Exploring the Impact of French Raw-Milk Cheeses on Oxidative Process Using Caenorhabditis elegans and Human Leukocyte Models.

Fermented foods, including cheeses, have garnered increased interest in recent years for their potential health benefits. This study explores the biological properties of eight French raw-milk cheeses-goat cheese, Saint-Nectaire, Cantal, Bleu d'Auvergne, Roquefort, Comté, Brie de Meaux, and Epoisses-on oxidative processes using both in vivo (Caenorhabditis elegans) and in vitro (human leukocytes) models. A cheese fractionation protocol was adapted to study four fractions for each cheese: a freeze-dried fraction (FDC) corresponding to whole cheese, an apolar (ApE), and two polar extracts (W40 and W70). We showed that all cheese fractions significantly improved Caenorhabditis elegans (C. elegans) survival rates when exposed to oxidative conditions by up to five times compared to the control, regardless of the fractionation protocol and the cheese type. They were also all able to reduce the in vivo accumulation of reactive oxygen species (ROS) by up to 70% under oxidative conditions, thereby safeguarding C. elegans from oxidative damage. These beneficial effects were explained by a reduction in ROS production up to 50% in vitro in human leukocytes and overexpression of antioxidant factor-encoding genes (daf-16, skn-1, ctl-2, and sod-3) in C. elegans.

A Mechanistic Study of the Antiaging Effect of Raw-Milk Cheese Extracts.

Many studies have highlighted the relationship between food and health status, with the aim of improving both disease prevention and life expectancy. Among the different food groups, fermented foods a have huge microbial biodiversity, making them an interesting source of metabolites that could exhibit health benefits. Our previous study highlighted the capacity of raw goat milk cheese, and some of the extracts recovered by the means of chemical fractionation, to increase the longevity of the nematode Caenorhabditis elegans. In this article, we pursued the investigation with a view toward understanding the biological mechanisms involved in this phenomenon. Using mutant nematode strains, we evaluated the implication of the insulin-like DAF-2/DAF-16 and the p38 MAPK pathways in the phenomenon of increased longevity and oxidative-stress resistance mechanisms. Our results demonstrated that freeze-dried raw goat milk cheese, and its extracts, induced the activation of the DAF-2/DAF-16 pathway, increasing longevity. Concerning oxidative-stress resistance, all the extracts increased the survival of the worms, but no evidence of the implication of both of the pathways was highlighted, except for the cheese-lipid extract that did seem to require both pathways to improve the survival rate. Simultaneously, the cheese-lipid extract and the dried extract W70, obtained with water, were able to reduce the reactive oxygen species (ROS) production in human leukocytes. This result is in good correlation with the results obtained with the nematode.

Development of an innovative methodology combining chemical fractionation and in vivo analysis to investigate the biological properties of cheese.

With the ever-increasing human lifespan, age-related affections have become a public health issue. The health sector is looking for new bioactive compounds to respond to this demand. The unexplored microbial biodiversity and its metabolites represent a major source of innovative bioactive molecules with health potential. Fermented foods, such as raw-milk cheese, have already been investigated for their rich microbial environment, especially for their organoleptic qualities. But studies remain limited regarding their effects on health and few metabolites of microbial origin have been identified. An efficient methodology was developed in this study to investigate the biological effect of raw-milk cheese, combining a chemical fractionation, to isolate the most metabolites from the cheese matrix, and an in vivo biological test using Caenorhabditis elegans. C. elegans was brought into contact with cheese extracts, obtained by means of chemical fractionation, and with freeze-dried whole cheese by supplementing the nematode growth medium. A longevity assay was performed to evaluate the effects of the extracts on the worms. Our results demonstrate the feasibility of the method developed to bring the worms into contact of the cheese extracts. The evaluation of the effects of the extracts on the longevity was possible. Some extracts showed a beneficial effect as extract W70 for example, obtained with water, which increases the mean lifespan by 16% and extends the longevity by 73% (p < 0.0001).

Pomegranate Peel Extract Prevents Bone Loss in a Preclinical Model of Osteoporosis and Stimulates Osteoblastic Differentiation in Vitro.

