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Nutrition 32 (2016) 716719

Contents lists available at ScienceDirect

Nutrition
journal homepage: www.nutritionjrnl.com

Brief report

Benecial effects of Bidobacterium lactis on lipid prole and


cytokines in patients with metabolic syndrome: A randomized
trial. Effects of probiotics on metabolic syndrome
Luciana Jesus Bernini M.Sc. a, Andrea Name Colado Sima ~o Ph.D. b,
Daniela Frizon Aleri M.Sc. , Marcell Alysson Batisti Lozovoy Ph.D. b,
b

Naiara Lourenco Mari M.Sc. b, Cnthia Hoch Batista de Souza Ph.D. a,


Isaas Dichi M.D., Ph.D. c, Giselle Nobre Costa Ph.D. a, *
a
Program of Master in Dairy Science and Technology, University of North ParanadUnopar, Londrina, Parana, Brazil
b
Department of Pathology, Clinical Analysis and Toxicology, University of Londrina, Londrina, Parana, Brazil
c
Department of Internal Medicine, University of Londrina, Londrina, Parana, Brazil

a r t i c l e i n f o a b s t r a c t

Article history: Objective: Human studies have shown the benecial effects of probiotic microorganisms on the
Received 25 July 2015 parameters of metabolic syndrome (MetS) and other cardiovascular risks, but to our knowledge the
Accepted 2 November 2015 effect of Bidobacterium lactis has not yet been reported. The aim of this study was to evaluate the
effect of consumption of milk containing the probiotic B. lactis HN019 on the classical parameters of
Keywords: MetS and other related cardiovascular risk factors.
Metabolic syndrome
Methods: Fifty-one patients with MetS were selected and divided into a control group (n 25) and
Probiotic
a probiotic group (n 26). The probiotic group consumed fermented milk with probiotics over the
Bidobacterium lactis
Cytokines course of 45 d. The effects of B. lactis on lipid prole, glucose metabolism, and proinammatory
Cardiovascular risk factors cytokines (tumor necrosis factor-a and interleukin-6) were assessed in blood samples of the in-
dividuals at the baseline and after 45 d.
Results: Daily ingestion of 80 mL fermented milk with 2.72  1010 colony-forming units of B. lactis
HN019 showed signicant reduction in body mass index (P 0.017), total cholesterol (P 0.009),
and low-density lipoprotein (P 0.008) compared with baseline and control group values.
Furthermore, a signicant decrease in tumor necrosis factor-a (P 0.033) and interleukin-6
(P 0.044) proinammatory cytokines was observed.
Conclusion: These data showed potential effects of B. lactis HN019 in reducing obesity, blood lipids,
and some inammatory markers, which may reduce cardiovascular risk in patients with MetS.
2016 Elsevier Inc. All rights reserved.

Introduction

Metabolic syndrome (MetS) is a set of disorders, whose risk


factors are atherogenic dyslipidemia (hypertriacylglycerols, high
LJB was responsible for recruiting the patients, interpretation of the results, levels of apolipoprotein B, small, dense particles of low-density
and writing the manuscript. CHBS was responsible for writing of the manuscript. lipoprotein cholesterol, and low high-density lipoprotein
DFA and MABL were responsible for recruiting the patients and the laboratory
cholesterol levels), hypertension, hyperglycemia, and a proin-
analysis. ANCS and ID were responsible for interpretation of the results and
writing the manuscript. GNC was responsible for the original concept of the ammatory state [1].
study, the study design, interpretation of the results, and writing of the Recent studies have associated the development of obesity in
manuscript. All authors read and approved the nal manuscript. The authors humans to the relative proportions of some major phyla of bac-
acknowledge the nancial support of the Development National Foundation of
teria of the intestinal microbiota, such as Bacteroidetes, Firmi-
Private Universities of Brazil.
* Corresponding author. Tel.: 55 433 371 7723; fax: 55 433 371 7834. cutes, and Actinobacteria, suggesting that the metabolic activity
E-mail address: gcnobre@gmail.com (G. N. Costa). of these intestinal microbiota facilitates the extraction and/or

http://dx.doi.org/10.1016/j.nut.2015.11.001
0899-9007/ 2016 Elsevier Inc. All rights reserved.
L. J. Bernini et al. / Nutrition 32 (2016) 716719 717

