Role of superoxide radicals in cytotoxic effects of Fe-NTA on cultured normal liver epithelial cells

We studied the cytotoxic effects of ferric nitrilotriacetate (Fe-NTA) on normal rat liver epithelial cells (RL34) cultured in medium containing 10% fetal calf serum. Marked cytolysis was present in cells exposed to greater than or equal to 25 micrograms/ml iron of Fe-NTA, but not all the cells exposed to 50 micrograms/ml iron were lethally injured. The remaining cells showed anomalous growth, namely cell pile-up and aggregation. Superoxide dismutase inhibited this iron-induced cytotoxicity, whereas catalase, mannitol, dimethyl sulfoxide, and 1,4-diazabicyclo-[2.2.2.] octane did not. RL34 cells exposed to Fe-NTA actually produced a large amount of superoxide radicals (O2-.), whereas unexposed control cells produced none. Allopurinol inhibited O2-. production and prevented cell injury by Fe-NTA. These results show that the injury to cells produced by Fe-NTA depends on the generation of O2-., the source of which may be xanthine oxidase.     

Albumin inhibits cytotoxic activity of lysophosphatidylcholine by direct binding

Fetal bovine serum (FBS) was found to protect Jurkat T cells from LPC-induced cytotoxicity. Twenty micromolar LPC-induced cytotoxicity of 80-90% of the cells in media without FBS for 3 h, whereas 50-70% in media with 0.5% FBS. However, LPC-induced cytotoxicity was not observed in the presence of 5% FBS in media. The cytotoxicity was specific for LPC among lysophospholipids tested and significantly observed with palmitoyl (C16:0) LPC, stearoyl (C18:0) LPC, and oleoyl (C18:1) LPC among 11 synthetic LPCs. Furthermore, the cytoprotective effect of FBS was observed only when it was added before the treatment, but not after the treatment of LPC, and premixing of FBS and LPC before addition to the cells ameliorated LPC-induced cytotoxicity. Finally, albumin, a major constituent of FBS, prevented completely LPC-induced cytotoxicity even at as low as 3 microM concentration. We also found that five molecules of LPC could sequentially bind to one BSA using isothermal titration calorimetry. The above results suggest that the cytotoxic activity of LPC could be attenuated by albumin in blood. Finally, it should be cautioned that, when experiments are conducted with LPC dissolved in assay buffers containing albumin, the albumin in the buffer could influence the results.     

Influence of albumin on granulocyte and macrophage colony-forming efficiency

Mouse granulocyte and macrophage precursors were assayed in plasma clot and fibrin clot cultures, and the effect of bovine serum albumin (BSA) on colony formation was investigated. The number of granulocyte colonies (CFU-g) and clusters increased as the albumin concentration was increased and the number of macrophage colonies (CFU-m) and clusters concomitantly decreased. The albumin-mediated suppression of macrophage colony formation was overcome by the addition of more than 10% fetal bovine serum (FBS) to the plasma clot culture. The effect of BSA and fatty-acid-free BSA on colony-forming efficiency was also tested in fibrin clot cultures containing 10% FBS. Both BSA and fatty-acid-free BSA at a final concentration of 0.5-2% enhanced CFU-g colony formation, while both forms of BSA reduced the number of CFU-m colonies. However, neither BSA nor fatty-acid-free BSA had any effect on colony formation in FBS-free fibrin clot cultures, and only BSA enhanced colony formation when transferrin, linoleic acid, alpha-thioglycerol and dextran were added to the culture. The number of CFU-g (15.6 +/- 3.1) was higher in cultures containing BSA, transferrin, etc., than the number (9.8 +/- 2.5) in cultures without BSA and including transferrin, etc., than the number (9.8 +/- 2.5) in cultures without BSA and including transferrin, etc. (p less than 0.01). The number of CFU-m (32.0 +/- 6.8) in cultures containing BSA and the other four factors was lower than the number (72.2 +/- 5.6) in the culture without BSA (p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Insulin insensitivity and altered glucose utilization in cultured rat adipose tissue

