Search

The construction of a small-size, magnetic sector, single focusing mass spectrometer (He-MS) for the continuous, on-site monitoring of He isotope ratios ((3)He/(4)He) is described. The instrument is capable of measuring (4)He/(20)Ne ratios dissolved in several different types of natural fluids of geochemical interest, such as groundwater and gas from hot springs, volcanoes and gas well fields. The ion optics of He-MS was designed using an ion trajectory simulation program "TRIO," which permits the simultaneous measurement of (3)He and (4)He with a double collector system under a mass resolution power (M/ΔM) of >700. The presently attained specifications of He-MS are; (1) a mass resolving power of ca. 430, sufficient to separate (3)He(+) from interfering ions, HD(+) and H3 (+), (2) ultra-high vacuum conditions down to 3×10(-8) Pa, and (3) a sufficiently high sensitivity to permit amounts of (3)He to be detected at levels as small as 10(-13) cm(3) STP (3×10(6) atoms). Long term stability for (3)He/(4)He analysis was examined by measuring the (3)He/(4)He standard gas (HESJ) and atmospheric He, resulting in ∼3% reproducibility and ≤5% experimental error for various amounts of atmospheric He from 0.3 to 2.3×10(-6) cm(3) STP introduced into the instrument. A dynamic range of measurable (3)He/(4)He ratios with He-MS is greater than 10(3) which was determined by measuring various types of natural fluid samples from continental gas (with a low (3)He/(4)He ratio down to 2×10(-8)) to volcanic gas (with a high (3)He/(4)He ratio up to 3×10(-5)). The accuracy and precision of (3)He/(4)He and (4)He/(20)Ne ratios were evaluated by comparing the values with those measured using well established noble gas mass spectrometers (modified VG5400/MS-III and -IV) in our laboratory, and were found to be in good agreement within analytical errors. Usefulness of the selective extraction of He from water/gas using a high permeability of He through a silica glass wall at high temperature (700°C) is demonstrated.

Seventy pluriparous high-yielding cows were used to investigate the impact of metabolic traits and body condition score (BCS) during early lactation on subclinical endometritis diagnosed at weeks 5, 6 and 7 postpartum (pp). Blood samples were collected from animals with no peripartum problems from the second (W2) to seventh (W7) weeks pp to estimate some blood metabolites including non-esterified fatty acids (NEFA), β-hydroxybutyric acid (BHBA), blood glucose, total cholesterol (T-chol) and blood urea nitrogen (BUN). Reproductive tract examination was carried out at weeks 5, 6 and 7 pp by endometrial cytology (percentage of polymorphonuclear cells; PMN%). Based on PMN%, animals having5% were defined as subclinical endometritis group (ENDM group) while animals unaffected by endometritis were defined as no subclinical endometritis group (NOENDM group). Animals with endometritis during week 5 were identified as ENDM5, during week 6 identified as ENDM6 and during week 7 identified as ENDM7 or animals with no endometritis during weeks 5 (NOENDM5), 6 (NOENDM6) and 7 (NOENDM7) pp. Animals diagnosed at week 5; BUN and BCS were lower p0.05 in ENDM 5 than NOENDM5 group at W2, W4, W6 and W7. Cows diagnosed at week 6; T-chol was significantly higher (p = 0.05) in ENDM6 group (279.2 ± 12.5 mg/dl) than NOENDM6 group (246 ± 9.5 mg/dl) at W7. Moreover, blood glucose was significantly low (p0.05) in ENDM6 group when compared to NOENDM group at W4 pp (49.2 ± 1.8 vs 53.8 ± 1.3 mg/dl). BCS was significantly lower (p0.001) in animals suffered from endometritis during week 7 when compared to NOENDM7 cows at W3, W4, W5, W6 and W7. In conclusion, lower blood glucose, BUN and BCS could be a risk factor for cytologically diagnosed endometritis at weeks 5, 6 and 7 pp.

