Do Pomegranate Hydrolyzable Tannins and Their Derived Metabolites Provide Relief in Osteoarthritis? Findings from a Scoping Review.

Osteoarthritis (OA) is the most common form of arthritis affecting both the elderly and the middle-aged population. Although various therapeutics have been developed to arrest the structural deterioration of cartilage, the current treatments are limited to delay the progress of OA clinically. Therefore, it is pivotal to study new therapeutic agents for chondroprotection and the prevention of cartilage degeneration. Hydrolyzable tannin (HT)-containing foods aroused considerable interest in recent years for their relevant anti-inflammatory effects. The focus of this scoping review is to provide an overview of the evidence of the therapeutic potential of HTs and their metabolites in preventing or alleviating the course of OA. A broad search of PubMed and Scopus databases on this topic resulted in 156 articles. After the exclusion of reviews and not relevant records, 31 articles were retrieved. Although only some papers did not consider the biotransformation of HTs, most recent studies also have investigated the effect of HT metabolites. Further larger clinical trials, with an in-deep analysis of HT metabolization, are still needed to unravel the potential benefits of these compounds in OA, paving the way towards the development of a dietary strategy for the improvement of pro-inflammatory cytokine-induced chondrocyte dysfunctions and injuries.

Co-Administration of Propionate or Protocatechuic Acid Does Not Affect DHA-Specific Transcriptional Effects on Lipid Metabolism in Cultured Hepatic Cells.

Long-chain n-3 polyunsaturated fatty acids (n-3 LC-PUFAs) are collectively recognized triglyceride-lowering agents, and their preventive action is likely mediated by changes in gene expression. However, as most studies employ fish oil, which contains a mixture of n-3 LC-PUFAs, the docosahexaenoic acid (DHA)-specific transcriptional effects on lipid metabolism are still unclear. The aim of the present study was to further elucidate the DHA-induced transcriptional effects on lipid metabolism in the liver, and to investigate the effects of co-administration with other bioactive compounds having effects on lipid metabolism. To this purpose, HepG2 cells were treated for 6 or 24 h with DHA, the short-chain fatty acid propionate (PRO), and protocatechuic acid (PCA), the main human metabolite of cyanidin-glucosides. Following supplementation, we mapped the global transcriptional changes. PRO and PCA alone had a very slight effect on the transcriptome; on the contrary, supplementation of DHA highly repressed the steroid and fatty acid biosynthesis pathways, this transcriptional modulation being not affected by co-supplementation. Our results confirm that DHA effect on lipid metabolism are mediated at least in part by modulation of the expression of specific genes. PRO and PCA could contribute to counteracting dyslipidemia through other mechanisms.

Pomegranate juice to reduce fecal calprotectin levels in inflammatory bowel disease patients with a high risk of clinical relapse: Study protocol for a randomized controlled trial.

BACKGROUND: Inflammatory bowel disease (IBD) is a chronic condition characterized by recurrent episodes of intestinal inflammation and is thought to be related to an autoimmune reaction to genetic and environmental factors. Although evidence indicates that a polyphenolic-rich diet plays an important role in modulating aspects of chronic inflammation, few studies have focused on the effect of ellagitannin (ET)-rich food consumption on long-term remission maintenance in IBD patients with a high risk of clinical relapse. Therefore, we hypothesize that supplementation with a pomegranate juice, a naturally rich source of ETs, could significantly modulate the markers of mucosal and systemic inflammation relative to a control group receiving a placebo. METHODS/DESIGN: This double-blind, randomized controlled trial includes patients with IBD involving the colorectum who have been in stable therapy for at least the three previous months and have a high risk of clinical relapse. Participants are randomly allocated to one of two groups: active supplementation (125 mL of cv. Wonderful pomegranate juice) or placebo (125 mL) taken twice daily for 12 weeks. The primary outcome is changes in the fecal neutrophil-derived protein calprotectin, a surrogate marker of mucosal improvement, between the two groups from baseline to 12 weeks later. The secondary outcomes include transcriptomic changes in peripheral blood mononuclear cells and intestinal biopsies and changes in circulating inflammatory markers and trimethylamine-N-oxide levels. Pomegranate ET-derived metabolites are identified and quantified in plasma and urine samples. DISCUSSION: The results will provide information on the possible reduction of fecal calprotectin levels following the consumption of pomegranate juice. The findings will also show the in vivo metabolism of pomegranate ETs. Finally, the effect of 12-week pomegranate juice consumption on local and systemic inflammatory markers will be elucidated, which will likely provide additional insights into the maintenance of remission in IBD patients. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03000101 . Registered on 21 December 2016.

