CLB-08704; No. of pages: 4; 4C: Clinical Biochemistry xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Clinical Biochemistry journal homepage: www.elsevier.com/locate/clinbiochem
Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects☆ Tatsuya Suzuki a,⁎, Junya Aoyama a, Masao Hashimoto a, Makoto Ohara b, Shoko Futami-Suda a, Kazunari Suzuki a, Motoshi Ouchi a, Yoshimasa Igari a, Kentaro Watanabe c, Hiroshi Nakano a a b c
Department of Internal Medicine, Division of Geriatric Medicine, Nippon Medical School, Tokyo, Japan Division of Diabetes, Metabolism, and Endocrinology, Department of Internal Medicine, Showa University School of Medicine, Tokyo, Japan Department of Neurology, Hematology, Metabolism, Endocrinology and Diabetology, Yamagata University Faculty of Medicine, Yamagata, Japan
a r t i c l e
i n f o
Article history: Received 10 January 2014 Received in revised form 19 April 2014 Accepted 23 April 2014 Available online xxxx Keywords: Bile acid Conjugation Normal weight Body compositions Body fat mass
a b s t r a c t Background: Bile acids (BAs) play important roles in glucose regulation and energy homeostasis via G protein-coupled receptors, such as enteroendocrine L cell TGR5. The aim of the present study was to investigate the relationship between postprandial BA levels and body composition after ingestion of a standard test meal. Methods: Eleven healthy subjects of normal weight (body-mass index, 22.0 ± 1.6 kg/m2 [mean ± SD]), ingested a 400-kcal test meal, and blood samples were obtained from them before ingestion and every 30 min for 120 min after ingestion. The BA fractions were measured with high-performance liquid chromatography. To evaluate body composition, body impedance analysis was performed 1 h before ingestion of the test meal. Results: Concentrations of both total BA and total glycine-conjugated BA (GCBA) at 30, 60, 90, and 120 min after test-meal ingestion were significantly higher than those at baseline. The body-mass index was correlated with total GCBA at baseline. Moreover, body fat mass was correlated with total GCBA at 30 min (r = –0.688, P = 0.019) and 60 min (r = – 0.642, P = 0.033) and with total BA at 30 min (r = – 0.688, P = 0.019) and 60 min (r = –0.642, P = 0.033). Conclusion: The postprandial BA response is inversely related with body fat mass in healthy subjects of normal weight. © 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.
Introduction Bile consists of bile acids (BAs), cholesterol, phosphatidylcholine, and bilirubin and is secreted from hepatocytes into the bile canaliculi. After being released by the gallbladder into the intestines, the BAs are almost completely reabsorbed in the terminal ileum, thereby decreasing the need for de novo bile synthesis. The synthesis of BAs from cholesterol is the primary pathway for cholesterol catabolism. In humans, the primary BAs cholic acid (CA) and chenodeoxycholic acid (CDCA) are synthesized from hepatic cholesterol. In the intestine, gut microbiota deconjugates part of BAs and converts primary BAs to secondary BAs. Both CA and CDCA are conjugated to taurine or glycine and transported into the biliary ductile system. Recently, BAs have been found to play important roles in glucose regulation and energy homeostasis [1]. For example, CA increases
☆ None of the authors have any competing interests to declare and have not received any financial support from pharmaceutical companies. ⁎ Corresponding author at: Department of Internal Medicine, Division of Geriatric Medicine, Nippon Medical School 1-1-5 Sendagi, Bunkyo-ku, Tokyo 113-8603, Japan. Fax: +81 3 5685 3066. E-mail address: [email protected] (T. Suzuki).
energy expenditure and prevents the development of high-fat-induced obesity and insulin resistance in mice [2]. These metabolic effects of CA are mediated by the G-protein-coupled receptor TGR5, leading to the induction of type 2 iodothyronine deiodinase. Moreover, modulation of BA metabolism by BA sequestrants has been shown to lower plasma glucose levels [3]. The postprandial BA response in obese subjects is significantly lower than that in subjects of normal weight [4]. In contrast, Vincent et al. have found that the postprandial BA response in obese patients with type 2 diabetes mellitus is greater than that in normoglycemic individuals [5]. However, to the best of our knowledge, no study has examined, according to body composition, whether subjects of normal weight have an altered postprandial BA response. The aim of the present study was to investigate the relationship between postprandial BAs and body composition after the ingestion of a standard 400-kcal test meal. Methods All subjects provided informed consent. The present study was designed in compliance with the ethics regulations set out by the Declaration of Helsinki.
http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025 0009-9120/© 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.
Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
T. Suzuki et al. / Clinical Biochemistry xxx (2014) xxx–xxx
The subjects were 11 healthy persons (5 men and 6 women). After an overnight fast, subjects ingested a 400-kcal test meal (41.0 g carbohydrate, 22.2 g fat, and 8.4 g protein) at 8:30 am. The test meal was a commercial energy-supplement food (CalorieMate®, Otsuka Pharmaceutical Co., Ltd., Tokyo, Japan). Samples of peripheral venous blood were obtained before ingestion of the test meal and every 30 min for 120 min after ingestion. The fasting plasma glucose (FPG) and glycated hemoglobin (HbA1C) were measured. Values for HbA1C (%) were estimated using the National Glycohemoglobin Standardization Program (NGSP) equivalent values (%), which were calculated from the formula HbA1C (%) = 1.02 × HbA1C (Japanese Diabetes Society) (%) + 0.25%. This formula takes into consideration the relation between HbA1C (Japanese Diabetes Society) (%) determined using the previous Japanese standard measurement method and HbA1C (NGSP) [6]. Furthermore, body mass index [BMI; body weight (kg) ÷ height (m2)] was calculated. All samples (EDTA-plasma) were stored at − 80 °C until analyzed. BA measurements The BA conjugate fractions were measured with high-performance liquid chromatography [7]. The fractions were compared by considering ursodioxycholic acid, CA, CDCA, deoxycholic acid, and lithocholic acid as totals of glycine- and taurine-conjugated forms of the respective BAs. The measurement method was linear from 0.1 to 10 μmol/L for all BAs and their conjugates, with coefficients of variation (CV) of 3.1% to 7.5% at the lower limit of quantitation (0.1 μmol/L). The intra-assay CV was 5.0% to 7.5%, and the interassay CV was 3.1% to 4.9%. The method was based on that of Okuyama [8,9] and HPLC is suitable for the analysis of major bile acids in normal serum [10]. According to its method, the separation of the individual bile acids including unconjugated, glycine-and taurine-conjugated bile acids was investigated by using HPLC and immobilized enzyme in column form. The separation is very clear except glycodeoxycholic acid and taurochenodeoxycholic acid. Each peak is equivalent to 100 ng of bile acid. The estimated detection limit is 10 ng approximately. After EDTA blood collection, all samples were centrifuged at 3000 rpm for 8 min and immediately frozen at −80 °C. The time between collection and centrifugation was approximately 30 min. Stability of EDTA-plasma was assessed immediately and after storage for 1, 4, 8 and 24 h at room temperature. Immediately frozen samples did not differ in BA levels compared with samples stored for 1, 4, 8 and 24 h at room temperature [11].
HbA1C levels at baseline were 84.5 ± 6.7 mg/dL and 4.9 ± 0.3%, respectively. The mean body fat mass (BFM) was 15.2 ± 3.1 kg, the mean skeletal muscle mass was 25.3 ± 6.2 kg, and the visceral fat area was 60.9 ± 20.6 cm2. The total BA concentrations 30 min, 60 min, 90 min, and 120 min after ingestion of a 400-kcal test meal were significantly higher than that of the baseline (from 1.3 ± 0.9 to 3.0 ± 2.4 μmol/L, 3.1 ± 2.5 μmol/L, 4.4 ± 2.7 μmol/L, and 5.4 ± 3.3 μmol/L; P = 0.024, P = 0.039, P = 0.004, and P = 0.002, respectively) (Fig. 1). Similarly, the total GCBA concentrations 30 min, 60 min, 90 min, and 120 min after ingestion were significantly higher than that of the baseline (from 0.7 ± 0.6 μmol/L to 2.5 ± 2.1 μmol/L, 2.3 ± 1.6 μmol/L, 3.6 ± 2.1 μmol/L, and 4.6 ± 2.6 μmol/L; P = 0.008, P = 0.005, P b 0.001, and P b 0.001, respectively) (Fig. 1). The total taurine-conjugated BA concentration 120 min after ingestion was significantly higher than that at baseline (P = 0.005). The BMI was correlated with total GCBA at baseline (r = − 0.721, P = 0.012) (Fig. 2). The BFM was negatively correlated with total GCBA at 30 min (r = –0.688, P = 0.019) and 60 min (r = –0.642, P = 0.033). In addition, BFM was negatively correlated with total BA at 30 min (r = –0.688, P = 0.019) and 60 min (r = –0.642, P = 0.033). However, other BA variables, including total BAs and total GCBA, were not significantly correlated with either skeletal muscle mass or visceral fat area. The BA fractions did not differ by sex or BFM.
