PLoS ONE. 2014 Dec 12;9(12):e114790. doi:10.1371/journal.pone.0114790

Wild blueberries (Vaccinium myrtillus) alleviate inflammation and hypertension associated with developing obesity in mice fed with a high-fat diet.

Otto T. Mykkänen1, Anne Huotari2, Karl-Heinz Herzig3, Thomas W. Dunlop2, Hannu Mykkänen1 and Pirkka V. Kirjavainen1,4


1Institute of Public Health and Clinical Nutrition, Department of Clinical Nutrition, Food and Health Research Centre, University of Eastern Finland, FI-70211 Kuopio, Finland

2Institute of Biomedicine, University of Eastern Finland, FI-70211 Kuopio, Finland

3Institute of Biomedicine and Biocenter of Oulu, University of Oulu, FI-90014 Oulu, Finland, Medical Center Oulu and Oulu University Hospital, FI-90220 Oulu, Finland.

4Department of Environmental Health, National Institute for Health and Welfare P.O. Box 95. FI-70701 Kuopio



Background: Lowgrade metabolic inflammation and hypertension are primary mechanisms involved in obesity-associated adverse health effects. Berries, especially Nordic wild blueberries (hereafter referred to as bilberries), represent an important source of dietary anthocyanins, a group of polyphenols with potential beneficial effects to combat obesity-associated metabolic disturbances.  Methods: The effects of 5% or 10% (w/w) of whole bilberries (BB) were studied on the development of obesity and its metabolic disturbances in C57BL mice fed with a high-fat diet (HFD) for three months. Cytokines, inflammatory cells, systolic blood pressure, glucose tolerance, insulin sensitivity, weight gain, body fat, food consumption and energy metabolism were assessed. Results: Bilberries ameliorated type 1 pro-inflammatory responsiveness induced by HFD. This was indicated by altered cytokine profile and reduced prevalence of interferon gamma -producing T-cells, in particular T helper type 1 cells. Bilberries also prevented the progression of obesity associated long term increase in systolic blood pressure in mice. Conclusions: Bilberries reduce the development of systemic inflammation and prevent the progression of chronic hypertension, thus supporting their potential role in alleviating the adverse health effects associated with developing obesity.

PMID: 25501421

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Obesity is associated with a high number of comorbidities, e.g. metabolic syndrome (MetS), type 2 diabetes (T2D) and cardiovascular diseases. Obesity has been well linked to low-grade inflammation and hypertension (1,2). Berries, especially bilberries (BB) and blueberries have been shown to have potential in the preventive management of these complications. Bilberries (wild European blueberries, Vaccinium myrtillus) are rich in anthocyanins (ANCs), making them a unique dietary source of flavonoids (3-5). Whilst fruits and berry peels (3) in general are rich in ANCs, bilberries contain also other flavonoids than anthocyanins (flavonols, phenolic acids and pro-anthocyanidins), and they contain vitamins C and E which all may be partly responsible for their biological effects. Previous studies on the anti-inflammatory effects of whole blueberries and bilberries of Vaccinium species have concluded that the health effects of these berries could be mediated by modulation of immune responses (6-9), but these effects are poorly characterized and virtually nothing is known about the systemic effects of bilberries at the cellular level. Earlier studies have also shown that ANCs or their sources reduce blood pressure in rodent models of hypertension (10-12), but these studies did not utilize whole berries.

Since prolonged feeding of C57BL “black” mice with a high-fat diet (HFD) is known to increase adiposity and production of inflammatory cytokines, we examined the effects of bilberries in a high-fat diet mouse model of obesity. The C57BL mice fed with HFD become obese within weeks and with prolonged feeding they develop insulin resistance and glucose intolerance, hypertension and low grade (metabolic, meta-) inflammation (13-18). This was the first study to utilize the HFD mouse model to comprehensively examine the effects of whole bilberries on the key features of MetS and obesity (energy and glucose metabolism, blood pressure and body fat content) together with an extensive concomitant analysis of their immunomodulatory effects.



Figure 1. Weight gain and fat percentage of mice fed HFD and bilberries. The diets used were normal control diet (NCD), high-fat diet (HFD), 5% and 10% (w/w) BB in HFD fed for 12–14 weeks. The time points for testing systolic blood pressure (SBP), glucose tolerance (GTT), insulin sensitivity (IST) and respiratory exchange ratio (RER) in four repeated experiments are displayed in the figure. The bars represent the means and SEM of n = 22–37 mice per diet group in measured weight gain and n = 8–9 mice in measured fat %. Asterisk (*) indicates the significant differences of HFD group from NCD and 5% BB and 10% BB in HFD groups (p<0.05, Mann Whitney U-Test with Bonferroni’s correction).


