Probiotics Blunt the Anti-Hypertensive Effect of Blueberry Feeding in Hypertensive Rats without Altering Hippuric Acid Production

<div><p>Previously we showed that feeding polyphenol-rich wild blueberries to hypertensive rats lowered systolic blood pressure. Since probiotic bacteria produce bioactive metabolites from berry polyphenols that enhance the health benefits of berry consumption, we hypothesized that adding probiotics to a blueberry-enriched diet would augment the anti-hypertensive effects of blueberry consumption. Groups (<i>n</i> = 8) of male spontaneously hypertensive rats were fed one of four AIN ‘93G-based diets for 8 weeks: Control (CON); 3% freeze-dried wild blueberry (BB); 1% probiotic bacteria (PRO); or 3% BB + 1% PRO (BB+PRO). Blood pressure was measured at weeks 0, 2, 4, 6, and 8 by the tail-cuff method, and urine was collected at weeks 4 and 8 to determine markers of oxidative stress (F2-isoprostanes), nitric oxide synthesis (nitrites), and polyphenol metabolism (hippuric acid). Data were analyzed using mixed models ANOVA with repeated measures. Diet had a significant main effect on diastolic blood pressure (<i>p</i> = 0.046), with significantly lower measurements in the BB- vs. CON-fed rats (<i>p</i> = 0.035). Systolic blood pressure showed a similar but less pronounced response to diet (<i>p</i> = 0.220), again with the largest difference between the BB and CON groups. Absolute increase in blood pressure between weeks 0 and 8 tended to be smaller in the BB and PRO vs. CON and BB+PRO groups (systolic increase, <i>p</i> = 0.074; diastolic increase, <i>p</i> = 0.185). Diet had a significant main effect on hippuric acid excretion (<i>p</i><0.0001), with 2- and ~1.5-fold higher levels at weeks 4 and 8, respectively, in the BB and BB+PRO vs. PRO and CON groups. Diet did not have a significant main effect on F2-isoprostane (<i>p</i> = 0.159) or nitrite excretion (<i>p</i> = 0.670). Our findings show that adding probiotics to a blueberry-enriched diet does not enhance and actually may impair the anti-hypertensive effect of blueberry consumption. However, probiotic bacteria are not interfering with blueberry polyphenol metabolism into hippuric acid.</p></div

Diets containing blueberry extract lower blood pressure in spontaneously hypertensive stroke-prone rats

abstract: Oxidative stress in the vasculature and kidneys contributes to hypertension, a major risk factor for cardiovascular disease. Blueberries (BB) are rich in antioxidants, and so we hypothesized that feeding diets enriched with BB would slow the development of hypertension in spontaneously hypertensive stroke-prone rats (SHRSP). Eight-week-old normotensive rats and SHRSP were fed either a control diet (Con) or a diet enriched with 3% freeze-dried BB for 8 weeks. Systolic blood pressure (SBP) was measured at weeks 2, 4, 6, 7, and 8 by the tail cuff method, and urine was collected at weeks 4 and 8. The SBP was elevated in SHRSP relative to normotensive rats over the entire 8-week feeding period. In SHRSP consuming BB, SBP was 19% lower at week 4 and 30% lower at week 6, relative to SHRSP on Con. Maximum SBP was 216 ± 11 mm Hg in SHRSP consuming Con vs 178 ± 15 mm Hg in the BB-fed group (P = .036). Spontaneously hypertensive stroke-prone rats had elevated levels of urine F2-isoprostanes/creatinine relative to normotensive rats, indicating systemic oxidative stress in this strain. Blueberry feeding had no effect on urinary excretion of F2-isoprostanes; therefore, it is unlikely that a systemic antioxidant effect of BB is responsible for the antihypertensive effects at weeks 4 and 6. Blueberry-fed rats had reduced markers of renal oxidative stress, such as proteinuria and kidney nitrites. Thus, a 3% BB diet may be capable of protecting the kidneys from oxidative damage in SHRSP, thereby reducing the magnitude of hypertension

Composition of AIN’93G diets.

