Impact of Glucoraphanin-Mediated Activation of Nrf2 on Non-Alcoholic Fatty Liver Disease with a Focus on Mitochondrial Dysfunction.

Non-alcoholic fatty liver disease (NAFLD) is a common disease in Western nations and ranges in severity from steatosis to steatohepatitis (NASH). NAFLD is a genetic-environmental-metabolic stress-related disease of unclear pathogenesis. NAFLD is triggered by caloric overconsumption and physical inactivity, which lead to insulin resistance and oxidative stress. A growing body of evidence indicates that mitochondrial dysfunction plays a critical role in the pathogenesis of NAFLD. Mitochondrial dysfunction not only promotes fat accumulation, but also leads to generation of reactive oxygen species (ROS) and lipid peroxidation, resulting in oxidative stress in hepatocytes. Nuclear factor erythroid 2-related factor 2 (Nrf2) is an important modulator of antioxidant signaling that serves as a primary cellular defense against the cytotoxic effects of oxidative stress. The pharmacological induction of Nrf2 ameliorates obesity-associated insulin resistance and NAFLD in a mouse model. Sulforaphane and its precursor glucoraphanin are derived from broccoli sprouts and are the most potent natural Nrf2 inducers-they may protect mitochondrial function, thus suppressing the development of NASH. In this review, we briefly describe the role of mitochondrial dysfunction in the pathogenesis of NASH and the effects of glucoraphanin on its development.

Bioavailability of Sulforaphane Following Ingestion of Glucoraphanin-Rich Broccoli Sprout and Seed Extracts with Active Myrosinase: A Pilot Study of the Effects of Proton Pump Inhibitor Administration.

We examined whether gastric acidity would affect the activity of myrosinase, co-delivered with glucoraphanin (GR), to convert GR to sulforaphane (SF). A broccoli seed and sprout extract (BSE) rich in GR and active myrosinase was delivered before and after participants began taking the anti-acid omeprazole, a potent proton pump inhibitor. Gastric acidity appears to attenuate GR bioavailability, as evidenced by more SF and its metabolites being excreted after participants started taking omeprazole. Enteric coating enhanced conversion of GR to SF, perhaps by sparing myrosinase from the acidity of the stomach. There were negligible effects of age, sex, ethnicity, BMI, vegetable consumption, and bowel movement frequency and quality. Greater body mass correlated with reduced conversion efficiency. Changes in the expression of 20 genes in peripheral blood mononuclear cells were evaluated as possible pharmacodynamic indicators. When grouped by their primary functions based on a priori knowledge, expression of genes associated with inflammation decreased non-significantly, and those genes associated with cytoprotection, detoxification and antioxidant functions increased significantly with bioavailability. Using principal components analysis, component loadings of the changes in gene expression confirmed these groupings in a sensitivity analysis.

Dietary glucosinolates and risk of type 2 diabetes in 3 prospective cohort studies.

Background: Glucosinolates are a group of phytochemicals that are abundant in cruciferous vegetables and precursors of the potentially chemopreventive isothiocyanates. Isothiocyanates may reduce oxidative stress and inflammation, but little is known regarding the association between glucosinolate intake and risk of type 2 diabetes (T2D). Objective: To evaluate the association between the intake of glucosinolates and the incidence of T2D in US men and women. Design: This prospective cohort study investigated 200,907 women and men [71,256 women from the Nurses' Health Study (NHS; 1984-2012), 88,293 women from the NHS II (1991-2013), and 41,358 men from the Health Professionals Follow-Up Study (1986-2012)] who were free of diabetes, cardiovascular disease, and cancer at baseline. Diet was assessed using validated semiquantitative food frequency questionnaires. Self-reported T2D incidence was confirmed by a supplementary questionnaire. Results: During follow-up in the 3 cohorts, we accumulated 4,303,750 person-years and 16,567 incident cases of T2D. After adjustment for major lifestyle and dietary risk factors for T2D, participants in the highest quintile of total glucosinolate intake had a 19% higher risk (95% CI: 13%, 25%; Ptrend < 0.001) of T2D than did those in the lowest quintile. The intake of 3 major glucosinolate subtypes was consistently and significantly associated with T2D risk, with pooled HRs ranging from 1.13 to 1.18 (all Ptrend < 0.001). A significant association was also observed between total cruciferous vegetable consumption and T2D (HR: 1.16; 95% CI :1.07, 1.25; Ptrend < 0.001). These associations persisted in subgroups defined by demographic, lifestyle, and other dietary factors. Conclusions: Dietary glucosinolate intake was associated with a moderately higher risk of T2D in US adults. These results need to be replicated in further investigations, including biomarker-based studies. Mechanistic research is also needed to understand the relation between exposures to glucosinolates, isothiocyanates, and other metabolites with T2D risk. This trial was registered at clinicaltrials.gov as NCT03366532.

