Resveratrol Metabolites Modify Adipokine Expression and Secretion in 3T3-L1 Pre-Adipocytes and Mature Adipocytes

Portillo MP (2013) Resveratrol Metabolites Modify Adipokine Expression and Secretion in 3T3-L1 Pre-Adipocytes and Mature Adipocytes. PLoS ONE 8(5): e63918. doi:10.1371/journal.pone.0063918 Resveratrol Metabolites Modify Adipokine Expression and Secretion in 3T3-L1 Pre-Adipocytes and Mature Adipocytes Itziar Eseberri 0 Arrate Lasa 0 Itziar Churruca 0 Mara P. Portillo 0 Pratibha V. Nerurkar, College of Tropical Agriculture and Human Resources, University of Hawaii, United States of America 0 1 Nutrition and Obesity Group, Department of Nutrition and Food Science, University of the Basque Country (UPV/EHU) , Vitoria, Spain, 2 CIBER de Fisiopatolog a de la Obesidad y Nutricio n (CIBEROBN) , Instituto de Salud Carlos III , Madrid , Spain Objective: Due to the low bioavailability of resveratrol, determining whether its metabolites exert any beneficial effect is an interesting issue. Methods: 3T3-L1 maturing pre-adipocytes were treated during differentiation with 25 mM of resveratrol or with its metabolites and 3T3-L1 mature adipocytes were treated for 24 hours with 10 mM resveratrol or its metabolites. The gene expression of adiponectin, leptin, visfatin and apelin was assessed by Real Time RT-PCR and their concentration in the incubation medium was quantified by ELISA. Results: Resveratrol reduced mRNA levels of leptin and increased those of adiponectin. It induced the same changes in leptin secretion. Trans-resveratrol-3-O-glucuronide and trans-resveratrol-49-O-glucuronide increased apelin and visfatin mRNA levels. Trans-resveratrol-3-O-sulfate reduced leptin mRNA levels and increased those of apelin and visfatin. Conclusions: The present study shows for the first time that resveratrol metabolites have a regulatory effect on adipokine expression and secretion. Since resveratrol has been reported to reduce body-fat accumulation and to improve insulin sensitivity, and considering that these effects are mediated in part by changes in the analyzed adipokines, it may be proposed that resveratrol metabolites play a part in these beneficial effects of resveratrol. - Funding: This study was supported by grants from the Ministerio de Economa y Competitividad (AGL2011-27406-ALI), Instituto de Salud Carlos III (RETIC PREDIMED and CiberObn), Government of the Basque Country (IT-512-13) and University of the Basque Country (UPV/EHU) (ELDUNANOTEK UFI11/32). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. In recent years, a remarkable range of biological functions has been ascribed to resveratrol. It shows chemopreventive, antiinflammatory and antioxidant properties [1], [2]. Beneficial cardiovascular effects have also been described [3]. More recently, resveratrol has been proposed as a potential anti-obesity compound [4-11], which also improves insulin sensitivity [4], [6], [11], [12]. Several studies have attributed the beneficial effects of resveratrol on body fat accumulation and glycemic control [13 16] in part to the changes induced by this polyphenol in adipokines production. Leptin increases energy expenditure and reduces food intake [17], [18]. This adipokine is also related to glycemic control. A physiological increase in plasma leptin levels has been shown to inhibit insulin secretion [19]. Adiponectin increases glucose uptake in muscles and insulin sensitivity, suppresses gluconeogenesis in hepatocytes [20] and increases fatty acid oxidation [21]. With regard to these adipokines, it has been reported that resveratrol reduces leptin expression and secretion and increases adiponectin expression in both in vitro and in vivo studies [13], [22]. Other adipokines, more recently discovered, such as visfatin and apelin, have been partially involved in obesity and glucose homeostasis. Apelin increases the expression of uncoupling proteins UCP1 and 3, increases energy expenditure and decreases respiratory quotient, resulting in increased fat oxidation [23]. Both of them improve insulin sensitivity and maintain glucose homeostasis in rodents and humans. Visfatin has been shown to play an important role in pancreatic b-cell function by acting as an intra and an extracellular NAD biosynthetic enzyme and regulating glucose-stimulated insulin secretion [24]. Apelin has a close relationship with insulin because it enhances glucose uptake in insulin responsive tissues, such as skeletal muscle [2528]. Derdemezis et al. [29] observed decreased visfatin secretion in SGBS adipocytes treated with resveratrol. No data have been reported to date concerning the effect of resveratrol on apelin production. One of the main concerns of scientists working in the field of resveratrol is its low bioavailability. It has been described that only a small proportion of this molecule reaches plasma and tissues 59- ATT TAA GGA CAC GCT GAT CAA AGG-39 59-GCT CTT CAG TTG TAG TAA CGT CAT C-39 after its oral intake [30], [31]. The concentrations of glucuronide and sulfate metabolites are relatively higher [3234]. In order to increase resveratrol bioavailability, and thus its probable effectiveness as a functional ingredient for prevention and treatment of several diseases, different strategies are under evaluation: combination with other molecules which inhibit its metabolization [35], [36], encapsulation with different excipients (microencapsulation or nanoencapsulation), or searching for more resistant structural analogues (pterostilbene). However, to better determine the target for these research strategies, it is very important to know whether these metabolites exert any effect, and to compare the magnitude of these effects with those of resveratrol. In this line of research it has been reported that several metabolites show biological activities, such as cancer chemoprotection [37], [38] and anti-inflammation [38], [39]. With regard to the effects of resveratrol metabolites on lipid metabolism there is only one study, reported recently by our group [40]. In this context, the aim of the present study was to determine the effect of resveratrol and the following resveratrol phase II metabolites: trans-resveratrol-3-O-glucuronide, trans-resveratrol-49O-glucuronide and trans-resveratrol-3-O-sulfate, on adipokine production in 3T3-L1 maturing pre-adipocytes and mature adipocytes. Materials and Methods Reagents Dulbeccos modified Eagles medium (DMEM) was purchased from GIBCO (BRL Life Technologies, Grand Island, NY). TransResveratrol (98% purity), trans-resveratrol-3-O-glucuronide (95% purity), trans-resveratrol-49-O-glucuronide (95% purity) and transresveratrol-3-O-sulfate (98% purity) were provided by Bertin Pharma (Montigny le Bretonneux, France). Experimental Design 3T3-L1 pre-adipocytes, supplied by American Type Culture Collection (Manassas, VA, USA), were cultured in DMEM containing 10% foetal calf serum (FCS). Two days after confluence (...truncated)