Phenethyl isothiocyanate inhibits hypoxia-induced accumulation of HIF-1α and VEGF expression in human glioma cells

Phenethyl isothiocyanate (PEITC), a natural dietary isothiocyanate, inhibits angiogenesis but the molecular mechanisms that underlie this effect are not known. In this study, under hypoxic conditions (1% O₂), we examined the effect of PEITC on the intracellular level of the hypoxia inducible factor (HIF-1α) and extracellular level of the vascular endothelial growth factor (VEGF) in a variety of human cancer cell lines. Surprisingly, we observed that PEITC suppressed the HIF-1α accumulation during hypoxia in human glioma U87, human prostate cancer DU145, colon cancer HCT116, liver cancer HepG2, and breast cancer SkBr3 cells. PEITC treatment also significantly reduced the hypoxia-induced secretion of VEGF. Suppression of HIF-1α accumulation during treatment with PEITC in hypoxia was related to PI3K and MAPK pathways. Taken together, these results suggest that PEITC inhibits the HIF-1α expression through inhibiting the PI3K and MAPK signalling pathway and provide a new insight into a potential mechanism of the anticancer properties of PEITC.     

Effect of supplementation with vitamin D₃ on glucose production pathways in human subjects

Scope: Research reports suggest that vitamin D affects glucose and insulin metabolism; however, the exact mechanisms are unclear. ²H NMR analysis of monoacetone glucose (MAG) after tracer administration provides a non‐invasive method of profiling hepatic glucose metabolism. This study examined the effects of supplementation with vitamin D₃ on contribution of glycogenolysis to glucose production. Methods and results: Tracer administration and biofluid collections were performed with eight healthy females before and following a 4‐wk vitamin D₃ administration period. Following an overnight fast subjects ingested deuterated water and acetaminophen. Full void urine samples were collected after 4 h. ²H NMR spectra of urinary monoacetone glucose were acquired to determine the contribution of glycogenolysis to glucose production. The mean contribution of glycogenolysis to glucose production was 60±13%. Supplementation with vitamin D₃ had no effect on hepatic glucose production. Regression analysis revealed a significant relationship between carbohydrate intake and the contribution of glycogenolysis (β=0.914, p=0.004). Conclusion: In conclusion, we saw no changes in the percentage contribution of glycogenolysis following supplementation with vitamin D₃. The reproducibility of our results and the non‐invasive nature of the method highlight the potential for this method in assessing mechanistic modes of action in future nutritional interventions.     

Effects of mixed feeder cells on the expansion of CD34⁺ cells

Synergistic effects of mesenchymal stem cells (MSCs) isolated from bone marrow (BM), umbilical cord blood (UCB) and periosteum, and fibroblasts as mixed feeder cells (MFCs) on the expansion of hematopoietic progenitor cells (HPCs) were investigated in serum- and exogenous cytokine-free conditions. Enriched CD34(+) cells were cultured for 2weeks over the cell lines alone, individually, or selected combinations of them. When the cells were cultured over MFCs, the maximum increase in expansion of total nucleated cells and CD34(+)/CD38(-) cells was 157.3- and 128.6-fold, respectively. Furthermore, hematopoietic cytokine such as IL-6 and chemokines (e.g., IL-8, growth related oncogene (GRO), GRO-alpha, matrix metalloproteinase (MMP)-1, and MMP-3) were significantly increased in mixed feeder cells. Based on these results, MFCs can be more efficient for the ex vivo expansion of HPCs. These results strongly suggest that MFCs are more suitable for HPCs mass production.     

Effects of quercetin metabolites on the enhancing effect of β-carotene on DNA damage and cytochrome P1A1/2 expression in benzo[a]pyrene-exposed A549 cells

A549 cells were pre-incubated with β-carotene (BC) alone or in combination with quercetin or three major quercetin metabolites in human plasma, quercetin 3-glucuronide (Q3G), quercetin 3′-sulphate (Q3′S) and isorhamnetin, followed by incubation with benzo[a]pyrene (BaP), to investigate the effects of these compounds on the BaP-induced harmful effects of BC. All the quercetin metabolites at 10 μM inhibited BaP + BC-induced cell death. Q3′S, Q3G and isorhamnetin also significantly decreased BaP ± BC-induced DNA damage by 64%, 60% and 24%, respectively. In a similar order, these compounds suppressed BaP + BC-induced cytochrome P450 (CYP)1A1/1A2 expression by 10–50%. Q3G and Q3′S significantly decreased the intracellular reactive oxygen species formation induced by BaP + BC; however, Q3G had the best effect on decreasing the loss of BC induced by Fe/NTA. The combined effects of quercetin metabolites were additive. This study indicates that quercetin metabolites decrease the BaP-induced harmful effect of β-carotene in A549 cells by downregulating the expression of CYP1A1/1A2, at least in part.     