a Concentratinons (µ µmol/L)
2
Bioelectric impedance analysis
Statistical analysis The SPSS software package (version 11.0; SPSS Inc., Chicago, IL, USA) was used for all statistical analyses. Spearman's correlation was used to evaluate correlations. Data were tested for normality with the Shapiro– Wilk W test. Continuous variables are expressed as means ± standard deviation (SD) and correlation coefficients (r). A P-value of b0.05 was considered to indicate statistical significance. Results The mean age of the subjects was 28.6 ± 1.6 years, with a bodymass index [BMI], 22.0 ± 1.6 (19.2–24.1) kg/m2 [men; 22.2 ± 1.8 (19.9–24.0), women; 21.8 ± 1.6 (21.8–24.1)]. The mean FPG and
b Concentratinons (mmol/L)
Subjects underwent bioelectric impedance analysis 1 h before they ingested the test meal. Body composition was analyzed with a bioelectric impedance device (InBody 720, BioSpace Co., Ltd., Seoul, Korea) and associated software (InBody 3.0, BioSpace) as previously described [12,13]. This device employs an 8-point tactile electrode system and measurements at 6 frequencies (1, 5, 50, 250, 500, and 1000 kHz).
Time (min)
Time (min) Fig. 1. Changes in postprandial total BA concentrations (a) and total GCBA concentrations (b) after ingestion of a test meal (mean ± SD). Total BA: total bile acid; and total GCBA: total glycine-conjugated bile acid.
Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
T. Suzuki et al. / Clinical Biochemistry xxx (2014) xxx–xxx
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Fig. 2. The correlation between FV and postprandial total GCBA, and total BA concentrations (mean ± SD) after ingestion of a test meal. (a) BFM and total GCBA at 30 min, (b) BFM and total GCBA at 60 min, (c) BFM and total BA at 30 min, and (d) BFM and total BA at 60 min. BFM: body fat mass; total GCBA: total glycine-conjugated bile acid; and total BA: total bile acid.
Discussion Both total BA concentrations and total GCBA concentrations increased after ingestion of a 400-kcal test meal. These results are comparable with those of a previous report [5]. Moreover, BMI was negatively correlated with total GCBA. In addition, BFM was negatively correlated with total GCBA and total BA. Postprandial levels of BAs may change according to body composition, such as BFM, even if subjects are healthy and of normal weight. In fact, the postprandial BA response in obese subjects has been shown to be lower than that in subjects of normal weight [4]. BAs are important biochemical modulators that activate certain nuclear receptors, such as farnesoid X receptor and vitamin D and G protein-coupled TGR5, which regulate GLP-1 and the gut hormone peptide YY. Both GLP-1 and peptide YY are secreted by L-cells of the small intestine in response to BAs. Rectal administration of BA increases GLP-1 and peptide YY by increasing the sensation of fullness and decreasing subsequent consumption compared with control [14]. In addition, the responses of GLP-1 and peptide YY in healthy male subjects are correlated with levels of CDCA [15]. To date, 2 hormones have been found to fulfill the definition of incretin hormones: GLP-1 and gastric inhibitory peptide (GIP). GIP is a
physiological gut peptide secreted from intestinal K-cells and has insulin-releasing actions, whereas GLP-1 is produced and released from L-cells. Both fat and protein markedly stimulate GIP secretion. We speculate that the postprandial BA response in healthy subjects of normal weight is associated with GLP-1 and peptide YY and with GIP, a type of incretin, which directly links overnutrition to obesity. In the present study, levels of BAs were negatively correlated with BFM but not with visceral fat area. However, these correlations did not differ between men and women. Therefore, further investigations, including studies of incretin hormones, such as GLP-1 and GIP, will be needed to clarify the association between BFM and BAs. Our study had several limitations. The subjects were few and were young volunteers; therefore, the findings might not be applicable to middle-aged or elderly persons. We believe that a case–control study with a greater number of subjects should be performed.
Conclusion The current results show that postprandial BA response is inversely related with body fat mass in healthy subjects of normal weight.
Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
4
T. Suzuki et al. / Clinical Biochemistry xxx (2014) xxx–xxx
Conflict of interest statement The authors declare that there are no conflicts of interest.
Acknowledgment This study received no specific grant from any funding agency in the public and commercial sections. We are grateful to Masao Okazaki, MD, of the Academic Information Center, The Jikei University School of Medicine, for his careful revision of the English language of the manuscript.
References [1] Lefebvre P, Cariou B, Lien F, Kuipers F, Staels B. Role of bile acids and bile acid receptors in metabolic regulation. Physiol Rev 2009;89:147–9. [2] Watanabe M, Houten SM, Mataki C, Christoffolete MA, Kim BW, Sato H, et al. Bile acids induce energy expenditure by promoting intracellular thyroid hormone activation. Nature 2006;439:484–9. [3] Suzuki T, Oba K, Futami S, Suzuki K, Ouchi M, Igari Y, et al. Blood glucose-lowering activity of colestimide in patients with type 2 diabetes and hypercholesterolemia: a case–control study comparing colestimide with acarbose. J Nippon Med Sch 2006;73:277–84. [4] Glicksman C, Pournaras DJ, Wright M, Roberts R, Mahon R, Welbourn R, et al. Postprandial plasma bile acid responses in normal weight and obese subjects. Ann Clin Biochem 2010;47:482–4.
[5] Vincent RP, Omar S, Ghozlan S, Taylor DR, Cross G, Sherwood RA, et al. Higher circulating bile acid concentrations in obese patients with type 2 diabetes. Ann Clin Biochem 2013;50(4):360–4. [6] Kashiwagi A, Kasuga M, Araki E, Oka Y, Hanafusa T, Ito H, et al. Committee on the Standardization of Diabetes Mellitus-Related Laboratory Testing of Japan Diabetes Society: International clinical harmonization of glycated hemoglobin in Japan: from Japan Diabetes Society to National Glycohemoglobin Standardization Program values. J Diabetes Invest 2012;3:39–40. [7] Sakakura H, Suzuki M, Kimura N, Takeda H, Nagata S, Maeda M. Simultaneous determination of bile acids in rat bile and serum by high-performance liquid chromatography. J Chromatogr 1993;62(2):123–31 [24]. [8] Okuyama S, Kokubun N, Higashidate S, Uemura D, Hirata Y. A new analytical method of individual bile acid using high performance liquid chromatography and immobilized 3α-hydroxysteroid dehydrogenase in column form. Chem Lett 1979:1443–6. [9] Okuyama S, Uemura D, Hirata Y. High performance liquid chromatography analysis of individual bile acids: free, glycine- and taurine-conjugated bile acids. Bull Chem Soc Jpn 1979;52(1):124–6. [10] Nakayama F. Quantitative microanalysis of bile acids in biological samples. Collaborative study. J Chromatogr 1988;452:399–408. [11] Scherer M, Gnewuch C, Schmitz G, Liebisch G. Rapid quatification of bile acids and their conjugates in serum by liquid chromatography-tandem mass spectrometry. J Chromatogr B 2009;877:3920–5. [12] Lorenzo AD, Andreoli A. Segmental bioelectrical impedance analysis. Curr Opin Clin Nutr Metab Care 2003;6:551–5. [13] Demura S, Sato S, Kitabayashi T. Percentage of total body fat as estimated by three automatic bioelectrical impedance analyzers. J Physiol Anthropol Appl Human Sci 2004;23:93–9. [14] Wu T, Bound MJ, Standfield SD, et al. Effects of rectal administration of taurocholic acid on glucagon-like peptide-1 and peptide YY in healthy humans. Diabetes Obes Metab 2013;15:474–7. [15] Roberts RE, Glicksman C, Zadeh JA, et al. The relationship between postprandial bile acid concentration, GLP-1, PYY and ghrelin. Clin Endocrinol (Oxf) 2011;74(1):67–72.
Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
Contents lists available at ScienceDirect
Clinical Biochemistry journal homepage: www.elsevier.com/locate/clinbiochem
Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects☆ Tatsuya Suzuki a,⁎, Junya Aoyama a, Masao Hashimoto a, Makoto Ohara b, Shoko Futami-Suda a, Kazunari Suzuki a, Motoshi Ouchi a, Yoshimasa Igari a, Kentaro Watanabe c, Hiroshi Nakano a a b c
Department of Internal Medicine, Division of Geriatric Medicine, Nippon Medical School, Tokyo, Japan Division of Diabetes, Metabolism, and Endocrinology, Department of Internal Medicine, Showa University School of Medicine, Tokyo, Japan Department of Neurology, Hematology, Metabolism, Endocrinology and Diabetology, Yamagata University Faculty of Medicine, Yamagata, Japan
a r t i c l e
i n f o
Article history: Received 10 January 2014 Received in revised form 19 April 2014 Accepted 23 April 2014 Available online xxxx Keywords: Bile acid Conjugation Normal weight Body compositions Body fat mass
a b s t r a c t Background: Bile acids (BAs) play important roles in glucose regulation and energy homeostasis via G protein-coupled receptors, such as enteroendocrine L cell TGR5. The aim of the present study was to investigate the relationship between postprandial BA levels and body composition after ingestion of a standard test meal. Methods: Eleven healthy subjects of normal weight (body-mass index, 22.0 ± 1.6 kg/m2 [mean ± SD]), ingested a 400-kcal test meal, and blood samples were obtained from them before ingestion and every 30 min for 120 min after ingestion. The BA fractions were measured with high-performance liquid chromatography. To evaluate body composition, body impedance analysis was performed 1 h before ingestion of the test meal. Results: Concentrations of both total BA and total glycine-conjugated BA (GCBA) at 30, 60, 90, and 120 min after test-meal ingestion were significantly higher than those at baseline. The body-mass index was correlated with total GCBA at baseline. Moreover, body fat mass was correlated with total GCBA at 30 min (r = –0.688, P = 0.019) and 60 min (r = – 0.642, P = 0.033) and with total BA at 30 min (r = – 0.688, P = 0.019) and 60 min (r = –0.642, P = 0.033). Conclusion: The postprandial BA response is inversely related with body fat mass in healthy subjects of normal weight. © 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.
Introduction Bile consists of bile acids (BAs), cholesterol, phosphatidylcholine, and bilirubin and is secreted from hepatocytes into the bile canaliculi. After being released by the gallbladder into the intestines, the BAs are almost completely reabsorbed in the terminal ileum, thereby decreasing the need for de novo bile synthesis. The synthesis of BAs from cholesterol is the primary pathway for cholesterol catabolism. In humans, the primary BAs cholic acid (CA) and chenodeoxycholic acid (CDCA) are synthesized from hepatic cholesterol. In the intestine, gut microbiota deconjugates part of BAs and converts primary BAs to secondary BAs. Both CA and CDCA are conjugated to taurine or glycine and transported into the biliary ductile system. Recently, BAs have been found to play important roles in glucose regulation and energy homeostasis [1]. For example, CA increases
☆ None of the authors have any competing interests to declare and have not received any financial support from pharmaceutical companies. ⁎ Corresponding author at: Department of Internal Medicine, Division of Geriatric Medicine, Nippon Medical School 1-1-5 Sendagi, Bunkyo-ku, Tokyo 113-8603, Japan. Fax: +81 3 5685 3066. E-mail address: [email protected] (T. Suzuki).
energy expenditure and prevents the development of high-fat-induced obesity and insulin resistance in mice [2]. These metabolic effects of CA are mediated by the G-protein-coupled receptor TGR5, leading to the induction of type 2 iodothyronine deiodinase. Moreover, modulation of BA metabolism by BA sequestrants has been shown to lower plasma glucose levels [3]. The postprandial BA response in obese subjects is significantly lower than that in subjects of normal weight [4]. In contrast, Vincent et al. have found that the postprandial BA response in obese patients with type 2 diabetes mellitus is greater than that in normoglycemic individuals [5]. However, to the best of our knowledge, no study has examined, according to body composition, whether subjects of normal weight have an altered postprandial BA response. The aim of the present study was to investigate the relationship between postprandial BAs and body composition after the ingestion of a standard 400-kcal test meal. Methods All subjects provided informed consent. The present study was designed in compliance with the ethics regulations set out by the Declaration of Helsinki.
http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025 0009-9120/© 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.
Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
T. Suzuki et al. / Clinical Biochemistry xxx (2014) xxx–xxx
The subjects were 11 healthy persons (5 men and 6 women). After an overnight fast, subjects ingested a 400-kcal test meal (41.0 g carbohydrate, 22.2 g fat, and 8.4 g protein) at 8:30 am. The test meal was a commercial energy-supplement food (CalorieMate®, Otsuka Pharmaceutical Co., Ltd., Tokyo, Japan). Samples of peripheral venous blood were obtained before ingestion of the test meal and every 30 min for 120 min after ingestion. The fasting plasma glucose (FPG) and glycated hemoglobin (HbA1C) were measured. Values for HbA1C (%) were estimated using the National Glycohemoglobin Standardization Program (NGSP) equivalent values (%), which were calculated from the formula HbA1C (%) = 1.02 × HbA1C (Japanese Diabetes Society) (%) + 0.25%. This formula takes into consideration the relation between HbA1C (Japanese Diabetes Society) (%) determined using the previous Japanese standard measurement method and HbA1C (NGSP) [6]. Furthermore, body mass index [BMI; body weight (kg) ÷ height (m2)] was calculated. All samples (EDTA-plasma) were stored at − 80 °C until analyzed. BA measurements The BA conjugate fractions were measured with high-performance liquid chromatography [7]. The fractions were compared by considering ursodioxycholic acid, CA, CDCA, deoxycholic acid, and lithocholic acid as totals of glycine- and taurine-conjugated forms of the respective BAs. The measurement method was linear from 0.1 to 10 μmol/L for all BAs and their conjugates, with coefficients of variation (CV) of 3.1% to 7.5% at the lower limit of quantitation (0.1 μmol/L). The intra-assay CV was 5.0% to 7.5%, and the interassay CV was 3.1% to 4.9%. The method was based on that of Okuyama [8,9] and HPLC is suitable for the analysis of major bile acids in normal serum [10]. According to its method, the separation of the individual bile acids including unconjugated, glycine-and taurine-conjugated bile acids was investigated by using HPLC and immobilized enzyme in column form. The separation is very clear except glycodeoxycholic acid and taurochenodeoxycholic acid. Each peak is equivalent to 100 ng of bile acid. The estimated detection limit is 10 ng approximately. After EDTA blood collection, all samples were centrifuged at 3000 rpm for 8 min and immediately frozen at −80 °C. The time between collection and centrifugation was approximately 30 min. Stability of EDTA-plasma was assessed immediately and after storage for 1, 4, 8 and 24 h at room temperature. Immediately frozen samples did not differ in BA levels compared with samples stored for 1, 4, 8 and 24 h at room temperature [11].
HbA1C levels at baseline were 84.5 ± 6.7 mg/dL and 4.9 ± 0.3%, respectively. The mean body fat mass (BFM) was 15.2 ± 3.1 kg, the mean skeletal muscle mass was 25.3 ± 6.2 kg, and the visceral fat area was 60.9 ± 20.6 cm2. The total BA concentrations 30 min, 60 min, 90 min, and 120 min after ingestion of a 400-kcal test meal were significantly higher than that of the baseline (from 1.3 ± 0.9 to 3.0 ± 2.4 μmol/L, 3.1 ± 2.5 μmol/L, 4.4 ± 2.7 μmol/L, and 5.4 ± 3.3 μmol/L; P = 0.024, P = 0.039, P = 0.004, and P = 0.002, respectively) (Fig. 1). Similarly, the total GCBA concentrations 30 min, 60 min, 90 min, and 120 min after ingestion were significantly higher than that of the baseline (from 0.7 ± 0.6 μmol/L to 2.5 ± 2.1 μmol/L, 2.3 ± 1.6 μmol/L, 3.6 ± 2.1 μmol/L, and 4.6 ± 2.6 μmol/L; P = 0.008, P = 0.005, P b 0.001, and P b 0.001, respectively) (Fig. 1). The total taurine-conjugated BA concentration 120 min after ingestion was significantly higher than that at baseline (P = 0.005). The BMI was correlated with total GCBA at baseline (r = − 0.721, P = 0.012) (Fig. 2). The BFM was negatively correlated with total GCBA at 30 min (r = –0.688, P = 0.019) and 60 min (r = –0.642, P = 0.033). In addition, BFM was negatively correlated with total BA at 30 min (r = –0.688, P = 0.019) and 60 min (r = –0.642, P = 0.033). However, other BA variables, including total BAs and total GCBA, were not significantly correlated with either skeletal muscle mass or visceral fat area. The BA fractions did not differ by sex or BFM.