Nutrigenomic effects of glucosinolates on liver, muscle and distal kidney in parasite-free and salmon louse infected Atlantic salmon.

BACKGROUND: Reduction of Lepeophtheirus salmonis infection in Atlantic salmon achieved by glucosinolates (GLs) from Brassica plants was recently reported. However, wider application of functional feeds based on GLs requires better knowledge of their positive and adverse effects. METHODS: Liver, distal kidney and muscle transcriptomes of salmon exposed to the extreme dose of GLs were profiled by microarray, while qPCR analysis followed up selected hepatic and renal responses under the extreme and moderate GLs dose during the L. salmonis challenge. Transcriptional analysis were complemented with measurements of organ indices, liver steatosis and plasma profiling, including indicators of cytolysis and bilirubin. Finally, the third trial was performed to quantify the effect of lower GLs doses on growth. RESULTS: The extreme GLs dose caused a decrease in hepatic fat deposition and growth, in line with microarray findings, which suggested tissue remodeling and reduction of cellular proliferation in the skeletal muscle and liver. Lower GLs inclusion levels in a follow-up trial did not show negative effects on growth. Microarray analysis of the distal kidney pointed to activation of anti-fibrotic responses under the overexposure. However, analyses of ALT, CK and AST enzymes in plasma provided no evidence of increased cytolysis and organ damage. Prevalent activation of phase-2 detoxification genes that occurred in all three tissues could be considered part of beneficial effects caused by the extreme dose of GLs. In addition, transcriptomic evidence suggested GLs-mediated iron and heme withdrawal response, including increased heme degradation in muscle (upregulation of heme oxygenase-1), decrease of its synthesis in liver (downregulation of porphobilinogen deaminase) and increased iron sequestration from blood (hepatic induction of hepcidin-1 and renal induction of intracellular storage protein ferritin). This response could be advantageous for salmon upon encountering lice, which depend on the host for the provision of iron carrying heme. Most of the hepatic genes studied by qPCR showed similar expression levels in fish exposed to GLs, lice and their combination, while renal induction of leptin suggested heightened stress by the combination of extreme dose of GLs and lice. High expression of interferon γ (cytokine considered organ-protective in mammalian kidney) was detected at the moderate GLs level. This fish also showed highest plasma bilirubin levels (degradation product of heme), and had lowest number of attached lice, further supporting hypothesis that making heme unavailable to lice could be part of an effective anti-parasitic strategy. CONCLUSIONS: Modulation of detoxification and iron metabolism in Atlantic salmon tissues could be beneficial prior and during lice infestations. Investigation of anti-lice functional feeds based on low and moderate GLs inclusion levels thus deserves further attention.

Concentrations of thiocyanate and goitrin in human plasma, their precursor concentrations in brassica vegetables, and associated potential risk for hypothyroidism.

