Fumonisin B1 protects against long-chained polyunsaturated fatty acid-induced cell death in HepG2 cells - implications for cancer promotion.

Fumonisin B1 (FB1), a food-borne mycotoxin, is a cancer promoter in rodent liver and augments proliferation of initiated cells while inhibiting the growth of normal hepatocytes by disrupting lipid biosynthesis at various levels. HepG2 cancer cells exhibited resistance to FB1-induced toxic effects presumably due to their low content of polyunsaturated fatty acids (PUFA) even though FB1-typical lipid changes were observed, e.g. significantly increased phosphatidylethanolamine (PE), decreased sphingomyelin and cholesterol content, increased sphinganine (Sa) and sphinganine/sphingosine ratio, increased C18:1ω-9, decreased C20:4ω-6 content in PE and decreased C20:4ω-6_PC/PE ratio. Increasing PUFA content of HepG2 cells with phosphatidylcholine (PC) vesicles containing C20:4ω-6 (SAPC) or C22:6ω-3 (SDPC) disrupted cell survival, cellular redox status and induced oxidative stress and apoptosis. A partially protective effect of FB1 was evident in PUFA-enriched HepG2 cells which may be related to the FB1-induced reduction in oxidative stress and the disruption of key cell membrane constituents indicative of a resistant lipid phenotype. Interactions between different ω-6 and ω-3 PUFA, membrane constituents including cholesterol, and the glycerophospho- and sphingolipids and FB1 in this cell model provide further support for the resistant lipid phenotype and its role in the complex cellular effects underlying the cancer promoting potential of the fumonisins.

Biocontrol of Fusarium Species Utilizing Indigenous Rooibos and Honeybush Extracts.

Mycotoxins produced by several Fusarium species have a significant effect on reducing maize yield and grain quality and have led to food safety concerns. The antifungal activities of rooibos (Aspalathus linearis) and honeybush (Cyclopia species) tea extracts reduced the growth of plant pathogen Botrytis cinerea, but their efficacy against Fusarium spp. is unknown. In this study, we examined the effects of fermented and unfermented rooibos (A. linearis) and honeybush (Cyclopia subternata) aqueous extracts as well as green tea (Camellia sinensis) against 10 Fusarium species. Conidial viability was assessed by fluorescence microscopy dyes, ATP production was determined using the BacTiter-Glo assay, the mode of action was analyzed by scanning electron microscopy (SEM), and quantification of polyphenols was done using high-performance liquid chromatography with diode array detection (HPLC-DAD). Fermented rooibos extract demonstrated the highest antifungal activity (P < 0.0001) against Fusarium verticillioides MRC 826-E, Fusarium subglutinans MRC 8553, Fusarium proliferatum MRC 8549, and Fusarium globosum MRC 6647, with only 9.53%, 9.26%, 11.0%, and 12.7% ATP production, respectively, followed by antifungal activity of the fermented C. subternata extract against F. subglutinans MRC 8553, F. subglutinans MRC 8554, F. proliferatum MRC 8550, and F. verticillioides MRC 826-E with 3.79%, 6.04%, 6.04%, and 8.40% ATP production, respectively. Extract-treated conidia examined by SEM exhibited disruption of conidial hyphae and collapsed spores. Overall, the fermented rooibos and C. subternata extracts showed higher antifungal activity against the Fusarium species than the unfermented extracts. IMPORTANCE In maize subsistence farming areas in South Africa, daily consumption of maize contaminated by high level of mycotoxins contributes to long-term health effects such as immune deficiency and cancer. Biocontrol methods that are safe and cost-effective are critical to addressing this public health problem. Plant extracts known as biocides or green pesticides are alternatives to chemical pesticides due to their safety and eco-friendly properties. In South Africa, rooibos (Aspalathus linearis) and honeybush (Cyclopia species) contain polyphenols with significant antioxidant and antimicrobial properties. These indigenous herbal teas are widely available and consumed in South Africa and have potential as an innovative approach to reduce mycotoxin levels and, subsequently, human and animal exposure to these toxins. This study evaluates the efficacy of the antifungal activities of several aqueous extracts prepared from fermented and unfermented rooibos (A. linearis), honeybush (Cyclopia subternata), and green tea (Camellia sinensis) on 10 Fusarium strains.