In our study mice became obese and had increased adiposity as expected when fed with HFD. Bilberries reduced the weight gain and showed a trend in the reduction of adiposity (Figure 1). We also showed that bilberries reduced several cytokines related to pro-inflammatory conditions. Cytokines such as IL-13, IL-15 and interferon gamma (IFN-γ) were increased by HFD and reduced below detection in mice fed with bilberries. Also significant reduction of monocyte chemoattractant protein-1 (MCP-1) in both groups of mice receiving bilberries attests that these berries ameliorate type 1-proinflammatory markers in circulation. Furthermore, T helper type 1 cells (Th 1) type cytokines (IL-2, IL-12, IFN-γ and TNF-α) and the ratio of T helper type 1 and type 2 cells (Th1/Th2) were reduced in bilberry fed animals (Figure 2). In addition, the percentage of NKT of all cytotoxic cells was significantly lower in mice fed 10% bilberries fed than in HFD fed mice (2.5 ± 0.2 vs.7.0 ± 1.2, p<0.05). Furthermore, mice fed with BB in HFD did not develop hypertension, while systolic blood pressure (SBP) in mice receiving HFD was moderately increased at 7-8 weeks and significantly increased at 11-12 weeks (Figure 3).


Figure 2Proportion of pro-inflammatory cells and levels of cytokines in serum from mice fed NCD, HFD or bilberries in HFD. The ratio of T helper type 1 cells (Th1, CD3+CD4+INFg+IL4-) to T helper type 2 cells (Th2, ) in spleen and Th1 type cytokines (IL-2, IL-12, INF-γ and TNF-α) in blood. The bars represent the mean and SEM of n = 6-11 mice per diet group. Asterisc (*) indicates the significant differences of HFD group from NCD and 5% BB and 10% BB in HFD groups (p<0.05, Mann Whitney U-Test with Bonferroni’s correction).



Figure 3 Effects of feeding NCD, HFD, or bilberries in HFD on the systolic blood pressure in mice. Systolic blood pressure (SBP) was measured by the non-invasive Tail Cuffs photoelectric method at 7-8 and 11-12 weeks after the start of the diets. The bars represent the means and SEM of n = 10-14 mice per diet group. Asterisk (*) indicates the significant differences of HFD group from NCD and 5% BB and 10% BB in HFD groups (*p<0.05, **p<0.001, Mann Whitney U-Test with Bonferroni’s correction).


In conclusion, HFD tended to increase pro-inflammatory cytokines in parallel with increased weight gain and body fat. However, addition of bilberries to HFD reduced several parameters of inflammation, approaching the levels found in NCD fed mice. Bilberry intake appeared to consistently direct the HFD induced immune profile away from the type 1 cell-mediated immune responses. Similar findings have been reported with blueberries (Vaccinium angustifolium, Vaccinium ashei and V. corymbosum) in rodents (6,9) as well as in human studies using bilberries (7,8).

Our current and previous findings (19) are in line with reported results on Vaccinium species berries (11,20) and other sources of ANCs (10) showing reduced blood pressure in rodents. Bilberry or blueberry intake has been associated with reduction of blood pressure also in humans (21), but the findings from intervention studies have thus far been inconsistent (8,22,23). Nevertheless, in this study we demonstrated that bilberries can prevent the development of hypertension in a dose dependent manner.

Importance of the study: We have shown here with the HFD mouse model that bilberries can ameliorate or prevent metabolic disturbances associated with developing obesity, especially systemic low-grade inflammation and hypertension. Since the obesity associated pathologies (IR-T2D-hypertension) are usually clustered (24), dietary approaches influencing all of these pathologies may be more beneficial than treating hypertension, inflammation or abnormal glucose levels alone. Our findings attest to the epidemiological (21,25,26) and clinical (27) evidence supporting bilberries as a promising candidate for the prevention of obesity related hypertension and low-grade inflammation.



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Acknowledgements: This study was supported The work was supported by the Academy of Finland project #133431 (Effects of berry consumption on obesity-associated metabolic disorders), the Graduate School of Applied Biosciences, Yrjö Jahnsson Foundation, The Northern Savo Fund of Finnish Cultural Foundation, Olvi Foundation, the University of Eastern Finland and in part by grants from the Academy of Finland (#108478 and #129568). Bilberries were kindly provided as gift from a local company, Joswola, Iisalmi, Finland and freeze dried within European networks project collaboration.



Otto T. Mykkänen, M.Sc.

Institute of Public Health and Clinical Nutrition, University of Eastern Finland, FI-70211 Kuopio, Finland.



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