Brassica vegetables are common components of the diet and have beneficial as well as potentially adverse health effects. Following enzymatic breakdown, some glucosinolates in brassica vegetables produce sulforaphane, phenethyl, and indolylic isothiocyanates that possess anticarcinogenic activity. In contrast, progoitrin and indolylic glucosinolates degrade to goitrin and thiocyanate, respectively, and may decrease thyroid hormone production. Radioiodine uptake to the thyroid is inhibited by 194 µmol of goitrin, but not by 77 µmol of goitrin. Collards, Brussels sprouts, and some Russian kale (Brassica napus) contain sufficient goitrin to potentially decrease iodine uptake by the thyroid. However, turnip tops, commercial broccoli, broccoli rabe, and kale belonging to Brassica oleracae contain less than 10 µmol of goitrin per 100-g serving and can be considered of minimal risk. Using sulforaphane plasma levels following glucoraphanin ingestion as a surrogate for thiocyanate plasma concentrations after indole glucosinolate ingestion, the maximum thiocyanate contribution from indole glucosinolate degradation is estimated to be 10 µM, which is significantly lower than background plasma thiocyanate concentrations (40-69 µM). Thiocyanate generated from consumption of indole glucosinolate can be assumed to have minimal adverse risks for thyroid health.

Antioxidant capacity of broccoli sprouts subjected to gastrointestinal digestion.

BACKGROUND: Broccoli is a common vegetable recognized as a rich source of antioxidants. To date, research on the antioxidant properties of broccoli, predominantly conducted on extracts, has not considered the lesions of composition and this activity after gastrointestinal digestion. Here the stability of antioxidants during gastrointestinal digestion was evaluated in conjunction with the protective effects of broccoli sprouts (BS) against oxidative stress in human colon cells. RESULTS: The obtained data suggest that, among the biocompounds identified in BS, glucosinolates were mainly degraded under gastrointestinal digestion, while phenolics, particularly hydroxycinnamic acid derivatives, were the most resistant constituents. The antioxidant capacity of BS extract subjected to gastrointestinal digestion was similar to or higher than that determined for non-digested BS. Gastrointestinal digested BS extract exhibited reactive oxygen species (ROS)-inhibitory capacity in NCM460 human colon cells, with 1 mg mL(-1) showing an ROS clearance of 76.59%. A 57.33% reduction in oxidative DNA damage in NCM460 cells due to treatment with digested BS extract was observed. CONCLUSION: The results lend support to the possible application of BS as a rich source of antioxidants to improve the defensive system against oxidative stress in the human colon mucosa.

Evaluation of Camelina sativa (L.) Crantz meal as an alternative protein source in ruminant rations.

BACKGROUND: Camelina sativa (CS) is an oilseed crop used for biofuel production. By-products from oil extraction are high in protein and can be used in ruminant rations; more information about their nutritive value is required also considering the antinutrional factor content of the by-products. The aim of this study was to evaluate the nutritive value of CS meal genotypes in comparison with canola. RESULTS: Ten CS genotypes and one canola cultivar were evaluated. Meals were obtained from seeds after solvent oil extraction. CS average crude protein (CP) content (g kg⁻¹ dry matter) was 457. Numerical differences in lysine and sulfur amino acid content were observed among CS genotypes. Glucosinolate (mmol kg⁻¹) content was higher for CS (23.1) than canola (7.2). Sinapine content (g kg⁻¹) was lower for CS (2.79) than for canola (4.32). Differences were observed among CS genotypes for rumen undegraded protein (RUP). Average RUP (g kg⁻¹ CP) was 316 for CS and 275 for canola. CONCLUSIONS: CS meal has potential for use in ruminant rations as a high-quality protein source. In vivo studies are needed to compare CS with other protein sources used in cattle rations. Implementation of breeding programs for improved meal quality is recommend.

Healthy and adverse effects of plant-derived functional metabolites: the need of revealing their content and bioactivity in a complex food matrix.