The electromechanical properties of porous polypropylene electrets were studied by means of linear and nonlinear dielectric measurements. The second and the third order nonlinear permittivity e 2 and e 3 were measured by two measurement methods. The e 2 and e 3 depend on the piezoelectric constant d 33 and the elastic compliance s 33 respectively when the nonlinear permittivity is induced by the electrostriction generated by the Maxwell force. The value of s 33 determined by e 3 was about two times larger than the value determined by the dielectric resonance. The result suggests an existence of frequency dependence of s 33 . On the other hand, the value of d 33 determined by e 2 showed a good agreement with the result of the dielectric resonance. This result means that the piezoelectric response of the p-PP electret is mainly induced by the size effect.

Clinically normal Holstein-Friesian cows (n = 43) were used to establish a relationship between the evaluations of the uterine condition by vaginoscopy, Metricheck, endometrial cytology (EC) and ultrasonography (US) during post-partum period (pp), and subsequent reproductive performance. The uterine status of the selected cows was evaluated by EC, Metricheck, US and vaginoscopy on a weekly basis from the third week (W3) to the seventh week (W7) pp. The animals were bred after a voluntary waiting period of 45-60 days pp and diagnosed for pregnancy status at 45 days after breeding by US. First service conception rate (FSCR) of bred animals was 32.6% (14/43). There was no significant difference in polymorphonuclear leucocyte percentage between conceived and non-conceived cows during different weeks post-partum. First service conception rate in animals with normal vaginal discharge as detected by Metricheck was significantly low when compared to those of abnormal discharge during W3 (2.3% vs 30.3%; p 0.001) and W4 (4.7% vs 29.7%; p0.01) post-partum. Using US, the number of pregnant cows having fluid in uterus (FIU) during W6 (p 0.01) and W7 (p 0.001) post-partum was significantly lower (2/34, 4.7% and 1/43, 2.3%, respectively) than those that had no FIU (12/43, 27.9% and 13/43, 30.3%, respectively). Using vaginoscopy, FSCR in animals having abnormal discharge was significantly (p 0.01) low when compared to that of normal discharge (2.4% vs 30.2%) during W5 and W6 post-partum. In conclusion, Metricheck during W3 and W4 and detection of FIU by US and vaginoscopy during W6 and W7 pp can be used as good predictive tools to anticipate the future reproductive performance of dairy cattle.

The effect of ovarian predominating structures and uterine condition on the result of some diagnostic tools for the evaluation of endometritis was studied in postpartum (pp) Holstein-Friesian dairy cows (n = 58). Endometrial cytology (EC) and the evaluation of vaginal mucus by vaginoscopy or Metricheck were performed weekly from week 3 to 7 pp. The ovarian studies involved the predominating structures including cystic follicles with plasma progesterone (P(4) ; more or1 ng/ml;23 mm), corpus luteum (CL), pre-ovulatory follicles (10-23 mm) and small follicles (10 mm). The uterine conditions comprised uterine involution, tonicity and fluid in uterus (FIU) regarding echogenicity extent. During week 5, the percentage of polymorphonuclear neutrophils (PMN%) was higher (p0.05) in animals with pre-ovulatory follicles (mean ± SEM, 26.3 ± 7.6%) than animals having CL (11.0 ± 3.6%). In cystic ovaries, during week 5, PMN% was higher (p0.05) in follicular cysts with low progesterone (P(4)1 ng/ml; 9.3 ± 2.6%) than those with high P(4) (P(4) ≥ 1 ng/ml; 1.5 ± 1.1%). Moreover, PMN% was higher (p0.01) in animals with non-involuted uterus (11.5 ± 7.4%) than those with involuted uterus (2.7 ± 0.6%) during week 7 pp. The animals that had abnormal mucus determined by Metricheck was higher in animals with atonic uterus than those with tonic uterus during week 6 (82.6% vs 51.5%; p0.05) and 7 (71.4% vs 25.7%; p0.01) pp. In addition, by vaginoscopy, the prevalence of animals with abnormal discharge showing small follicles (100%, 5/5) during week 3 pp and pre-ovulatory follicles (40.0%, 8/20) during week 5 pp was higher (p0.05) when compared to those having CL during week 3 (33.3%; 1/3) and week 5 pp (7.7%; 2/26), respectively. In conclusion, endometrial cytology, Metricheck and vaginoscopy were influenced by the predominating various ovarian structures and uterine condition in early pp high-yielding dairy cows.