The nutritional benefits of pomegranate have attracted great scientific interest. The pomegranate, including the pomegranate peel, has been used worldwide for many years as a fruit with medicinal activity, mostly antioxidant properties. Among chronic diseases, osteoporosis, which is associated with bone remodelling impairment leading to progressive bone loss, could eventually benefit from antioxidant compounds because of the involvement of oxidative stress in the pathogenesis of osteopenia. In this study, with in vivo and ex vivo experiments, we investigated whether the consumption of pomegranate peel extract (PGPE) could limit the process of osteopenia. We demonstrated that in ovariectomized (OVX) C57BL/6J mice, PGPE consumption was able to significantly prevent the decrease in bone mineral density (-31.9%; p < 0.001 vs. OVX mice) and bone microarchitecture impairment. Moreover, the exposure of RAW264.7 cells to serum harvested from mice that had been given a PGPE-enriched diet elicited reduced osteoclast differentiation and bone resorption, as shown by the inhibition of the major osteoclast markers. In addition, PGPE appeared to substantially stimulate osteoblastic MC3T3-E1 alkaline phosphatase (ALP) activity at day 7, mineralization at day 21 and the transcription level of osteogenic markers. PGPE may be effective in preventing the bone loss associated with ovariectomy in mice, and offers a promising alternative for the nutritional management of this disease.

Autumnalamide, a prenylated cyclic peptide from the cyanobacterium Phormidium autumnale, acts on SH-SY5Y cells at the mitochondrial level.

Filamentous cyanobacteria of the genus Phormidium have been rarely studied for their chemical diversity. For the first time, the cultivable Phormidium autumnale was shown to produce a prenylated cyclic peptide named autumnalamide (1). The structure of this peptide was fully determined after a deep exploration of the spectroscopic data, including NMR and HRMS. Interestingly, a prenyl moiety was located on the guanidine end of the arginine amino acid. The absolute configurations of most amino acids were assessed using enantioselective GC/MS analysis, with (13)C NMR modeling being used for the determination of d-arginine and d-proline. The effects of 1 on sodium and calcium fluxes were studied in SH-SY5Y and hNav 1.6 HEK cells. When the Ca(2+) influx was stimulated by thapsigargin, strong inhibition was observed in the presence of 1. As a consequence, this compound may act by disrupting the normal calcium uptake of this organelle, inducing the opening of the mitochondrial permeability transition pore, which results in the indirect blockade of store-operated channels.

Pomegranate and its derivatives can improve bone health through decreased inflammation and oxidative stress in an animal model of postmenopausal osteoporosis.

PURPOSE: Recently, nutritional and pharmaceutical benefits of pomegranate (PG) have raised a growing scientific interest. Since PG is endowed with anti-inflammatory and antioxidant activities, we hypothesized that it may have beneficial effects on osteoporosis. METHODS: We used ovariectomized (OVX) mice as a well-described model of postmenopausal osteoporosis to study the influence of PG consumption on bone health. Mice were divided into five groups as following: two control groups sham-operated and ovariectomized (OVX CT) mice fed a standard diet, versus three treated groups OVX mice given a modified diet from the AIN-93G diet, containing 5.7% of PG lyophilized mashed totum (OVX PGt), or 9.6% of PG fresh juice (OVX PGj) or 2.9% of PG lyophilized mashed peel (OVX PGp). RESULTS: As expected, ovariectomy was associated with a decreased femoral bone mineral density (BMD) and impaired bone micro-architecture parameters. Consumption of PGj, PGp, or PGt induced bone-sparing effects in those OVX mice, both on femoral BMD and bone micro-architecture parameters. In addition, PG (whatever the part) up-regulated osteoblast activity and decreased the expression of osteoclast markers, when compared to what was observed in OVX CT animals. Consistent with the data related to bone parameters, PG consumption elicited a lower expression of pro-inflammatory makers and of enzymes involved in ROS generation, whereas the expression of anti-inflammatory markers and anti-oxidant actors was enhanced. CONCLUSION: These results indicate that all PG parts are effective in preventing the development of bone loss induced by ovariectomy in mice. Such an effect could be partially explained by an improved inflammatory and oxidative status.

Bioprospecting marine plankton.