In recent years, both food quality and its effect on human health have become a fundamental issue all over the world. As a consequence of this new and increased awareness, American, European, and Asian policymakers have strongly encouraged the research programs on food quality and safety thematic. Attempts to improve human health and to satisfy people's desire for healthcare without intake of pharmaceuticals, has led the food industry to focus attention on functional or nutraceutical food. For a long time, compounds with nutraceutical activity have been produced chemically, but the new demands for a sustainable life have gradually led the food industry to move towards natural compounds, mainly those derived from plants. Many phytochemicals are known to promote good health, but, sometimes, undesirable effects are also reported. Furthermore, several products present on the market show few benefits and sometimes even the reverse - unhealthy effects; the evidence of efficacy is often unconvincing and epidemiological studies are necessary to prove the truth of their claims. Therefore, there is a need for reliable analytical control systems to measure the bioactivity, content, and quality of these additives in the complex food matrix. This review describes the most widespread nutraceutics and an analytical control of the same using recently developed biosensors which are promising candidates for routine control of functional foods.

Sulforaphane and related mustard oils in focus of cancer prevention and therapy.

The plant family Brassicaceae, formerly Cruciferae, contains mustard oil glycosides, from which mustard oils are enzymatically hydrolyzed. Mustard oils offer protection from pests, microorganisms and fungi. More than 120 different mustard oils with various biological functions are known. Since ancient times, these substances are used as natural antibiotics, antiviral drugs and antimycotics. The antioxidative effect of mustard oils contributes to protection from DNA damage. Epidemiological and experimental studies have shown preventive and therapeutic effects of crucifers or isolated substances thereof. Particularly well studied is the mustard oil sulforaphane, which is contained in high concentrations in broccoli and its sprouts. As has been shown in mice recently, sulforaphane also targets the most malignant cancer stem cells, which are not affected by conventional cancer treatments. Based on these promising results, the first prospective clinical studies with cancer patients and sulforaphane-enriched broccoli sprouts have now been initiated in the United States.

Multidirectional time-dependent effect of sinigrin and allyl isothiocyanate on metabolic parameters in rats.

Sinigrin (SIN) and allyl isothiocyanate (AITC) are compounds found in high concentrations in Brassica family vegetables, especially in Brussels sprouts. Recently, they have been used as a nutrition supplement for their preventive and medicinal effect on some types of cancer and other diseases. In this research, nutritional significance of parent glucosinolate sinigrin 50 µmol/kg b. w./day and its degradation product allyl isothiocyanate 25 µmol/kg b. w./day and 50 µmol/kg b. w./day was studied by the evaluation of their influence on some parameters of carbohydrate and lipid metabolism in an animal rat model in vivo after their single (4 h) and 2 weeks oral administration. Additionally, the aim of this trial was to evaluate the direct action of AITC on basal and epinephrine-induced lipolysis in isolated rat adipocytes at concentration 1 µM, 10 µM and 100 µM in vitro. Sole AITC after 4 h of its ingestion caused liver triacylglycerols increment at both doses and glycaemia only at the higher dose. Multiple SIN treatment showed its putative bioconversion into AITC. It was found that SIN and AITC multiple administration in the same way strongly disturbed lipid and carbohydrate homeostasis, increasing esterified and total cholesterol, free fatty acids and lowering tracylglycerols in the blood serum. Additionally, AITC at both doses elevated insulinaemia and liver glycogen enhancement. The in vitro experiment revealed that AITC potentiated basal lipolysis process at 10 µM, and had stimulatory effect on epinephrine action at 1 µM and 10 µM. The results of this study demonstrated that the effect of SIN and AITC is multidirectional, indicating its impact on many organs like liver as well as pancreas, intestine in vivo action and rat adipocytes in vitro. Whilst consumption of cruciferous vegetables at levels currently considered "normal" seems to be beneficial to human health, this data suggest that any large increase in intake could conceivably lead to undesirable effect. This effect is potentiated with time of action of the examined compounds, whose influence is rather adverse for the majority of metabolic pathways (liver steatosis at short duration and insulinaemia, cholesterolaemia at long time treatment). Beneficial action of AITC concerned intensified hydrolysis of TG in the blood serum with a simultaneous lipolysis in adipocytes.

Genetic and molecular approaches to improve nutritional value of Brassica napus L. seed.