storage of calories ingested [2,3]. Moreover, imbalance in this Anthropometric measurements and biochemical parameters were assessed
ecosystem can lead not only to obesity, as well as the develop- at the beginning of the study (T0) and after 45 d (T45). Waist circumference (WC),
body weight, and height were measured, and body mass index (BMI) was
ment of insulin resistance [46]. calculated (kg/m2). Three blood pressure measurements taken at 1-min intervals
In this context, probiotic microorganisms have demonstrated after the participant had been seated were recorded. The mean of these mea-
health-improving effects on the hosts. The Firmicutes and Acti- surements was used.
nobacteria include the Lactobacilli and Bidobacteria, respec- After fasting for 12 h, serum or plasma samples were obtained and patients
underwent the following laboratory blood analysis: glucose, total cholesterol
tively, whose genera are the most used and often are related to
(TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein
these benecial probiotic effects [7,8]. cholesterol (LDL-C), and triacylglycerols (TGs). Levels were evaluated by a
Although some studies have reported the benecial effects of biochemical auto-analyzer (Dimension Dade AR Dade Behring, Deereld, IL,
species of Lactobacilli in patients with MetS, we are not aware of USA), using Dade Behring kits; plasma insulin level was determined by chem-
any report on the effects of Bidobacterium lactis in this condition iluminescence microparticle immunoassay (Architect, Abbott Laboratory, Abbott
Park, IL, USA). Homeostasis model assessment insulin resistance (HOMA-IR) was
and it is important to emphasize that the effects of probiotics are used as a surrogate measure of insulin sensitivity [11]. Tumor necrosis factor
highly strain-dependent [9]. (TNF)-a levels were measured by a sandwich enzyme-linked immunosorbent
Therefore, the aim of this study was to evaluate the inuence assay (ELISA) using a commercial immunoassay ELISA (Ready-Set Go Set, e-
of fermented milk with B. lactis HN019 on lipid prole, glucose Bioscience, San Diego, CA, USA).
The sample size was calculated to obtain a statistically signicant result when
metabolism, and cytokines in patients with MetS.
changes of the parameters evaluated were at 10% level. This study was per-
formed to obtain 80% statistic power. Mann-Whitney test was performed to
compare differences between parameters of groups at baseline and differences
Participants and methods across treatment groups (intergroup changes). Wilcoxon matched pairs test was
performed to verify changes from baseline (intragroup changes). Data are pre-
The study was conducted according to the guidelines in the Declaration of sented as median (25%75%). A statistical analysis program (Graph Pad Prism
Helsinki and approved by the Ethical Committee involving humans of the Uni- version 4.0) was used for evaluations. Signicance was set at P < 0.05.
versity of Londrina, Parana, Brazil, protocol number 185/2013. Patients were
recruited between March and May 2013 in the city of Jataizinho, Parana State,
Brazil. Inclusion criteria were being between ages 18 and 60 y, and meeting MetS
diagnosis criteria, according to the National Cholesterol Education Program, Results
Adult Treatment Panel III (NCEP/ATP III) [1]. Fifty-one patients with MetS were
randomized by drawing and began to participate in the study Exclusion criteria During the proposed study period, only one of the partici-
involved patients with chronic diseases or individuals with use of drugs that
pants failed to join the group, claiming typical effects of lactose
interfere in the lipids and/or glycemic prole, antibiotics, and anti-inammatory
agents, vitamins and foods with probiotic microorganisms, besides lactose- intolerance, although the recruitment survey excluded in-
intolerant individuals. The patients were instructed to maintain their usual di- dividuals with this characteristic.
ets, level of physical activity or other lifestyle factors throughout the intervention There were no differences between control and probiotic
period. groups with respect to age, sex, medication, and antihyperten-
Fifty-one patients with MetS agreed to participate in the study by written
consent. The participants were divided into a control group or untreated (n 25)
sive parameters at baseline.
and a probiotic group (n 26). The diet of the probiotic groups included the daily No signicant differences were observed intra- or intergroup
ingestion of 80 mL of the probiotic milk containing on average 3.4  108 colony- for WC, systolic and diastolic blood pressure levels, TGs, HDL-C,
forming units (CFU)/mL of B. animalis ssp. lactis ssp. nov. HN019. glucose, insulin, and the HOMA (Table 1). The probiotic group
The probiotic milk was produced and packaged aseptically at the Laboratory
showed signicant decreases in BMI (P 0.017), TC (P 0.009),
of Microbiology of the University of North Parana, according the current Brazilian
legislation. The probiotic milk ready for consumption showed high probiotics and LDL-C (P 0.008) compared with baseline values. Regarding
counts with 3.4  108 CFU/mL, 0.38% acidity, pH 5.15, 2.80% protein, 7.54% car- intergroup changes, there was a signicant decrease (P < 0.05) in
bohydrates, 0.0% fat, 12.08% total solids, and 0.48% ash that results in 33.09 cal- BMI, TC, and LDL-C veried after 45 d in the probiotic group
ories per bottles. Additionally, the nal product contained 0.01% (v/v) orange, compared with the control group. The control group had a sig-
vanilla, or both avors. Pathogens were not detected, as required by legislation
[10], thus the product was characterized as suitable for consumption and seven
nicant increase in BMI and HDL-C, and a signicant decrease in
bottles of 80 mL were distributed weekly to each volunteer until they reached TC and LDL-C compared with the baseline values of the probiotic
45 d of intake. Patients were asked to drink one bottle every morning. group (Table 1; P < 0.05).