Differential Cytotoxicity of Rooibos and Green Tea Extracts against Primary Rat Hepatocytes and Human Liver and Colon Cancer Cells - Causal Role of Major Flavonoids.

Differential anti-proliferative and pro-apoptotic effects of aqueous extracts of green rooibos (Rg; Aspalathus linearis) and green tea (GT; Camellia sinensis) and an aspalathin-enriched extract of green rooibos (GRE), were investigated in primary rat hepatocytes (PH) and human liver (HepG2) and colon (HT-29) cancer cells. Rooibos flavonoids, aspalathin and luteolin, and the green tea flavanol, epigallocatechin gallate (EGCG), were included to assess their contribution relative to their extract concentrations. GRE was the most effective in reducing cell growth parameters which was associated with a high total polyphenol content and high ferric reducing potential. Differential cell responses were noticed with HepG2 cells more sensitive than PH toward the induction of apoptosis by GRE. Luteolin induced apoptosis in PH and HepG2 cells while aspalathin lacked any effect. EGCG induced apoptosis in HepG2 cells while PH were resistant. HT-29 cells were resistant to apoptosis induction by the tea and pure flavonoids. Differences existed in the individual effects of the major rooibos and GT flavonoids against cell growth parameters compared to their equivalent concentrations in the extract mixtures. Diversity of the flavonoid constituents, physicochemical properties and cellular redox status governing cell survival are likely to explain the differential cell responses.

The protective role of liver X receptor (LXR) during fumonisin B1-induced hepatotoxicity.

Fumonisin B1 (FB1), a congener of fumonisins produced by Fusarium species, is the most abundant and most toxicologically active fumonisin. FB1 causes severe mycotoxicosis in animals, including nephrotoxicity, hepatotoxicity, and disruption of the intestinal barrier. However, mechanisms associated with FB1 toxicity are still unclear. Preliminary studies have highlighted the role of liver X receptors (LXRs) during FB1 exposure. LXRs belong to the nuclear receptor family and control the expression of genes involved in cholesterol and lipid homeostasis. In this context, the toxicity of FB1 was compared in female wild-type (LXR+/+) and LXRα,ß double knockout (LXR-/-) mice in the absence or presence of FB1 (10 mg/kg body weight/day) for 28 days. Exposure to FB1 supplemented in the mice's drinking water resulted in more pronounced hepatotoxicity in LXR-/- mice compared to LXR+/+ mice, as indicated by hepatic transaminase levels (ALT, AST) and hepatic inflammatory and fibrotic lesions. Next, the effect of FB1 exposure on the liver transcriptome was investigated. FB1 exposure led to a specific transcriptional response in LXR-/- mice that included altered cholesterol and bile acid homeostasis. ELISA showed that these effects were associated with an elevated FB1 concentration in the plasma of LXR-/- mice, suggesting that LXRs participate in intestinal absorption and/or clearance of the toxin. In summary, this study demonstrates an important role of LXRs in protecting the liver against FB1-induced toxicity, suggesting an alternative mechanism not related to the inhibition of sphingolipid synthesis for mycotoxin toxicity.

The mutagenic and antimutagenic activity of Sutherlandia frutescens extracts and marker compounds.