Oilseed rape (Brassica napus L.) is a major oil crop that also supplies proteins for the feed industry. In order to reduce total cost production, the objective is to increase oil yield while reducing crop inputs (especially nitrogen and pesticides). Concomitantly, it is necessary to anticipate specific uses (e.g., fatty acid composition) and to ensure the valorisation of the by-products (rapeseed meal). By the past, improvement of seed quality focused on fatty acid balance and low seed glucosinolate content. Current goals include the breeding of yellow-seeded rapeseed lines with high content of seed oil. The use of molecular tools and the exploitation of Arabidopsis knowledge will be presented and discussed.

Evaluation of the safety and bioactivity of purified and semi-purified glucoraphanin.

The anti-carcinogenic effects of broccoli have been attributed to sulforaphane, the hydrolysis product of glucoraphanin (GRP). Here we determined if purified GRP, in the absence of the plant-derived hydrolyzing enzyme myrosinase, could affect pulmonary and hepatic ethoxyresorufin O-deethylase (EROD) and/or NAD(P)H-quinone oxidoreductase 1 (NQO1) activity. Male F344 rats were administered semi-synthetic, semi-purified or purified GRP (240 mg/kg: 550 micromol/kg rat daily for 4 days) by gavage. Hepatic and pulmonary NQO1 activity increased ( approximately 20%), but not EROD. Varying doses of semi-purified GRP (30, 60, or 120 mg/kg rat daily for 4 days) again caused no change in EROD activity, although a dose-dependent increase in NQO1 was seen. Urinary excretion of mercapturic acids showed no difference between preparations, and recovery increased with decreasing dose. Histopathologic examination revealed no abnormal tissues other than cecum, where inflammation was dose dependent; mild at 120 mg/kg and severe at 240 mg/kg, a greatly supra-physiological dose. We conclude that GRP 30-60 mg/kg p.o. is safe and effectively enhances NQO1 in all tissues evaluated.

Sensitivity to allyl isothiocyanate, dimethyl trisulfide, sinigrin, and cooked cauliflower consumption.

The consumption of cauliflower consumers has been related to the olfactory and gustatory sensitivities to potentially objectionable flavor compounds in this vegetable. Based on the ascending concentration series method of limits, a first experiment was designed to develop rapid tests dedicated to estimate individuals' olfactory thresholds for allyl isothiocyanate (AITC) and dimethyl trisulfide (DMTS) and gustatory thresholds for sinigrin (SIN). The best compromise between rapidity and reliability was obtained with two replications of a four-alternative forced choice (AFC) at six ascending concentrations (6x2x4-AFC) for both AITC and DMTS, and with a 6x1x4-AFC for SIN. In a second experiment, sensitivity to SIN, AITC and DMTS was determined on 267 participants divided into three cauliflower consumer target groups: non-, medium- or high consumers. The non-consumers were significantly more sensitive to SIN and AITC than were the medium and high consumers. No effect of consumer's sensitivity to DMTS was observed.

[Biologically-active phytochemicals in vegetable food]. / Sostanze biologicamente attive presenti negli alimenti di origine vegetale.

Phytochemicals constitute a heterogeneous group of substances and evidence for their role in the protective effect on human health, when their dietary intake is significant, is emerging. These compounds have biological properties as antioxidant activity, modulation of detoxification enzymes, stimulation of the immune system, decrease of platelet aggregation and modulation of hormone metabolism. Glucosinolates and widely group of polyphenols, including the main category of flavonoids, are examined. Since their weak estrogenic activity is of interest at present, isoflavones are discussed at length. Numerous factors affect the phytochemical content and availability in plants; these factors include environmental conditions and industrial or domestic processing. The bioavailability of phytochemicals can be influenced by intrinsic factors in food and/or in human, in general the substances are little adsorbed, largely metabolized and rapidly eliminated. For this reason it is advisable that their consumption is constant in the time to maintain high concentrations of metabolites in the blood.

Studies on the toxic effects of crambe meal and two of its constituents, 1-cyano-2-hydroxy-3-butene (CHB) and epi-progoitrin, in broiler chick diets.

