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OBJECTIVETransient neurological events (TNEs) occur frequently in the acute phase after direct bypass surgery for moyamoya disease (MMD), but there is currently no way to predict them. FlowInsight is a specialized software for analyzing indocyanine green (ICG) videoangiography taken with a surgical microscope. The purpose of this study was to investigate whether intraoperative evaluation of local hemodynamic changes around anastomotic sites using FlowInsight could predict the incidence and duration of TNEs.METHODSFrom patients who were diagnosed with MMD in our hospital between August 2014 and March 2017 and who underwent superficial temporal artery–middle cerebral artery bypass surgery, we investigated 25 hemispheres (in 22 patients) in which intraoperative ICG analysis was performed using FlowInsight. To evaluate the local cerebral hemodynamics before and after anastomosis, regions of interest were set at 3 locations on the brain surface around the anastomotic site, and the mean cerebral blood flow (CBF), mean gradation (Grad), mean transit time (MTT), and mean time to peak (TTP) were calculated from the 3 regions of interest. Furthermore, the change rate in CBF (ΔCBF [%]) was calculated using the formula (postanastomosis mean CBF − preanastomosis mean CBF)/preanastomosis mean CBF. ΔGrad (%), ΔMTT (%), and ΔTTP (%) were similarly calculated.RESULTSPostoperative stroke without TNE occurred in 2 of the 25 hemispheres. These 2 hemispheres (in 2 patients) were excluded from the study, and data from the remaining 23 hemispheres (in 20 patients) were analyzed. For each parameter (ΔCBF, ΔGrad, ΔMTT, and ΔTTP) calculated by FlowInsight, the difference between the groups with and without TNEs was significant. The median values for ΔCBF and ΔGrad were significantly higher in the TNE group than in the no-TNE group (ΔCBF 30.13 vs 3.54, p = 0.0106; ΔGrad 62.05 vs 10.78, p = 0.00435), whereas the median values for ΔMTT and ΔTTP were significantly lower in the TNE group (ΔMTT −16.90 vs −7.393, p = 0.023; ΔTTP −29.07 vs −7.02, p = 0.00342). Comparison of the area under the curve (AUC) for each parameter showed that ΔTTP had the highest AUC and was the parameter with the highest diagnostic accuracy (AUC 0.857). The Youden index revealed that the optimal cutoff value of ΔTTP was −11.61 (sensitivity 77.8%, specificity 71.4%) as a predictor of TNEs. In addition, Spearman’s rank correlation coefficients were calculated, and ΔCBF, ΔGrad, ΔMTT, and ΔTTP each showed a strong correlation with the duration of TNEs. The larger the change in each parameter, the longer the TNEs persisted.CONCLUSIONSIntraoperative ICG videoangiography findings were correlated with the occurrence and duration of TNEs after direct bypass surgery for MMD. Screening for cases at high risk of TNEs can be achieved by ICG analysis using FlowInsight.

Introduction We previously demonstrated that cyclic AMP-dependent protein kinase (PKA) phosphorylates neuronal nitric oxide synthase (nNOS) at Ser1412 in the hippocampal dentate gyrus after forebrain ischemia; this phosphorylation event activates NOS activity and might contribute to depression after cerebral ischemia. In this study, we revealed chronological and topographical changes in the phosphorylation of nNOS at Ser1412 immediately after subarachnoid hemorrhage (SAH). Methods In a rat single-hemorrhage model of SAH, the hippocampus and adjacent cortex were collected up to 24 h after SAH. Samples from rats that were not injected with autologous blood were used as controls. NOS was partially purified from crude samples via an ADP-agarose gel. Levels of nNOS, nNOS phosphorylated at Ser1412 (p-nNOS), PKA, and p-PKA at Thr197 were studied in the rat hippocampus and cortex using Western blot analyses and immunohistochemistry. Results According to the Western blot analysis, levels of p-nNOS at Ser1412 were significantly increased in the hippocampus, but not in the cortex, between 1 and 3 h after SAH. Immunohistochemistry revealed the phosphorylation of nNOS at Ser1412 and PKA at Thr197 in the dentate gyrus, but not in the CA1 area, 1 h after SAH. An injection of saline instead of blood also significantly increased levels of p-nNOS at Ser1412 in the hippocampus 1 h after the injection. Conclusions An immediate increase in intracranial pressure (ICP) might induce transient cerebral ischemia and promote the PKA-mediated phosphorylation of nNOS at Ser1412 in the dentate gyrus. This signal transduction pathway induces the excessive production of nitric oxide (NO) and might be involved in cognitive dysfunction after SAH.

Background: In moyamoya disease (MMD), the causes of differences in clinical features between children and adults and of the dramatic temporal changes in moyamoya vessels are poorly understood. We previously discovered elevated levels of m/z 4588 and m/z 4473 peptides in cerebrospinal fluid (CSF) in patients with MMD. This study examined the amino acid sequences of these peptides and quantified in specimens. Methods: The m/z 4588 and m/z 4473 peptides in CSF from patients with MMD were purified and concentrated by high-performance liquid chromatography and ultrafiltration. Liquid chromatography coupled with tandem mass spectrometry analysis was performed to identify the amino acid sequences of these peptides. We quantified these peptides in samples using sandwich enzyme-linked immunosorbent assay, and concentrations in CSF were compared between MMD (n = 40, 19 male; median age, 37 years) and non-MMD intracranial disease (n = 40, 19 male; median age, 39 years) as controls. Results: These peptides were identified as proenkephalin 143-183 (PENK 143–183). The concentration of PENK 143–183 was significantly greater in patients with MMD (median, 8,270 pmol/L) than control patients (median, 3,760 pmol/L; P < 0.001) and decreased in an age-dependent manner in MMD (r = −0.57; P < 0.001). The area under the receiver operating characteristic curve in children (age, ファイル公開日: 2019/01/01

In the present study, we developed a novel, simple, and specific detection method using an RP-HPLC at UV 285 nm for the separation and quantification of N-nitroso-bile acids. First, we found that N-nitroso-bile acids have a specific spectrophotometric absorbance at 285 nm. Using this 285 nm detection system, we could especially measure N-nitroso-bile acids, even in co-existence of non-N-nitroso-bile acids. Next, we observed the decomposition of N-nitroso-glychocholate under alkaline, acidic, and neutral conditions. N-nitroso-glychocholate rapidly decomposed under alkaline conditions (pH 9) (t(1/2) = 0.96 h), but remained fairly stable under acidic (pH 2) (t(1/2) = 12.8 h) and neutral (pH 7) (t(1/2) = 7.8 h) conditions. This study is the first report, which simply and specifically analyzes N-nitroso-bile acids using an RP-HPLC system.

