Anthocyanin enhances adipocytokine secretion and adipocyte-specific gene expression in isolated rat adipocytes q Takanori Tsuda, a, * Yuki Ueno, a Hiromitsu Aoki, b Takatoshi Koda, b Fumihiko Horio, d Nobuyuki Takahashi, c Teruo Kawada, c and Toshihiko Osawa d a Research Center for Biomarkers of Preventive Medicine, Doshisha University, Kamigyo-ku, Kyoto 602-8580, Japan b San-Ei Gen F.F.I., Inc., Toyonaka 561-8588, Japan c Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan d Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan Received 3 February 2004 Abstract Adipocyte dysfunction is strongly associated with the development of obesity and insulin resistance. It is accepted that the regulation of adipocytokine secretion or the adipocyte-specific gene expression is one of the most important targets for the pre- vention of obesity and amelioration of insulin sensitivity. In this study, we demonstrated that anthocyanins (cyanidin or cyanidin 3- glucoside) have the potency of a unique pharmacological function in isolated rat adipocytes. Treated adipocytes with anthocyanins enhanced adipocytokine (adiponectin and leptin) secretion and up-regulated the adipocyte specific gene expression without acti- vation of PPARc in isolated rat adipocytes. The gene expression of adiponectin was also up-regulated in white adipose tissue in mice fed an anthocyanin supplemented diet. As one of the possible mechanisms, AMP-activated protein kinase activation would be associated with these changes, nevertheless, the AMP:ATP ratio was significantly decreased by administration of the anthocyanins. These data suggest that anthocyanins have a potency of unique therapeutic advantage and also have important implications for preventing obesity and diabetes. Ó 2004 Elsevier Inc. All rights reserved. Keywords: Anthocyanin; Cyanidin; Adipocyte; Adipocytokine; Adiponectin; Adipocyte-specific gene; Peroxisome proliferator-activated receptor Adipocyte is the primary site of energy storage and accumulates triacylglycerol during nutritional excess. In recent years, it is well known that adipocyte dysfunction plays an important role in the development of obesity and insulin resistance. Adipocyte synthesizes and secretes biologically active molecules called adipocytokines [1]. For example, leptin is the product of the ob gene and is secreted from adipocytes, and reduces food intake and increases energy expenditure [2]. Adiponectin is one of the most important adipocytokines, and is specifically and highly expressed in adipocytes. The plasma adipo- nectin concentration and mRNA expression level are decreased in the obese and insulin resistant state [3,4]. The administration of adiponectin improves insulin ac- tion accompanied by increases in fatty acid oxidation and a decreased triacylglycerol level in muscle [5,6]. There are some drugs which are the target for regu- lation of the adipocyte function to improve insulin sensitivity or glucose homeostasis. The peroxisome proliferator-activated receptor (PPAR)c is the master regulator of adipocyte differentiation and controls var- ious kinds of adipocyte specific gene expressions. Thia- zolidinediones (TZD) are some of the synthetic PPAR ligands, and used for antidiabetic drugs through their q Abbreviations: AMPK, AMP-activated protein kinase; aP2, adipocyte fatty acid binding protein; BSA, bovine serum albumin; C3G, cyanidin 3-O-b-D-glucoside; Cy, cyanidin; ELISA, enzyme-linked immunosorbent assay; FFA, free fatty acid; HPLC, high-performance liquid chromatography; KRBH, Krebs–Ringer bicarbonate buffer; LPL, lipoprotein lipase; MAPK, mitogen-activated protein kinase; PCC, purple corn color; PPAR, peroxisome proliferator-activated receptor; ROS, reactive oxygen species; RT-PCR, reverse transcription- polymerase chain reaction; TTBS, Tris–HCl-buffered saline containing 0.05% Tween 20; TZD, thiazolidinediones; UCP, uncoupling protein. * Corresponding author. Fax: +81-75-251-4418. E-mail address: ttsuda@mail.doshisha.ac.jp (T. Tsuda). 0006-291X/$ - see front matter Ó 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2004.02.031 Biochemical and Biophysical Research Communications 316 (2004) 149–157 BBRC www.elsevier.com/locate/ybbrc