1. Studies were undertaken to determine a safe inclusion rate for crambe (Crambe abyssinica) meal in broiler chick diets, and to determine the mechanism for adverse effects by investigating its constituents; 1-cyano-2-hydroxy-3-butene (CHB) and 3-butenyl glucosinolate (epi-progoitrin, E-PG). 2. Crambe meals were prepared to differ in E-PG (19, 36 and 40 g/kg) and CHB contents (0.1, 0.7 and 1.9 g/kg), and with either active or inactive thioglucosidase. 3. Meals were fed to 7-d-old broiler chicks at 50 or 100 g/kg of the diet for 12 or 13 d. In separate studies, isolated E-PG or CHB were mixed into the diet or administered by gavage to 7-d-old broiler chicks in amounts equivalent to 50 or 100 g/kg crambe meal diets for 10 and 12 d, respectively. 4. Weight gain decreased (P < 0.05) in chicks fed on the high glucosinolate crambe diets or isolated E-PG. Food consumption decreased (P < 0.05) in chicks fed on the diet containing the high E-PG meal with active enzyme. 5. Mild liver lesions and increased serum aspartate aminotransferase were found in chicks fed on the diet containing the high glucosinolate meal with active enzyme. Other organs, including thyroids, were normal. 6. Commercially-processed crambe meal appeared safe at an inclusion rate of 50 or 100 g/kg diet, but could not be recommended at this point for long term feeding.

Glucosinolatos de la dieta, un posible factor causal de neuropatías con modificaciones del trnasporte axonal / Diet glucosinolates, a possible causal factor of neuropathies with modifications of the axonal transport

Los glucosinolatos son compuestos de origen vegetal cuyos productos catabólicos, los isotiocianatos, reaccionan con los ggrupos amino de la lisina y modofocan la estructura proteica. Esta modificación a nivel de tubulina de los axones, similar a la que se produce por otras vías en las neuropatías diabéyica, alcohólica y por agentes químicos, puede fundamentar bioquímicamente un posible efecto neurotóxico por la ingestión sostenida de glucosinolatos con la dieta (AU)

Glucosinolatos de la dieta, un posible factor causal de neuropatias con modificaciones del trnasporte axonal / Diet glucosinolates, a possible causal factor of neuropathies with modifications of the axonal transport

Los glucosinolatos son compuestos de origen vegetal cuyos productos catabolicos, los isotiocianatos, reaccionan con los ggrupos amino de la lisina y modofocan la estructura proteica. Esta modificacion a nivel de tubulina de los axones, similar a la que se produce por otras vias en las neuropatias diabeyica, alcoholica y por agentes quimicos, puede fundamentar bioquimicamente un posible efecto neurotoxico por la ingestion sostenida de glucosinolatos con la dieta

Rapeseed meal-glucosinolates and their antinutritional effects. Part 5. Animal reproduction.

Although there is no clear evidence to fully describe the mechanism involved in glucosinolate-related effects on animal reproduction, lowered fertility in animals fed diets with rapeseed meal (RSM) inclusion is related to glucosinolate content in the diet. Negative effects can result both from multidirectional action of glucosinolates, malnutrition of mother due to the thyroid dysfunction, transfer of goitrogenic compounds to foetus and reduced transfer of nutritive compounds (e.g. iodine) through the placenta. The degree of reproduction impairment depends both on glucosinolate content and on the type of animal. Pregnant female rats are sensitive to the presence of glucosinolates, and the first symptoms of fertility impairment and lowering of offspring survival rate may occur at low glucosinolate levels, e.g. with diets containing low glucosinolate rapeseed meal (LG-RSM). In the case of swine, the limiting value above which sows fertility may be impaired is 4 mumol of total glucosinolates per g diet and 8 mmol of daily intake of these compounds. Opinions on the permitted RSM inclusion in diet for hens are very divergent. Some authors recommend limiting of the RSM inclusion to 10%, while others did not find any lowering of egg production in hens fed with two- or three-fold greater RSM inclusion rates. Most long-term experiments with heifers, cows and ewes indicate that ruminants tolerate the level of glucosinolate in LG-RSM although feeding this fodder as the only high-protein concentrate component may lead to impaired fertility.