The dietary components responsible for the development of Barrett's esophagus (BE) and esophageal adenocarcinoma (EAC) remain unclear. Wistar rats were divided into four groups based on their diet: a low soybean-oil diet, a low cow-fat diet, a high soybean-oil diet, and a high cow-fat diet. First, we evaluated the bile acid composition of the bile juice in each group without operation, using high-performance liquid chromatography. Because only high cow-fat intake induced changes in the composition of bile acids in bile juice, we then selected animals fed with a low soybean-oil diet and those with a high cow-fat diet to carry out esophago-jejunostomy for reflux of the duodenal contents, and compared sequential morphological changes between these groups up to 30 weeks after surgery. At 30 weeks after surgery, the reflux animals in the high cow-fat group showed a significantly higher incidence of BE and Barrett's dysplasia than those in the low soybean-oil group, and the incidence of EAC in the high cow-fat group was also slightly higher than that in the low soybean-oil group. High dietary animal fat changed the bile-acid composition and increased the concentration of taurine conjugates in the bile juice. These increased bile acids promoted the development of BE and Barrett's dysplasia leading to EAC.

Curcumin is a phenolic natural product isolated from the rhizome of Curcuma longa (turmeric). We evaluated the effects of curcumin on the development of dextran sulfate sodium (DSS)-induced experimental colitis. BALB/c mice were fed a chow containing either 3.5% (wt/wt) DSS or 3.5% DSS + 2.0% (wt/wt) curcumin. The body weight loss was more apparent in DSS-treated mice than in DSS + curcumin-treated mice. The disease activity index, histological colitis score, and MPO activity were all significantly higher in DSS-treated mice than in DSS plus curcumin-treated mice. Microscopically, mucosal edema, cellular infiltration, and epithelial disruption were much more severe in DSS-treated mice than in DSS + curcumin-treated mice. In DSS + curcumin-treated mice, NF-kappaB activation was blocked in the mucosa. In conclusion, the development of DSS-induced colitis was significantly attenuated by curcumin. Being a nontoxic natural dietary product, curcumin could be useful in treatment of IBD patients.

Interleukin (IL)-17 is a newly identified T-cell-specific cytokine. In this study, we investigated the effects of IL-17 on colony-stimulating factor (CSF) release in human colonic subepithelial myofibroblasts (SEMFs).CSF release and mRNA expression were determined by enzyme-linked immunosorbent assay (ELISA) and Northern blotting, respectively. Nuclear factor (NF)-kappaB- and activating protein (AP-1)-DNA binding activities were evaluated by electrophoretic gel mobility shift assays (EMSAs).Unstimulated cells secreted a small amount of granulocyte G- and granulocyte/macrophage (GM)-CSF, and a considerable amount of M-CSF. IL-17 weakly enhanced G-CSF release, but did not affect GM- and M-CSF release. IL-17 selectively enhanced tumor necrosis factor (TNF)-alpha-induced G- and GM-CSF release. The combination of IL-17 plus TNF-alpha induced a marked increase in NF-kappaB- and AP-1-DNA binding activities. The adenovirus-mediated transfer of a stable form of IkappaBalpha and/or a dominant negative mutant of c-Jun markedly inhibited the IL-17 plus TNF-alpha-induced G- and GM-CSF mRNA expression. Furthermore, a stability study showed that IL-17 plus TNF-alpha markedly enhanced the stability of G- and GM-CSF mRNA.IL-17 augments TNF-alpha-induced G- and GM-CSF release via transcriptional and posttranscriptional mechanisms.

The barrier functions in epithelial and endothelial cells seem to be very important for maintaining normal biological homeostasis. However, it is unclear whether or how bile acids affect the epithelial barrier. We examined the bile acid-induced disruption of the epithelial barrier. We measured the transepithelial electrical resistance (TEER) of Caco-2 cells as a marker of disruption of the epithelial barrier. Reactive oxygen species (ROS) generation was also measured. Cholic acid (CA) decreased the TEER and increased intracellular ROS generation. PLA2 (phospholipase A2), COX (cyclooxygenase), PKC (protein kinase), ERK 1/2 (extracellular signal-regulated kinase 1/2), PI 3 K (phosphatidylinositol 3-kinase), p38 MAPK (p38 mitogen-activated protein kinase), MLCK (myosin light-chain kinase), NADH dehydrogenase, and XO (xanthine oxidase) inhibitors or ROS scavengers prevented the CA-induced TEER decrease. PLA2, COX, PKC, NADH dehydrogenase, and XO inhibitors prevented the CA-induced ROS generation but not ERK 1/2, PI 3 K, p38 MAPK, and MLCK inhibitors. If the cells were treated with ROS generators such as superoxide dismutase, the TEER decreased. ERK 1/2, PI 3 K, p38 MAPK, and MLCK inhibitors prevent these ROS generators from inducing the TEER decrease. These results suggest that ROS play an important role. In addition, PLA2, COX, PKC, NADH dehydrogenase, and XO are located upstream of the ROS generation, but ERK 1/2, PI 3 K, p38 MAPK, and MLCK are downstream during the signaling of CA-induced TEER alterations.

In human pancreatic myofibroblasts, interleukin (IL)-17 markedly enhances tumor necrosis factor (TNF)-α-induced IL-6 secretion through the induction of IL-6 mRNA stabilization. Induced stability of IL-6 mRNA was markedly decreased by the inhibitors of extracellular signal-regulated kinase (ERKs), PD98059 and U0216. This indicates that activation of the ERK pathway is involved in the induction of IL-6 mRNA stabilization by IL-17 plus TNF-α.

Decay-accelerating factor (DAF) expressed on the surface of colonic cancer cells presents a barrier to complement-mediated clearance by contributing to the ineffectiveness of the humoral immune response. In this study, to investigate the mechanisms responsible for the anti-tumor effects of butyrate, we evaluated how butyrate modulates DAF expression in colonic cancer cells. Three colonic cancer cell lines (HT-29, Caco-2, and T84 cells) were studied. DAF protein expression was assessed by western blot, and DAF mRNA expression was evaluated by northern blot. Complement C3 deposition on the surface of colonic cancer cells was determined by enzyme-linked immunosorbent assay (ELISA). The promoter activity of the DAF gene was assessed by a reporter gene-luciferase assay. Butyrate reduced the basal and interleukin-4 (IL-4)- and tumor necrosis factor-alpha (TNF-alpha)-induced expression of DAF protein and mRNA in HT-29 cells. It increased the susceptibility to complement attack and enhanced C3 deposition on HT-29 cells. The inhibitory effect of butyrate on DAF mRNA expression was also observed in T84 and Caco-2 cells. Butyrate decreased basal DAF expression at both transcriptional and post-transcriptional levels. The inhibitory effect of butyrate on IL-4-induced DAF expression was closely associated with a blockade of IL-4-induced DAF mRNA stability. TNF-alpha-induced transcriptional activation and the increased stability of the DAF gene were also blocked by butyrate. Similar but weak effects were induced by trichostatin A, a potent histone deacetylase inhibitor, suggesting that histone acetylation might participate in butyrate activity. These observations indicate that both a down-regulation of DAF expression and the induction of susceptibility to complement attack contribute to the anti-tumor effects of butyrate in colonic cancer.

