A Cocoa Peptide Protects <i>Caenorhabditis elegans</i> from Oxidative Stress and β-Amyloid Peptide Toxicity

<div><p>Background</p><p>Cocoa and cocoa-based products contain different compounds with beneficial properties for human health. Polyphenols are the most frequently studied, and display antioxidant properties. Moreover, protein content is a very interesting source of antioxidant bioactive peptides, which can be used therapeutically for the prevention of age-related diseases.</p><p>Methodology/Principal Findings</p><p>A bioactive peptide, 13L (DNYDNSAGKWWVT), was obtained from a hydrolyzed cocoa by-product by chromatography. The <i>in vitro</i> inhibition of prolyl endopeptidase (PEP) was used as screening method to select the suitable fraction for peptide identification. Functional analysis of 13L peptide was achieved using the transgenic <i>Caenorhabditis elegans</i> strain CL4176 expressing the human Aβ_1–42 peptide as a pre-clinical <i>in vivo</i> model for Alzheimer's disease. Among the peptides isolated, peptide 13L (1 µg/mL) showed the highest antioxidant activity (<i>P</i>≤0.001) in the wild-type strain (N2). Furthermore, 13L produced a significant delay in body paralysis in strain CL4176, especially in the 24–47 h period after Aβ_1–42 peptide induction (<i>P</i>≤0.0001). This observation is in accordance with the reduction of Aβ deposits in CL4176 by western blot. Finally, transcriptomic analysis in wild-type nematodes treated with 13L revealed modulation of the proteosomal and synaptic functions as the main metabolic targets of the peptide.</p><p>Conclusions/Significance</p><p>These findings suggest that the cocoa 13L peptide has antioxidant activity and may reduce Aβ deposition in a <i>C. elegans</i> model of Alzheimer's disease; and therefore has a putative therapeutic potential for prevention of age-related diseases. Further studies in murine models and humans will be essential to analyze the effectiveness of the 13L peptide in higher animals.</p></div

Probiotic Strain <i>Bifidobacterium animalis</i> subsp. <i>lactis</i> CECT 8145 Reduces Fat Content and Modulates Lipid Metabolism and Antioxidant Response in <i>Caenorhabditis elegans</i>

Recently, microbial changes in the human gut have been proposed as a possible cause of obesity. Therefore, modulation of microbiota through probiotic supplements is of great interest to support obesity therapeutics. The present study examines the functional effect and metabolic targets of a bacterial strain, <i>Bifidobacterium animalis</i> subsp. <i>lactis</i> CECT 8145, selected from a screening in <i>Caenorhabditis elegans</i>. This strain significantly reduced total lipids (40.5% ± 2.4) and triglycerides (27.6% ± 0.5), exerting antioxidant effects in the nematode (30% ± 2.8 increase in survival vs control); activities were also preserved in a final food matrix (milk). Furthermore, transcriptomic and metabolomic analyses in nematodes fed with strain CECT 8145 revealed modulation of the energy and lipid metabolism, as well as the tryptophan metabolism (satiety), as the main metabolic targets of the probiotic. In conclusion, our study describes for the first time a new <i>B. animalis</i> subsp. <i>lactis</i> strain, CECT 8145, as a promising probiotic for obesity disorders. Furthermore, the data support future studies in obesity murine models

<i>Lactobacillus rhamnosus</i> CNCM I-3690 has an anti-inflammatory effect <i>in vitro</i>.

<p>A. Relative IL-8, production and I-κB-luciferase detection from the HT-29-bacteria interaction assays. Each bar represent the mean value of three replicate samples and and error bars depict corresponding standard deviation. Black bars: I-kB-luciferase, grey bars: IL-8. **p-value ≤0.05. B. Phenotypical analysis of monocyte-derived DCs co-cultured with HT-29-NF-κB-luciferase cells. Cells were incubated with LPS or LPS+bacteria. CNCM I-3690 down-regulated the expression of HLA-DR and CD86 surface markers. Results were expressed according to the following equation [(bacteria+LPS)-LPS]/LPS-basal*100. Experiments were performed with two different donors. C. Cytokine ratios (IL-8/IL-10, IL-6/IL-10, IL-12/IL-10 and TNF-α/IL-10) of DCs from donor 2 in co-culture with HT-29-NF-κB-luciferase cells and bacteria w or w/o LPS. Experiments were performed with two different donors (data shown for donor 2). *p-value≤0.05.</p

Screening for antioxidant bacteria in <i>C. elegans</i>.

<p>Survival upon a 5 mM H₂O₂ treatment for 5 h of <i>C. elegans</i> BA 17 strain fed for 3 days with <i>Bifidobacteria, Lactobacilli</i> and <i>Streptococci</i> strains in liquid medium. Worms were fed with <i>E. coli</i> OP 50, for larvae 1^st stage synchronization. E. coli OP 50 strain was inhibited with antibiotics prior of LAB and <i>Bifidobacteria</i> feeding. Protection by <i>E. coli</i> OP50 is fixed at 100%.</p

Protective effect of <i>Lactobacillus rhamnosus</i> CNCM I-3690 in a TNBS-induced murine model of colitis.

<p>A. Design of the interventional animal study. B. 3-day post colitis body-weight losses (% of initial) for healthy control mice, vehicle-TNBS-treated animal and either <i>L. rhamnosus</i> CNCM I-3690-fed TNBS-treated mice or Prednisolone-TNBS-treated mice. Data represent the mean+/−SEM, (number of mice n = 10); ***: P<0.001. C: Individual inflammatory macroscopic (Wallace score, left panel), and histological damage scores (Ameho score, right panel) in Control, probiotic and drug treatment conditions respectively, means +/− SD are indicated, **: P<0.01, ***: P<0.001. D: Representative histological colon sections from mice treated in various conditions after induction of TNBS colitis; May-Grünwald and Giemsa-stainings of 5 µm paraffin sections, original magnification×40.</p

Transcriptomic analysis of <i>C. elegans</i> N2 fed with CNCM I-3690 vs CNCM I-4317.

<p>Transcriptomic analysis of <i>C. elegans</i> N2 fed with CNCM I-3690 vs CNCM I-4317.</p

<i>Lactobacillus rhamnosus</i> CNCM I-3690 increases lifespan of <i>C. elegans</i> wild-type N2 strain and this effect is at least partially dependent of the DAF2-DAF16 pathway.

<p>A. <i>C. elegans</i> N2 wild-type strain was fed with <i>L. rhamnosus</i> CNCM I-3690, CNCM I-4317 and <i>E. coli</i> OP 50 and survival of worms was followed for 21 days. Mean lifespan, indicating the time in days where half of the worm population is still alive, is shown on the X-axis for the three situations. Curves comparisons are indicated (P-values) CNCM I-3690/CNCM I-4317 vs <i>E. coli</i> OP50 (red) and CNCM I-4317 vs <i>E. coli</i> OP50 (blue). NS: Non statistical difference. B, C, D. <i>C. elegans daf-16</i> (GR1307), <i>daf-2</i> (CB1370) and <i>skn-1</i> (LG333) loss-of-function mutant strains were fed with <i>L. </i><i>rhamnosus</i> CNCM I-3690 and <i>E. coli</i> OP 50. Survival of worms was followed up for 21 days. B. Curves comparisons vs <i>E. coli</i> OP50 are indicated in red (P-values).</p