Table 1
Anthropometric, blood pressure, and laboratory parameters in control and probiotic groups of patients with metabolic syndrome

Parameters Control (n 25) Probiotic (n 26)

T0 T45 P value T0 T45 P value


BMI (kg/m2)*,y 35.8 (33.444.5) 35.5 (32.240.7) 0.966 30.8 (27.233.7) 29.5 (25.933.3) 0.017
WC (cm) 107 (101128) 109 (99.5124.5) 0.084 105 (98.5112.5) 107 (98.5115) 0.208
SBP (mm Hg) 125 (112.8139) 121 (110131) 0.135 130 (120150) 140 (127.5150) 0.974
DBP (mm Hg) 77.5 (7090) 72.5 (63.582.3) 0.191 80 (8097.5) 90 (80100) 0.842
TG (mg/dL) 199 (103.3251.3) 168.5 (113.5221.3) 0.597 163 (128314.5) 174 (124296) 0.808
TC (mg/dL)*,y 199 (166.5208.5) 205 (173220.5) 0.506 209 (183.8249.8) 194 (168251) 0.009
HDL (mg/dL)* 42 (3547.5) 39 (35.547) 0.617 40.5 (3049.5) 38.5 (31.346) 0.820
LDL (mg/dL)*,y 117 (83142) 115 (95146) 0.820 128.5 (103152.8) 111 (93.6134.6) 0.008
Glucose (mg/dL) 98 (94115.5) 99 (91113.0) 1.000 94 (82.5115.5) 97 (88.3124.3) 0.424
Insulin (uU/mL) 13.90 (9.9317.78) 13.65 (9.4621.78) 0.107 13.4 (8.1817.3) 15.3 (10.217.4) 0.171
HOMA 3.63 (2.094.44) 3.23 (2.335.55) 0.119 3.43 (2.294.77) 4.09 (3.375.87) 0.196

BMI, body mass index; DBP, diastolic blood pressure; HDL, high-density lipoprotein; HOMA, homeostatic model assessment; LDL, low-density lipoprotein; SBP, systolic
blood pressure; TC, total cholesterol; TG, triacylglycerol; WC, waist circumference
MannWhitney test was performed to compare differences between the baseline values and across treatment groups (intergroup changes). Wilcoxon matched pairs test
was performed to verify changes from baseline (intragroup changes). Data are expressed as median (25%75%)
* Differences of baseline groups, P < 0.05.
y
Percentage of change from baseline (P < 0.05).
718 L. J. Bernini et al. / Nutrition 32 (2016) 716719