Microorganisms are reported to use siderophores as iron carriers in their high-affinity type iron-uptake systems. Since siderophores form rather stable complexes with ferric iron, the release of iron from internalized complexes is probably accomplished through reduction to ferrous forms, being catalyzed by ferric reductases. In the present study, one of the ferric reductases from the soluble fraction of Vibrio anguillarum E6-5 was partially purified and characterized. This was an NADH-dependent ferric reductase with ability to reduce different iron complexes such as ferric citrate, ferric-EDTA, and ferric chloride. Effects of flavin derivatives, metal ions, dithiothreitol (DTT), and β-mercaptoethanol were also studied. In gel filtration chromatography, the enzyme showed an apparent molecular mass of 26 kDa. Upon native-PAGE analysis, the partially purified preparation gave a single band with ferric reductase activity.

In this study, we isolated three kinds of antigenic polysaccharide components (tentatively designed as AP-1∼3) from cells of Leptospira biflexa serovar patoc strain Patoc I ( L. biflexa patoc Patoc I) by the hot phenol–water procedure, followed by treatment with mild acid and column chromatography. Two of them (AP-1 and AP-2) were recovered from the phenol-soluble fraction whereas another (AP-3) was recovered from the aqueous fraction. All of them reacted toward an anti- L. biflexa serum and also cross-reacted in similar extents toward most of the other leptospiral antisera tested. Such immunoreactions were specifically inhibited by a β-(1→4)-linked mannobiose, but were not by any mono- and oligosaccharide tested. From their structural analyses including 1 H and 13 C NMR spectrometry, Smith degradation and methylation analysis, it was revealed that all of these antigenic polysaccharides had the same disaccharide unit →3)-β- d -Man p -(1→4)-β- d -Man p -(1→ in their major polysaccharide parts, but they differed in the acyl substituents. Therefore it is most likely that such mannobiose unit is a candidate for the antigenic epitopes of L. biflexa polysaccharides.

Germinated barley foodstuff (GBF), which is derived from the aleurone and scutellum fractions of germinated barley, is rich in water-insoluble dietary fiber, which reportedly modulates mineral absorption in vivo. This study was conducted to examine whether GBF affects mineral absorption. We performed three experiments. In experiment 1, female ICR mice (4 weeks old) were fed a diet supplemented with 10% GBF or cellulose (CE; as a control) for 24 weeks. Body weight and food intake were recorded twice weekly. On the last experimental day, serum biochemical parameters, including electrolyte parameters, were determined, and major digestive organs were examined histopathologically. In experiment 2, male Sprague-Dawley rats (5 weeks old) were fed the CE and GBF diets, with the mineral content of the CE diet adjusted to the same level as that in the GBF diet, for 14 days, and the intake and fecal excretion of minerals (calcium, Ca; magnesium, Mg: Iron, Fe; Phosphate, P) were determined for the last 3 days. In experiment 3, the binding of Ca and Mg to GBF or cellulose and the release of Ca and Mg from GBF or cellulose was measured in phosphate buffer (pH 6.8). Long-term administration of GBF did not affect the growth rate of mice. GBF did not inhibit the absorption of Ca and Mg, and rather promoted the absorption of Ca and Mg by the gastrointestinal tract. The absorption of Fe and P was not attenuated by GBF administration, and GBF contained only a low level of phytic acid. GBF did not inhibit mineral absorption, differing from the effect of other dietary fibers, and is expected to be useful as a dietary source of Ca and Mg.

SUMMARY Ws/Ws rats have a small deletion of the c-kit gene, and are deficient in both mucosal-type mast cells (MMC) and connective tissue-type mast cells (CTMC). In the present study we investigated the role of intestinal MMC in the development of dextran sulphate sodium (DSS)-induced experimental colitis using Ws/Ws rats. Ws/Ws and control (+/+) rats were given a 3% DSS aqueous solution orally for 10 days, and the subsequent mucosal damage was evaluated macroscopically and histologically. The mucosal myeloperoxidase (MPO) activities and histamine levels were also measured. (i) DSS induced severe oedema and hyperaemia with sporadic erosions in the control (+/+) rats, but these changes were significantly attenuated in the Ws/Ws rats (P

Elemental diet (ED) therapy has been established as primary therapy for Crohn's disease, and home enteral nutrition (HEN) has been reported to control relapse at a dose of more than 30 kcal/kg of ideal body weight. However, a decrease in ED compliance with long-term use is becoming problem. We developed an n-3 fatty acid-rich diet and carried out nutritional education specifically for Crohn's disease patients using HEN to facilitate compliance and to improve their nutritional status. After the introduction of this n-3 rich diet, disease activity was not altered, and nutritional status, especially serum n-3 fatty acid levels, improved. The remission periods in patients with poor compliance seemed to be prolonged by the nutritional education. Thus, a n-3 rich diet in combination with nutritional education specific for Crohn's disease patients is very important for the in maintenance of high compliance and for maintaining nutritional balance.

Bacillus cereus CH was shown to excrete chitinases into the culture supernatant when cultivated in a medium containing 0.2% colloidal chitin, whereas the removal of colloidal chitin resulted in a low activity. After concentration of the culture supernatant by precipitation with ammonium sulfate, the induced chitinases were purified by sequential chromatography. Four different chitinases, A, B1, B2, and B3 with molecular masses of 35, 47, 58, and 64 kDa, respectively, were separated. All chitinases showed similarities in their kinetic parameters when observed with colloidal chitin, including an optimal pH of 5.0-7.5, and an optimal temperature between 50-60 degrees C. Chitinase A hydrolyzed glycol chitin and p-nitrophenyl-di-N-acetyl-beta-chitobioside at similar rates to that of colloidal chitin, whereas group B chitinases hydrolyzed both substrates in much lower rates. From analyses of the reaction products, it is most likely that chitinase A and all group B chitinases hydrolyze the substrates tested in an endo-fashion. However, group B chitinases were distinct from chitinase A in possessing high transglycosylation activity. From amino terminal sequencing, chitinases B1, B2, and B3 were shown to have almost identical sequences, which differed from that of chitinase A. The similarities in the reaction modes and amino terminal sequences among chitinases B1, B2, and B3 suggest that these chitinases may be derived from a presumptive precursor protein through C-terminal processing.

As far as the flame-retardant polyester fibers are concerned, the copolymerization of phosphorus retardants is the most common method. But a serious problem is that the phosphorus-containing polymer is easily hydrolyzed. We investigated the flame retardancy and the hydrolysis properties of two poly(ethylene terephthalate) (PET) fibers, one with a phosphorus compound as a side chain (side-chain type: HEIM® Toyobo Co., Ltd.), and one with a phosphorus compound inserted in the polymer backbone (main-chain type). Both types had almost the same properties of fibers and flame retardancy, but the main-chain type was hydrolyzed about two times faster than the side-chain type, and led to a decrease of toughness immediately. This difference of hydrolysis properties between main-chain type and side-chain type depends on whether a phosphonate ester bond is placed in the polymer backbone or the pendant site. In the case of the main-chain type, the scission of the polymer backbone chain occurs by hydrolysis of phosphonate ester bonds; however, in the case of the side-chain type, this does not occur. These results demonstrate that the flame-retardant polyester fiber with the side-chain type modifier gives sufficient flame retardancy and excellent hydrolysis resistance. © 2000 John Wiley & Sons, Inc. J Appl Polym Sci 78: 1134–1138, 2000

SUMMARY Ws/Ws rats have a small deletion of the c-kit gene, and are deficient in both mucosal and connective tissue-type mast cells. In this study, the role of mucosal type mast cells (MMC) in the development of intestinal ischaemia-reperfusion injury was investigated in Ws/Ws rats. Autoperfused segments of the jejunum were exposed to 60 min of ischaemia, followed by reperfusion for various time periods. The epithelial permeability was then assessed by the 51Cr-EDTA clearance rate. In the control (+/+) rats, the maximal increase in mucosal permeability was achieved at 45 min of reperfusion. In contrast, this increase was significantly and potently attenuated in the Ws/Ws rats. Mucosal alkaline phosphatase activity decreased in the control (+/+) rats, but was not altered in the Ws/Ws rats. There were no differences in mucosal myeloperoxidase activity, indicating that granulocytes did not contribute to tissue injury. These results provide direct evidence for the role of mast cells in the pathogenesis of intestinal ischaemia-reperfusion injury.