BACKGROUND: Sutherlandia frutescens (L.) R. Br is endemic to Southern Africa where it has been traditionally used for cancer and diabetes. In recent times it has been marketed for its reputed (but not proven) anticancer, antidiabetic and anti-HIV properties. Little is known about the mutagenic and antimutagenic potential of extracts and common marker compounds of Sutherlandia frutescens. Therefore this study aimed to investigate the putative efficacy and possible long-term adverse effects of using this herb. METHODS: Ethylacetate (EA) and 50% Methanol (MeOH) extracts were screened for mutagenic and antimutagenic activity using the Ames assay utilising TA97a, TA98, TA100 and TA102 in the presence and absence of metabolic activation. Four compounds, L-arginine, L-canavanine, GABA and D-pinitol known to occur in sutherlandia were also included. The total polyphenolic content of the both extracts was determined using the Folin-Ciocalteau method and FRAP and ABTS were used to determine the anti-oxidant potential of the extracts. RESULTS: The extracts and the standards did not show any cytotoxicity except in TA97a. The EA extract exhibited antimutagenicity against all the bacterial strains at all concentrations tested. The MeOH extract showed both pro-mutagenic and antimutagenic activities with 2-acetamidofluorene and aflatoxin B1 in the presence of metabolic activation of TA98 and TA100, respectively. All compounds, except L-canavanine exhibited antimutagenic activity against all strains. L-canavanine, on the other hand showed co-mutagenicity with 9-aminoacridine on TA97a, at all test concentrations. The extracts and pure compounds exhibited their antimutagenic activity in a dose response manner. L-arginine and GABA showed an some antimutagenic response. EA extract had three times the total phenolic content (12.56 µg GE / mg) observed in the MeOH extract. There was correlation between total phenolic content, antioxidant potential and antimutagenicity. CONCLUSION: Both extracts exhibited a protective effect, with the EA extract exhibiting greater potency. L-canavanine acted as a co-mutagen in a dose response manner without metabolic activation. It is suggested that the EA extract be priotized for future development work as it showed a better risk profile and activity.

Modulation of pre-neoplastic biomarkers induced by sequential aflatoxin B1 and fumonisin B1 exposure in F344 rats treated with UPSN clay.

Populations consuming aflatoxin (AF) and fumonisin (FN)-contaminated foods may be at increased risk for hepatocellular carcinoma (HCC) and developmental disorders; consequently, development of intervention strategies to reduce AF/FN-induced liver disease and adverse health effects in humans could be very useful. Calcium montmorillonite clay (NovaSil) has been shown to absorb AF in vitro, in multiple animal models, as well as in human studies. In the present study, we aimed to evaluate whether uniform particle size NovaSil (UPSN) possessed an ability to modulate the co-carcinogenic potentials of aflatoxin B1 (AFB1) and fumonisin B1 (FB1) in F344 rats. Sequential treatment of FB1 following AFB1 synergistically induces preneoplastic alterations as well as liver damage, indicating that AFB1 acts as an initiator while FB1 as a promoter in the carcinogenesis model, confirming findings from previous studies. The enterosorbent agent UPSN clay at dose of up to 0.5% in diet was shown to be effective in modulating the toxicity and carcinogenicity of co-exposure to AFB1 and FB1, as demonstrated by significant reduction in number and size of hepatic GST-P+ foci, in alterations indicative of liver toxicity, and in levels of AFB1 and FB1 biomarkers.

In Vitro Chemopreventive Properties of Green Tea, Rooibos and Honeybush Extracts in Skin Cells.

The chemopreventive properties of the herbal teas rooibos (Aspalathus linearis) and honeybush (Cyclopia spp.) have been demonstrated on mouse skin in vivo but the underlying mechanisms are not clear. The aim of the current study was to determine the anti-proliferative and pro-apoptotic activity of methanol and aqueous extracts of rooibos and two Cyclopia species in different skin cells, using green tea (Camellia sinensis) as a benchmark. Extracts were also characterised for their major individual polyphenols by high performance liquid chromatography and spectroscopically for the total polyphenol (TP) groups. The methanol extract of rooibos, containing higher levels of polyphenols than its aqueous extract, displayed similar activity to green tea as it selectively targeted premalignant cells by inhibiting cell proliferation at lower concentrations whilst inducing apoptosis via membrane depolarisation at higher concentrations. Specific roles of the major rooibos dihydrochalcones and flavanol/proanthocyanidin-type (FLAVA) compounds are likely to be involved. The aqueous extracts of the Cyclopia species were more active against cell proliferation and at inducing apoptosis which was associated with a higher FLAVA content and a reduced TP/FLAVA ratio. In contrast, their methanol extracts exhibited a cytoprotective effect against apoptosis which was related to their monomeric xanthone and flavanone content. The underlying chemopreventive properties of green tea and the herbal teas appear to be associated with diverse and complex monomeric/polymeric polyphenolic cell interactions.

Return-to-play rates following arthroscopic treatment of femoroacetabular impingement in competitive baseball players.

