Glucose-lowering effects of a synbiotic combination containing Pediococcus acidilactici in C. elegans and mice.

AIMS/HYPOTHESIS: Modulation of gut microbiota has emerged as a promising strategy to treat or prevent the development of different metabolic diseases, including type 2 diabetes and obesity. Previous data from our group suggest that the strain Pediococcus acidilactici CECT9879 (pA1c) could be an effective probiotic for regulating glucose metabolism. Hence, the objectives of this study were to verify the effectiveness of pA1c on glycaemic regulation in diet-induced obese mice and to evaluate whether the combination of pA1c with other normoglycaemic ingredients, such as chromium picolinate (PC) and oat ß-glucans (BGC), could increase the efficacy of this probiotic on the regulation of glucose and lipid metabolism. METHODS: Caenorhabditis elegans was used as a screening model to describe the potential synbiotic activities, together with the underlying mechanisms of action. In addition, 4-week-old male C57BL/6J mice were fed with a high-fat/high-sucrose diet (HFS) for 6 weeks to induce hyperglycaemia and obesity. Mice were then divided into eight groups (n=12 mice/group) according to dietary supplementation: control-diet group; HFS group; pA1c group (1010 colony-forming units/day); PC; BGC; pA1c+PC+BGC; pA1c+PC; and pA1c+BGC. Supplementations were maintained for 10 weeks. Fasting blood glucose was determined and an IPGTT was performed prior to euthanasia. Fat depots, liver and other organs were weighed, and serum biochemical variables were analysed. Gene expression analyses were conducted by real-time quantitative PCR. Sequencing of the V3-V4 region of the 16S rRNA gene from faecal samples of each group was performed, and differential abundance for family, genera and species was analysed by ALDEx2R package. RESULTS: Supplementation with the synbiotic (pA1c+PC+BGC) counteracted the effect of the high glucose by modulating the insulin-IGF-1 signalling pathway in C. elegans, through the reversal of the glucose nuclear localisation of daf-16. In diet-induced obese mice, all groups supplemented with the probiotic significantly ameliorated glucose tolerance after an IPGTT, demonstrating the glycaemia-regulating effect of pA1c. Further, mice supplemented with pA1c+PC+BGC exhibited lower fasting blood glucose, a reduced proportion of visceral adiposity and a higher proportion of muscle tissue, together with an improvement in the brown adipose tissue in comparison with the HFS group. Besides, the effect of the HFS diet on steatosis and liver damage was normalised by the synbiotic. Gene expression analyses demonstrated that the synbiotic activity was mediated not only by modulation of the insulin-IGF-1 signalling pathway, through the overexpression of GLUT-1 and GLUT-4 mediators, but also by a decreased expression of proinflammatory cytokines such as monocyte chemotactic protein-1. 16S metagenomics demonstrated that the synbiotic combinations allowed an increase in the concentration of P. acidilactici, together with improvements in the intestinal microbiota such as a reduction in Prevotella and an increase in Akkermansia muciniphila. CONCLUSIONS/INTERPRETATION: Our data suggest that the combination of pA1c with PC and BGC could be a potential synbiotic for blood glucose regulation and may help to fight insulin resistance, diabetes and obesity.

Grifola frondosa (Maitake) Extract Reduces Fat Accumulation and Improves Health Span in C. elegans through the DAF-16/FOXO and SKN-1/NRF2 Signalling Pathways.

In recent years, food ingredients rich in bioactive compounds have emerged as candidates to prevent excess adiposity and other metabolic complications characteristic of obesity, such as low-grade inflammation and oxidative status. Among them, fungi have gained popularity for their high polysaccharide content and other bioactive components with beneficial activities. Here, we use the C. elegans model to investigate the potential activities of a Grifola frondosa extract (GE), together with the underlying mechanisms of action. Our study revealed that GE represents an important source of polysaccharides and phenolic compounds with in vitro antioxidant activity. Treatment with our GE extract, which was found to be nongenotoxic through a SOS/umu test, significantly reduced the fat content of C. elegans, decreased the production of intracellular ROS and aging-lipofuscin pigment, and increased the lifespan of nematodes. Gene expression and mutant analyses demonstrated that the in vivo anti-obesity and antioxidant activities of GE were mediated through the daf-2/daf-16 and skn-1/nrf-2 signalling pathways, respectively. Taken together, our results suggest that our GE extract could be considered a potential functional ingredient for the prevention of obesity-related disturbances.

Bioactive extracts from persimmon waste: influence of extraction conditions and ripeness.

In this work, a bioactive persimmon extract was produced from discarded fruits. A central composite design was used to evaluate the effect of different extraction parameters and ripeness stages of persimmon fruits on the total phenolic content and antioxidant activity of the resulting extracts. Significantly greater phenolic contents were obtained from immature persimmon (IP) fruits. The optimum IP extract with the conditions set by the experimental design was industrially up-scaled and its composition and functional properties were evaluated and compared with those obtained under lab-scale conditions. Both extracts contained significant protein (>20%) and phenolic contents (∼11-27 mg GA/g dry extract) and displayed significant antiviral activity against murine norovirus and hepatitis A virus. Moreover, the extract showed no toxicity and significantly reduced the fat content and the cellular ageing of Caenorhabditis elegans (C. elegans) without affecting the worm development. These effects were mediated by down-regulation of fat-7, suggesting an anti-lipogenic activity of this extract.