We reported previously a method for the separation and quantification of sulfated polysaccharides, dextran sulfate sodium (DSS), using fluorometric labeling and size-exclusion chromatography (SEC). In the present study, we evaluated chemical depolymerization of DSS. In fluorometric analyses using SEC, pyridylamino-DSS (PA-DSS) was depolymerized under acidic conditions, but not under alkaline conditions. In addition, we used ion-pair liquid chromatography (IPLC) and UV analyses for the separation and quantification of sulfate. Sulfate was considerably depleted from DSS under alkaline conditions, but little depleted under acidic conditions. Spectrophotometric studies using toluidine blue revealed that molecular mass is an important factor and a minimum molecular mass of 2500 is needed to induce metachromasia. Fluorometric analyses using SEC and UV analyses using IPLC allow for the detection of DSS, reactants and sulfate.

Objective Hemangioblastomas remain a surgical challenge because of their arteriovenous malformation–like character. Recently, indocyanine green (ICG) videoangiography has been applied to neurosurgical vascular surgery. The aim of this study was to evaluate the usefulness of tumor blood flow imaging by intraoperative ICG videoangiography in surgery for hemangioblastomas. Methods Twenty intraoperative ICG videoangiography procedures were performed in 12 patients with hemangioblastomas. Seven lesions were located in the cerebellum, two lesions were in the medulla oblongata, and three lesions were in the spinal cord. Results Ten procedures were performed before or during dissection, and 10 procedures were performed after tumor resection. ICG videoangiography could provide dynamic images of blood flow in the tumor and its related vessels under surgical view. Interpretation of these dynamic images of tumor blood flow was useful for discrimination of transit feeders (feeders en passage) and also for estimation of unexposed feeders covered with brain parenchyma. Postresection ICG videoangiography could confirm complete tumor resection and normalized blood flow in surrounding vessels. Conclusions In surgery for hemangioblastomas, careful interpretation of dynamic ICG images can provide useful information on transit feeders and unexposed hidden vessels that cannot be directly visualized by ICG.

Paranitrophenylphosphate (pNPP) induced fluorescence changes in fluorescence isothiocyanate (FITC)-labeled Na+,K(+)-ATPase preparations. The extents of changes were similar to those induced by acetylphosphate (AcP) accompanying accumulation of a K(+)-sensitive phosphoenzyme (E2P) and an ouabain bound phosphoenzyme in the presence of Mg2+ and 16 mM Na+. Phosphoenzymes formed from [32P]pNPP were shown to turn over. The ratio of the maximum amount of the phosphoenzyme formed from pNPP to that of the phosphoenzyme formed from ATP and that of the ouabain-enzyme complex under steady-state conditions was shown to be close to 1:0.5:1. Such extra phosphorylation has hitherto only been observed in a transient state with the additions of high concentrations of ATP [Peluffo, R.D., Garrahan, P.J., and Rega, A. (1992) J. Biol. Chem. 267, 6596-6601]. Our data are compatible with the simultaneous presence of high and low affinity ATP-binding sites in Na+,K(+)-ATPase [Hamer, E. and Schoner, W. (1993) Eur. J. Biochem. 213, 743-748]. The maximum amount of paranitrophenol-sensitive fraction to synthesize [32P]pNPP in fully accumulated ADP-sensitive phosphoenzyme (E1P) from [32P]ATP was around 1/4 of the amount of ouabain-enzyme complex. These data and others indicate that a much higher degree of oligomerization, rather than (alpha beta)2, may be the functional unit of the enzyme in the membranes.

We hypothesized that the severity of dextran sodium sulfate (DSS)-induced colitis could differ between DSS preparations of the same molecular weight, and that this difference may be affected by the sulfur content. To test this, we used three DSS preparations of similar molecular weights but with different sulfur contents. Three DSS preparations with molecular weights of 40,000 to 50,000 were tested: MP Biomedicals (MP Bio), USB (USB), and The Lab Depot (The Lab). Epithelial cell lines were used to assess the levels of poly (ADP-ribose) polymerase (PARP) in the presence of 2.0% DSS in vitro. Eight-week-old female C57/B6 mice were fed 2.0% DSS in water for 1 week, and then sacrificed to investigate the effects of the DSS preparations in vivo. In vitro experiments using CaCo-2 and CMT-93 cells revealed decreased PARP levels from all DSS preparations. Notably, the PARP level was significantly decreased in CaCo-2 cells treated with DSS from USB as compared to The Lab Mice treated with The Lab DSS had significantly decreased body weight losses on day 7 as compared to mice receiving DSS from MP Bio and USB. This result was supported by their DAI score, colon weight/length ratio, and histological scores. The severity of colitis can differ between similar DSS preparations of the same molecular weight range. This difference in colitogenic properties may be affected by the total sulfur content of each DSS preparation.

The O-polysaccharide chain (PS-1), released by mild acidic treatment of the LPS of V. anguillarum V-123 (serogroup JO-2), a pathogenic bacterium of marine and estuarine fish, consists of 2-amino-2-deoxy-D-galacturonic acid, 2-amino-2,6-dideoxy-D-glucose (D-quinovosamine), and 4-amino-4,6-dideoxy-D-glucose (D-viosamine) N-acylated with 2,4-dihydroxy-3,3,4-trimethylpyroglutamic acid. Strong-acid hydrolysis of PS-1 afforded alpha-GalNA-(1----4)-alpha-GalNA-(1----3)-QuiN (A1) and alpha-GalNA-(1----3)-QuiN (A2), and hydrolysis with hydrogen fluoride gave N-acetylated A1 and 4-amino-4,6-dideoxy-D-glucose N-acylated by 2,4-dihydroxy-3,3,4-trimethylpyroglutamic acid. Mild treatment of PS-1 with alkali removed the N-formyl substituents and Smith degradation of the product gave alpha-QuiNAc-(1----3)-beta-VioNAcyl-(1----3)-alpha-GalNAc A-(1----3)-2,3,4- trihydroxybutanoic acid (S1) and S2 in which the carboxyl group of the GalNAcA residue was amidated. Thus, the repeating unit of the O-polysaccharide is----3)-alpha-GalNAcA(amino)-(1----4)-alpha-GalNFoA-(1----3 )- alpha-QuiNAc-(1----3)-beta-VioNAcyl-(1----in which the N-Acyl group is 2,4-dihydroxy-3,3,4-trimethylpyroglutamic acid and Fo is formyl.