Resveratrol and inflammatory bowel disease: the evidence so far.

Despite the fact that inflammatory bowel disease (IBD) has still no recognised therapy, treatments which have proven at least mildly successful in improving IBD symptoms include anti-inflammatory drugs and monoclonal antibodies targeting pro-inflammatory cytokines. Resveratrol, a natural (poly)phenol found in grapes, red wine, grape juice and several species of berries, has been shown to prevent and ameliorate intestinal inflammation. Here, we discuss the role of resveratrol in the improvement of inflammatory disorders involving the intestinal mucosa. The present review covers three specific aspects of resveratrol in the framework of inflammation: (i) its content in food; (ii) its intestinal absorption and metabolism; and (iii) its anti-inflammatory effects in the intestinal mucosa in vitro and in the very few in vivo studies present to date. Actually, if several studies have shown that resveratrol may down-regulate mediators of intestinal immunity in rodent models, only two groups have performed intervention studies in human subjects using resveratrol as an agent to improve IBD conditions. The effects of resveratrol should be further investigated by conducting well-designed clinical trials, also taking into account different formulations for the delivery of the bioactive compound.

Could Pomegranate Juice Help in the Control of Inflammatory Diseases?

Fruits rich in polyphenols, such as pomegranates, have been shown to have health benefits relating to their antioxidant and anti-inflammatory properties. Using data obtained from PubMed and Scopus, this article provides a brief overview of the therapeutic effects of pomegranate on chronic inflammatory diseases (CID) such as inflammatory bowel disease (IBD), rheumatoid arthritis (RA), metabolic and cardiovascular disorders, and other inflammatory-associated conditions, with an emphasis on fruit-derived juices. Most studies regarding the effects of pomegranate juice have focused on its ability to treat prostate cancer, diabetes, and atherosclerosis. However, pomegranate juice has shown therapeutic potential for many other illnesses. For instance, a small number of human clinical trials have highlighted the positive effects of pomegranate juice and extract consumption on cardiovascular health. The beneficial effects of pomegranate components have also been observed in animal models for respiratory diseases, RA, neurodegenerative disease, and hyperlipidaemia. Furthermore, there exists strong evidence from rodent models suggesting that pomegranate juice can be used to effectively treat IBD, and as an anti-inflammatory agent to treat CID. The effects of pomegranate intake should be further investigated by conducting larger and more well-defined human trials.

Is cytotoxicity a determinant of the different in vitro and in vivo effects of bioactives?

BACKGROUND: Foodstuffs of both plant and animal origin contain a wide range of bioactive compounds. Although human intervention studies are mandatory to assess the health effects of bioactives, the in vitro approach is often used to select the most promising molecules to be studied in vivo. To avoid misleading results, concentration and chemical form, exposure time, and potential cytotoxicity of the tested bioactives should be carefully set prior to any other experiments. METHODS: In this study the possible cytotoxicity of different bioactives (docosahexaenoic acid, propionate, cyanidin-3-O-glucoside, protocatechuic acid), was investigated in HepG2 cells using different methods. Bioactives were supplemented to cells at different concentrations within the physiological range in human blood, alone or in combination, considering two different exposure times. RESULTS: Reported data clearly evidence that in vitro cytotoxicity is tightly related to the exposure time, and it varies among bioactives, which could exert a cytotoxic effect even at a concentration within the in vivo physiological blood concentration range. Furthermore, co-supplementation of different bioactives can increase the cytotoxic effect. CONCLUSIONS: Our results underline the importance of in vitro cytotoxicity screening that should be considered mandatory before performing studies aimed to evaluate the effect of bioactives on other cellular parameters. Although this study is far from the demonstration of a toxic effect of the tested bioactives when administered to humans, it represents a starting point for future research aimed at verifying the existence of a potential hazard due to the wide use of high doses of multiple bioactives.