The ocean dominates the surface of our planet and plays a major role in regulating the biosphere. For example, the microscopic photosynthetic organisms living within provide 50% of the oxygen we breathe, and much of our food and mineral resources are extracted from the ocean. In a time of ecological crisis and major changes in our society, it is essential to turn our attention towards the sea to find additional solutions for a sustainable future. Remarkably, while we are overexploiting many marine resources, particularly the fisheries, the planktonic compartment composed of zooplankton, phytoplankton, bacteria and viruses, represents 95% of marine biomass and yet the extent of its diversity remains largely unknown and underexploited. Consequently, the potential of plankton as a bioresource for humanity is largely untapped. Due to their diverse evolutionary backgrounds, planktonic organisms offer immense opportunities: new resources for medicine, cosmetics and food, renewable energy, and long-term solutions to mitigate climate change. Research programs aiming to exploit culture collections of marine micro-organisms as well as to prospect the huge resources of marine planktonic biodiversity in the oceans are now underway, and several bioactive extracts and purified compounds have already been identified. This review will survey and assess the current state-of-the-art and will propose methodologies to better exploit the potential of marine plankton for drug discovery and for dermocosmetics.

Pomegranate seed oil prevents bone loss in a mice model of osteoporosis, through osteoblastic stimulation, osteoclastic inhibition and decreased inflammatory status.

In the current context of longer life expectancy, the prevalence of osteoporosis is increasingly important. This is why development of new strategies of prevention is highly suitable. Pomegranate seed oil (PSO) and its major component, punicic acid (a conjugated linolenic acid), have potent anti-inflammatory and anti-oxidative properties both in vitro and in vivo, two processes strongly involved in osteoporosis establishment. In this study, we demonstrated that PSO consumption (5% of the diet) improved significantly bone mineral density (240.24±11.85 vs. 203.04±34.19 mg/cm(3)) and prevented trabecular microarchitecture impairment in ovariectomized (OVX) mice C57BL/6J, compared to OVX control animals. Those findings are associated with transcriptional changes in bone tissue, suggesting involvement of both osteoclastogenesis inhibition and osteoblastogenesis improvement. In addition, thanks to an ex vivo experiment, we provided evidence that serum from mice fed PSO (5% by gavage) had the ability to significantly down-regulate the expression of specific osteoclast differentiation markers and RANK-RANKL downstream signaling targets in osteoclast-like cells (RAW264.7) (RANK: negative 0.49-fold vs. control conditions). Moreover, in osteoblast-like cells (MC3T3-E1), it elicited significant increase in alkaline phosphatase activity (+159% at day 7), matrix mineralization (+271% on day 21) and transcriptional levels of major osteoblast lineage markers involving the Wnt/ß-catenin signaling pathways. Our data also reveal that PSO inhibited pro-inflammatory factors expression while stimulating anti-inflammatory ones. These results demonstrate that PSO is highly relevant regarding osteoporosis. Indeed, it offers promising alternatives in the design of new strategies in nutritional management of age-related bone complications.

Balibalosides, an original family of glucosylated sesterterpenes produced by the Mediterranean sponge Oscarella balibaloi.

The chemical investigation of the recently described Mediterranean Homoscleromorpha sponge Oscarella balibaloi revealed an original family of five closely related glucosylated sesterterpenes 1-4, named balibalosides. Their structure elucidation was mainly inferred from NMR and HRMS data analyses. Balibalosides differ by the pattern of acetyl substitutions on the three sugar residues linked to the same aglycone sesterterpenoid core. From a biosynthetic perspective, these compounds may represent intermediates in the pathways leading to more complex sesterterpenes frequently found in Dictyoceratida, a sponge Order belonging to Demospongiae, a clade which is phylogenetically distinct from the Homoscleromorpha. While steroid and triterpenoid saponins were already well known from marine sponges, balibalosides are the first examples of glycosilated sesterterpenes.

Novel liquid chromatography-electrospray ionization mass spectrometry method for the quantification in human urine of microbial aromatic acid metabolites derived from dietary polyphenols.

An HPLC-ESI-MS-MS method was developed to quantify in human urine fourteen aromatic acids known as metabolites of dietary polyphenols. These metabolites were determined simultaneously in a single 20-min chromatographic analysis with multiple reaction monitoring detection. The inter- and intra-day precisions, calculated from quality control samples were 8.8 and 5.3%, respectively, and the mean accuracy was 2.3%. The method was tested on urine samples collected from one healthy volunteer who consumed a polyphenol-rich diet for 3 days. Increased levels of several aromatic acid metabolites were observed, demonstrating that the method can be used to detect changes in the excretion of microbial metabolites induced by the consumption of polyphenol-containing foods in humans.