Sake yeast can produce high levels of ethanol in concentrated rice mash. While both sake and laboratory yeast strains belong to the species Saccharomyces cerevisiae, the laboratory strains produce much less ethanol. This disparity in fermentation activity may be due to the strains' different responses to environmental stresses, including ethanol accumulation. To obtain more insight into the stress response of yeast cells under sake brewing conditions, we carried out small-scale sake brewing tests using laboratory yeast strains disrupted in specific stress-related genes. Surprisingly, yeast strains with disrupted ubiquitin-related genes produced more ethanol than the parental strain during sake brewing. The elevated fermentation ability conferred by disruption of the ubiquitin-coding gene UBI4 was confined to laboratory strains, and the ubi4 disruptant of a sake yeast strain did not demonstrate a comparable increase in ethanol production. These findings suggest different roles for ubiquitin in sake and laboratory yeast strains.

The pathogenic mechanisms responsible for inflammatory bowel disease (IBD) are poorly understood. In an IBD animal model, the oral administration of polysaccharides such as dextran sulfate sodium (DSS) induced colitis, which exhibited several clinical and histological features similar to IBD. However, pathogenic factors in the development of colitis remain unclear, as do the chemical properties of DSS. Therefore, we investigated the chemical properties of DSS in this study. DSS reacted under 1 N acidic or alkaline conditions, and the reactants were analyzed using size exclusion and ion chromatography. The reactants were also analyzed in terms of metachromasia as a peculiarity of sulfated polysaccharides. DSS was depolymerized to a Mr 2500 moiety under acidic, but not under alkaline conditions. Sulfate was depleted from DSS under alkaline conditions (30%), but less so under acidic conditions (10%). In addition, autoclave treatment induced depolymerization to a Mr 1800 moiety and depleted the sulfate. This depolymerized DSS under acidic conditions has as many sulfates as a native DSS molecule with a Mr of 5000, exhibiting metachromasia as a peculiarity of sulfated polysaccharides. Therefore, these Mr 2500 or 1800 DSS molecules may be useful in investigating the mechanisms of DSS-induced colitis.

Recent studies have suggested that the enhanced release of reactive oxygen species (ROS) plays an important role in the pathogenesis of clinical inflammatory bowel disease (IBD), such as ulcerative colitis and Crohn's disease. In the present study, we investigated the effects of the free radical scavengers edaravone and tempol in the development of experimental dextran sulfate sodium (DSS)-induced colitis in mice. Male BALB/cA mice were fed 4% (w/w of diet) DSS in standard powder chow for 8 days. Edaravone, tempol, or vehicle saline were then injected subcutaneously twice per day. After the experimental period, the colonic length, histological damage score, and mucosal myeloperoxidase (MPO) and serum interleukin-6 (IL-6) levels were measured. Edaravone (15 mg/kg/day) and tempol (5-15 mg/kg/day) suppressed the colonic shortening and the damage score. In particular, tempol at 15 mg/kg/day significantly attenuated the colonic shortening and damage score. Edaravone and tempol suppressed the serum IL-6 levels, and significantly suppressed the increased colonic MPO levels. These results strongly support the involvement of ROS in the pathogenesis of DSS-induced colitis. A clinical effect for edaravone and tempol in IBD patients is strongly expected.

[structure: see text] The jewel pendant ligand has multiple chromogenic units combined in a single molecule with the dyes linked to a semiselective binding site by three heteroatoms (O, N, S) having different HSAB characteristics, to indicate diverse response to individual transition metal ions. Using a single-molecular multianalyte sensor, multiple analytes could be determined with a minimal sensing system.

Dextran sulfate sodium (DSS) is a strong negatively charged heparin-like polysaccharide and has anti-immunodeficiency virus, anti-carcinogenesis, or occasionally tumor-promotion effects. The biological metabolism of DSS, however, remains unclear. In a previous study, we reported a novel method for the separation and quantification of DSS, using fluorometric labeling with 2-aminopyridine and a combination of size-exclusion and reverse phase high-performance liquid chromatography. In the present study, we have applied this method for analyses of in vitro chemical or enzymatic depolymerization of pyridylamino-DSS (PA-DSS). PA-DSS was depolymerized by specific enzymes such as alpha-amylase and alpha-glycosidase, but not by dextranase or heparinase. Unknown enzymes derived from cultured intestinal cells also strongly depolymerized PA-DSS as did alkaline substances. On the other hand, we have established a novel detection system using a post-column reaction. This method utilizes the spectrophotometrically metachromatic reaction of toluidine blue solution with DSS. This novel detection system may be specific and may potentially provide useful information in the analyses of sulfated polysaccharides, which are present in environmental and biological materials.

Bile acids, especially those with hydrophobic properties, are known to possess cytotoxicity. However, the mechanisms responsible for the cytotoxicity of bile acids are still under investigation. On the other hand, the hydrophilic bile acid, ursodeoxycholic acid has been reported to exhibit therapeutic effects against cytotoxic hydrophobic bile acids. The aim of the present study was to investigate the cytotoxicity of individual bile acids and combinations of bile acids using the intestinal cell lines IEC-6 and Caco-2 cells. The cytotoxicities of individual bile acids and the effects of various bile acid combinations were evaluated using the MTS [3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay. The bile acids induced cytotoxic effects depending on their hydrophobicity except for hyodeoxycholic acid. In the study for the effects of combined bile acids, not only ursodeoxycholic acid but other hydrophilic bile salts such as cholic acid and hyocholic acid exhibited cytoprotection against deoxycholic acid-induced cytotoxicity. Moreover, even some hydrophobic bile acids, such as chenodeoxycholic acid also exhibited cytoprotection. It is possible that the cytotoxicity of hydrophobic bile acids is ameliorated by more hydrophilic bile acids under certain conditions. The understanding of the precise mechanism of this phenomenon remains to be determined.

Dietary fat is an important factor involved in the pathogenesis of inflammatory bowel disease (IBD). It remains unclear how n-3 and n-6 fatty acids modulate intestinal inflammation. In this study, we investigated the effects of n-3 and n-6 fatty acid-rich diets on trinitrobenzene sulfonic acid (TNBS)-induced enteritis. The rats were fed an n-3 or n-6 rich-diet for 12 days, and then starved for the following 2 days. An intraileal injection of TNBS was administered, and TNBS-enteritis subsequently developed. Macroscopic and histological examination was performed after 24 h. Serum cytokine levels were determined by ELISA. The n-6 fatty acid-rich diet markedly enhanced mucosal damage as compared to the n-3 fatty acid-rich diet. The damage score was significantly higher in the n-6 fatty acid-rich diet group (P

Recent studies suggest that the enhanced release of reactive oxygen species (ROS) plays an important role in the pathogenesis of clinical inflammatory bowel diseases (IBD) such as ulcerative colitis and Crohn's disease. In the present study, we investigated the effects of the free radical scavenger edaravone, which is used clinically as an anti-stroke agent, in the development of experimental dextran sulfate sodium (DSS)-induced colitis in rats. The rats were fed 4% (w/w of diet) DSS in standard powder chow for 8 days. The edaravone and vehicle saline were injected subcutaneously twice a day. After the experimental period, the wet colonic weight, macroscopic mucosal damaged area, histological damage score, mucosal myeloperoxidase (MPO) activity, mucosal tissue lipid peroxidate and serum interleukin-6 (IL-6) levels were measured. In the DSS-induced colitis model, edaravone treatment (1-20 mg/kg day) significantly reduced the wet colonic weight, macroscopic damaged area, and the histological damage score. Edaravone treatment also reduced mucosal MPO activity, mucosal tissue lipid peroxidate level and serum IL-6 level. In particular, edaravone at a dose of 20 mg/kg day significantly reduced mucosal MPO activity and serum IL-6 level. These results strongly support the involvement of ROS in the pathogenesis of DSS-induced colitis. A clinical effect for edaravone against IBD patients is strongly expected.