OBJECTIVE: Femoroacetabular impingement (FAI) has been increasingly recognized in cutting sports including soccer, hockey and football. More recently, the prevalence among overhead athletes has also been recognized. The purpose of this study was to review impingement patterns, return-to-play rates and clinical outcome following arthroscopic treatment of FAI among high-level baseball players. METHODS: Between 2010 and 2014, 70 competitive baseball players (86 hips; age 22.4 ± 4.5 years) were identified. Demographics and return-to-play rates were recorded. Patient-reported outcome scores, including the Modified Harris Hip Score (mHHS), the Hip Outcome Score-Activity of Daily Living (HOS-ADL), the Sport-specific Subscale (HOS-SSS), and the International Hip Outcome Tool (iHOT-33), were collected pre-operatively at 6 months and 1year (n = 34, 49% of cohort). RESULTS: The cohort included professional (27.1%), college (57.1%), high-school (8.6%) and club-team athletes (7.1%). Infielder (37.5%), pitcher (22.9%) and catcher (16.7%) were the most common positions. Average follow-up was 16.8 months (range 12.1-34.2). There was no relationship between playing position and impingement pattern (p ≥ 0.459), or between symptom laterality and handedness, batting position or playing position (p ≥ 0.179). One patient required revision surgery (infection). Return to sport rate was 88%, at a mean of 8.6 ± 4.2 months, with 97.7% returning at/above their pre-injury level of play. There was significant improvement in all outcome measures: mHHS (60.1 ± 11.9 to 93 ± 9.5), HOS-ADL (71.3 ± 16.7 to 96.3 ± 3.6), HOS-SSS (51.3 ± 24.8 to 92.3 ± 8.2) and iHOT-33 (40.7 ± 19.9 to 85.9 ± 14) (p < 0.001). CONCLUSION: Arthroscopic treatment of FAI in competitive baseball players resulted in high return-to-play rates at short-term follow-up, with significant improvements in clinical outcome scores.

Active control of the visual field in the starfish Acanthaster planci.

Photoreception in echinoderms has been studied for several years with a focus on the dermal photoreceptors of echinoids. Even though spatial vision has been proposed for this dermal photosystem, by far the most advanced system is found in a number of asteroids where an unpaired tube foot at the tip of each arm carries a proper eye, also known as the optical cushion. The eyes resemble compound eyes, except for the lack of true optics, and they typically have between 50 and 250 ommatidia each. These eyes have been known for two centuries but no visually guided behaviors were known in starfish until recently when it was shown that both Linckia laevigata and Acanthaster planci navigate their coral reef habitat using vision. Here we investigate the visual system of A. planci and find that they have active control of their visual field. The distalmost tube foot holding the eye is situated on a movable knob, which bends to adjust the vertical angle of the visual field. On the leading arms the visual field is directed 33° above the horizon, whereas the eyes on the trailing arms are directed 44° above horizontal on average. When the animal traverses an obstacle the knob bends and counteracts most of the arm bending. Further, we examined a previously described behavior, rhythmic arm elevation, and suggest that it allows the animal to scan the surroundings while preventing photoreceptor adaptation and optimizing image contrast.

Sequential dietary exposure to aflatoxin B1 and fumonisin B1 in F344 rats increases liver preneoplastic changes indicative of a synergistic interaction.

Dietary co-exposure to aflatoxin B1 (AFB1) and fumonisin B1 (FB1) and their interaction on hepatocellular carcinogenesis is of particular concern in toxicology and public health. In this study we evaluated the liver preneoplastic effects of single and sequential dietary exposure to AFB1 and FB1 in the F344 rat carcinogenesis model. Serum biochemical alterations, liver histopathological changes, and the formation of liver glutathione S transferase positive (GST-P+) foci were the major outcome parameters examined. Compared to the AFB1-only treatment, the FB1-only treatment induced less dysplasia, and more apoptosis and mitoses. Sequential AFB1 and FB1 treatment lead to increased numbers of dysplasia, apoptosis and foci of altered hepatocytes, as compared to either mycotoxin treatment alone. More importantly, sequential exposure to AFB1 and FB1 synergistically increased the numbers of liver GTP-P+ foci by approximately 7.3-and 12.9-fold and increased the mean sizes of GST-P+ foci by 6- and 7.5-fold, respectively, as compared to AFB1- or FB1-only treatment groups. In addition, liver ALT and AST levels were significantly increased after sequential treatment as compared to single treatment groups. The results demonstrate the interactive effect of dietary AFB1 and FB1 in inducing liver GST-P+ foci formation and provide information to model future intervention studies.