Evidence of a DHA Signature in the Lipidome and Metabolome of Human Hepatocytes.

Cell supplementation with bioactive molecules often causes a perturbation in the whole intracellular environment. Omics techniques can be applied for the assessment of this perturbation. In this study, the overall effect of docosahexaenoic acid (DHA) supplementation on cultured human hepatocyte lipidome and metabolome has been investigated using nuclear magnetic resonance (NMR) in combination with traditional techniques. The effect of two additional bioactives sharing with DHA the lipid-lowering effect-propionic acid (PRO) and protocatechuic acid (PCA)-has also been evaluated in the context of possible synergism. NMR analysis of the cell lipid extracts showed that DHA supplementation, alone or in combination with PCA or PRO, strongly altered the cell lipid profile. The perfect discrimination between cells receiving DHA (alone or in combination) and the other cells reinforced the idea of a global rearrangement of the lipid environment induced by DHA. Notably, gas chromatography and fluorimetric analyses confirmed the strong discrimination obtained by NMR. The DHA signature was evidenced not only in the cell lipidome, but also in the metabolome. Results reported herein indicate that NMR, combined with other techniques, represents a fundamental approach to studying the effect of bioactive supplementation, particularly in the case of molecules with a broad spectrum of mechanisms of action.

Mixed pro- and anti-oxidative effects of pomegranate polyphenols in cultured cells.

In recent years, the number of scientific papers concerning pomegranate (Punica granatum L.) and its health properties has increased greatly, and there is great potential for the use of bioactive-rich pomegranate extracts as ingredients in functional foods and nutraceuticals. To translate this potential into effective strategies it is essential to further elucidate the mechanisms of the reported bioactivity. In this study HepG2 cells were supplemented with a pomegranate fruit extract or with the corresponding amount of pure punicalagin, and then subjected to an exogenous oxidative stress. Overall, upon the oxidative stress the gene expression and activity of the main antioxidant enzymes appeared reduced in supplemented cells, which were more prone to the detrimental effects than unsupplemented ones. No differences were detected between cells supplemented with the pomegranate juice or the pure punicalagin. Although further studies are needed due to the gaps existing between in vitro and in vivo studies, our results suggest caution in the administration of high concentrations of nutraceutical molecules, particularly when they are administered in concentrated form.

Bioactive-rich Sideritis scardica tea (mountain tea) is as potent as Camellia sinensis tea at inducing cellular antioxidant defences and preventing oxidative stress.

BACKGROUND: In several countries, tea (hot-water infusions of dried Camellia sinensis (CS) leaves) is a major source of antioxidant flavonoids, and its consumption has been associated with several favourable outcomes. Other plants used for the preparation of herbal teas are sources of phenolic antioxidant compounds; among them Sideritis scardica (SS) is used for the preparation of a popular drink throughout Eastern and Central Europe. We have compared the effects of an SS extract to a CS extract in HepG2 cells to set the scientific basis for the exploitation of other herbal teas in counteraction of oxidative stress. RESULTS: Although SS extract had a lower phenolic concentration and total antioxidant capacity than CS extract, their cellular antioxidant effects were similar. The different phenolic pattern of the extracts suggests that the protective activity is not limited to catechins. CONCLUSION: Although further research is needed, our data represent a first contribution for the evaluation of the potential effect of SS in increasing antioxidant defences. © 2013 Society of Chemical Industry.

Sugar cane and sugar beet molasses, antioxidant-rich alternatives to refined sugar.