Recent studies suggest that the enhanced release of reactive oxygen species (ROS) plays an important role in the pathogenesis of clinical acute pancreatitis. In the present study, we investigated the effects of the free radical scavenger edaravone, which is used clinically as an anti-stroke agent, in the development of experimental closed duodenal loop (CDL)-induced acute pancreatitis. In the CDL-pancreatitis model, after edaravone and vehicle saline were injected intravenously, pancreatitis was induced for 7 h by the CDL technique. The subsequent ascites volume, wet pancreatic weight, serum amylase levels, and pancreatic tissue lipid peroxide levels were evaluated. Pancreatic tissue damage was also evaluated histologically. In this CDL-induced pancreatitis model, edaravone treatment tended to reduce the ascites volume and inhibit the increases in the wet pancreatic weight. Edaravone also tended to reduced the microscopic mucosal damage scores and pancreatic tissue lipid peroxide levels. In particular, the serum amylase levels in the edaravone-treated rats (1-20 mg/kg i.v.) were significantly reduced as compared to the vehicle-treated rats. These results strongly support the involvement of ROS in the pathogenesis of CDL-induced acute pancreatitis and cytoprotective effects of free radical scavender against pancreatic acinar cells. A clinical effect for edaravone against acute pancreatitis is strongly expected.

Germinated barley foodstuff (GBF), which contains glutamine-rich protein and hemicellulose-rich fiber, exhibits therapeutic effects in ulcerative colitis; however, its mechanism is still under investigation. The aim of this study was to evaluate the anti-inflammatory effects of GBF on colitis in terms of the epithelial inflammatory response. Methods: Mice with dextran sulfate sodium-induced colitis were used. The effects of GBF on the colitis were evaluated by measuring the body weight; disease activity; mucosal damage (histology, mucosal inflammatory parameters, nuclear factor kappa B [NFkB] activation, and signal transducer and activator of transcription 3 [STAT3]); serum interleukin 6 (IL-6) level; cecal short-chain fatty acids (SCFAs); and bile acid contents. Results: GBF significantly prevented disease activity and body weight loss after induction of colitis. Serum IL-6 level and mucosal STAT3 expression were also significantly attenuated, with a conspicuous reduction of mucosal damage; NFkB activity showed the same tendency. Cecal butyrate content was significantly higher and, interestingly, GBF mice had lower bile acid concentrations than the control group. Conclusions: GBF has the potential to reduce the epithelial inflammatory response by depressing STAT-3 expression and inhibiting NFkB binding activity. These effects may be brought about by an increase of butyrate production and adsorption of bile acids.

IL-17 enhances the TNF-α-induced IL-6 and IL-8 secretion in human colonic subepithelial myofibroblasts. In this study, we investigated how IL-17 modulates RANTES secretion in these cells. TNF-α potently induced RANTES secretion, but IL-17 dose-dependently inhibited the TNF-α-induced RANTES secretion. This was also observed at the mRNA level. Even after pretreatment with TNF-α for 12 h, the inhibitory effect of IL-17 was detectable. IL-17 did not affect the TNF-α-induced stability of the RANTES gene. IL-17 significantly decreased the TNF-α-induced increase in RANTES promoter activity, and IL-17 actually blocked the TNF-α-induced RANTES gene transcription. EMSAs demonstrated that IL-17 did not modulate the TNF-α-induced NF-κB DNA-binding activity, but markedly decreased TNF-α-induced IFN regulatory factor-1 (IRF-1) DNA-binding activity. Because cooperation between NF-κB and IRF-1 is important in the TNF-α-induced RANTES gene expression, the major mechanism mediating the inhibitory effect of IL-17 may be achieved by the inhibition of IRF-1 DNA-binding activity.

The pathogenesis of inflammatory bowel disease (IBD) remains unknown. It has been suggested that luminal factors such as the microflora, short chain fatty acids (SCFAs), food antigens and so on play important roles in the disease progression. Many reports have revealed alterations in the SCFA and organic acid concentration of the colon, especially increased lactate and decreased butyrate, in IBD patients. The mechanisms responsible for these alterations, however, remain unclear. Therefore, the effects of aerobic conditions on the alterations in the SCFA and organic anion levels in the feces was evaluated. Fecal specimens were collected from 5 healthy volunteers. Under aerobic condition, a mixture of feces and distilled water was incubated in 37 degrees C for 1 and 3 h. The pH, osmotic pressures, the concentrations of potassium, bicarbonate, SCFA and organic anion, and the activities of alpha-amylase and lactate dehydrogenase (LDH) in the mixture were then measured. We also examined any changes in the microscopic microflora under the hypotonic and aerobic conditions. The results showed the osmotic pressure, and the concentrations of lactate and SCFAs (formate, acetate, propionate and n-valerate) were progressively increased with longer incubation times, and reached a statistically significant difference. In particular, the ratios of lactate, succinate and n-valerate after 1 and 3 h of incubation increased remarkably. In contrast, the electrolyte levels and both alpha-amylase and LDH activities were not altered significantly. Microscopically, the microflora in the mixture decreased with prolonged incubation times. These data suggest that under these in vitro conditions, the organic anion and SCFA levels in the feces easily increased. It is probable that the alterations in the SCFA and organic anion levels in IBD patients may be partly due to intracellular components derived from microflora destroyed under hypotonic and aerobic conditions in the colonic lumen, for example caused by mucosal bleeding. These alterations may influence the pathogenesis and progression of IBD.

Recent studies have suggested that short chain fatty acids (SCFAs) exert a therapeutic effect on some human and experimental animal diseases. Clostridium butyricum produces high levels of SCFAs in the gut lumen. The aim of the present study was to analyze the product derived from Clostridium butyricum in a culture system, and to develop methods to eliminate the odor derived from SCFAs in the product. Clostridium butyricum was incubated in CS medium for 24 h and subsequently in CS broth for 24 h. The suspension of Clostridium butyricum in the broth was centrifugated and the supernatant was analyzed. The results showed this product contained high levels of SCFAs, especially acetic acid and n-butyric acid. Many food materials were tested in order to eliminate the odor derived from SCFAs in the product. Of the food materials tested, yogurt was shown to most effectively eliminate the odor. Using a yogurt base, we prepared a special food additive. Use of the additive completely eliminated the odor of the product derived from Clostridium butyricum. Finally, we administered the product with the additive to Sprague-Dawley rats for 14 days. The rats grew normally for the duration of the experimental period. It is possible that this novel product with the additive exerts therapeutic effects on some gastointestinal disorders.

