatty acid C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
oxidation. Mice treated with Ex or GTE + Ex had significantly reduced expression of stearoyl-CoA desaturase 1 (Scd1) (0.5and 0.5-fold, respectively) compared to HF-fed mice (Fig. 2B). This decrease in hepatic expression of Scd1 suggests a decrease in hepatic de novo lipogenesis, but further studies at the enzyme level are required. The expression of sterol regulatory element binding transcription factor 1 (Srebf1) gene was significantly increased in the GTE + Ex-treated mice (1.4-fold) compared to HF-fed mice but not in mice treated with GTE or Ex (Fig. 2B). Although hepatic expression of Srebf1 increased, the ratio of Ppara to Srebf1 mRNA expression was increased by 139.2, 143.5, and 158.6% in mice treated with GTE, Ex or GTE + Ex, respectively, compared to HF-fed mice. This increased ratio suggests that all treatments modulate the fatty acid metabolism in the liver by favoring fatty acid oxidation rather than de novo lipogenesis. A previous study found that obese patients had higher ratio of Srebf1/Ppara [22]. In summary, we report for the first time that Ex in combination with GTE reduced symptoms of metabolic syndrome related to HF-fed mice more significantly than either treatment alone. These phenotypic changes may result in part from enhanced energy metabolism and decreased de novo lipogenesis, and involve peroxisome proliferator-activated receptor signaling. The authors thank KAG, DD, ALB, TX, and SCF for technical assistance. This study was supported by the National Institutes of Health (AT004678) to JDL. The authors have declared no conflict of interest. www.mnf-journal.com
Mol. Nutr. Food Res. 2014, 58, 1156–1159
References
1159 [12] Yang, C. S., Maliakal, P., Meng, X., Inhibition of carcinogenesis by tea. Ann. Rev. Pharmacol. Toxicol. 2002, 42, 25–54.
[1] Flegal, K. M., Carroll, M. D., Ogden, C. L., Curtin, L. R., Prevalence and trends in obesity among US adults, 1999–2008. J. Am. Med. Assoc. 2010, 303, 235–241.
[13] Hursel, R., Viechtbauer, W., Westerterp-Plantenga, M. S., The effects of green tea on weight loss and weight maintenance: a meta-analysis. Int. J. Obesity 2009, 33, 956–961.
[2] Main, M. L., Rao, S. C., O’Keefe, J. H., Trends in obesity and extreme obesity among US adults. J. Am. Med. Assoc. 2010, 303, 1695–1696.
[14] Murase, T., Haramizu, S., Shimotoyodome, A., Nagasawa, A. et al., Green tea extract improves endurance capacity and increases muscle lipid oxidation in mice. Am. J. Physiol. 2005, 288, R708–R715.
[3] Sae-tan, S., Grove, K. A., Lambert, J. D., Weight control and prevention of metabolic syndrome by green tea. Pharmacol. Res. 2011, 64, 146–154. ´ J. P., St-Jean, L. et al., [4] Tremblay, A., Nadeau, A., Despres, Long-term exercise training with constant energy intake. 2. Effect of glucose metabolism and resting energy expenditure. Int. J. Obesity 1990, 14, 75–84. [5] Gauthier, M. S., Couturier, K., Charbonneau, A., Lavoie, J. M., Effects of introducing physical training in the course of a 16-week high-fat diet regimen on hepatic steatosis, adipose tissue fat accumulation, and plasma lipid profile. Int. J. Obesity Rel. Metabol. Disord. 2004, 28, 1064–1071. [6] Narath, E., Skalicky, M., Viidik, A., Voluntary and forced exercise influence the survival and body composition of ageing male rats differently. Exp. Gerontol. 2001, 36, 1699–1711. [7] Kawashima, H., Saito, T., Yoshizato, H., Fujikawa, T. et al., Endurance treadmill training in rats alters CRH activity in the hypothalamic paraventricular nucleus at rest and during acute running according to its period. Life Sci. 2004, 76, 763– 774. [8] Hayes, K., Sprague, S., Guo, M., Davis, W. et al., Forced, not voluntary, exercise effectively induces neuroprotection in stroke. Acta Neuropathol. 2008, 115, 289–296. [9] Liu, Y. F., Chen, H. I., Wu, C. L., Kuo, Y. M. et al., Differential effects of treadmill running and wheel running on spatial or aversive learning and memory: roles of amygdalar brainderived neurotrophic factor and synaptotagmin I. J. Physiol. 2009, 587, 3221–3231. [10] Rogers, C. J., Zaharoff, D. A., Hance, K. W., Perkins, S. N. et al., Exercise enhances vaccine-induced antigen-specific T cell responses. Vaccine 2008, 26, 5407–5415. [11] Bradley, R. L., Jeon, J. Y., Liu, F.-F., Maratos-Flier, E., Voluntary exercise improves insulin sensitivity and adipose tissue inflammation in diet-induced obese mice. Am. J. Physiol. 2008, 295, E586–E594.
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[15] Shimotoyadome, A., Haramizu, S., Inaba, M., Murase, T. et al., Exercise and green tea extract stimulate fat oxidation and prevent obesity in mice. Med. Sci. Sports Exerc. 2005, 37, 1884–1892. [16] Bose, M., Lambert, J. D., Ju, J., Reuhl, K. R. et al., The major green tea polyphenol, (-)-epigallocatechin-3-gallate, inhibits obesity, metabolic syndrome, and fatty liver disease in highfat-fed mice. J. Nutr. 2008, 138, 1677–1683. [17] Huang, P., Li, S., Shao, M., Qi, Q. et al., Calorie restriction and endurance exercise share potent anti-inflammatory function in adipose tissues in ameliorating diet-induced obesity and insulin resistance in mice. Nutr. Metab. 2010, 7, 59–67. [18] Gao, C. L., Liu, G. L., Liu, S., Chen, X. H. et al., Overexpression of PGC-1beta improves insulin sensitivity and mitochondrial function in 3T3-L1 adipocytes. Mol. Cell Biochem. 2011, 353, 215–223. [19] Li, L., Pan, R., Li, R., Niemann, B. et al., Mitochondrial biogenesis and peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) deacetylation by physical activity: intact adipocytokine signaling is required. Diabetes 2011, 60, 157–167. [20] Chance, D. S., Wu, S. M., McIntosh, M. K., Inverse relationship between peroxisomal and mitochondrial beta-oxidation in HepG2 cells treated with dehydroepiandrosterone and clofibric acid. Proc. Soc. Exp. Biol. Med. 1995, 208, 378–384. [21] van Raalte, D. H., Li, M., Pritchard, P. H., Wasan, K. M., Peroxisome proliferator-activated receptor (PPAR)-alpha: a pharmacological target with a promising future. Pharm. Res. 2004, 21, 1531–1538. [22] Pettinelli, P., Del Pozo, T., Araya, J., Rodrigo, R. et al., Enhancement in liver SREBP-1c/PPAR-alpha ratio and steatosis in obese patients: correlations with insulin resistance and n3 long-chain polyunsaturated fatty acid depletion. Biochim. Biophys. Acta 2009, 1792, 1080–1086.
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