Correction: Van der Merwe, J.D., et al. Short-Term and Sub-Chronic Dietary Exposure to Aspalathin-Enriched Green Rooibos (Aspalathus linearis) Extract Affects Rat Liver Function and Antioxidant Status. Molecules 2015, 20, 22674-22690.

The authors wish to make the following corrections to their published paper [1].[...].

Biologically Based Methods for Control of Fumonisin-Producing Fusarium Species and Reduction of the Fumonisins.

Infection by the fumonisin-producing Fusarium spp. and subsequent fumonisin contamination of maize adversely affect international trade and economy with deleterious effects on human and animal health. In developed countries high standards of the major food suppliers and retailers are upheld and regulatory controls deter the importation and local marketing of fumonisin-contaminated food products. In developing countries regulatory measures are either lacking or poorly enforced, due to food insecurity, resulting in an increased mycotoxin exposure. The lack and poor accessibility of effective and environmentally safe control methods have led to an increased interest in practical and biological alternatives to reduce fumonisin intake. These include the application of natural resources, including plants, microbial cultures, genetic material thereof, or clay minerals pre- and post-harvest. Pre-harvest approaches include breeding for resistant maize cultivars, introduction of biocontrol microorganisms, application of phenolic plant extracts, and expression of antifungal proteins and fumonisin degrading enzymes in transgenic maize cultivars. Post-harvest approaches include the removal of fumonisins by natural clay adsorbents and enzymatic degradation of fumonisins through decarboxylation and deamination by recombinant carboxylesterase and aminotransferase enzymes. Although, the knowledge base on biological control methods has expanded, only a limited number of authorized decontamination products and methods are commercially available. As many studies detailed the use of natural compounds in vitro, concepts in reducing fumonisin contamination should be developed further for application in planta and in the field pre-harvest, post-harvest, and during storage and food-processing. In developed countries an integrated approach, involving good agricultural management practices, hazard analysis and critical control point (HACCP) production, and storage management, together with selected biologically based treatments, mild chemical and physical treatments could reduce fumonisin contamination effectively. In rural subsistence farming communities, simple, practical, and culturally acceptable hand-sorting, maize kernel washing, and dehulling intervention methods proved to be effective as a last line of defense for reducing fumonisin exposure. Biologically based methods for control of fumonisin-producing Fusarium spp. and decontamination of the fumonisins could have potential commercial application, while simple and practical intervention strategies could also impact positively on food safety and security, especially in rural populations reliant on maize as a dietary staple.

Simultaneous Versus Staged Bilateral Hip Arthroscopy in the Treatment of Femoroacetabular Impingement.

PURPOSE: To compare the clinical outcomes and complication rates of patients undergoing simultaneous versus staged bilateral hip arthroscopy for bilateral symptomatic femoroacetabular impingement (FAI). METHODS: Between 2010 and 2013, a total of 1,800 hip arthroscopy cases were retrospectively reviewed for cases of simultaneous bilateral hip arthroscopy. All patients with minimum 1-year follow-up were included. This group was matched 1:2 for age, sex, and alpha angle, to a control group of patients who underwent staged, bilateral procedures. Patient-reported outcome scores, including the Modified Harris Hip Score (mHHS), the Hip Outcome Score-Activity of Daily Living (HOS-ADL), and the Hip Outcome Score-Sport-Specific Subscale (HOS-SSS) were obtained preoperatively at 6 months and 1 and 2 years postoperatively. RESULTS: Eighty-one patients (162 hips) were identified who underwent bilateral hip arthroscopy for symptomatic FAI. Twelve patients (24 hips) who underwent simultaneous bilateral arthroscopy with minimum 1-year follow-up were compared with a matched cohort of 24 patients (48 hips) who underwent staged bilateral procedures. Mean preoperative alpha angle was 65.3° ± 9.6° in the simultaneous group and 65.9° ± 11.2° in the staged group (P = .6). At a mean follow-up of 17.8 months (range, 12 to 33 months), there were comparable improvements in simultaneous versus staged patient-reported outcome scores (mHHS 90.8 ± 11 v 88.9 ± 12.5, P = .662; HOS-ADL 97.3 ± 3.8 v 92.6 ± 10.3, P = .057; HOS-SSS 93.3 ± 10.2 v 86.5 ± 16.6, P = .203). The mean single anesthetic traction time was 90.8 ± 21.9 minutes (sum of both hips) in the simultaneous group, compared with a combined 2-anesthetic traction time of 85.7 ± 27.2 minutes in the staged group (P = .579). There were no traction-related complications in either group. No patients in the simultaneous group required revision surgery, whereas 1 patient in the staged group required lysis of adhesions at 24 months postoperatively. CONCLUSIONS: In a small sample, simultaneous bilateral hip arthroscopy is shown to be safe and effective, resulting in similar improvements in patient-reported outcomes at 1-year follow-up compared with staged bilateral procedures. LEVEL OF EVIDENCE: Level III, case-control study.