The structure of the title compound, gallium sodium silicate, is similar to that of jadeite (NaAlSi 2 O 6 ). Comparison of the new crystal-structure refinement for NaGaSi 2 O 6 with published refinements for eleven NaM 3+ Si 2 O 6 pyroxenes suggests that there are two different electronic states, Ga(α) and Ga(β), for the octahedral Ga 3+ ion. Ga(α) occurs in the Mn-Fe-Ga(α) series and Ga(β) in the Al-Ga(β)-In series.

Ca(2+)-dependent carbohydrate-binding proteins were purified from bovine kidney extracts. Upon SDS-polyacrylamide gel electrophoresis under nonreducing conditions, the purified fraction gave doublet protein bands corresponding to 33 kDa (p33) and 41 kDa (p41). Under reducing conditions, a single protein band (p33) was observed. p33 and p41 were submitted to proteolytic digestion with endoproteinase Lys-C, the peptides produced were separated by reversed-phase high performance liquid chromatography, and their amino acid sequences were determined by an automated gas-phase protein sequenator. Most of the resulting partial amino acid sequences of these proteins were strikingly homologous to annexin IV, an annexin family protein, i.e. Ca2+/phospholipid-binding proteins, especially in the consensus sequences. In the presence of Ca2+, both proteins bound to vesicles composed of phosphatidylserine and phosphatidylethanolamine, but not phosphatidylcholine. These results indicated that p33 and p41 are members of annexin family proteins.

Adult rats which received repeated electroshock convulsions (ESCs) during postnatal development underwent electrical kindling of the amygdala (AM) and hippocampus (HIPP). Completion of kindling, especially AM kindling, was significantly faster in the groups with ESCs from the neonatal to late infantile age than in the control group. It is discussed that especially the late infantile age in the postnatal development was supposed to play an important role in the faster kindling brought about by postnatal convulsions and that the faster kindling was due in part to abbreviation of the partial seizure process. However, the neonatal ESC group showed significantly delayed completion of HIPP kindling.

Articular cartilage has the depth-dependent laminar structure, and the mechanical behavior of articular cartilage is strongly dependent of its microstructure characterized by three-dimensional, depth-dependent anisotropic arrangement of collagen fibre, chondrocyte and so on. In this study, a method identifying the mechanical properties of cartilage is described by using a viscoelastic model of cartilage considering its anisotropic and inhomogeneous microstructure and the time history of reaction force observed in the indentation test. The method is examined with bovine femoral cartilage at the medial and lateral regions.

Epidemiological and experimental studies have suggested that diesel exhaust particles (DEPs), which generate reactive oxygen species, may be involved in the recent increase in the prevalence of lung diseases. Cacao liquor proanthocyanidins (CPs) are naturally occurring polyphenols with antioxidative activities. We carried out a study in mice to investigate the effects of dietary supplementation of CPs on lung injury induced by intratracheal administration of DEPs (500 μg/body). Dietary supplementation with 1.0% CPs inhibited DEP-induced lung injury, characterized by neutrophil sequestration and edema. Immunohistochemical analyses showed that CPs prevented enhanced expression of vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 caused by DEPs in the lung injury. Numerous adducts of nitrotyrosine, N-(hexanonyl) lysine, 4-hydroxy-2-nonenal, and 8-OHdG were also observed immunohistochemically in the lungs of mice treated with DEPs. However, these indicators of oxidative stress were barely visible in mice pretreated with CP supplementation. In addition, the level of thiobarbituric acid reactive substances in the lung was decreased by CP supplementation in the presence of DEPs. These results suggest that CPs inhibit DEP-induced lung injury by reducing oxidative stress, in association with a reduction in the expression of adhesion molecules.