Glucose utilization was studied in isolated fat cells prepared from rat adipose tissue which had been cultured for 18 hr in TC 199 medium. When 1% bovine serum albumin (BSA) was in the culture medium, basal rates of (14)CO(2) and [(14)C]triglyceride production from [1-(14)C]glucose were markedly depressed and there was no effect of insulin. With 4% BSA, basal (14)CO(2) production was the same as in cells prepared from fresh tissue and basal triglyceride production was greatly increased. Insulin effect on these cells was minimal. One-minute uptake of [(14)C]2-deoxyglucose was stimulated by 800-1000% in fresh cells and 300-500% in cells cultured with either 1% or 4% BSA. Oxidation of [U-(14)C]glucose showed a much smaller impairment in cultured cells than for [1-(14)C]glucose, suggesting that the pentose phosphate shunt was more severely impaired than glycolysis. Glyceride-glycerol production was increased in cultured cells relative to preculture (fresh) cells. There was no effect of insulin in the culture medium in any of these systems. Rates of free fatty acid and glycerol release were markedly increased in cultured cells, especially when insulin was present in the culture medium. The acute antilipolytic effect of insulin was retained, so that insulin in the test incubation decreased lipolysis by 40-80%. Nevertheless, cell-associated fatty acids were increased in cultured cells and FFA/albumin ratios in the medium often reached potentially toxic levels. The reduction in pentose phosphate shunt activity, lipogenesis, and insulin effect resembles other models of insulin insensitivity. The impaired metabolism is probably due to an intracellular defect. A possible toxic role of either intracellular or extracellular fatty acids cannot be excluded. This system should be a useful model in which to study the cellular mechanisms of insulin insensitivity in adipocytes.-Bernstein, R. S. Insulin insensitivity and altered glucose utilization in cultured rat adipose tissue.     

Effect of albumin on the in vitro conjugation of bilirubin by rat liver microsomes

These studies were carried out to determine whether bovine serum albumin (BSA), which is usually included in the incubation mixture for the in vitro determination of bilirubin-UDP-glucuronyl transferase (GT) activity, affects GT activity. Using bilirubin as substrate, addition of BSA to the enzyme reaction mixture at concentrations varying from 2 to 30 mg/ml resulted in a dose-related inhibition of "native" GT activity of rat liver microsomes. When detergent-activated enzyme was employed, increasing concentrations of BSA also required higher concentrations of deoxycholate, digitonin, or Triton X-100 to produce maximal bilirubin conjugation. Low BSA concentrations (2 mg/ml) prevented enzyme activation by both detergents and UDP-N-acetyl glucosamine. When BSA was omitted and bilirubin dissolved in dimethyl sulfoxide, UDP-N-acetyl glucosamine failed to enhance GT activity, and activation by detergents was only 15-25% of that observed in the presence of optimal concentrations of BSA. When rat albumin was substituted for BSA, a similar dose-related inhibition of in vitro bilirubin conjugation by untreated microsomes was observed, although at any given albumin concentration, GT activity was lower with rat than with bovine albumin. Additionally, both detergents and UDP-N-acetyl glucosamine produced similar GT activation regardless of the rat albumin concentration. Finally, these effects of BSA and rat albumin could not be reproduced when beta-lactoglobulin was employed and/or when p-nitrophenol was the acceptor substrate of GT. These findings indicate that albumin, in particular BSA, profoundly and selectively influences the in vitro activity of microsomal GT toward bilirubin as the acceptor substrate.     

The fatty acid analogue 11-(dansylamino)undecanoic acid is a fluorescent probe for the bilirubin-binding sites of albumin and not for the high-affinity fatty acid-binding sites.

