Elsevier

Journal of Functional Foods

Volume 54, March 2019, Pages 371-380
Journal of Functional Foods

Inhibitory effects of lingonberry (Vaccinium vitis-idaea L.) fruit extract on obesity-induced inflammation in 3T3-L1 adipocytes and RAW 264.7 macrophages

https://doi.org/10.1016/j.jff.2019.01.040Get rights and content

Highlights

  • Lingonberries suppress pro-inflammatory genes expression in adipocytes and macrophages.

  • Lingonberries counteract the decrease in adiponectin and IL-10 level in adipocytes.

  • Lingonberries decrease ROS overproduction in TNF-α-induced adipocytes.

  • Lingonberries affect the expression of antioxidant and oxidant enzymes in adipocytes.

Abstract

In this study, the lingonberry fruit extract (LGBE) has been investigated for the ability to suppress the inflammatory response and mitigate oxidative stress in activated 3T3-L1 adipocytes and RAW 264.7 macrophages. The obtained results showed that LGBE significantly reduced IL-6, MCP-1 and IL-1β production, and counteracted the decrease in adiponectin and IL-10 expression in TNF-α-induced adipocytes. Moreover, LGBE exhibited a high anti-inflammatory potential in the macrophage cell culture by down-regulation the expression of pro-inflammatory mediators (IL-6, TNF-α, IL-1β, MCP-1, COX-2, iNOS). Furthermore, LGBE decreased intracellular ROS generation in inflamed adipocytes by enhancing expression of antioxidant defense enzymes (SOD, catalase, GPx) and inhibiting an oxidant enzyme (NADPH oxidase 4), which is one of the primary sources of ROS.

In conclusion, we have demonstrated that lingonberry fruit may reduce adipose tissue inflammation and support immune cell homeostasis, and thus can be considered a natural tool for inflammation control.

Introduction

Obesity-induced inflammation characterized by the increased cytokine expression and immune cell infiltration is one of the complications of metabolic syndrome and insulin resistance as a result of over-nutrition and stress pathways that drive abnormal metabolic homeostasis (e.g., high levels of lipid, glucose and reactive oxygen species) (Lee & Lee, 2014). Adipose tissue is an endocrine organ and produces of various hormones, and cytokines include leptin, adiponectin, resistin, and cytokines such as TNF-α and IL-6. In obesity state, adipose tissue expresses very high levels of many inflammatory mediators and is considered to be the main inflammatory organ that mediates obesity-induced inflammation (Ouchi, Parker, Lugus, & Walsh, 2011). While obesity-induced inflammation resembles the inflammation in classical immunity, it also differs from the latter in that it is a low-grade inflammation that produces much lower levels of circulating cytokines and is also considered to be chronic inflammation (Lee & Lee, 2014).

The adipose tissue consists of adipocytes and non-adipocyte cells, including endothelial cells, preadipocytes and immune cells (Shoelson, Herrero, & Naaz, 2007). Many different types of immune cells are found in adipose tissue, however, adipose-specific macrophages play a crucial role in the regulation of obesity-induced inflammation (Weisberg et al., 2003). Adipocytes and macrophages are the primary sources of inflammatory proteins (Ouchi et al., 2011). In lean mice and humans, macrophages constitute around 5% of the cells in adipose tissue, during obesity their number increases to 50% of all adipose tissue cells (Weisberg et al., 2003). Thus, the immune cell-adipocyte crosstalk is an essential regulator of adipose tissue function and systemic metabolism (Chung, Nati, Chavakis, & Chatzigeorgiou, 2018). Besides, considering the broad spectrum of pro- and anti-inflammatory adipokines, which are altered in obesity, it is likely that crosstalk of many adipokines rather than a single adipokine in adipose tissue may be involved in the metabolic dysregulation (Jung & Choi, 2014). There are two types of inflammation-related proteins: pro-inflammatory, including MCP-1, TNF-α, IL-6, IL-1β, leptin, and anti-inflammatories, such as IL-10 and adiponectin. Pro-inflammatory cytokines such as IL-6, IL-1β, and TNF-α are elevated in most, if not all, inflammatory states and have been recognized as targets for therapeutic intervention (Scheller, Chalaris, Schmidt-Arras, & Rose-John, 2011).

Recent studies have shown the beneficial health effect of berry fruits in attenuating adipose tissue inflammation and insulin resistance in experimental models of metabolic syndrome (Kowalska & Olejnik, 2016b). Importantly, berries contain a variety of bioactive compounds that may act synergistically in the prevention of adipose tissue inflammation and insulin resistance (Kowalska and Olejnik, 2016a, Kowalska and Olejnik, 2016b, Kowalska et al., 2017). Lingonberry fruit has been found to exhibit high antioxidant and anti-inflammatory activities (Grace, Esposito, Dunlap, & Lila, 2014). The potential health benefits of lingonberries are attributed to their diverse polyphenol composition. Polyphenolics such as flavonoids, phenolic acids, anthocyanins, and procyanidins as well as organic acids and vitamins (A, B1, B2, B3 and C), have been determined in lingonberries (Ek, Kartimo, Mattila, & Tolonen, 2006).