Effects of engineered methionine in the 8Sα globulin of mungbean on its physicochemical and functional properties and potential nutritional quality

The major storage protein of mungbean, 8Sα globulin or vicilin, was engineered using site-directed mutagenesis to increase the number of methionine (Met) residues in the molecule for improvement of functional and nutritional qualities. Eight Met-rich proteins were designed and prepared to have 2 to 10 Met residues introduced in disordered regions II and IV. The designed proteins were highly expressed as soluble form in Escherichia coli. Their production level of the modified proteins was estimated to be about 30%, and was almost the same as that of 8Sα globulin wild type (WT). The modified proteins formed stable native conformation similar to WT as shown by gel filtration chromatography. They demonstrated greater stability in terms of thermal denaturation temperature and greater emulsifying ability and emulsion stability, especially the 10-Met protein, compared to the wild type. Met-rich proteins with 3, 5, and 10 Met residues had 74, 96, and 145% of nutritional requirement for Met compared with 41% for WT. Based on allergenicity prediction programs, WT and all the modified proteins had no allergenic potential.     

Effects of nutritional level on pork quality and gene expression of μ-calpain and calpastatin in muscle of finishing pigs

The study was designed to investigate the effects of nutritional level (control diet (CD), 14.19% crude protein, 13.81 MJ of DE/kg; low nutritional level diet (LND), 11.08% crude protein, 12.55 MJ of DE/kg) on pork quality and gene expression of μ-calpain and calpastatin in muscle of finishing pigs. The LND treatment increased drip loss (P < 0.05), had a trend to increase intramuscular fat (IMF) content (P = 0.09), decreased Warner–Bratzler shear force (WBSF) of pork (P < 0.05), improved mRNA level of μ-calpain (P < 0.05) in skeletal muscle, but had no effect on gene expression of calpastatin, compared with the CD treatment. These data suggest that a moderately reduced energy and protein diet increased pork tenderness and intramuscular fat. The increase in tenderness by LND treatment may be partly due to increased gene expression of μ-calpain in muscle.     

Effects of processing and packaging on vitamin C and β-carotene content of ready-to-use (RTU) vegetables

High pressure liquid chromatographic methods were used for measurement of the concentration of vitamin C and β-carotene in broccoli and green pepper. The effects of processing, packaging and storage on the levels of these nutrients in both unprocessed and processed ready-to-use (RTU) vegetables were determined. Systems investigated included: (a) unpacked and pillow packaged broccoli, and (b) unpacked, pillow, partial vacuum, and total vacuum packaged green pepper. There was a statistically significant decrease in vitamin C over a 10 day storage period of unpacked and packaged vegetables including all four packaging systems (P<0.001, overall average decrease of 11%). The overall loss of β-carotene during the 10 day storage period was not statistically significant (P=0.14). Although there was a significant loss in vitamin C during storage, in most cases there was no difference in loss of vitamin C or β-carotene between the processed and unprocessed vegetables, and the packaging systems.     

Stabilizing vitamin D3 using the molten globule state of α-lactalbumin

α-Lactalbumin (α-LA) is the second most abundant bovine whey protein. It has been intensively studied because of its readiness to populate the molten globular (MG) state, a partially folded state with native levels of secondary structure but loss of tertiary structure. The MG state of α-LA exposes a significant number of hydrophobic patches that could be used to bind and stabilize small hydrophobic molecules such as vitamin D₃ (vitD). Accordingly, we tested the ability of α-LA to stabilize vitD in a pH interval from 7.4 to 2; over this pH interval, α-LA transitions from the folded state to the MG state. The MG state stabilized vitD better than the folded state and was superior to the major bovine whey protein β-lactoglobulin (β-LG), which is known to stabilize vitD. At pH 7.4, β-LG and α-LA stabilized vitD to the same extent. Tryptophan fluorescence quenching measurements indicated that α-LA has one binding site at pH 7.4 but acquires an additional binding site when the pH is lowered to pH 2 to 4. Stability measurements of the vitD in the α-LA–vitD complex at different temperatures suggest that UHT processing would lead to little loss of vitD. This study demonstrates the potential of α-LA as a component in vitD fortification, particularly for low pH applications.     