<p>^aIngredients purchased from Dyets, Inc. (Bethlehem, PA) except for blueberry powder (donated by the Wild Blueberry Association of North America) and probiotic (VSL#3® purchased from <a href="http://www.vsl3.com/" target="_blank">www.vsl3.com</a>).</p><p>^bNinety percent tetrasaccharides and higher.</p><p>^cComposition (g/kg mineral mix): CaCO₃, 357.0; KH₂PO₄, 196.0; K Citrate•H₂O, 70.78; NaCl, 74.0; K₂SO₄, 46.6; MgO, 24.3; Fe citrate, 6.06; ZnCO₃, 1.65; MnCO₃, 0.63; CuCO_3, 0.31; KIO₃, 0.01; Na₂SeO₄, 0.01025; (NH₄) ₆ Mo₇O₂₄•4H₂O, 0.00795; Na₂SiO₃•9H₂O, 1.45; CrK(SO₄) ₂•12H₂O, 0.275; LiCl, 0.0174; H₃BO₃, 0.0815; NaF, 0.0635; 2NiCO₃•3Ni(OH) ₂•4H₂O, 0.0318; NH₄VO₃, 0.0066.</p><p>^d Composition (g/kg vitamin mix): thiamin HCl, 0.6; riboflavin, 0.6; pyridoxine HCl, 0.7; nicotinic acid, 3.0; Ca pantothenate, 1.6; folic acid, 0.2; D-biotin, 0.02; vitamin B12 (0.1% in mannitol), 2.5; vitamin A palmitate (500 000 IU/g), 0.8; DL-α-tocopheryl acetate (500 IU/g), 15; vitamin D3 (400 000 IU/g), 0.25; vitamin K/dextrose 10 mg/g (phylloquinone), 7.5.</p><p>*Nutritional analysis of blueberry powder: 397 kcalories/100 g, 77.2% by weight available carbohydrate, 3.15% protein, 2.3% fat, 13.8% insoluble fiber, 3.6% soluble fiber.</p><p>Composition of AIN’93G diets.</p

Urine levels of hippuric acid.

<p>Urine levels of hippuric acid normalized to creatinine at weeks 4 and 8 in spontaneously hypertensive rats fed control (CON), 3% blueberry (BB), 1% probiotic (PRO), or 3% blueberry + 1% probiotic (BB+PRO) diet for 8 weeks. Data points are presented as means ± SE bars for 8 rats in each diet group except CON week 8, where <i>n</i> = 7. Statistical analysis was performed using mixed models ANOVA with repeated measures and post-hoc comparisons with Tukey’s adjustment. Diet exerted a significant main effect on hippuric acid level, <i>p</i><0.0001. Within week, means with different letters are significantly different at <i>p</i><0.0001 except BB vs. PRO week 8 at <i>p</i> = 0.001 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142036#pone.0142036.s003" target="_blank">S3 Dataset</a>).</p

Systolic blood pressure.

<p>Systolic blood pressure of spontaneously hypertensive rats fed control (CON), 3% blueberry (BB), 1% probiotic (PRO), or 3% blueberry + 1% probiotic (BB+PRO) diet for 8 weeks. Data points are presented as means ± SEM for 8 rats in each diet group, with statistical analysis performed by mixed models ANOVA with repeated measures. Diet did not exert a significant main effect on systolic blood pressure across time points (<i>p</i> = 0.220) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142036#pone.0142036.s002" target="_blank">S2 Dataset</a>).</p

Inhibition of matrix metalloproteinase activity in DU145 human prostate cancer cells by flavonoids from lowbush blueberry (Vaccinium angustifolium): possible roles for protein kinase C and mitogen-activated protein-kinase-mediated events