The cytokines TNF-a (P < 0.05) and interleukin (IL)-6 coprecipitation of cholesterol together with the deconjugated
(P < 0.05) decreased after 45 d of probiotic intake compared with bile acids, which are less effective in the solubilization and ab-
baseline values. Differences between treatment groups veried a sorption of lipids from diet [18]. Additionally, the use of pro-
statistically signicant decrease in TNF-a and IL-6 (P < 0.05) biotics has been associated with increased levels of short-chain
when the probiotic group was compared with the control group fatty acids (SCFA) such as propionate, which improve lipid
(Fig. 1A, B). metabolism by acting directly on the liver by inhibiting
hydroxymethylglutaryl coenzyme A reductase, a rate-limiting
Discussion step of the cholesterol synthesis pathway [19].
Studies evaluating probiotic intake and markers of inam-
The main ndings of the present study are related to the mation are controversial. Some studies with Lactobacilli showed
benecial effects of B. lactis on BMI, lipid prole, and cytokine a decrease in inammatory markers [20], whereas others did not
levels in patients with MetS. A recent report demonstrated that report such ndings [21,22]. In a recent study, the effects of a
fermented milk with Lactobacillus plantarum decreased TC, multispecies probiotic supplement containing L. acidophilus, L.
glucose, and IL-6 levels. Meanwhile, there was a trend for de- casei, L. rhamnosus, L. bulgaricus, B. breve, B. longum, and Strep-
creases in LDL-C, although there were no signicant differences tococcus thermophilus on high-sensitivity C-reactive protein
in BMI [12]. (hs-CRP) in patients with diabetes have been reported. The au-
Several studies have associated phyla and species of intestinal thors found a signicant decrease in serum hs-CRP levels [20].
microbiota to obesity or weight loss in humans and high levels of Results from another study demonstrated the effects of L. plan-
bidobacteria in the gastrointestinal tract are related to weight tarum on the classical parameters related to the MetS and other
loss [46]. Although the fecal microbiota was not characterized parameters related to cardiovascular risk [12]. The authors found
in the present study, the high daily intake of B. lactis as a signicant decrease in IL-6 between the control (P 0.032) and
allochthonous microbiota can positively affect the microbial the intervention (P 0.001) groups. Therefore, it is difcult with
balance [13] leading to an improvement of the lipid metabolism these sparse data to support a strain specicity related to in-
and weight loss. The mechanisms involved in body weight ammatory status. The metabolic activity of gut microbiota is
reduction are not clear, but studies have pointed to the reduction associated with the inammatory responses originated from
of adipocyte size, inhibition of adipogenesis, and suppression of high-fat diets [14]. Bacterial lipopolysaccharides derived from
energy intake [9,14]. intestinal microorganisms can act in this process triggering in-
The reduction of TC and LDL-C in the probiotic group cor- ammatory response and consequently may predispose to the
roborates others reports [9,15]. A meta-analysis of randomized occurrence of MetS.
controlled trials to evaluate the effects of probiotics observed a In this study, we observed the reduction on levels of TNF-a
decrease LDL and TC concentrations in individuals with normal, and IL-6, cytokines that have been widely studied and related to
borderline high, and high cholesterol levels [15]. The mecha- obesity. Fat accumulation caused by obesity provokes increased
nisms proposed for these effects are associated with the assim- production of these adipokines, which stimulate the generation
ilation of cholesterol during bacterial multiplication or of hepatic acute-phase proteins, leading to a proinammatory
cholesterol binding to the surface of the cell wall of these bac- condition associated with the development of obesity comor-
teria, preventing the absorption by the intestinal tract into the bidities as insulin resistance [23].
bloodstream [16,17]. There are mechanisms related to the ability In this study, the small number of participants (N 51) and
of some probiotic microorganisms to produce hydrolase that the differences at the baseline values in some parameters be-
deconjugate bile acids. These compounds, when deconjugated, tween the groups limited the interpretation of the results. For
are absorbed less efciently, resulting in increased excretion in example, decreases in TC, LDL-C, TNF-a, and IL-6 levels can be
the feces. Another hypothesis is the reduction of serum choles- related to reduction in BMI. Nevertheless, the percentage of
terol as a function of lower lipid absorption, due to the changes from baseline (intergroup differences) and intragroup

Fig. 1. Concentration of tumor necrosis factor (TNF)-a and interleukin (IL)-6 in the control (CONT) and probiotic (PROB) groups evaluated at the beginning of the study (T0)
and after 45 d (T45) of milk ingestion. MannWhitney test was performed to compare differences between the baseline values and across treatment groups (inter-group
changes). Wilcoxon matched pairs test was performed to verify changes from baseline (intragroup changes). Data are expressed as median (25%75%). *Differences of baseline
groups, P < 0.05. yPercentage of change from baseline (P < 0.05).
L. J. Bernini et al. / Nutrition 32 (2016) 716719 719

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