Short-Term and Sub-Chronic Dietary Exposure to Aspalathin-Enriched Green Rooibos (Aspalathus linearis) Extract Affects Rat Liver Function and Antioxidant Status.

An aspalathin-enriched green rooibos (Aspalathus linearis) extract (GRE) was fed to male Fischer rats in two independent studies for 28 and 90 days. The average dietary total polyphenol (TP) intake was 756 and 627 mg Gallic acid equivalents (GAE)/kg body weight (bw)/day over 28 and 90 days, respectively, equaling human equivalent doses (HEDs) of 123 and 102 GAE mg/kg bw/day. Aspalathin intake of 295 mg/kg bw/day represents a HED of 48 mg/kg bw/day (90 day study). Consumption of GRE increased feed intake significantly (p < 0.05) compared to the control after 90 days, but no effect on body and organ weight parameters was observed. GRE significantly (p < 0.05) reduced serum total cholesterol and iron levels, whilst significantly (p < 0.05) increasing alkaline phosphatase enzyme activity after 90 days. Endogenous antioxidant enzyme activity in the liver, i.e., catalase and superoxide dismutase activity, was not adversely affected. Glutathione reductase activity significantly (p < 0.05) increased after 28 days, while glutathione (GSH) content was decreased after 90 days, suggesting an altered glutathione redox cycle. Quantitative Real Time polymerase chain reaction (PCR) analysis showed altered expression of certain antioxidant defense and oxidative stress related genes, indicative, among others, of an underlying oxidative stress related to changes in the GSH redox pathway and possible biliary dysfunction.

Development and evaluation of a sensitive mycotoxin risk assessment model (MYCORAM).

The differential risk of exposure to fumonisin (FB), deoxynivalenol (DON), and zearalenone (ZEA) mycotoxins to the South African population, residing in the nine Provinces was assessed during a cross-sectional grain consumer survey. The relative per capita maize intake (g/day) was stratified by gender, ethnicity, and Province and the probable daily intake (PDI) for each mycotoxin (ng/kg body weight/day) calculated utilizing SPECIAL and SUPER dry milled maize fractions representing different exposure scenarios. Men consumed on an average more maize (173 g/day) than women (142 g/day) whereas the black African ethnic group had the highest intake (279 g/day) followed by the Colored group (169 g/day) with the Asian/Indian and White groups consuming lower quantities of 101 and 80 g/day, respectively. The estimated mean PDIs for the various subgroups and Provinces, utilizing the different dry milled maize fractions, were below the provisional maximum tolerable daily intake (PMTDI) for each mycotoxin. A distinct and more sensitive mycotoxin risk assessment model (MYCORAM) for exposure, stratified by Province and ethnicity were developed utilizing specific maize intake increments (g/kg body weight/day) that provides information on the percentage of the population exposed above the PMTDI for each mycotoxin. Evaluation of the MYCORAM utilizing commercial and EXPERIMENTALLY DERIVED: SPECIAL milling fractions, containing predefined mycotoxins levels, predicts the percentage of maize consumers exposed above the respective PMTDI. Safety modeling using the MYCORAM could also predict a maximum tolerated level adequate to safeguard all South African maize consumers including the most vulnerable groups.