Molasses, the main byproduct of sugar production, is a well-known source of antioxidants. In this study sugar cane molasses (SCM) and sugar beet molasses (SBM) were investigated for their phenolic profile and in vitro antioxidant capacity and for their protective effect in human HepG2 cells submitted to oxidative stress. According to its higher phenolic concentration and antioxidant capacity in vitro, SCM exhibited an effective protection in cells, comparable to or even greater than that of α-tocopherol. Data herein reported emphasize the potential health effects of molasses and the possibility of using byproducts for their antioxidant activity. This is particularly important for consumers in developing countries, as it highlights the importance of consuming a low-price, yet very nutritious, commodity.

EPA or DHA supplementation increases triacylglycerol, but not phospholipid, levels in isolated rat cardiomyocytes.

It is well recognized that a high dietary intake of long-chain polyunsaturated fatty acids (LC-PUFA) has profound benefits on health and prevention of chronic diseases. In particular, in recent years there has been a dramatic surge of interest in the health effects of n-3 LC-PUFA derived from fish, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids. Notwithstanding, the metabolic fate and the effects of these fatty acids once inside the cell has seldom been comprehensively investigated. Using cultured neonatal rat cardiomyocytes as model system we have investigated for the first time, by means of high-resolution magic-angle spinning nuclear magnetic resonance (HR-MAS NMR) spectroscopy in combination with gas chromatography (GC), the modification occurring in the cell lipid environment after EPA and DHA supplementation. The most important difference between control and n-3 LC-PUFA-supplemented cardiomyocytes highlighted by HR-MAS NMR spectroscopy is the increase of signals from mobile lipids, identified as triacylglycerols (TAG). The observed increase of mobile TAG is a metabolic response to n-3 LC-PUFA supplementation, which leads to an increased lipid storage. The sequestration of mobile lipids in lipid bodies provides a deposit of stored energy that can be accessed in a regulated fashion according to metabolic need. Interestingly, while n-3 LC-PUFA supplementation to neonatal rat cardiomyocytes causes a huge variation in the cell lipid environment, it does not induce detectable modifications in water-soluble metabolites, suggesting negligible interference with normal metabolic processes.

Food-derived bioactives as potential regulators of the IL-12/IL-23 pathway implicated in inflammatory bowel diseases.

The gene-specific modulation of inflammatory cytokines by food bioactives represents a possible approach to the nutritional or pharmaceutical prevention and treatment of inflammatory bowel disease (IBD). There is evidence for a key role of the interleukin-12beta1/23 receptor (IL-12 Rbeta1/23 R) pathway in IBD, and that reduction of the normal expression of the IL-23 R gene may provide a therapeutic target for this disease. The binding of interleukin-23 (IL-23) to its receptor IL-23 R regulates a newly defined effector T-cell subset, Th17 cells, characterised by the production of interleukin-17 (IL-17) and other cytokines, including tumour necrosis factor-alpha (TNF-alpha). In this study we developed an assay that measured IL-17 and TNF-alpha expression after incubation with specific dietary bioactives in the human T-cell Kit 225. It is anticipated that these changes will reflect differences in IL-23 R production, albeit indirectly. The cell line Kit 225 has similarities to Th17 cells, a subset of T cells producing IL-17 and TNF-alpha, and in initial experiments we demonstrated that the cells express both IL-23 receptor subunits, as well as IL-17 and TNF-alpha genes. Upon verification that stimulation of Kit 225 cells with 1ng/mL IL-23 significantly upregulated IL-17 and TNF-alpha gene expression, and IL-17 production, we supplemented cells with selected food bioactives, caffeic acid phenethyl ester (CAPE), epigallocatechin gallate (EGCG), docosahexaenoic acid (DHA), and linoleic acid (LA), and with phorbol myristate acetate (PMA) and sodium salicylate, used as pro-inflammatory and anti-inflammatory controls, respectively. In both unstimulated cells and after IL-23 stimulation, bioactives modulated the pro-inflammatory cytokines involved in IBD, underlining the possible role of foods in this disease. EGCG and DHA, which significantly inhibited both IL-17 and TNF-alpha expression, appeared particularly interesting.

n-3 PUFA as regulators of cardiac gene transcription: a new link between PPAR activation and fatty acid composition.