1. The fluorescent fatty acid probe 11-(dansylamino)undecanoic acid (DAUDA) binds with high affinity to bovine and human serum albumin (BSA and HSA) at three sites. 2. The Kd of the primary binding site could not be determined; however, the two secondary sites appeared to be equivalent, with an apparent Kd of 8 x 10(-7) M for both BSA and HSA. 3. The spectral characteristics of DAUDA when bound to the primary site of the two albumins were different, with HSA producing a greater fluorescence enhancement and emission maximum at a shorter wavelength (480 nm) than for BSA (495 nm). 4. Displacement studies indicated that the DAUDA-binding sites were not equivalent to the primary long-chain fatty acid-binding sites on albumin, but corresponded to the bilirubin sites. Fatty acyl-CoAs also bind to the bilirubin sites, as do medium-chain fatty acids. 5. The solubility, stability and spectral properties of DAUDA make it an excellent probe for investigating the bilirubin-binding sites of albumin, particularly HSA.     

Cellular oxidant stress and advanced glycation endproducts of albumin: caveats of the dichlorofluorescein assay

In order to understand the mechanism by which advanced glycation endproducts (AGEs) elicit oxidative stress, macrophage-like RAW264.7 cells were exposed to various AGE-albumins, and oxidant stress was estimated from the fluorescence of oxidized dichlorofluorescein using the microtiter plate assay. Strongest fluorescence was observed with methylglyoxal modified albumin (MGO-BSA) compared with native albumin. Similar effects that were prevented by arginine coincubation were seen with phenylglyoxal-BSA. MGO-BSA had increased affinity for Cu(2+) and Ca(2+), but was conformationally similar to native albumin. Surprisingly, the mere addition of unmodified albumin to cells suppressed the fluorescence of oxidized DCF. While, several site-directed mutants of human serum albumin (HSA), including C34S and recombinant domains II and III retained fluorescence suppressing activity, proteolytic digests, recombinant domain I, and several nonalbumin proteins failed to suppress. Kinetic and ANS binding studies suggested albumin quenches DCF fluorescence by binding to hydrophobic pockets in domains II and III and that MGO-BSA is less hydrophobic than BSA. Finally, BSA also prevented H(2)O(2) catalyzed DCF fluorescence more potently than MGO-BSA. These studies reveal important caveats of the widely used dichlorofluorescein assay and suggest methods other than the microtiter plate assay are needed to accurately assess cellular oxidant stress in presence of native or modified albumin.     

A serum-free medium for use in a cumulus cell co-culture system for bovine embryos derived from in vitro maturation and in vitro fertilization

The objective of this study was to develop a serum-free medium for the co-culture of bovine embryos that would yield a percentage of blastocysts equal to that obtained with fetal bovine serum (FBS)-supplemented medium. Cumulus cell-enclosed oocytes (CEO) matured and inseminated in vitro were cultured in a tissue culture medium (TCM)-199 or in a serum-free medium (bovine embryo culture medium; BECM) until 240 h post insemination. Replacement of 10% (v/v) FBS with either 3 mg crystallized bovine serum albumin (BSA)/ml or 3 mg fatty acid-free BSA/ml in TCM-199 had no effect (P > 0.14) on embryo development to the >or= 2-cell (51 to 60%), >or= 8-cell (24 to 33%), blastocyst (16 to 19%) and hatched-blastocyst (7 to 10%) stages at 48, 96, 192 and 240 h post insemination, respectively. Oocyte-enclosing cumulus cells in BSA-supplemented medium grew in clusters rather than in layers, as was noted in FBS-supplemented medium. When CEO were cultured in fatty acid-free BSA-supplemented media (TCM-199 and BECM), a significantly (P < 0.001) higher percentage of oocytes developed to blastocysts after culture with (22%) or without (18%) a cumulus cell monolayer than after denuding the oocytes (7%). Glucose in concentrations of 0 to 5.56 mM added for periods of 18 and 120 h post-insemination had neither a stimulatory nor a deleterious effect on preimplantation development. In conclusion, a serum-free medium supplemented with BSA can be successfully used in a cumulus cell co-culture system for bovine embryos.     

