Effect of processed and red meat on endogenous nitrosation and DNA damage

Haem in red meat (RM) stimulates the endogenous production of mutagenic nitroso compounds (NOC). Processed (nitrite-preserved red) meat additionally contains high concentrations of preformed NOC. In two studies, of a fresh RM versus a vegetarian (VEG) diet (six males and six females) and of a nitrite-preserved red meat (PM) versus a VEG diet (5 males and 11 females), we investigated whether processing of meat might increase colorectal cancer risk by stimulating nitrosation and DNA damage. Meat diets contained 420 g (males) or 366 g (females) meat/per day. Faecal homogenates from day 10 onwards were analysed for haem and NOC and asso- ciated supernatants for genotoxicity. Means are adjusted for differ- ences in male to female ratios between studies. Faecal NOC concentrations on VEG diets were low (2.6 and 3.5 mmol/g) but significantly higher on meat diets (PM 175 ± 19 nmol/g versus RM 185 ± 22 nmol/g; P 5 0.75). The RM diet resulted in a larger pro- portion of nitrosyl iron (RM 78% versus PM 54%; P < 0.0001) and less nitrosothiols (RM 12% versus PM 19%; P < 0.01) and other NOC (RM 10% versus PM 27%; P < 0.0001). There was no statis- tically significant difference in DNA breaks induced by faecal water (FW) following PM and RM diets (P 5 0.80). However, PM re- sulted in higher levels of oxidized pyrimidines (P < 0.05). Surpris- ingly, VEG diets resulted in significantly more FW-induced DNA strand breaks than the meat diets (P < 0.05), which needs to be clarified in further studies. Meats cured with nitrite have the same effect as fresh RM on endogenous nitrosation but show increased FW-induced oxidative DNA damage.

Detection of transgenic and endogenous plant DNA fragments in the blood, tissues, and digesta of broilers

The aim was to determine the fate of transgenic and endogenous plant DNA fragments in the blood, tissues, and digesta of broilers. Male broiler chicks (n = 24) were allocated at 1 day old to each of four treatment diets designated T1-T4. T1 and T2 contained the near isogenic nongenetically modified (GM) maize grain, whereas T3 and T4 contained GM maize grain [cry1a(b) gene]; T1 and T3 also contained the near isogenic non-GM soybean meal, whereas T2 and T4 contained GM soybean meal (cp4epsps gene). Four days prior to slaughter at 39-42 days old, 50% of the broilers on T2-T4 had the source(s) of GM ingredients replaced by their non-GM counterparts. Detection of specific DNA sequences in feed, tissue, and digesta samples was completed by polymerase chain reaction analysis. Seven primer pairs were used to amplify fragments (similar to 200 bp) from single copy genes (maize high mobility protein, soya lectin, and transgenes in the GM feeds) and multicopy genes (poultry mitochondrial cytochrome b, maize, and soya rubisco). There was no effect of treatment on the measured growth performance parameters. Except for a single detection of lectin (nontransgenic single copy gene; unsubstantiated) in the extracted DNA from one bursa tissue sample, there was no positive detection of any endogenous or transgenic single copy genes in either blood or tissue DNA samples. However, the multicopy rubisco gene was detected in a proportion of samples from all tissue types (23% of total across all tissues studied) and in low numbers in blood. Feed-derived DNA was found to survive complete degradation up to the large intestine. Transgenic DNA was detected in gizzard digesta but not in intestinal digesta 96 h after the last feeding of treatment diets containing a source of GM maize and/or soybean meal.

Effect of corn silage from an herbicide-tolerant genetically modified variety on milk production and absence of transgenic DNA in milk