Chocolate intake increases urinary excretion of polyphenol-derived phenolic acids in healthy human subjects.

BACKGROUND: Proanthocyanidins, the most abundant polyphenols in chocolate, are not depolymerized in the stomach and reach the small intestine intact, where they are hardly absorbed because of their high molecular weight. In vitro and in vivo studies using pure compounds as substrates suggest that proanthocyanidins and the related catechin monomers may be degraded into more bioavailable low-molecular-weight phenolic acids by the microflora in the colon. OBJECTIVE: The aim of the study was to estimate the amounts of phenolic acids formed by the microflora and excreted in the urine of human subjects after consumption of polyphenol-rich chocolate. DESIGN: After consumption of a polyphenol-free diet for 2 d and a subsequent overnight fast, 11 healthy subjects (7 men and 4 women) consumed 80 g chocolate containing 439 mg proanthocyanidins and 147 mg catechin monomers. All urine was collected during the 24 h before chocolate consumption and at 3, 6, 9, 24, and 48 h after chocolate consumption. Aromatic acids were identified in urine by gas chromatography-mass spectrometry and were quantified by HPLC-electrospray ionization tandem mass spectrometry. RESULTS: Chocolate intake increased the urinary excretion of the 6 following phenolic acids: m-hydroxyphenylpropionic acid, ferulic acid, 3,4-dihydroxyphenylacetic acid, m-hydroxyphenylacetic acid, vanillic acid, and m-hydroxybenzoic acid. CONCLUSION: The antioxidant and biological effects of chocolate may be explained not solely by the established absorption of catechin monomers but also by the absorption of microbial phenolic acid metabolites.

Cocoa procyanidins are stable during gastric transit in humans.

BACKGROUND: Polyphenolic procyanidins are abundant flavonoid polymers in Western diets. In vitro biological activity has been reported for these compounds, but activity in vivo depends on the amount and chemical nature of the flavonoids reaching the gastrointestinal tract. Degradation of procyanidins under simulated gastric conditions at pH 2.0 has been reported in vitro. OBJECTIVE: The objective was to examine whether depolymerization of procyanidins occurs in the stomach of human subjects in vivo. DESIGN: After an overnight fast, 6 healthy subjects (3 men and 3 women) consumed 500 mL of a cocoa beverage containing 733 mg procyanidin polymers and 351 mg structurally related flavanol monomers. With the use of a nasogastric tube, stomach contents were collected every 10 min after beverage ingestion until the stomach was emptied. Flavanols and procyanidins (up to pentamers) were quantified by normal and reversed-phase HPLC. RESULTS: In all subjects, gastric transit lasted approximately 50-60 min. No change in the HPLC profile of procyanidins was observed during this period, showing that procyanidins were remarkably stable in the stomach environment. CONCLUSION: The results suggest that most ingested procyanidins reach the small intestine intact and are available for absorption or metabolism.

Procyanidins are not bioavailable in rats fed a single meal containing a grapeseed extract or the procyanidin dimer B3.

Flavanols are the most abundant flavonoids in the human diet where they exist as monomers, oligomers and polymers. In the present study, catechin, the procyanidin dimer B3 and a grape-seed extract containing catechin, epicatechin and a mixture of procyanidins were fed to rats in a single meal. After the meals, catechin and epicatechin were present in conjugated forms in both plasma and urine. In contrast, no procyanidins or conjugates were detected in the plasma or urine of any rats. Procyanidins were not cleaved into bioavailable monomers and had no significant effects on the plasma levels or urinary excretion of the monomers when supplied together in the grapeseed extract. We conclude that the nutritional effects of dietary procyanidins are unlikely to be due to procyanidins themselves or monomeric metabolites with the intact flavonoid-ring structure, as they do not exist at detectable concentrations in vivo. Future research should focus on other procyanidin metabolites such as phenolic acids and on the effects of the unabsorbed oligomers and polymers on the human gastrointestinal tract.