We examined the cytolytic mechanisms of activated macrophages by using proteose peptone- or thioglycollate broth-induced mouse peritoneal macrophages or mouse macrophage hybridomas as effector cells, L.P3 cells, a clone of L929 cells, and P815 cells as target cells, and IFN-gamma and LPS as activators. It was determined that TNF is the main cytolytic molecule against L.P3 cells from the following results: 1) activated macrophages can produce TNF; 2) TNF shows cytotoxic activity against L.P3 cells; 3) the addition of anti-TNF antibody inhibited most of the cytolytic activity of activated macrophages against L.P3 cells. On the other hand, it was concluded that the main cytolytic mechanism against P815 cells is the production of NO2-/NO3- from L-arginine, from the following results: 1) activated macrophages can produce NO2-; 2) NaNO2 shows high cytotoxic activity against P815 cells; 3) the depletion of L-arginine from the medium inhibited most of the cytolytic activity of activated macrophages against P815 cells and NO2- production by activated macrophages. In this study, however, cytostatic effects of L-arginine-dependent effector mechanism were not studied. Thus, these results show that activated macrophages can express at least two cytolytic mechanisms independently, namely, the one that appears to be mediated by the L-arginine-dependent effector mechanism and the second that appears to be mediated directly by TNF. Furthermore, it was demonstrated that TNF and L-arginine-dependent NO2- production act synergistically as killing mechanisms of activated macrophages. These mechanisms can explain the cytolytic activity of activated macrophages against a variety of target cells.

A 52-year-old man stabbed in his neck and abdomen by a kitchen knife for the purpose of suicide. He was immediately transferred by ambulance car. Subcutaneous emphysema was not observed. Stabbed wounds were observed in the neck and the abdomen. Radiography and computed tomography showed subcutaneous and mediastinal emphysema. The patient was diagnosed as having trachea injury. After admission, emphysema progressed rapidly and respiratory distress appeared. The patient was orally intubated and emergency surgery was performed. The stabbed wound was found to extend to thyroid grand and cricotracheal ligament. After thyroidectomy, the injured trachea was repaired primarily. The course of the patient was almost uneventful, and was discharged on the 23th operative day. He was diagnosed as having alcoholism by a psychiatrist. The patient died due to suicide by hanging 16 months after surgery. Orotracheal intubation and primary suture were effective in a patient with trachea injury. Prevention of suicide is also important after leaving hospital in patients with history of suicide.

In Japan, the degree of marbling in ribeye (M. longissimus thoracis) is evaluated in the beef meat grading process. However, other muscles (e.g., M. trapezius) are also important in determining the meat quality and carcass market prices. The purpose of this study was to estimate genetic parameters for M. longissimus thoracis (M-LONG) and M. trapezius (M-TRAP) of carcass cross section of Japanese Black steers by computer image analysis. The number of records of Japanese Black steers and the number of pedigree records were 2,925 and 10,889, respectively. Digital images of the carcass cross section were taken between the sixth and seventh ribs by photographing equipment. Muscle area (MA), fat area ratio (FAR), overall coarseness of marbling particles (OCM), and coarseness of maximum marbling particle (MMC) in M-LONG and M-TRAP were calculated by image analysis. Genetic parameters for these traits were estimated using the AIREMLF90 program with an animal model. Fixed effects that were included in the model were dates of arrival at the carcass market and slaughter age (mo), and random effects of fattening farms, additive genetic effects and residuals were included in the model. For M-LONG, heritability estimates (+/-SE) were 0.46 +/- 0.06, 0.59 +/- 0.06, 0.47 +/- 0.06, and 0.20 +/- 0.05 for MA, FAR, OCM, and MMC, respectively. Heritability estimates (+/-SE) in M-TRAP were 0.47 +/- 0.06, 0.57 +/- 0.07, 0.49 +/- 0.07, and 0.13 +/- 0.04 for the same traits. Genetic correlations between subcutaneous fat thickness and FAR for M-LONG and M-TRAP were negative (-0.21 and -0.19, respectively). Those correlations between M-LONG and M-TRAP were moderate to high for MA, FAR, OCM, and MMC (0.38, 0.52, 0.39, and 0.60, respectively). These results indicate that other muscles including M-LONG should be evaluated for more efficient genetic improvement.