Structural studies were carried out on an O-antigenic polysaccharide moiety derived from Porphyromonas circumdentaria NCTC 12469, a reference strain of Porphyromonas species. The polysaccharide chain was composed of D-glucose, D-galactose, N-acetyl-D-glucosamine, and N-acetyl-D-galactosamine in a molar ratio of 1:2:1:1. On the basis of results from 1H- and 13C-NMR spectroscopic analyses including COSY, TOCSY, and HMQC experiments together with results of Smith degradation, methylation analysis, and partial acid hydrolysis, it is concluded that the polysaccharide chain has a pentasaccharide repeating unit of -->6)-beta-D-Glcp-(1-->6)-beta-D-Galp-(1-->3)-beta-D-GlcpNAc-(1-->3)-beta-D-GalpNAc-(1-->. The immunoreaction between P. circumdentaria LPS and the corresponding antiserum was strongly inhibited by the pentasaccharide fragment (Glc-Gal-Gal-GlcNAc-GalNAc) isolated from partial acid hydrolysis of the above polysaccharide, suggestive of O-antigen specific antibodies in the used antiserum.

Bile acids have been shown to exhibit varying degrees of cytotoxicity, depending on their hydrophobic-hydrophilic balance. We have recently reported the strong cytotoxicity of hyodeoxycholic acid (HDCA), and the aim of the present study is to investigate the mechanisms underlying the cytotoxicity of HDCA.The intestinal cell lines IEC-6 and Caco-2 cells were used. The cytotoxicities of various bile acids were evaluated using the MTS assay; their cytolytic effects were measured using the LDH release assay. The induction of apoptosis was determined by the specific figure changes in the cellular cytoplasm and nucleus, including DNA ladder formations. IL-8 synthesis induced by the bile acids was measured using an ELISA assay.The bile acids induced cytotoxic effects, LDH release, IL-8 synthesis and apoptosis, depending on their hydrophobic properties. On the other hand, HDCA induced strong cytotoxicity, apoptosis and IL-8 synthesis but not cytolysis, although HDCA has a hydrophilic nature. In addition, HDCA exerted the strongest effects on dispersing monolayer cells.These results strongly suggest that HDCA induces cytotoxicity through its ability to induce apoptosis rather than its detergent effect.

The present study describes a size-exclusion high-performance liquid chromatographic method for the separation and quantification of sulfated polysaccharides, such as dextran sulfate sodium (DSS). Pyridylamination of DSS was achieved without difficulty using 2-aminopyridine as a fluorometric label. In addition, 0.1-0.2 M phosphate buffer (pH 3.0) was found to be the mobile phase which produced the best separation. In vitro enzymatic degradation of the pyridylamino-DSS (PA-DSS5000, Mr 5000) using alpha-amylase and the in vivo metabolism in the rat feces after oral administration of PA-DSS5000 were then evaluated. Two small peaks of approximately Mr 380 and 600 appeared after co-incubation with alpha-amylase, indicating PA-DSS5000 may be considerably depolymerized. In vivo, however, PA-DSS5000 excreted in the feces was mainly of PA-DSS5000 polymer. No peaks of less than Mr 5000 were not clearly detectable in the feces because of background fluorescence attributable to gut lumen contents. This method of fluorometric analysis allows fairly selective detection of sulfated polysaccharides in biological materials.

BACKGROUND The physiological effects on faecal bile acids and short chain fatty acids (SCFAs) or intestinal microflora in dextran sulphate sodium (DSS)-induced colitis remain unknown and are an area of interest DESIGN ALTERATIONS: of these parameters in DSS-induced colitis in rats were evaluated. METHODS Male Sprague-Dawley rats (n = 10) were given a 3% DSS aqueous solution orally for 7 days. The concentrations of bile acids and SCFAs in the faeces were measured using gas chromatography and high-performance liquid chromatography. Intestinal microflora, especially anaerobes, were investigated by microbiological methods. RESULTS On day 7, the concentrations of lithocholic acid and alpha-muricholic acid were significantly decreased and that of cholic acid was significantly increased. There was a strong correlation between the concentration of cholic acid and the macroscopic area of damaged tissue in the colon (R = 0.74, P < 0.05). With respect to SCFAs, DSS administration significantly decreased the concentrations of acetic acid and n-butyric acid. There was also some correlation between the concentration of acetic acid and macroscopic damaged area in the colon (R = -0.60, P = 0.07). Bacteriological studies revealed significantly decreased eubacteria, bifidobacteria and total anaerobes after the administration of DSS. In contrast, lactobacilli were significantly increased. CONCLUSIONS With the progression of DSS-induced colitis, faecal bile acids, SCFAs and intestinal microflora were altered. It is possible that these alterations contribute in part to the progression of DSS-induced colitis.

Background. The pathogenesis of inflammatory bowel disease is still unknown. However, it is possible that faecal bile acids influence the clinical course. Aims. To evaluate the eliminating effects of faecal bile acids by the oral adsorbent on dextran sulphate sodium-induced rat colitis. Methods. Rats were given 3% dextran sulphate sodium aqueous solution for 7 days, with or without concomitant administration of oral adsorbent, or the rats were given dextran sulphate sodium for 7 days, followed with or without oral adsorbent for 5 days. Macroscopic and microscopic examinations of the colons and measurement of faecal bile acids were performed. The cytotoxicity of bile salts on Caco-2 cells was also evaluated. Results. Oral adsorbent tended to attenuate the dextran sulphate sodium-induced colitis. Oral adsorbent was fairly effective in reducing faecal hyodeoxycholic acid concentration. A positive correlation was found between the size of the ulcer area and the faecal hyodeoxycholic acid concentration. In a cell culture study, cytotoxicity of bile acid was parallel with increasing hydrophobicity of the bile acid. However, hyodeoxycholate exhibited severe cytotoxicity, despite its hydrophilic properties. Conclusions. Oral adsorbent tended to attenuate the dextran sulphate sodium-induced colitis and tended to promote the recovery process. It is possible that bile acids in the gut lumen influence the progression of dextran sulphate sodium-induced colitis and its repair process.

Recent studies have suggested that dietary fiber exerts a therapeutic effect on IBD patients. The aim of this study was to evaluate the effects of a dietary combination of germinated barley foodstuff (GBF), derived from the aleurone and scutellum fraction of germinated barley, plus Clostridium butyricum against dextran sulfate sodium (DSS)-induced experimental colitis in rats.Sprague-Dawley rats were fed a 3% DSS diet containing GBF only, GBF plus C. butyricum, cellulose only (control) or cellulose plus C. butyricum for 8 days. The mucosal damage (macroscopic and microscopic inflammation) and fecal short-chain fatty acid (SCFA) levels were then determined.The combination of GBF plus C. butyricum most effectively prevented bloody diarrhea and mucosal damage. The GBF-only diet also showed some preventive effects. With respect to fecal SCFAs, the combination of GBF plus C. butyricum most effectively increased the fecal SCFA level.The dietary combination of GBF plus C. butyricum most effectively suppressed DSS-induced experimental colitis in rats. These effects may be closely associated with its high activity to increase SCFA levels in the gut lumen. The potential clinical efficacy of GBF in IBD patients is also discussed.