The fatty acids regulate gene expression directly binding to nuclear receptors or affecting the protein content of transcription factors. In this work, supplementing primary cultures of neonatal rat cardiomyocytes with 60 microM EPA or DHA, we demonstrated by an ELISA assay an increased PPAR beta/delta binding to DNA. n-3 PUFA supplementation deeply changed the acyl composition of both cytosolic and nuclear fractions. The high content of total fatty acids, particularly EPA and DHA, and its increase following supplementation suggested a selective accumulation of n-3 PUFAs in the nucleus, supporting the direct interaction of n-3 PUFA with PPAR. The activity of acyl-CoA thioesterase (ACOT), catalyzing the reaction leading to NEFA from acyl-CoA, increased in n-3 PUFA supplemented cells. The NEFA/acyl-CoA ratio is an important regulator of the fatty acid transport to the nucleus and consequent modulation of gene transcription, and although ACOT activity is not the only parameter of this ratio, it is important for the control of the NEFA pool composition. Our data further clarify what happens in cardiomyocytes following n-3 PUFA supplementation, linking the modification of acyl composition to ACOT activity and PPAR activation.

Green tea extract selectively activates peroxisome proliferator-activated receptor beta/delta in cultured cardiomyocytes.

Hypoxia/reoxygenation is one of the causes of the increased expression of inducible NO synthase in cardiomyocytes. In a recent study we demonstrated that a single, high dose of green tea extract (GT) supplemented to the medium of cultured cardiomyocytes just before hypoxia/reoxygenation is able to prevent the increased expression of inducible NO synthase, therefore reducing NO overproduction. In the present study we investigated the mechanism by which GT reduces NO production. Since a molecular mechanism for polyphenol activity has been postulated, and PPAR activation is related to the transcription of the inducible NO synthase gene, we evaluated the activation of PPAR by GT. A moderate GT concentration, supplemented to the cardiomyocyte medium since the initial seeding, selectively activated the PPAR-beta/delta isoform. Furthermore, we observed a reduction in NO production and an increase in total antioxidant activity, indicating that GT components may act on both reactive oxygen species, via an antioxidant mechanism, and NO overproduction. PPAR-beta/delta activation could represent the key event in the reduction of NO production by GT. Although PPAR activation by GT was lower than activation by fenofibrate, it is very interesting to note that it was selective for the beta/delta isoform, at least in neonatal cardiomyocytes.

Effect of cultivar on the protection of cardiomyocytes from oxidative stress by essential oils and aqueous extracts of basil (Ocimum basilicum L.).

Notwithstanding the wide range of biological and pharmacological activities reported for sweet basil (Ocimum basilicum L.), many discrepancies are still present in the evaluation of its health-promoting properties. These discordances could be at least in part due to insufficient details of qualitative and quantitative composition, connected to the ample variability of this species. Furthermore, many investigations have been carried out in vitro, with few data available on the effectiveness in biological systems. In this study, the protective effect of essential oils and water-soluble extracts derived from three different cultivars of sweet basil has been evaluated in cultured cardiomyocytes. To verify the effectiveness of supplemented oils/extracts in counteracting oxidative damage, cardiomyocytes were stressed by the addition of hydrogen peroxide. The results indicate that (a) in vitro antioxidant activity is not predictive of biological activity and (b) basil can yield extracts with substantially different protective effects, in relation to composition and extraction techniques. Variation among different cultivars has also been detected.

Dietary Selenium for the counteraction of oxidative damage: fortified foods or supplements?

Since any significant modification in the Se status, leading to changes in the activity of the seleno-enzymes, may have important consequences on the susceptibility of tissues to oxidative stress, considerable efforts have been made upon increasing Se dietary intake. In this respect, an important debate is still open about the bioavailability and the effectiveness of Se, and more generally nutrients, in supplements compared with foods. Using male Wistar rats, we have compared the effectiveness of two different diets in which an adequate Se content (0.1 mg/kg) was achieved by adding the element as sodium selenite or as component of a lyophilized Se-enriched food, in the counteraction of an oxidative stress induced by intraperitoneal administration of adriamycin. Both Se-enriched diets were able to reduce the consequences of the oxidative stress in liver, mainly by increasing glutathione peroxidase activity. This increase was more evident in rats fed on the diet enriched with the lyophilized food, probably due to the different chemical forms of Se, or to other components of the food itself. Although further studies are needed, data herein presented may contribute to the characterization of the effectiveness of Se from different sources, foods or supplements, in the light of dietary advice to the population concerning improvement of Se intake.