Calcium montmorillonite clay reduces AFB1 and FB1 biomarkers in rats exposed to single and co-exposures of aflatoxin and fumonisin.

Aflatoxins (AFs) and fumonisins (FBs) can co-contaminate foodstuffs and have been associated with hepatocellular and esophageal carcinomas in humans at high risk for exposure. One strategy to reduce exposure (and toxicity) from contaminated foodstuffs is the dietary inclusion of a montmorillonite clay (UPSN) that binds AFs and FBs in the gastrointestinal tract. In this study, the binding capacity of UPSN was evaluated for AFB1, FB1 and a combination thereof in Fischer 344 rats. Rats were pre-treated with different dietary levels of UPSN (0.25% or 2%) for 1 week. Rats were gavaged with a single dose of either 0.125 mg AFB1 or 25 mg FB1 per kg body weight and a combination thereof in the presence and absence of an aqueous solution of UPSN. The kinetics of mycotoxin excretion were monitored by analyzing serum AFB1 -albumin, urinary AF (AFM1) and FB1 biomarkers over a period of 72 h. UPSN decreased AFM1 excretion by 88-97%, indicating highly effective binding. FB1 excretion was reduced, to a lesser extent, ranging from 45% to 85%. When in combination, both AFB1 and FB1 binding occurred, but capacity was decreased by almost half. In the absence of UPSN, the combined AFB1 and FB1 treatment decreased the urinary biomarkers by 67% and 45% respectively, but increased levels of AFB1 -albumin, presumably by modulating its cytochrome metabolism. UPSN significantly reduced bioavailability of both AFB1 and FB1 when in combination; suggesting that it can be utilized to reduce levels below their respective thresholds for affecting adverse biological effects.

The influence of Aspalathus linearis (Rooibos) and dihydrochalcones on adrenal steroidogenesis: quantification of steroid intermediates and end products in H295R cells.

The steroid hormone output of the adrenal gland is crucial in the maintenance of hormonal homeostasis, with hormonal imbalances being associated with numerous clinical conditions which include, amongst others, hypertension, metabolic syndrome, cardiovascular disease, insulin resistance and type 2 diabetes. Aspalathus linearis (Rooibos), which has been reported to aid stress-related symptoms linked to metabolic diseases, contains a wide spectrum of bioactive phenolic compounds of which aspalathin is unique. In this study the inhibitory effects of Rooibos and the dihydrochalcones, aspalathin and nothofagin, were investigated on adrenal steroidogenesis. The activities of both cytochrome P450 17α-hydroxylase/17,20 lyase and cytochrome P450 21-hydroxylase were significantly inhibited in COS-1 cells. In order to study the effect of these compounds in H295R cells, a human adrenal carcinoma cell line, a novel UPLC-MS/MS method was developed for the detection and quantification of twenty-one steroid metabolites using a single chromatographic separation. Under both basal and forskolin-stimulated conditions, the total amount of steroids produced in H295R cells significantly decreased in the presence of Rooibos, aspalathin and nothofagin. Under stimulated conditions, Rooibos decreased the total steroid output 4-fold and resulted in a significant reduction of aldosterone and cortisol precursors. Dehydroepiandrosterone-sulfate levels were unchanged, while the levels of androstenedione (A4) and 11ß-hydroxyandrostenedione (11ßOH-A4) were inhibited 5.5 and 2.3-fold, respectively. Quantification of 11ßOH-A4 showed this metabolite to be a major product of steroidogenesis in H295R cells and we confirm, for the first time, that this steroid metabolite is the product of the hydroxylation of A4 by human cytochrome P450 11ß-hydroxylase. Taken together our results demonstrate that Rooibos, aspalathin and nothofagin influence steroid hormone biosynthesis and the flux through the mineralocorticoid, glucocorticoid and androgen pathways, thus possibly contributing to the alleviation of negative effects arising from elevated glucocorticoid levels.

Mangiferin glucuronidation: important hepatic modulation of antioxidant activity.