A combination of borage seed oil and quercetin reduces fat accumulation and improves insulin sensitivity in obese rats.

The metabolic properties of omega-6 fatty acid consumption are being increasingly accepted. We had previously observed that supplementation with a borage seed oil (BSO), as a source of linoleic (18:2n-6; LA) and gamma-linolenic (18:3n-6; GLA) acids, reduces body weight and visceral adiposity and improves insulin sensitivity in a diet-induced obesity model of Wistar rats. Here, it was investigated whether the anti-obesogenic properties of BSO could be maintained in a pre-obese model of rats, and if these effects are enhanced by a combination with low doses of quercetin, together with its potential role in the regulation of the adipocyte biology. The combination of BSO and quercetin during 8 weeks was able to ameliorate glucose intolerance and insulin resistance, and to improve liver steatosis. Although no effects were observed on body weight, animals supplemented with this combination exhibited a lower proportion of visceral adiposity. In addition, in vitro differentiation of epididymal adipose-precursor cells of the BSO-treated animals exhibited a down-regulation of Fasn, Glut4, Pparg and Srebp1 genes, in comparison with the control group. Finally, in vitro evaluation of the components of BSO demonstrated that the anti-adipogenic activity of quercetin was significantly potentiated by the combination with both LA and GLA through the down-regulation of different adipogenesis-key genes in 3T3-L1 cells. All these data suggest that omega-6 fatty acids LA and GLA, and their natural sources such as BSO, could be combined with quercetin to potentiate their effects in the prevention of the excess of adiposity and the insulin resistance.

Low doses of cocoa extract supplementation ameliorate diet-induced obesity and insulin resistance in rats.

Cocoa polyphenols exhibit high antioxidant activity and have been proposed as a potential adjuvant for the treatment of metabolic disturbances. Here, we demonstrate that supplementation with low doses (14 and 140 mg per kg per rat) of a complete cocoa extract induces metabolic benefits in a diet-induced obesity (DIO) model of Wistar rats. After 10 weeks, cocoa extract-supplemented animals exhibited significantly lower body weight gain and food efficiency, with no differences in energy intake. Cocoa significantly reduced visceral (epididymal and retroperitoneal) and subcutaneous fat accumulation accompanied by a significant reduction in the adipocyte size, which was mediated by downregulation of the adipocyte-specific genes Cebpa, Fasn and Adipoq. Additionally, cocoa extract supplementation reduced the triacylglycerol/high density lipoprotein (TAG/HDL) ratio, decreased hepatic triglyceride accumulation, improved insulin sensitivity by reducing HOMA-IR, and significantly ameliorated glucose tolerance after an intraperitoneal glucose tolerance test. Finally, no adverse effect was observed in an in vivo toxicity evaluation of our cocoa extract at doses up to 500 mg kg-1 day-1. Our data demonstrate that low doses of cocoa extract supplementation (14 and 140 mg kg-1 day-1) are safe and sufficient to counteract obesity and type-2 diabetes in rats and provide new insights into the potential application of cocoa supplements in the management of the metabolic syndrome.

Borago officinalis seed oil (BSO), a natural source of omega-6 fatty acids, attenuates fat accumulation by activating peroxisomal beta-oxidation both in C. elegans and in diet-induced obese rats.

Obesity is a medical condition with increasing prevalence, characterized by an accumulation of excess fat that could be improved using some bioactive compounds. However, many of these compounds with in vitro activity fail to respond in vivo, probably due to the sophistication of the physiological energy regulatory networks. In this context, C. elegans has emerged as a plausible model for the identification and characterization of the effect of such compounds on fat storage in a complete organism. However, the results obtained in such a simple model are not easily extrapolated to more complex organisms such as mammals, which hinders its application in the short term. Therefore, it is necessary to obtain new experimental data about the evolutionary conservation of the mechanisms of fat loss between worms and mammals. Previously, we found that some omega-6 fatty acids promote fat loss in C. elegans by up-regulation of peroxisomal fatty acid ß-oxidation in an omega-3 independent manner. In this work, we prove that the omega-6 fatty acids' effects on worms are also seen when they are supplemented with a natural omega-6 source (borage seed oil, BSO). Additionally, we explore the anti-obesity effects of two doses of BSO in a diet-induced obesity rat model, validating the up-regulation of peroxisomal fatty acid ß-oxidation. The supplementation with BSO significantly reduces body weight gain and energy efficiency and prevents white adipose tissue accumulation without affecting food intake. Moreover, BSO also increases serum HDL-cholesterol levels, improves insulin resistance and promotes the down-regulation of Cebpa, an adipogenesis-related gene. Therefore, we conclude that the effects of omega-6 fatty acids are highly conserved between worms and obesity-induced mammals, so these compounds could be considered to treat or prevent obesity-related disorders.