N-3 PUFAs modulate global gene expression profile in cultured rat cardiomyocytes. Implications in cardiac hypertrophy and heart failure.

In cardiac cells the effects of n-3 PUFAs on the whole genome are still unknown despite their recognized cardioprotective effects and ability to modulate gene expression. We have evaluated the effects of n-3 PUFAs supplementation on the global gene expression profile in cultured neonatal rat cardiomyocytes, detecting many genes related to lipid transport and metabolism among the upregulated ones. Many of the downregulated genes appeared related to inflammation, cell growth, extracellular and cardiac matrix remodelling, calcium movements and ROS generation. Our data allow to speculate that the cardioprotective effect of n-3 PUFAs is related to a direct modulation of genes in cardiac cells.

Vitamin B6 deficiency and dietary fats: effects on lipid composition and glutathione peroxidase activity in rat liver.

Dietary selenium, vitamin B6 and fatty acids modulate both tissue acyl composition by regulating polyunsaturated fatty acid metabolism and antioxidant defences by influencing glutathione peroxidase activity. Alteration in the intake of one of them could therefore lead to different results depending on the intake of the others. To clarify this complex relationship, in the present study we have evaluated the modifications occurring in fatty acid composition and glutathione peroxidase activity in total liver and liver microsomes of rats fed diets containing the same amount of selenium, but different vitamin B6 content and fatty acid composition. Our data indicate that both acyl composition and glutathione peroxidase activity are greatly influenced not only by vitamin B6 deficiency, but also by the diet unsaturation degree. This study underlines that not only selenium availability but also other nutrients can modulate glutathione peroxidase activity.

Counteraction of adriamycin-induced oxidative damage in rat heart by selenium dietary supplementation.

Many reports indicate that dietary selenium, potentially increasing the activity of glutathione peroxidase, could offer protection against free-radical-induced damage. The effects of diets moderately enriched in selenium, as sodium selenite or as a lyophilized selenium-rich food, were studied in rats. Adriamycin, an anticancer drug causing a free-radical-mediated cardiotoxicity, was administered intraperitoneally to some rats. The onset of an oxidative damage was indicated by the increase in the plasma level of reactive oxygen metabolites coupled to a decrease in the total antioxidant activity but without modification of glutathione peroxidase activity, which were observed in all rats, independent of the dietary treatment. On the contrary, in the heart, selenium supplementation caused an increase in the total antioxidant activity, glutathione concentration, and glutathione peroxidase and catalase activities leading to a decreased generation of reactive oxygen metabolites. These results clearly indicate that a moderate Se dietary supplementation counteracts adriamycin-induced cardiotoxicity by preservation of endogenous antioxidants.

Susceptibility to hypoxia/reoxygenation of aged rat cardiomyocytes and its modulation by selenium supplementation.

Since in the aged heart an increased basal production of reactive oxygen species (ROS) has been demonstrated, and the resistance to ROS attack could be ameliorated by antioxidant supplementation, we verified the protective effect of selenium, as sodium selenite (SS) or seleno methionine (SM), in cultured rat cardiomyocytes aged in vitro. In normoxia, glutathione peroxidase (GPx) activity and total antioxidant activity were higher in old than in young cardiomyocytes, suggesting the existence of a compensatory increase of antioxidant defenses. When aged cells were submitted to hypoxia/reoxygenation, GPx activity was not modified; while total antioxidant activity decreased, conjugated diene level increased. Selenium supplementation, particularly as SM, was able to increase GPx, and consequently total antioxidant activity, and to decrease conjugated diene production. The observed ability of selenium supplementation to protect aged cardiomyocytes from hypoxia/reoxygenation damage underlines the importance of an optimal selenium dietary intake, particularly in the elderly.