Data from 60 multiparous Holstein cows were used in a 12-wk continuous design feeding trial. Cows were allocated to 1 of 4 experimental treatments (T1 to T4). In T1 and T2, the total mixed ration (TMR) contained either corn silage from the genetically modified (GM) variety Chardon Liberty Link, which is tolerant to the herbicide glufosinate ammonium, or its near isogenic nonGM counterpart, whereas the TMR used in T3 and T4 contained corn silage from the commercially available nonGM varieties Fabius and Antares, respectively. The objectives of the study were to determine if the inserted gene produced a marked effect on chemical composition, nutritive value, feed intake, and milk production, and to determine if transgenic DNA and the protein expressed by the inserted gene could be detected in bovine milk. The nutritive value, fermentation characteristics, mineral content, and amino acid composition of all 4 silages were similar. There were no significant treatment effects on milk yield, milk composition, and yield of milk constituents, and the dry matter (DM) intake of the GM variety was not significantly different from the 2 commercial varieties. However, although the DM intake noted for the nonGM near-isogenic variety was similar to the commercial varieties, it was significantly lower when compared with the GM variety. Polymerase chain reaction analyses of milk samples collected at wk 1, 6, and 12 of the study showed that none of the 90 milk samples tested positive, above a detection limit of 2.5 ng of total genomic DNA/mL of milk, for either tDNA (event T25) or the single-copy endogenous Zea mays gene, alcohol dehydrogenase. Using ELISA assays, the protein expressed by the T25 gene was not detected in milk.

Effect of plasma insulin and branched-chain amino acids on skeletal muscle protein synthesis in fasted lambs

The increase in fractional rate of protein synthesis (K-s) in the skeletal muscle of growing rats during the transition from fasted to fed state has been explained by the synergistic action of a rise in plasma insulin and branched-chain amino acids (BCAA). Since growing lambs Also exhibit an increase in K-s with level of feed intake, the objective of the present study was to determine if this synergistic relationship between insulin and BCAA also occurs in ruminant animals. Six 30 kg fasted (72 h) lambs (8 months of age) received each of four treatments, which were based on continuous infusion into the jugular vein for 6 h of: (1) saline (155 mmol NaCl/l); (2) a mixture of BCAA (0.778 mumol leucine, 0.640 mumol isoleucine and 0.693 mumol valine/min.kg); (3) 18.7 mumol glucose/min.kg (to induce endogenous insulin secretion): (4) co-infusion of BCAA and glucose. Within each period all animals received the same isotope of phenylalanine, (Phe) as follows: (1) L-[1-C-13]Phe; (2) L-phenyl-[ring H-2(5)]-alanine; (3) L-[N-15]Phe; (4) L-[ring 2,6-H-3]Phe. Blood was sampled serially during infusions to measure plasma concentrations of insulin, glucose and amino acids, and plasma free Phe isotopic activity; biopsies were taken 6 h after the beginning of infusions to determine K-s in in. longissimus dorsi and vastus muscle. Compared with control (saline-infused) lambs, K-s was increased by an average of 40% at the end of glucose infusion, but this effect was not statistically significant in either of the muscles sampled. BCAA infusion, alone or in combination with glucose, also had no significant effect on K-s compared with control sheep. K-s was approximately 60% greater for vastus muscle than for m. longissimus dorsi (P<0.01), regardless of treatment. It is concluded that there are signals other than insulin and BCAA that are responsible for the feed-induced increase in K-s in muscle of growing ruminant animals.

Effect of corn silage from an herbicide-tolerant genetically modified variety on milk production and absence of transgenic DNA in milk

Data from 60 multiparous Holstein cows were used in a 12-wk continuous design feeding trial. Cows were allocated to 1 of 4 experimental treatments (T1 to T4). In T1 and T2, the total mixed ration (TMR) contained either corn silage from the genetically modified (GM) variety Chardon Liberty Link, which is tolerant to the herbicide glufosinate ammonium, or its near isogenic nonGM counterpart, whereas the TMR used in T3 and T4 contained corn silage from the commercially available nonGM varieties Fabius and Antares, respectively. The objectives of the study were to determine if the inserted gene produced a marked effect on chemical composition, nutritive value, feed intake, and milk production, and to determine if transgenic DNA and the protein expressed by the inserted gene could be detected in bovine milk. The nutritive value, fermentation characteristics, mineral content, and amino acid composition of all 4 silages were similar. There were no significant treatment effects on milk yield, milk composition, and yield of milk constituents, and the dry matter (DM) intake of the GM variety was not significantly different from the 2 commercial varieties. However, although the DM intake noted for the nonGM near-isogenic variety was similar to the commercial varieties, it was significantly lower when compared with the GM variety. Polymerase chain reaction analyses of milk samples collected at wk 1, 6, and 12 of the study showed that none of the 90 milk samples tested positive, above a detection limit of 2.5 ng of total genomic DNA/mL of milk, for either tDNA (event T25) or the single-copy endogenous Zea mays gene, alcohol dehydrogenase. Using ELISA assays, the protein expressed by the T25 gene was not detected in milk.