a Concentratinons (µ µmol/L)
2
Bioelectric impedance analysis
Statistical analysis The SPSS software package (version 11.0; SPSS Inc., Chicago, IL, USA) was used for all statistical analyses. Spearman's correlation was used to evaluate correlations. Data were tested for normality with the Shapiro– Wilk W test. Continuous variables are expressed as means ± standard deviation (SD) and correlation coefficients (r). A P-value of b0.05 was considered to indicate statistical significance. Results The mean age of the subjects was 28.6 ± 1.6 years, with a bodymass index [BMI], 22.0 ± 1.6 (19.2–24.1) kg/m2 [men; 22.2 ± 1.8 (19.9–24.0), women; 21.8 ± 1.6 (21.8–24.1)]. The mean FPG and
b Concentratinons (mmol/L)
Subjects underwent bioelectric impedance analysis 1 h before they ingested the test meal. Body composition was analyzed with a bioelectric impedance device (InBody 720, BioSpace Co., Ltd., Seoul, Korea) and associated software (InBody 3.0, BioSpace) as previously described [12,13]. This device employs an 8-point tactile electrode system and measurements at 6 frequencies (1, 5, 50, 250, 500, and 1000 kHz).
Time (min)
Time (min) Fig. 1. Changes in postprandial total BA concentrations (a) and total GCBA concentrations (b) after ingestion of a test meal (mean ± SD). Total BA: total bile acid; and total GCBA: total glycine-conjugated bile acid.
Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
T. Suzuki et al. / Clinical Biochemistry xxx (2014) xxx–xxx
3
Fig. 2. The correlation between FV and postprandial total GCBA, and total BA concentrations (mean ± SD) after ingestion of a test meal. (a) BFM and total GCBA at 30 min, (b) BFM and total GCBA at 60 min, (c) BFM and total BA at 30 min, and (d) BFM and total BA at 60 min. BFM: body fat mass; total GCBA: total glycine-conjugated bile acid; and total BA: total bile acid.
Discussion Both total BA concentrations and total GCBA concentrations increased after ingestion of a 400-kcal test meal. These results are comparable with those of a previous report [5]. Moreover, BMI was negatively correlated with total GCBA. In addition, BFM was negatively correlated with total GCBA and total BA. Postprandial levels of BAs may change according to body composition, such as BFM, even if subjects are healthy and of normal weight. In fact, the postprandial BA response in obese subjects has been shown to be lower than that in subjects of normal weight [4]. BAs are important biochemical modulators that activate certain nuclear receptors, such as farnesoid X receptor and vitamin D and G protein-coupled TGR5, which regulate GLP-1 and the gut hormone peptide YY. Both GLP-1 and peptide YY are secreted by L-cells of the small intestine in response to BAs. Rectal administration of BA increases GLP-1 and peptide YY by increasing the sensation of fullness and decreasing subsequent consumption compared with control [14]. In addition, the responses of GLP-1 and peptide YY in healthy male subjects are correlated with levels of CDCA [15]. To date, 2 hormones have been found to fulfill the definition of incretin hormones: GLP-1 and gastric inhibitory peptide (GIP). GIP is a
physiological gut peptide secreted from intestinal K-cells and has insulin-releasing actions, whereas GLP-1 is produced and released from L-cells. Both fat and protein markedly stimulate GIP secretion. We speculate that the postprandial BA response in healthy subjects of normal weight is associated with GLP-1 and peptide YY and with GIP, a type of incretin, which directly links overnutrition to obesity. In the present study, levels of BAs were negatively correlated with BFM but not with visceral fat area. However, these correlations did not differ between men and women. Therefore, further investigations, including studies of incretin hormones, such as GLP-1 and GIP, will be needed to clarify the association between BFM and BAs. Our study had several limitations. The subjects were few and were young volunteers; therefore, the findings might not be applicable to middle-aged or elderly persons. We believe that a case–control study with a greater number of subjects should be performed.
Conclusion The current results show that postprandial BA response is inversely related with body fat mass in healthy subjects of normal weight.
Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
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T. Suzuki et al. / Clinical Biochemistry xxx (2014) xxx–xxx
Conflict of interest statement The authors declare that there are no conflicts of interest.
Acknowledgment This study received no specific grant from any funding agency in the public and commercial sections. We are grateful to Masao Okazaki, MD, of the Academic Information Center, The Jikei University School of Medicine, for his careful revision of the English language of the manuscript.
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Please cite this article as: Suzuki T, et al, Correlation between postprandial bile acids and body fat mass in healthy normal-weight subjects, Clin Biochem (2014), http://dx.doi.org/10.1016/j.clinbiochem.2014.04.025