One of the goals of computational intelligence is to coordinate various distributed data sources and services to compose more complex services that cover more than provided by a single service. The semantic grid [4] is proposed that integrates grid technologies and semanticWeb technologies by providing semantic annotations of grid services and data sources. We argue that technologies for automated composition of services are also useful for constructing statically composed service cooperating system(called ‘services integration system’). In this paper, we propose a framework that supports the development of services integration systems. It helps portal site developers to discover, evaluate, and execute service chaining paths for services integration systems. Semi-automated source code generation will reduce the cost of services integration system development.

Constitutively active internal tandem duplication (ITD) in the juxtamembrane domain of Fms-like tyrosine kinase 3 (FLT3), a type III receptor tyrosine kinase, is the most common molecular defect associated with acute myeloid leukemia. Its presence confers a poor outcome in patients with acute myeloid leukemia who receive conventional chemotherapy. FLT3-ITD has therefore been considered to be an attractive molecular target for a novel therapeutic modality. We describe here the identification and characterization of Ki23819 as a novel FLT3 inhibitor. Ki23819 suppressed proliferation and induced apoptosis of FLT3-ITD-expressing human leukemia cell lines. The growth-inhibitory effect of Ki23819 on MV4-11 cells was superior to that of SU11248, another FLT3 inhibitor (IC(50)

This paper proposes a stereo method robust against occlusions that can reconstruct 3D building facades from images captured from a vehicle. When using ground-based views of facades, the occlusion problem caused by roadside trees and telegraph poles in urban areas is the one of the most difficult barriers. In this study, we show the stereo matching based on parallel images minimize the occlusion effect. To acquire the parallel images, we use two line scan cameras linked a rotary encoder mounted on a vehicle. Experimental results using outdoor scenes confirm the effectiveness of this method.

The structure of cobalt aluminium silicate garnet, Co 3 Al 2 Si 3 O 12 , has been redetermined from diffractometer data. Comparison of the present refinement with the published refinements for the M 3 2+ Al 2 Si 3 O 12 garnets suggests that two isostructural phases exist for the Co 3 Al 2 Si 3 O 12 garnet which are dependent on the pressure.

CD4+ and CD8+ T Iymphocytes from the peripheral blood of melanoma patients are prone to undergo spontaneous apoptosis when compared with healthy individuals. The rapid Iymphocyte turnover and the loss of CD4+ and CD8+ T cells may cause the weak antitumor response in cancer patients. Since CD4+ and CD8+ T cell mediated immune responses play an important role in controlling the antitumor response, enhancing the survival and function of these T cells may improve the disease outcome of melanoma patients. In our experiment, we tried to see the effect of interleukin 15 (IL-15) on the spontaneous apoptosis of peripheral blood mononuclear cells (PBMCs) from patients with melanoma. The PBMCs obtained from 16 melanoma patients and 16 healthy controls were incubated overnight in medium with or without IL-15. The PBMGs were then stained for CD4+ or CD8+ cells and evaluated for apoptosis with the Annexin V binding method using a multicolor flow cytometry. The results showed that IL-15 inhibits spontaneous apoptosis in CD4+ and CD8+ T cells in both melanoma patients and healthy individuals. IL-15 significantly reduced 6.3% (mean±spontaneous apoptosis of CD8+ T-cells in melanoma patients from 26 8.7% (n=16, P± SD) to 19 ±< 3.4% to±0.005) and in healthy controls from 15 3.4% (n=16, P±11