Effect of time of day and rabbit strain on patterns of aortic wall permeability

Lipid deposits occur more frequently downstream of branch points than upstream in immature rabbit and human aortas but the opposite pattern is seen in mature vessels. These distributions correlate spatially with age-related patterns of aortic permeability, observed in rabbits, and may be determined by them. The mature but not the immature pattern of permeability is dependent on endogenous nitric oxide synthesis. Although the transport patterns have hitherto seemed robust, recent studies have given the upstream pattern in some mature rabbits but the downstream pattern in others. Here we show that transport in mature rabbits is significantly skewed to the downstream pattern in the afternoon compared with the morning (P < 0.05), and switches from a downstream to an upstream pattern at around 21 months in rabbits of the Murex strain, but at twice this age in Highgate rabbits (P < 0.001). The effect of time of day was not explained by changes in nitric oxide production, assessed from plasma levels of nitrate and nitrate, nor did it correlate with conduit artery tone, assessed from the shape of the peripheral pulse wave. The effect of strain could not be explained by variation in nitric oxide production nor by differences in wall structure. The effects of time of day and rabbit strain on permeability patterns explain recent discrepancies, provide a useful tool for investigating underlying mechanisms and may have implications for human disease.

Effect of dietary fish oil on biohydrogenation of fatty acids and milk fatty acid content in cows

Mechanisms underlying milk fat conjugated linoleic acid (CLA) responses to supplements of fish oil were investigated using five lactating cows each fitted with a rumen cannula in a simple experiment consisting of two consecutive 14-day experimental periods. During the first period cows were offered 18 kg dry matter (DM) per day of a basal (B) diet formulated from grass silage and a cereal based-concentrate (0.6 : 0.4; forage : concentrate ratio, on a DM basis) followed by the same diet supplemented with 250 g fish oil per day (FO) in the second period. The flow of non-esterified fatty acids leaving the rumen was measured using the omasal sampling technique in combination with a triple indigestible marker method based on Li-Co-EDTA, Yb-acetate and Cr-mordanted straw. Fish oil decreased DM intake and milk yield, but had no effect on milk constituent content. Milk fat trans-11C(18:1), total trans-C-18:1, cis-9 trans-11 CLA, total CLA, C-18 :2 (n- 6) and total C-18:2 content were increased in response to fish oil from 1.80, 4.51, 0.39, 0. 56, 0.90 and 1.41 to 9.39, 14.39, 1.66, 1.85, 1.25 and 4.00 g/100 g total fatty acids, respectively. Increases in the cis-9, trans-11 isomer accounted for proportionately 0.89 of the CLA response to fish oil. Furthermore, fish oil decreased the flow of C-18:0 (283 and 47 g/day for B and FO, respectively) and increased that of trans-C-18:1 fatty acids entering the omasal canal (38 and 182 g/day). Omasal flows of trans-C-18:1 acids with double bonds in positions from delta-4 to -15 inclusive were enhanced, but the effects were isomer dependent and primarily associated with an increase in trans-11C(18:1) leaving the rumen (17.1 and 121.1 g/day for B and FO, respectively). Fish oil had no effect on total (4.36 and 3.50 g/day) or cis-9, trans-11 CLA (2.86 and 2.08 g/day) entering the omasal canal. Flows of cis-9, trans-11 CLA were lower than the secretion of this isomer in milk. Comparison with the transfer of the trans-9, trans-11 isomer synthesized in the rumen suggested that proportionately 0.66 and 0.97 of cis-9, trans-11 CLA was derived from endogenous conversion of trans-11 C-18:1 in the mammary gland for B and FO, respectively. It is concluded that fish oil enhances milk fat cis-9, trans-11 CLA content in response to increased supply of trans-11 C-18:1 that arises from an inhibition of trans C-18:1 reduction in the rumen.