In this study, we investigated the ability of lingonberry fruit extract to mitigate adipose tissue inflammation using TNF-α-induced 3T3-L1 adipocytes and activated RAW 264.7 macrophages. The anti-inflammatory effects were evaluated based on the expression of pro-inflammatory and anti-inflammatory cytokines. Since is well known that berries possess high antioxidant activity and reactive oxygen species (ROS) are involved in the development of inflammation we examined the effect of lingonberry extract on intracellular ROS production and antioxidant defense enzymes expression in 3T3-L1 adipocytes treated with TNF-α.

Section snippets

Reagents

Dulbecco’s Modified Eagle’s Medium (DMEM), insulin, dexamethasone (DEX), 3-isobutyl-1-methylxanthine (IBMX), lipopolysaccharides (LPS) from E. coli O127, TNF-α were purchased from Sigma (Sigma–Aldrich, Poland). Fetal bovine serum (FBS) was obtained from Gibco BRL (Grand Island, NY, USA). The total RNA was isolated using the Tri Reagent (Sigma–Aldrich, Poland). Template cDNA synthesis was performed using the Transcriptor First Strand cDNA Synthesis Kit (Roche Diagnostics, Poland). PCR was

Polyphenol composition in the lingonberry fruit extract

The results of HPLC-DAD-ESI-MS analysis of the polyphenolic compounds present in the aqueous extract of the freeze-dried lingonberry fruit are shown in Table 1. The identified compounds were classified as members of several groups, namely anthocyanins (peaks 1–5), flavanols (peaks 7–8) and B-type and A-type proanthocyanidins (peaks 6, 9–10, 12–13), phenolic acid derivatives (peaks 11, 14–19) and flavonols (peaks 20–26). The HPLC-DAD chromatograms obtained for the extract at the four wavelengths

Discussion

Obesity increases the risk of developing a variety of pathological metabolic dysfunction, including insulin resistance, type 2 diabetes, dyslipidemia, and hypertension. Growing evidence suggests that chronic inflammation in adipose tissue and adipose tissue macrophage-related inflammatory activities play a critical role in the development of obesity-related metabolic dysfunction (Xu et al., 2003). During obesity, inflammatory adipokines are overproduced, including leptin, TNF-α, IL-6, and

Conclusions

Increase in pro-inflammatory and a reduction in anti-inflammatory adipokines in obese adipose tissue contributes to local and systemic inflammation and disturbances in adipocytes homeostasis. Natural diet interventions which target adipokines and their intracellular signaling cascades to enhance immune function or ameliorate chronic inflammation in obese adipose tissue may be safe and promising therapies to counteract the diet-induced metabolic complications. In conclusion, we have demonstrated

Conflict of interest

The authors declare no conflicts of interest.

Ethics statements file

This study does not include any human subjects and animal experiments

Acknowledgment

The authors thank the company DANEX (Wieleń, Poland) for providing lingonberry fruit for experiments.

This work was supported by the National Science Centre, Poland (Grant No. 2015/19/B/NZ9/01054).

References (43)

  • J.M. Bruun et al.

    Regulation of adiponectin by adipose tissue-derived cytokines: In vivo and in vitro investigations in humans

    American Journal of Physiology Endocrinology and Metabolism

    (2003)
  • K.J. Chung et al.

    Innate immune cells in the adipose tissue

    Reviews in Endocrine and Metabolic Disorders

    (2018)
  • H. Dominguez et al.

    Metabolic and vascular effects of tumor necrosis factor-α blockade with etanercept in obese patients with type 2 diabetes

    Journal of Vascular Research

    (2005)
  • J.A. Ehses et al.

    IL-1 antagonism reduces hyperglycemia and tissue inflammation in the type 2 diabetic GK rat

    Proceedings of the National Academy of Sciences of the United States of America

    (2009)
  • S. Ek et al.

    Characterization of phenolic compounds from lingonberry (Vaccinum vitis-idaea)

    Journal of Agricultural and Food Chemistry

    (2006)
  • A. Fortuno et al.

    Is leptin involved in phagocytic NADPH oxidase overactivity in obesity? Potential clinical implications

    Journal of Hypertension

    (2010)
  • S. Fujisaka et al.

    Regulatory mechanisms for adipose tissue M1and M2 macrophages in diet-induced obese mice

    Diabetes

    (2009)
  • S. Furukawa et al.

    Increased oxidative stress in obesity and its impact on metabolic syndrome

    The Journal of Clinical Investigation

    (2004)
  • M.H. Grace et al.

    Comparative analysis of phenolic content and profile, antioxidant capacity and anti-inflammatory bioactivity in wild Alaskan and commercial Vaccinium berries

    Journal of Agricultural and Food Chemistry

    (2014)
  • L. Heyman-Lindén et al.

    Lingonberries alter the gut microbiota and prevent low-grade inflammation in high-fat diet fed mice

    Food & Nutrition Research

    (2016)
  • N. Hosogai et al.