In this study, the spontaneous apoptosis of CD4+ and CD8+ T-cells in patients with melanoma were examined. Fifty-seven patients enrolled in the study were participating in an experimental immunotherapy study and were administered vaccines consisting of irradiated, gene modified allogenic melanoma cells: MICH I H6/ GMCSF + MICH II H6/GMCSF. The control group consisted of 20 healthy volunteers. Venous blood was obtained on the day of the administration of the first vaccine, and then approximately once a month. Blood was obtained in EDTA tubes, after which the Iymphocytes were isolated by Ficoll-Hypaque gradient centrifugation. The Iymphocytes were then incubated for 24hrs in medium containing fetal calf serum at 37°C in 5% CO2 to allow for spontaneous apoptosis to take place. The Iymphocytes were then stained with PE-Iabeled CD4 or CD8 and next, stained for apoptosis using the FITC-labeled Annexin V binding method. A two color flow cytometry was used to measure the proportion of CD4+ and CD8+ Iymphocytes that underwent apoptosis. SD) of CD4+ T cells were Annexin±6.3% (mean ±In patients with melanoma 21 6.3% in healthy controls (P±V+ compared with 15 < 0.0001). For CD8+ T cells, 4.5%±8.1% of T cells were Annexin V+ and in healthy controls 19±in patients 31 of T cells were Annexin V+(P < 0.0001). There were no significant changes with time in the proportion of CD4+ and CD8+ Iymphocytes undergoing spontaneous apoptosis after the start of the gene modified tumor vaccine therapy.

We propose a new resource control algorithm, referred to as offset-ARP for cellular systems. In the proposed algorithm, a new threshold, referred to as offset C/I threshold, is introduced for channel allocation. By shifting a channel allocation criterion from the minimum link quality to a better quality, a large transmitting power reduction and a short reuse distance of the channels are realized. Thus, the proposed algorithm effectively integrates dynamic channel allocation with transmitting power control in the sense of two dimensional optimal control. By computer simulation, the proposed algorithm is evaluated through comparison with the conventional algorithm. The result shows that offset-ARP improves the handoff frequency and the forced termination probability is about ten times lower than those of the conventional algorithm.

We have proposed a Superconducting Fault Current Limiting Cable (SFCLC), which is an HTS cable with fault current limiting function. SFCLC is expected to limit the fault current and also immediately recover the cable function after the fault clearance. In the SFCLC operation, a longitudinal temperature distribution will exist due to heat penetration, AC loss, dielectric loss and the performance of cryocooling system, which will influence its current limitation characteristics. In this paper, we investigate the influence of the longitudinal temperature distribution on current limiting function and temperature rise after the current limitation of SFCLC. We suggested the effective measures of parameter control, i.e. decreasing the critical current (Ic@77K), n value at flux flow region (n1-0), increasing the coefficient of longitudinal temperature gradient (α), inflow temperature (Tin) to achieve both the higher current limiting function and the lower temperature rise.

The exposure of human lymphoid leukemia Molt 4B cells to sesaminol, a component of sesame oil led to both growth inhibition and the induction of apoptosis. Morphological change showing apoptotic bodies was observed in the cells treated with sesaminol. The fragmentation of DNA by sesaminol to oligonucleosomal-sized fragments that are characteristics of apoptosis was observed to be concentration- and time-dependent. These findings suggest that growth inhibition of Molt 4B cells by sesaminol results from the induction of apoptosis in the cells.