Effects of endogenous flour lipids on the quality of short-dough biscuits

Fractionation and reconstitution techniques were used to study the contribution of endogenous flour lipids to the quality of short-dough (shortcake type) biscuits. Biscuit flour was defatted with chloroform and baked with bakery fat, but without endogenous lipid. Short-dough biscuits baked from defatted flour had smaller diameters, and were flatter, denser and harder than control biscuits. Defatted flour shortcake doughs exhibited different rheological behaviour from the control samples, showing higher storage and loss moduli (G' and G" values), ie higher viscoelasticity. Functionality was restored when total non-starch flour lipids were added back to defatted flour. The polar lipid fraction had a positive effect in restoring flour quality whereas the non-polar lipid fraction had no effect. Both fractions were needed for complete restoration of both biscuit quality and dough rheological characteristics. A study of the microstructure of defatted biscuits revealed that their gluten protein was more hydrated and developed than the gluten of the control biscuits. This conclusion was supported by the higher water absorption of the defatted gluten. (C) 2004 Society of Chemical Industry.

Variability in fecal water genotoxicity, determined using the Comet assay, is independent of endogenous N-nitroso compound formation attributed to red meat consumption

Red meat consumption causes a dose-dependent increase in fecal apparent total N-nitroso compounds (ATNC). The genotoxic effects of these ATNCs were investigated using two different Comet assay protocols to determine the genotoxicity of fecal water samples. Fecal water samples were obtained from two studies of a total of 21 individuals fed diets containing different amounts of red meat, protein, heme, and iron. The first protocol incubated the samples with HT-29 cells for 5 min at 4 degrees C, whereas the second protocol used a longer exposure time of 30 min and a higher incubation temperature of 37 degrees C. DNA strand breaks were quantified by the tail moment (DNA in the comet tail multiplied by the comet tail length). The results of the two Comet assay protocols were significantly correlated (r = 0.35, P = 0.003), however, only the second protocol resulted in detectable levels of DNA damage. Inter-individual effects were variable and there was no effect on fecal water genotoxicity by diet (P > 0.20), mean transit time (P = 0.588), or weight (P = 0.705). However, there was a highly significant effect of age (P = 0.019). There was no significant correlation between concentrations of ATNCs in fecal homogenates and fecal water genotoxicity (r = 0.04, P = 0.74). ATNC levels were lower in fecal water samples (272 microg/kg) compared to that of fecal homogenate samples (895 microg/kg) (P < 0.0001). Failure to find dietary effects on fecal water genotoxicity may therefore be attributed to individual variability and low levels of ATNCs in fecal water samples.

The antithrombotic effect of dextran-40 in man is due to enhanced fibrinolysis in vivo

BACKGROUND: Dextran-40 is effective in reducing postoperative Doppler-detectable embolization in patients undergoing carotid endarterectomy (CEA). Dextrans are thought to have antithrombotic and antiplatelet effects. The mode of action is unclear. In rats, dextran blocks uptake of tissue plasminogen activator (tPA) by mannose-binding receptors. Because this would have the effect of enhancing endogenous fibrinolysis, we explored this effect of dextran-40 on fibrinolysis in man. METHODS: Twenty patients undergoing endovascular stenting for abdominal aortic aneurysm were randomized to receive 100 mL of 10% dextran-40 or saline, over 1 hour, during their operation in addition to heparin. Blood samples were taken preoperatively, intraoperatively (immediately after operative procedure), and 24 hours postoperatively. Thrombi were formed in a Chandler loop and used to assess endogenous fibrinolysis over 24 hours, measured as the fall in thrombus weight, and the release of fluorescently labelled fibrinogen from the thrombus. Plasma samples were analyzed for markers of fibrinolysis; plasmin-antiplasmin (PAP), PAI-1, and t-PA, and for functional von Willebrand factor (vWF). Platelet response to thrombin and other agonists was measured by flow cytometry. RESULTS: Thrombi formed ex vivo from the intraoperative blood samples from the dextran-treated patients exhibited significantly greater fibrinolysis vs preoperative samples, seen both as a significantly greater percentage reduction in thrombus weight (from 34.7% to 70.6% reduction) and as an 175% increase in the release of fluorescence (P < .05). Fibrinolysis returned to baseline levels the next day. No change was seen in the saline-treated group. Plasma levels of PAP and PAI-1 increased significantly postoperatively in the dextran-treated group vs the saline group (P < .05). The postoperative level of functional VWF was significantly lower in the dextran-treated group vs controls. A specific reduction occurred in the platelet response to thrombin, but not to other agonists, in the intraoperative samples from the dextran-treated group (11.1% vs 37.1%; P = .022), which was not seen in the controls. CONCLUSIONS: These data are consistent with a rise in plasmin due to dextran blockade of tPA uptake in vivo, leading to enhanced fibrinolysis, cleavage of vWF and of the platelet protease-activated receptor-1 (PAR-1) thrombin receptor. This suggests that dextran exerts a combined therapeutic effect, enhancing endogenous fibrinolysis, whilst also reducing platelet adhesion to vWF and platelet activation by thrombin. The proven antithrombotic efficacy of low-dose dextran in carotid surgery may be applicable to wider therapeutic use.