Effects of prepartum 2,4-thiazolidinedione on insulin sensitivity, plasma concentrations of tumor necrosis factor-α and leptin, and adipose tissue gene expression

Administration of peroxisome proliferator-activated receptor gamma (PPARγ) ligands, thiazolidinediones (TZD), to prepartum dairy cattle has been shown to improve dry matter intake and decrease circulating nonesterified fatty acids (NEFA) around the time of calving. The objective of this work was to elucidate mechanisms of TZD action in transition dairy cattle by investigating changes in plasma leptin, tumor necrosis factor-α (TNFα), the revised quantitative insulin sensitivity check index (RQUICKI), and adipose tissue gene expression of leptin, PPARγ, lipoprotein lipase (LPL), and fatty acid synthase (FAS). Multiparous Holstein cows (n = 40) were administered 0, 2.0, or 4.0 mg of TZD/kg of body weight (BW) by intrajugular infusion once daily from 21 d before expected parturition until parturition. Plasma samples collected daily from 22 d before expected parturition through 21 d postpartum were analyzed for glucose, NEFA, and insulin. Plasma samples collected on d −14, −3, −1, 1, 3, 7, 14, and 49 relative to parturition were also analyzed for leptin and TNFα. Adipose tissue was collected on d 7 before expected parturition from a subset of cows, and gene expression was examined via quantitative real-time PCR. A tendency for a treatment by time effect on plasma leptin prepartum was observed such that values were similar on d −14 but cows receiving 2.0 mg/kg of BW of TZD tended to have lower circulating leptin as calving approached. Postpartum leptin tended to be increased linearly (2.3, 2.4, and 2.5 ± 0.1 ng/mL for 0, 2.0, and 4.0 mg/kg treatments, respectively) in cows that received TZD prepartum. Plasma TNFα increased linearly (2.6, 3.7, and 4.0 ± 0.1 pg/mL) in response to TZD treatment and decreased through the first week postpartum. Calculation of RQUICKI 1/[log(glucose) + log(insulin) + log(NEFA)] suggested altered insulin sensitivity in cows administered TZD that may depend on day relative to calving. Administration of TZD increased adipose tissue expression of PPARγ mRNA (11.0, 13.3, and 12.8 ± 1.9). Administration of TZD had a quadratic effect on gene expression of leptin (16.2, 10.7, and 17.4 ± 1.6) and no effect on LPL expression, and expression of FAS was lower for TZD-treated cows than for controls (8.2, 4.2, and 6.1 ± 1.8, respectively). Results imply altered expression and plasma concentrations of leptin, increased plasma TNFα concentrations, and increased expression of PPARγ in adipose tissue as potential mechanisms for the effects of TZD administration on transition dairy cattle.     

Effects of the peroxisome proliferator-activated receptor-α agonists clofibrate and fish oil on hepatic fatty acid metabolism in weaned dairy calves

Peroxisome proliferator-activated receptor-α (PPARα) agonists increase fatty acid oxidation in liver of nonruminants. If similar effects occur in dairy cattle, enhanced hepatic oxidative capacity could decrease circulating nonesterified fatty acids and hepatic triacylglycerol accumulation in periparturient cows. The objectives of this study were 1) to determine whether partitioning of fatty acid metabolism by liver slices from weaned Holstein calves treated with PPARα agonists in vivo is altered compared with partitioning by liver slices from control (untreated) calves, and 2) to measure in vitro metabolism of palmitate and oleate by bovine liver slices and relate these to mRNA abundance for key enzymes. Weaned male Holstein calves (7 wk old; n = 15) were assigned to 1 of 3 groups for a 5-d treatment period: control (untreated), clofibrate (62.5 mg/kg of BW), or fish oil (250 mg/kg of BW). Calves treated with clofibrate consumed less dry matter. Body weight, liver weight, liver weight:body weight ratio, blood nonesterified fatty acids, β-hydroxybutyrate, and liver composition were not significantly different among treatments. Liver slices were incubated for 2, 4, and 8 h to determine in vitro conversion of [1-¹⁴C] palmitate and [1-¹⁴C] oleate to CO₂, acid-soluble products, esterified products, and total metabolism. In liver slices incubated for 8 h, conversion of palmitate to CO₂ was greater for calves treated with clofibrate compared with control calves or calves treated with fish oil. Conversion of palmitate to esterified products, total palmitate metabolism, and metabolism of oleate were not different among treatments. Conversion of palmitate to CO₂ was greater than that from oleate for all treatments, but rates of total metabolism did not differ. Clofibrate increased or tended to increase liver expression of several PPARα target genes involved in fatty acid oxidation (e.g., ACADVL, ACOX1, CPT1A), whereas fish oil did not significantly affect genes associated with fatty acid oxidation but tended to increase DGAT1. Overall, our data indicated that bovine liver responded to clofibrate treatment but not fish oil, although increases in hepatic lipid metabolism were much less than those reported in rodents treated with clofibrate or fish oil. Applications of PPARα agonists may be of interest to increase the rate of hepatic fatty acid oxidation and decrease triacylglycerol accumulation in periparturient dairy cows.     