Evaluation of polyvinyl alcohol for fatty acid supplementation in adipose tissue explant culture

Cultures of adipose tissue explants are a valuable tool for studying the intracellular mechanisms involving hormones and nutrients. However, testing how fatty acids affect cells requires a carrier molecule; bovine serum albumin (BSA) has been used for this purpose. However, contaminants can alter the cellular response. Our objectives were to: 1) test BSA as a fatty acid carrier and 2) evaluate polyvinyl alcohol (PVA) as a replacement for BSA. Adipose tissue explants from nine pigs were cultured in medium 199 for 4, 12, 24, and 48 h, with the following treatments: control, PVA (100 mM PVA added) and PVA + pGH (100 mM PVA plus 0.1 mg/mL porcine growth hormone). After each culture period, explants were collected and assayed for lipogenesis. After 48 h in culture, explants were assayed for lipolysis. A preliminary study with different commercial sources and high concentrations showed that BSA affected lipogenic rates. On the other hand, there were no effects of PVA on lipid synthesis, while pGH (positive control) reduced glucose incorporation into lipids (P < 0.01) when compared to both control and PVA (P < 0.05). There was no difference between control and PVA for lipolysis rates. However, pGH increased lipolysis when compared to control (P < 0.01) and PVA (P < 0.05). We demonstrated that BSA can alter lipogenesis, which precludes its use as a carrier molecule. On the other hand, addition of PVA had no effect on lipolysis or lipogenesis. We suggest the use of PVA instead of BSA for adding bioactive fatty acids to cultures of adipose tissue.     

Oviductal fluid and growth factors failed to enhance development of porcine embryos

Porcine embryos (1-, 2- and 4-cell) were cultured in a basal medium consisting of Krebs-Ringer bicarbonate buffer supplemented with oviductal fluid and several growth factors and observed for further development. Oviducts were flushed at either 48 h (Experiment 1) or 96 h (Experiment 2) after the onset of estrus. Observations were made every 48 h (Experiment 1) or 12 h (Experiment 2) until failure of the embryos to develop for 2 consecutive observations. Embryos were scored 0 = no development, 1 = cleavage, 2 = morula, 3 = blastocyst, or 4 = hatched blastocyst. In the first experiment, development of 1-, 2- and 4-cell embryos (n=282) in the basal medium supplemented with oviductal fluid (4:1) or 3 sets of growth factors, was less or equal to one cleavage stage. Those embryos cultured in the basal medium supplemented with bovine serum albumin (fatty acid free) (BSA) advanced to the blastocyst stage. In the second experiment, 96 h aged embryos (n=142) were cultured in the basal medium supplemented with IGF-1 and - 2 and EGF, or with BSA alone or with BSA and the three growth factors. In the treatments without BSA, the embryonic development was less than one cleavage, whereas in those treatments with BSA, embryos advanced beyond hatching and began to expand. We conclude that for culture of porcine embryos, supplementation with several growth factors or with oviductal fluid, in the concentration used in this study, was of little benefit at this stage of development. However, the type of BSA significantly affected development. More than 90% of the embryos reached the morula and blastocyst stages in medium than included BSA (fatty acid free).     

A low-serum medium with BSA and ferrous sulfate for the production of tissue plasminogen activator by human embryo lung cells

A low-serum medium containing bovine serum albumin (BSA) was investigated with respect to the growth of and tissue plasminogen activator (TPA) production by human embryo lung (HEL) cells on microcarrier beads and in collagen gel. BSA and ferrous sulfate were chosen as substitutes for fetal calf serum (FCS) through a simple screening test involving many substances. The growth promoting effects of BSA and ferrous sulfate were independent of each other and from the FCS concentration. Though BSA inhibited initial cell attachment to the carrier surface, it did promote the growth of cells attached to microcarrier beads. Cells grown on microcarrier beads in the low-serum medium containing BSA, ferrous sulfate and 3% FCS produced an amount of TPA similar to that produced by ones grown in the 10% FCS medium. Although cells on the dish surface did not grow at all on serum-free media containing BSA and ferrous sulfate, cells in the collagen gel were able to grow slightly on the serum-free medium. Cells grown on the low-serum medium in collagen gel produced more TPA over a long period than those in the microcarrier beads using the low-serum medium. The optimum concentration of proteose peptone in the TPA production medium for the collagen gel culture was similar to that for the dish surface culture.     