Effect of hypobaric hypoxia, simulating conditions during long-haul air travel, on coagulation, fibrinolysis, platelet function, and endothelial activation

CONTEXT: The link between long-haul air travel and venous thromboembolism is the subject of continuing debate. It remains unclear whether the reduced cabin pressure and oxygen tension in the airplane cabin create an increased risk compared with seated immobility at ground level. OBJECTIVE: To determine whether hypobaric hypoxia, which may be encountered during air travel, activates hemostasis. DESIGN, SETTING, AND PARTICIPANTS: A single-blind, crossover study, performed in a hypobaric chamber, to assess the effect of an 8-hour seated exposure to hypobaric hypoxia on hemostasis in 73 healthy volunteers, which was conducted in the United Kingdom from September 2003 to November 2005. Participants were screened for factor V Leiden G1691A and prothrombin G20210A mutation and were excluded if they tested positive. Blood was drawn before and after exposure to assess activation of hemostasis. INTERVENTIONS: Individuals were exposed alternately (> or =1 week apart) to hypobaric hypoxia, similar to the conditions of reduced cabin pressure during commercial air travel (equivalent to atmospheric pressure at an altitude of 2438 m), and normobaric normoxia (control condition; equivalent to atmospheric conditions at ground level, circa 70 m above sea level). MAIN OUTCOME MEASURES: Comparative changes in markers of coagulation activation, fibrinolysis, platelet activation, and endothelial cell activation. RESULTS: Changes were observed in some hemostatic markers during the normobaric exposure, attributed to prolonged sitting and circadian variation. However, there were no significant differences between the changes in the hypobaric and the normobaric exposures. For example, the median difference in change between the hypobaric and normobaric exposure was 0 ng/mL for thrombin-antithrombin complex (95% CI, -0.30 to 0.30 ng/mL); -0.02 [corrected] nmol/L for prothrombin fragment 1 + 2 (95% CI, -0.03 to 0.01 nmol/L); 1.38 ng/mL for D-dimer (95% CI, -3.63 to 9.72 ng/mL); and -2.00% for endogenous thrombin potential (95% CI, -4.00% to 1.00%). CONCLUSION: Our findings do not support the hypothesis that hypobaric hypoxia, of the degree that might be encountered during long-haul air travel, is associated with prothrombotic alterations in the hemostatic system in healthy individuals at low risk of venous thromboembolism.

Output sustainability to exogenous and endogenous shocks: evidence from emerging economies

This paper studies the impact of exogenous and endogenous shocks (exogenous shock is used interchangeably with external shock; endogenous shock is used interchangeably with domestic shock) on output fluctuations in post-communist countries during the 2000s. The first part presents the analytical framework and formulates a research hypothesis. The second part presents vector autoregressive estimation and analysis model proposed by Pesaran (2004) and Pesaran and Smith (2006) that relates bank real lending, the cyclical component of output and spreads and accounts for cross-sectional dependence (CD) across the countries. Impulse response functions show that exogenous positive shock lead to a drop in output sustainability for 9 over 12 Central Eastern European countries and Russia, when the endogenous shock is mild and ambiguous. Moreover, the effect of exogenous shock is more significant during the crises. Variance decompositions show that exogenous shock in the aftermath of crisis had a substantial impact on economic activity of emerging economies.