In the present study, we characterize the covalent modification of a protein by crotonaldehyde, a representative carcinogenic aldehyde, and describe the endogenous production of this aldehyde in vivo. The crotonaldehyde preferentially reacted with the lysine and histidine residues of bovine serum albumin and generated a protein-linked carbonyl derivative. Upon incubation with the histidine and lysine derivatives, crotonaldehyde predominantly generated beta-substituted butanal adducts of histidine and lysine and N(epsilon)-(2,5-dimethyl-3-formyl-3,4-dehydropiperidino)lysine (dimethyl-FDP-lysine) as the putative carbonyl derivatives generated in the crotonaldehyde-modified protein. To verify the endogenous formation of crotonaldehyde in vivo, we raised the monoclonal antibody (mAb82D3) against the crotonaldehyde-modified protein and found that it cross-reacted with the protein-bound 2-alkenals, such as crotonaldehyde, 2-pentenal, and 2-hexenal. The anti-2-alkenal antibody recognized multiple crotonaldehyde-lysine adducts, including dimethyl-FDP-lysine and an unknown product, which showed the greatest immunoreactivity with the antibody. On the basis of the chemical and spectroscopic evidence, the major antigenic product was determined to be a novel Schiff base-derived crotonaldehyde-lysine adduct, N(epsilon)-(5-ethyl-2-methylpyridinium)lysine (EMP-lysine). It was found that the lysine residues that had disappeared in the protein treated with crotonaldehyde were partially recovered by EMP-lysine. The presence of immunoreactive materials with mAb82D3 in vivo was demonstrated in the kidney of rats exposed to the renal carcinogen, ferric nitrilotriacetate. In addition, the observations that the metal-catalyzed oxidation of polyunsaturated fatty acids in the presence of proteins resulted in an increase in the antigenicity of the protein indicated that lipid peroxidation represents a potential pathway for the formation of crotonaldehyde/2-alkenals in vivo. These data suggest that the formation of carcinogenic aldehydes during lipid peroxidation may be causally involved in the pathophysiological effects associated with oxidative stress.

Exposure of cells to a wide variety of chemoprotective compounds confers resistance to a broad set of carcinogens. For a subset of the chemoprotective compounds, protection is generated by an increase in the abundance of protective enzymes, such as glutathione S-transferases (GSTs). In the present study, we developed a cell culture system that potently responds to phenolic antioxidants and found that antitumor prostaglandins (PGs) are potential inducers of GSTs. We screened primary hepatocytes and multiple cell lines for inducing GST activity upon incubation with the phenolic antioxidant (tert-butylhydroquinone) and found that rat liver epithelial RL34 cells most potently responded. Based on an extensive screening of diverse chemical agents on the induction of GST activity in RL34 cells, the J2 series of PGs, 15-deoxy-Delta(12,14)-prostaglandin J2 (15-deoxy-Delta(12,14)-PGJ2) in particular, were found to be potential inducers of GST. Enhanced gene expression of Class pi GST isozyme (GSTP1) by 15-deoxy-Delta(12,14)-PGJ2 was evident as a drastic elevation of the mRNA level. Hence, we examined the molecular mechanism underlying the 15-deoxy-Delta(12, 14)-PGJ2-induced GSTP1 gene expression. From functional analysis of various deletion mutant genes, we found that the 15-deoxy-Delta(12, 14)-PGJ2 reponse element was localized in a region containing a GSTP1 enhancer I (GPEI) that consists of two imperfect phorbol 12-O-tetradecanoylphorbol-13-acetate response elements. When the GPEI was combined with the minimum GSTP1 promoter, the element indeed showed an enhancer activity in response to 15-deoxy-Delta(12, 14)-PGJ2. Point mutations of either of the two imperfect 12-O-tetradecanoylphorbol-13-acetate response elements in GPEI completely abolished the enhancer activity. Gel mobility shift assays demonstrated that 15-deoxy-Delta(12,14)-PGJ2 specifically stimulated the binding of nuclear proteins including the transcription factor c-Jun, but not Nrf2, to GPEI. These results suggest that 15-deoxy-Delta(12,14)-PGJ2 induces the expression of the rat GSTP1 gene through binding of proteins, including c-Jun, to a specific GPEI.