    Adipose tissue hypoxia in obesity and its impact on adipocytokine dysregulation

    Diabetes

    (2007)
  • Cited by (29)

    • Exploring Vaccinium vitis-idaea L. as a potential source of therapeutic agents: antimicrobial, antioxidant, and anti-inflammatory activities of extracts and fractions

      2022, Journal of Ethnopharmacology
      Citation Excerpt :

      Complex and condensed proanthocyanidins were found to be more potent PKC inhibitors than their monomeric component units with the hypothesis that the activity of proanthocyanidins increase according to the number of hydroxyl groups and presence of galloyl moiety, which are involved in competitive actions towards phorbol ester (Kashiwada et al., 1992). More recent evidence (Kowalska et al., 2019; Limtrakul et al., 2016) underlined the strong anti-inflammatory potential of polymeric proanthocyanidin-rich extracts in macrophage cultures by down-regulation the expression of proinflammatory mediators. This study reports a comprehensive investigation on crude extracts and fractions from widely used lingonberry leaves and fruits, combining results from the specialized metabolite profiling and bioassays, namely agar well diffusion, hyaluronidase inhibiting, hydrogen peroxide scavenging, and its production in a macrophage culture J774 inhibiting properties.

    • The anti-obesogenic effects of dietary berry fruits: A review

      2021, Food Research International
      Citation Excerpt :

      In the recent years, some researches have demonstrated that mice that were exposed to HFD treated with freeze-dried lingonberry fruits exhibited significantly lower risks of inflammation and metabolic homeostasis as compared with those without consuming lingonberry fruits (Heyman-Lindén et al., 2016). Based on cell models, lingonberry fruits extracts rich in polyphenols with anthocyanins, phenolic acids and proanthocyanidins accounting for 16.4%, 31% and 23% of total phenolic compounds, respectively, were found to effectively alleviate inflammation and oxidative stress induced by obesity (Kowalska et al., 2019a). Moreover, consumption of anthocyanins extracted from lingonberry fruits could boost the metabolism of cholesterol and protect mice from developing overweight, hyperlipemia and hypercholesterolemia while reduce the lipid accumulation and damage in the liver induced by a high cholesterol diet (Zhang, Zhou, Huangfu, Wu, and Zhang, 2019) (Table 1).

    • Andean berry (Vaccinium meridionale Swartz) juice in combination with Aspirin modulated anti-inflammatory markers on LPS-stimulated RAW 264.7 macrophages

      2020, Food Research International
      Citation Excerpt :

      Similar effects were observed in another study for A. chilensis extracts on LPS-treated RAW 264.7 macrophages, where the anti-inflammatory effects were attributed to quercetin, myricetin, luteolin, and catechin, together with several anthocyanins such as delphinidin-3-glucoside, delphinidin-3,5-diglucoside, delphinidin-3-sambubioside, and cyanidin-3-sambubioside (Cespedes et al., 2017). Most of the antioxidant capacity derived from berries is involved in the reduction of ROS, contributing to the overall decrease of the inflammation process (Kowalska et al., 2019). Fig. 2 shows the effect of ABJ (Fig. 2A), Aspirin (Fig. 2B), and ABJ + Aspirin (Fig. 2C) in the production of ROS on RAW 264.7 macrophages.

    • Zero waste biorefining of lingonberry (Vaccinium vitis-idaea L.) pomace into functional ingredients by consecutive high pressure and enzyme assisted extractions with green solvents

      2020, Food Chemistry
      Citation Excerpt :

      V. vitis-idaea areal parts and fruits are rich in various nutrients and bioactive compounds, namely sugars glucose and fructose, various organic acids, ω-3 and ω-6 fatty acids, dietary fiber, vitamins B1, B2, B3, C, A, and E, minerals, and polyphenols, mainly anthocyanins, proanthocyanidins, flavonols, hydroxycinnamic acid, and arbutin derivatives (Bujor et al., 2018; Ek, Kartimo, Mattila, & Tolonen, 2006; Hurkova et al., 2019; Mikulic-Petkovsek, Schmitzer, Slatnar, Stampar, & Veberic, 2012; Tian et al., 2018). Due to the strong antioxidant, antiviral, antimicrobial, anti-inflammatory and neuroprotective potential (Grace, Esposito, Dunlap, & Lila, 2014; Heinonen, 2007; Kelly, Vyas, & Weber, 2017), dietary consumption of fresh lingonberries and lingonberry-based products may reduce the risk and development of metabolic (Eid, Ouchfoun, Brault, Vallerand, Musallam, Arnason, & Haddad, 2014; Kowalska, Olejnik, Zielińska-Wasielica, & Olkowicz, 2019), cardiovascular (Kivimäki, Siltari, Ehlers, Korpela, & Vapaatalo, 2013), gastrointestinal (Brown et al., 2014) and neurodegenerative (Vyas, Kalidindi, Chibrikova, Igamberdiev, & Weber, 2013) disorders. Commercially cultivated and collected from the natural habitats, mainly in Scandinavian countries and Canada, lingonberries are consumed either fresh or frozen; however, the major part of harvests is processed into various longer shelf-life food products and pharmaceutical preparations (Bujor et al., 2018; Jin et al., 2019).

    View all citing articles on Scopus
    View full text