Regulation of the matrix metalloproteinases (MMPs) is crucial to regulate extracellular matrix (ECM) proteolysis which is important in metastasis. This study investigated the mechanism(s) by which three flavonoid-enriched fractions from lowbush blueberry (Vaccinium angustifolium) down-regulate MMP activity in DU145 human prostate cancer cells. Metalloproteinase activity was evaluated from cells exposed to \u201ccrude,\u201d anthocyanin-enriched (AN) and proanthocyanidin-enriched (PAC) fractions. Differential down-regulation of MMPs was observed. The activity of the endogenous tissue inhibitors of metalloproteinases (TIMPs) from these cells was also evaluated. Increases in TIMP-1 and TIMP-2 activity were observed in response to these fractions. The possible involvement of protein kinase C (PKC) and mitogen-activated protein (MAP) kinase pathways in the flavonoid-mediated decreases in MMP activity was observed. These findings indicate that blueberry flavonoids may use multiple mechanisms in down-regulating MMP activity in these cells.Peer reviewed: YesNRC publication: Ye

Choice of diet impacts the incidence of stroke-related symptoms in the spontaneously hypertensive stroke-prone rat model

The spontaneously hypertensive stroke-prone (SHRSP) rat is a commonly used model of cerebrovascular disease and hypertension. SHRSP rats have been shown to develop stroke-related symptoms (SRS) by age 14 weeks when fed a purified diet, such as AIN-93G, supplemented with 1% NaCl. We conducted a pathology pilot study to compare the incidence of SRS in SHRSP rats fed either AIN-93G (with 1% NaCl in drinking water) or commercially available rat chow (with 4% NaCl in the diet), starting at 8 weeks of age. These results prompted us to analyze data from 5 earlier feeding trials using SHRSP rats. Overall, we found that SHRSP rats fed AIN-93G purified diet for 8 or 17 weeks did not demonstrate SRS (n = 18), whereas all SHRSP rats fed lab chow exhibited SRS at age 15.1 ± 0.6 weeks (n = 23). In addition, SHRSP rats fed lab chow had decreased mass gain starting at age 13 weeks, as well as decreased feed efficiencies after the first 5 weeks of feeding (p < 0.05). In conclusion, our data suggest that diet composition is a major contributor to the onset of stroke in SHRSP rats and that diet choice should be critically evaluated based on endpoint measures in the SHRSP model

Depression of the electroretinogram in rats deficient in zinc and taurine during prenatal and postnatal life

The objective of this study was to investigate whether zinc interacts with taurine to influence the development of the electroretinogram. Virgin female Sprague-Dawley rats were bred overnight and assigned to 1 of 4 treatments in a 2 × 2 factorial design with two levels of zinc (50 μg/g through gestation and 50 μg/g after parturition; 15 μg/g through gestation and 7.5 μg/g after parturition) and two levels of taurine (2 or 0 μmol/g). Guanidinoethyl sulfonate (10 g/L), a structural analogue of taurine, was added to the drinking water of the animals receiving 0 μmol/g taurine. At postnatal day 23, male pups (n = 10) were weaned onto their respective diets. Dark-adapted electroretinograms were recorded as a function of stimulus intensity on 7 1/2–8 1/2-week-old anesthetized pups. Two-factor analysis of variance demonstrated no interaction between zinc and taurine for a- or b-wave amplitudes or latencies (P < 0.05). Zinc and taurine deficiencies each independently depressed electroretinogram a-wave and b-wave amplitudes but not latencies. The amplitude of the b-wave was plotted as a function of log stimulus intensity, and an iterative curve-fitting procedure was used to determine the maximum response, slope, and half-saturation constant. No interaction was noted. A significant treatment effect on maximum response was demonstrated for zinc (P = 0.0498) and taurine (P = 0.0014). No treatment effects were evident for the half-saturation constant or slope. These findings indicate that zinc and taurine deficiencies are not synergistic in their depressing effects on the electroretinogram in this model