Mangiferin displays an extensive spectrum of pharmacological properties, including antioxidant activity. Its phase II metabolism in the presence of Aroclor 1254-induced and un-induced microsomal and cytosolic fractions from rat liver and the antioxidant potency of the glucuronidated conjugates were investigated. Mangiferin was not a substrate for the cytosolic sulphotransferases. Glucuronidation led to the formation of two monoglucuronidated metabolites of mangiferin and a monoglucuronidated metabolite of homomangiferin (a minor constituent of the mangiferin standard). Deconjugation utilising glucuronidase resulted in the disappearance of the metabolites, with the concomitant formation of the two parent compounds. Considering steric hinderance caused by the C-2 glucosyl moiety and the relative acidity of the xanthone OH groups, the 6-OH of mangiferin and, to a lesser degree the 7-OH, are likely to be the primary glucuronidation targets. The ferric iron reducing ability of the glucuronidated reaction mixture was reduced, while the free radical scavenging abilities of mangiferin, utilising on-line post-column HPLC-DAD-DPPH· and HPLC-DAD-ABTS·+ assays, were eliminated, providing further evidence that the catechol arrangement at C-6 and C-7 was the preferred site of conjugation. This paper provides the first evidence that the glucuronidated metabolites of mangiferin resulted in a loss in free radical scavenging and ferric iron reducing ability.

Fumonisin B1 as a urinary biomarker of exposure in a maize intervention study among South African subsistence farmers.

BACKGROUND: The consumption of maize highly contaminated with carcinogenic fumonisins has been linked to high oesophageal cancer rates. The aim of this study was to validate a urinary fumonisin B1 (UFB1) biomarker as a measure of fumonisin exposure and to investigate the reduction in exposure following a simple and culturally acceptable intervention. METHODS: At baseline home-grown maize, maize-based porridge, and first-void urine samples were collected from female participants (n=22), following their traditional food practices in Centane, South Africa. During intervention the participants were trained to recognize and remove visibly infected kernels, and to wash the remaining kernels. Participants consumed the porridge prepared from the sorted and washed maize on each day of the two-day intervention. Porridge, maize, and urine samples were collected for FB1 analyses. RESULTS: The geometric mean (95% confidence interval) for FB1 exposure based on porridge (dry weight) consumption at baseline and following intervention was 4.84 (2.87-8.14) and 1.87 (1.40-2.51) µg FB1/kg body weight/day, respectively, (62% reduction, P<0.05). UFB1C, UFB1 normalized for creatinine, was reduced from 470 (295-750) at baseline to 279 (202-386) pg/mg creatinine following intervention (41% reduction, P=0.06). The UFB1C biomarker was positively correlated with FB1 intake at the individual level (r=0.4972, P<0.01). Urinary excretion of FB1 was estimated to be 0.075% (0.054%-0.104%) of the FB1 intake. CONCLUSION: UFB1 reflects individual FB1 exposure and thus represents a valuable biomarker for future fumonisin risk assessment. IMPACT: The simple intervention method, hand sorting and washing, could positively impact on food safety and health in communities exposed to fumonisins.

Interstitial Deletions at 6q14.1-q15 Associated with Obesity, Developmental Delay and a Distinct Clinical Phenotype.

BACKGROUND: Interstitial deletions of the long arm of chromosome 6 have been described in several patients with obesity and a Prader-Willi-like phenotype. Haploinsufficiency of the SIM1 gene located at 6q16.3 is suggested as being responsible for the regulation of body weight. Here we report on 2 patients with interstitial deletions at 6q14.1-q15 presenting with obesity and symptoms strikingly similar to those reported for deletions involving the SIM1 gene despite not having a deletion of this gene. METHODS: Array comparative genomic hybridisation was used to diagnose 2 children with obesity and developmental delay, revealing 2 interstitial deletions at 6q14.1-q15 of 8.73 and 4.50 Mb, respectively, and a region of overlap of 4.2-Mb. RESULTS: The similar phenotype in the 2 patients was most likely due to a 4.2-Mb common microdeletion at 6q14.1-q15. Another patient has previously been described with an overlapping deletion. The 3 patients share several features, such as developmental delay, obesity, hernia, rounded face with full cheeks, epicanthal folds, short palpebral fissures, bulbous nose, large ears, and syndactyly between toes II and III. CONCLUSIONS: Together with a previously reported patient, our study suggests that the detected deletions may represent a novel clinically recognisable microdeletion syndrome caused by haploinsufficiency of dosage-sensitive genes in the 6q14.1-q15 region.