Inhibition of Choline Acetyltransferase Activity by Serum Albumin Modified with Octanoic Acid and Other Fatty Acids

In this study, we examined the effect of fatty acids on choline acetyltransferase (ChAT) activity. ChAT is unstable in a solution of low protein concentration, so serum albumin (BSA) is usually added to stabilize the enzyme. However, we found that ChAT from bovine caudate nucleus rapidly lost its activity when diluted with a buffer containing commercial preparations of BSA. This effect was caused by octanoic acid, which was found in the gas chromatography/mass spectrometry system of lipid extract in commercial BSAs. The inhibition of ChAT activity by octanoic acid depended on the concentrations of the octanoic acid and of the albumin. We also found that ChAT activity was decreased by some long-chain fatty acids, arachidonic acid having exhibited the strongest effect. The extent to which arachidonic acid inhibited ChAT activity depended on the molar ratio of arachidonic acid and albumin, rather than upon the concentration of arachidonic acid. The effect of octanoic acid and arachidonic acid on ChAT activity appeared to increase in the presence of albumin.     

Modifications of protein by polyunsaturated fatty acid ester peroxidation products

The ability of unsaturated fatty acid methyl esters to modify bovine serum albumin (BSA) in the presence of a metal-catalyzed oxidation system [ascorbic acid/Fe (II)/O2] was investigated. The exposure of BSA to PUFA esters led to the generation of carbonyl groups and the formation of high-molecular-weight proteins, which were strongly dependent on the degree of fatty acid unsaturation. The high-molecular-weight proteins were detected by Western blot analysis and were not recognized by five antibodies. The observation that levels of conjugated dienes and malondialdehyde formed in the presence of BSA were substantially lower in the presence than in the absence of BSA indicated that radicals formed during the degradation of lipid hydroperoxides are likely involved in the formation of protein derivatives. These results may be important in understanding the specific roles of different polyunsaturated fatty acids in the pathophysiological effects associated with oxidative stress.     

Hepatocyte or serum albumin protein carbonylation by oxidized fructose metabolites: Glyceraldehyde or glycolaldehyde as endogenous toxins?

Excessive sugar intake in animal models may cause tissue damage associated with oxidative and carbonyl stress cytotoxicity as well as inflammation. Fructose became a 100-fold more cytotoxic if hepatocytes were exposed to a non-toxic infusion of H₂O₂ so as to simulate H₂O₂ released by Kupffer cells or infiltrating immune cells. In order to determine the molecular mechanisms involved, protein carbonylation of fructose and its metabolites were determined using the 2,4-dinitrophenylhydrazine method. In a cell-free system, fructose was found to carbonylate bovine serum albumin (BSA) only if low concentrations of FeII/H₂O₂ were added. Protein carbonylation by the fructose metabolites glyceraldehyde or glycolaldehyde was also markedly increased by FeII/H₂O₂. The protein carbonylation may be attributed to glyoxal formation by hydroxyl radicals as the glyoxal trapping agent aminoguanidine or hydroxyl radical scavengers prevented protein carbonylation. Glyoxal was also much more effective than other carbonyls at causing protein carbonylation. When BSA was replaced by isolated rat hepatocytes, fructose metabolite glyceraldehyde in the presence of non-toxic 2 μM FeII:8-hydroxyquinoline (HQ) and a H₂O₂ generating system (glucose/glucose oxidase) markedly increased cytotoxicity, protein carbonylation and reactive oxygen species (ROS)/H₂O₂ formation. Furthermore this was prevented by hydroxyl radical scavengers or aminoguanidine, a glyoxal scavenger. CuII: 8-hydroxyquinoline increased H₂O₂ induced hepatocyte protein carbonylation less but was prevented by aminoguanidine. However, cytotoxicity and protein carbonylation induced by glyceraldehyde/CuII:HQ/H₂O₂ were not affected by hydroxyl radical scavengers. Although fatty liver induced by an excessive sugar diet in animal models has been proposed as the first hit for non-alcoholic steatohepatitis (NASH) we propose that oxidative stress induced by the oxidation of fructose or fructose metabolites catalysed by Fenton FeII/H₂O₂ could be a ‘second hit’. A perpetual cycle of oxidative stress in hepatocytes could lead to cytotoxicity and contribute to NASH development.     