Meta-analysis of the effects of dietary vitamin E supplementation on α-tocopherol concentration and lipid oxidation in pork

Meta-analyses have been carried out to quantify the effect of dietary vitamin E on α-tocopherol accumulation and on lipid oxidation in porcine M. longissimus. Published results of 13 (vitamin E accumulation) and 10 (lipid oxidation) experiments respectively were used for the analyses. After a number of standardization procedures, a nonlinear relationship was found between the supplementary vitamin E and the accumulation of α-tocopherol in pork which approached a maximum value of 6.4 μg/g tissue. Pork lipid oxidation levels were described in terms of Thiobarbituric Acid Reacting Substances (TBARS) values. The statistical analysis revealed significant effect of vitamin E dose, muscle α-tocopherol concentration and supplementation time on TBARS, resulting in two prediction models for lipid oxidation. Meta-analysis has proven to be a valuable tool for combining results from previous studies to quantify the effects of dietary vitamin E. Further studies, carried out with standardized experimental protocols would be beneficial for model validation and to increase the predictive power of the derived models.     

Vitamin B6 suppresses serine protease inhibitor 3 expression in the colon of rats and in TNF-α-stimulated HT-29 cells

Scope: Previous reports in the areas of animal studies and, recently epidemiology, have linked anti‐tumorigenic and anti‐inflammatory effects to dietary vitamin B6. This study investigated the molecular mechanism of these effects of vitamin B6. Methods and results: DNA microarray analysis was used to obtain information on changes in colon gene expression from vitamin B6 (pyridoxine) repletion in vitamin B6‐deficient rats. Pyridoxine supplementation down‐regulated the inflammatory molecule, serine protease inhibitor clade A member 3 (SPI‐3) mRNA expression in the colon. This study also showed that tumor necrosis factor α (TNF‐α) induced SPI‐3 mRNA expression in HT‐29 human colon cancer cells, and vitamin B6 (pyridoxal hydrochloride) pretreatment of HT‐29 cells inhibited TNF ‐induced mRNA expression of SPI‐3. Vitamin B6 inhibited TNF‐α‐induced NF‐κB activation via suppression of IκBα degradation in HT‐29 cells. HT‐29 cells stably expressing epitope‐tagged ubiquitin were generated and vitamin B6 pretreatment was shown to inhibit ubiquitination of the IkB protein in response to TNF‐α‐i. Conclusion: Vitamin B6 suppressed SPI‐3 expression in the colon of rats and in TNF‐α‐stimulated HT‐29 cells. Further, this study showed a possible role of vitamin B6 in the regulation of protein ubiquitination.     

Functional and heterologous expression of human protein O-linked mannose β-1,2-N-acetylglucosaminyltransferase 1 in zebrafish

Although membrane-associated proteins are related to many diseases and are important targets for drug discovery, their expression is often difficult in bacterial hosts such as Escherichia coli. To overcome this limitation, here, we focused on a novel host-vector system in zebrafish for the expression of human protein O-linked mannose β-1,2-N-acetylglucosaminyltransferase 1 (hPOMGnT1) which is related to muscle-eye-brain disease. For the expression of hPOMGnT1, the vector pZex-EGFP-pXI-hPOMGnT1 was constructed and injected into fertilized eggs. Using this system, we demonstrated that recombinant hPOMGnT1 was successfully expressed in the whole bodies of zebrafish embryos.     

A Review on the Fermentation of Foods and the Residues of Pesticides—Biotransformation of Pesticides and Effects on Fermentation and Food Quality

Residues of pesticides in food are influenced by processing such as fermentation. Reviewing the extensive literature showed that in most cases, this step leads to large reductions in original residue levels in the fermented food, with the formation of new pesticide by-products. The behavior of residues in fermentation can be rationalized in terms of the physical-chemical properties of the pesticide and the nature of the process. In addition, the presence of pesticides decrease the growth rate of fermentative microbiota (yeasts and bacterias), which provokes stuck and sluggish fermentations. These changes have in consequence repercussions on several aspects of food sensory quality (physical-chemical properties, polyphenolic content, and aromatic profile) of fermented food. The main aim of this review is to deal with all these topics to propose challenging needs in science-based quality management of pesticides residues in food.