Background: It is important to develop an appropriate animal model for further investigation into inflammatory bowel disease (IBD). We therefore investigated a trinitrobenzene sulfonic acid (TNBS) ileitis model. Dietary fat in Crohn’s disease is still a controversial risk factor for IBD. We therefore also studied the effects of medium-chain triglycerides (MCT) and long-chain triglycerides (LCT) on TNBS ileitis. Methods and Results: An intraileal injection of TNBS induced ulceration and inflammation with thickening of the intestinal wall, which were characterized histologically by infiltration of polymorphic nuclear leucocytes and by granuloma formation. The mucosal damage score and serum sialic acid levels reached their highest 7 days after the TNBS injection and then gradually decreased. The mucosal damage series in the MCT group was significantly lower than in the LCT group, and levels of tumour necrosis factor-α (TNF-α) and leukotriene B4 (LTB4) tended to be lower in the MCT group. Conclusions: These results suggested that TNBS enteritis might be useful as an IBD animal model and that MCT modulates intestinal inflammation and is less damaging than LCT. © 1999 Blackwell Science Asia Pty Ltd

SUMMARYThe various biological activities of butyrate have been well documented. In this study, we tested the effects of butyrate on TNF-α-induced complement C3 and factor B biosynthesis in human intestinal epithelial cells. The biosynthesis of C3, factor B and IL-8 was evaluated at the protein and mRNA levels. To evaluate transcriptional activation, the nuclear run-on assay was performed. The transcription factor–DNA binding activity was assessed by an electrophoretic gel mobility shift assay (EMSA). In the intestinal epithelial cell lines HT-29, T84 and Caco-2, sodium butyrate enhanced TNF-α-induced C3 secretion, but suppressed TNF-α-induced factor B and IL-8 secretion. Nuclear run-on assay revealed that transcriptional regulatory mechanisms are involved in the effects of sodium butyrate. The EMSAs indicated that sodium butyrate suppressed TNF-α-induced nuclear factor (NF)-κB– and activation protein (AP)-1–DNA binding activity, but enhanced TNF-α-induced activation of CCAAT/enhancer-binding protein (C/EBP)β (NF-IL-6)–DNA binding activity. Sodium butyrate induced a counter-regulatory effect on TNF-α-induced C3 and factor B biosynthesis in human intestinal epithelial cells. Butyrate action has been discussed with its activity to induce histone hyperacetylation, but its counter-regulatory effect on complement biosynthesis may be closely associated with the modulation of transcription factor activation.

S. Sawada and co-workers reported that a monoclonal antibody (MAb), E87, interacted with about 80% of Pseudomonas aeruginosa isolates, and they separated a rhamnose-rich polysaccharide as the probable antigen for MAb E87 from P. aeruginosa IFO 3080 (S. Sawada, T. Kawamura, Y. Masuho, and K. Tomibe, J. Infec. Dis. 152:1290-1299, 1985). In the present study, the rhamnose-rich polysaccharide was shown to be structurally and immunologically identical to the D-rhamnan of P. aeruginosa IID 1008 (S. Yokota, S. Kaya, S. Sawada, T. Kawamura, Y. Araki, and E. Ito, Eur. J. Biochem. 167:203-209, 1987). Furthermore, a set of enzymes responsible for the formation of GDP-rhamnose (probably in a D-form) from GDP-D-mannose was found in the 100,000 x g supernatant fractions obtained from all of nine P. aeruginosa strains reactive against MAb E87. The result strongly supports a possibility that lipopolysaccharides having a D-rhamnan chain widely occur as the common antigen among various P. aeruginosa isolates.

The DNA homology and cell wall sugar constituents of eight Streptococcus sanguis(-like) strains, three isolated from the patients with Behcet's disease (BD114-23, BD113-20, BD118-1), two from patients with Kawasaki disease (MCLS-1, MCLS-2), and three type and reference strains of ATCC (ATCC10556T: S. sanguis, ATCC10557: S. oralis, and ATCC10558T: S. gordonii) were analyzed. Strains BD114-23 and BD118-1 showed high DNA homology to ATCC10556T, and their cell wall constituents were identical. Conversely, BD113-20, MCLS-1, MCLS-2, and ATCC10557 showed little DNA homology to ATCC10556T and ATCC10558T, but showed approximately 50 to 60% homology to each other. The cell wall constituents of BD113-20, MCLS-1, MCLS-2, and ATCC10557, however, were somewhat different, indicating that some of the clinical isolates have different characters from those of the three ATCC strains.

A new and highly branched amino acid was found as an N -acyl substituent of the O-polysaccharide chain obtained from the lipopolysaccharide of Vibrio anguillarum V-123 (serogroup JO-2) and evidence is presented ot support the structure as 2,4-dihydroxy-3,3,4-trimethylpyroglutamic acid. Acid hydrolysis of the O-polysaccharide gave the lactone of 2,4-dihydroxy-3,3,4-trimethylglutamic acid, together with 2-amino-2-deoxy- d -galacturonic acid, 2-amino-2,6-dideoxy- d -glucose ( d -quinovosamine), and 4-amino-4,6-dideoxy- d -glucose ( d -viosamine). Degradation of the O-polysaccharide with hydrogen fluoride yielded a fragment (H1) that was indicated by the 1 H-NMR data to be 4-amino-4,6-dideoxy- d -glucose N -acetylated with 2,4-dihydroxy-3,3,4-trimethylpyroglutamic acid. The configuration of the amino acid was not determined.

Lipoteichoic acid-like substances, macroamphiphiles, were isolated from cell homogenates of Bifidobacterium bifidum YIT 4007 and YIT 4013, Bifidobacterium breve YIT 4010 and YIT 4014, and Bifidobacterium longum YIT 4021 by phenol extraction followed by nuclease digestion, gel chromatography, ion-exchange chromatography, and hydrophobic interaction chromatography. The macroamphiphile preparations from these five strains contained D-glucose, D-galactose, glycerol, phosphorus, L-alanine, and fatty acids in molar ratios of 1.00, 1.57 to 1.95, 1.02 to 1.99, 0.97 to 1.72, 0.15 to 0.46, and 0.16 to 0.43. Data from structural analyses including methylation, 1H nuclear magnetic resonance measurement, alkaline hydrolysis, mild acid hydrolysis, and hydrogen fluoride treatment led to the most likely common structure for the macroamphiphiles of the examined strains, (formula; see text) where Gro-P is glycerophosphate, m is the number of repeating units of galactofuranan, and n is the number of repeating units of glucan. Whereas the polymers from the respective strains differed in the numbers of repeating units of the galactofuranan and glucan moieties and in the number of fatty acid residues, the proposed structure is essentially the same as that reported previously for the macroamphiphile of B. bifidum subsp. pennsylvanicum DSM 20239 by W. Fischer (Eur. J. Biochem. 165:639-646, 1987).