Octanoate reduces very low-density lipoprotein secretion by decreasing the synthesis of apolipoprotein B in primary cultures of chicken hepatocytes

Fatty acids of varying lengths and saturation differentially affect plasma apolipoprotein B (apoB) levels. To identify the mechanisms underlying the effect of octanoate on very low-density lipoprotein (VLDL) secretion, chicken primary hepatocytes were incubated with either fatty acid-bovine serum albumin (BSA) complexes or BSA alone. Addition of octanoate to culture medium significantly reduced VLDL-triacylglycerol (TG), VLDL-cholesterol and apoB secretion from hepatocytes compared to both control cultures with BSA only and palmitate treatments, but did not modulate intracellular TG accumulation. However, no differences in cellular microsomal triglyceride transfer protein levels were observed in the cultures with saturated fatty acid. In pulse-chase studies, octanoate treatment resulted in reduced apoB-100 synthesis, in agreement with its promotion of secretion. This characteristic effect of octanoate was confirmed by addition of a protease inhibitor, N-acetyl-leucyl-leucyl-norleucinal (ALLN), to hepatocyte cultures. Analysis showed that the level of apoB mRNA was lower in cultures supplemented with octanoate than in the control cultures, but no significant changes were observed in the levels of apolipoprotein A-I, fatty acid synthase and 3-hydroxy-3-methylglutaryl-CoA reductase mRNA as a result of octanoate treatment. Time-course studies indicate that a 50% reduction in apoB mRNA levels requires 12 h of incubation with octanoate. We conclude that octanoate reduced VLDL secretion by the specific down-regulation of apoB gene expression and impairment of subsequent synthesis of apoB, not by the modulation of intracellular apoB degradation, which is known to be a major regulatory target of VLDL secretion of other fatty acids.     

Developmental responses of 2-cell embryos to oxygen tension and bovine serum albumin in Wistar rats.

To improve rat embryo culture conditions, responses of Wistar 2-cell embryos from 2 breeders to oxygen tension (5 vs 20%) and bovine serum albumin (BSA) (0 vs 3 mg/ml) were examined using rat 1-cell embryo culture medium (mR1ECM). Supplementation of 3 mg/ml BSA significantly stimulated and accelerated development to the blastocyst and expanded blastocyst stages during 72 and 96 h culture, while reduced oxygen tension stimulated cell division. Fetus development after transfer of blastocysts obtained from 72 h culture under 5% O2 with BSA was significantly higher than those cultured under atmospheric oxygen without BSA. However, the nuclear numbers of in vitro cultured blastocysts and fetus development after embryo transfer were still significantly lower than in vivo developed blastocysts, indicating the current culture condition is still suboptimal.     

Dietary docosahexaenoic acid enhances ferric nitrilotriacetate-induced oxidative damage in mice but not when additional alpha-tocopherol is supplemented

Weaning mice were fed a diet supplemented with beef tallow (BT) or BT plus docosahexaenoic acid (DHA) containing 100 mg alpha-tocopherol/kg (alpha-Toc100) or 500 mg alpha-tocopherol/kg (alpha-Toc500) for 4 wk to modify membrane fatty acid unsaturation, and then were administered ferric nitrilotriacetate (Fe-NTA). The mortality caused by Fe-NTA was higher in the group fed the DHA (alpha-Toc100) diet than in the BT diet groups but the DHA (alpha-Toc500) diet suppressed this increase. Serum and kidney alpha-tocopherol contents were slightly influenced by the dietary fatty acids but not significantly. These results indicate that the increased unsaturation of tissue lipids enhances oxidative damage induced by Fe-NTA in mice fed DHA (alpha-Toc100) but not when additional alpha-tocopherol is supplemented. The apparent discrepancy between the observed enhancement by dietary DHA of oxidative damage and the beneficial effects of dietary DHA on the so-called free radical diseases is discussed in terms of strong bolus oxidative stress and moderate chronic oxidative stress.     

Role of protein supplements in the culture of mouse embryos

The effect of various types of proteins used in single protein supplements for Bigger-Whitten-Whittingham (BWW) medium on the in vitro development of mouse preimplantation embryos was evaluated. Thioredoxin, superoxide dismutase (SOD), and apotransferrin showed prominent growth-promoting activity, whereas bovine serum albumin (BSA), fatty acid-free BSA, and catalase showed moderate promoting effects. beta-lipoprotein, ovalbumin and hemoglobin were ineffective, and holo-type transferrin and ceruloplasmin were actually toxic to the embryos. These results suggest that the growth-promotive effect of proteins on mouse pronuclear stage embryos is due to their antioxidative action, or to the removal of some free metal ion(s) such as Fe(3+). The mild growth promoting effect of both BSA and fatty acid free BSA suggest that the effect mediated by BSA is not dependent on bound fatty acids, but more likely is due to their antioxidative effect or chelating effect.     

Dietary glycated protein modulates the colonic microbiota towards a more detrimental composition in ulcerative colitis patients and non-ulcerative colitis subjects

Aim:  To investigate the effect of native, heated and glycated bovine serum albumin (BSA) on the ulcerative colitis (UC) and non-UC colonic microbiota in vitro .
Methods and Results:  Continuous flow culture (CFC) models of the human colonic microbiota inoculated with faeces from UC and non-UC volunteers were maintained on BSA as growth substrate. Changes in bacterial populations and short-chain fatty acids were determined. UC and non-UC microbiota differed significantly in microbial populations, with elevated numbers of sulfate-reducing bacteria (SRB) and clostridia in the microbiota from UC patients. Compared with native BSA, glycated BSA modulated the gut microbiota of UC patients in vitro towards a more detrimental community structure with significant increases in putatively harmful bacteria (clostridia, bacteroides and SRB; P  < 0·009) and decreases in dominant and putatively beneficial bacterial groups (eubacteria and bifidobacteria; P  < 0·0004). The levels of beneficial short-chain fatty acids were significantly decreased by heated or glycated BSA, but were increased significantly by native BSA.
Conclusion:  The UC colonic microbiota maintained in CFC was significantly modified by glycated BSA.
Significance and Impact of the Study:  Results suggest that dietary glycated protein may impact upon the composition and activity of the colonic microbiota, an important environmental variable in UC.     

Protective effect of capsinoid on lipid peroxidation in rat tissues induced by Fe-NTA

The activity of a single IP administration (15 or 30 mg/Kg body weight) of vanillyl nonanoate, a simplified analog of capsiate, on ferric nitrilotriacetate (Fe-NTA)-mediated oxidative damage was investigated. A sub-lethal dose of Fe-NTA (15 mg Fe/Kg body weight) was administered IP to rats; animals were sacrificed, and kidney and plasma were collected 1 h after injection. In response to the Fe-NTA administration, a reduction of the levels of total lipids, total unsaturated fatty acids and cholesterol was observed, accompanied by a rise in the concentrations of malondialdehyde (MDA), conjugated dienes fatty acids hydroperoxides and 7-ketocholesterol in plasma and kidney 1 h after administration. A pre-treatment with synthetic capsiate (SCPT) showed remarkable protective effect on the reduction of the levels of total lipids, total unsaturated fatty acids and cholesterol, and the cellular antioxidant vitamin E, inhibiting the increase of MDA, conjugated dienes fatty acids hydroperoxides and 7-ketocholesterol in the plasma and kidney. The protective effect of SCPT and two analogues (vanillyl alcohol and vanillin) during the linoleic acid and cholesterol oxidation was investigated in in vitro systems, providing evidence of definite structure-activity relationships.