Dithiothreitol effectively separated the laminae densae and the laminae fibroreticulares of the basement membranes of oral mucosal epithelia as it does the epidermis and dermis. Dithiothreitol-separated epithelial basement membranes of palate, tongue, and lip were immunoreacted with colloidal gold-conjugated anti-type VII collagen antibody. By transmission electron microscopy, gold particles were observed only on the anchoring fibrils. For three-dimensional observation of the distribution of type VII collagen by scanning electron microscopy, secondary and backscattered electron images were compared. The secondary image showed the fine structure of the laminae densae and anchoring fibrils, and the backscattered images showed the gold particles conjugated with the antibody. By using an osmium conductive metal coating under optimal conditions, secondary and backscattered electron images of sufficient quality could be obtained. With the osmium coating, the backscattered electron image could show not only the gold particles but also the general morphological outline, making possible a comparison of the two images, which revealed the three-dimensional distribution of type VII collagen. Type VII collagen was also seen only on the anchoring fibrils, as in the epidermal basement membrane.

Major photoproducts induced by carcinogenic ultraviolet (UV) radiation are the cylobutane pyrimidine dimers (CPDs) and pyrimidine-pyrimidone (6-4) photoproducts (6-4 PPs). 8-Hydroxy-2 -deoxyguanosine (8-OHdG) is also a DNA base-modified product generated by reactive oxygen species in conditions of ultraviolet stress, Although UVB-induced CPDs and 6-4 PPs have been investigated in animal and human skin, little is known about the role of 8-OHdG in UVB-induced human skin damage or carcinogenesis. Normal human skin from three volunteers was exposed to UV radiation, and the time course of induction and removal of 8-OHdG was examined by immunohistochemical analysis with catalysed signal amplification on formalin-fixed paraffin sections. Formation of CPDs and 6-4 PPs was also examined by immunostaining on the same skin specimens. Control epidermis with no exposure to UV radiation showed little nuclear staining of 8-OHdG, but an increased level of 8-OHdG was clearly observed in epidermis biopsied after irradiation. Induced 8-OHdG can rapidly be removed from nucleus during the first 24-48 h, as the staining intensity diminished gradually, almost reaching the control level by 72-96 h after irradiation. Staining for CPDs or 6-4 PPs revealed induction of these photoproducts in human skin, although 6-4 PP-positive cells disappeared more rapidly than those that stained for CPDs or 8-OHdG. Together with protective effect of antioxidants, our results indicate that not only CPDs and 6-4 PPs but also 8-OHdG may play a significant part in UV carcinogenesis.

Here we report the molecular mechanism underlying the induction of glutathione S-transferase (GST) in rat liver epithelial RL34 cells treated with a cancer chemopreventive isothiocyanate compound, benzylisothiocyanate (BITC). BITC was found to significantly induce GST activity in RL34 cells. Northern and Western blot analyses demonstrated that BITC specifically enhanced the production of the class pi GST isozyme (GSTP1). Our studies demonstrated for the first time that the addition of BITC to the cells resulted in an immediate increase in the reactive oxygen intermediates (ROIs) detected by a fluorescence probe, 2',7'-dichlorofluorescin diacetate. The level of the ROIs in the cells treated with BITC (10 microM) was approximately 50-fold higher than those in the control cells. Furthermore, glutathione depletion by diethyl maleate significantly enhanced BITC-induced ROI production and accelerated the BITC-induced elevation of the GST activity, whereas pretreatment of the cells with glutathione inhibited both the ROI production and GST induction. The structure-activity relationship of the isothiocyanates also indicated that the ROI-producing activities closely correlated with their GST-inducing potencies. Moreover, the GSTP1 enhancer I-containing region was found to be essential for induction of the GSTP1 gene by intracellular ROI inducers such as BITC and diethyl maleate. These data suggest the involvement of the redox regulation on the induction of GSTP1 by BITC.

Ten Thoroughbred yearlings (5 females and 5 males) were used to examine the effect of time of grazing on pasture forage and digestible energy (DE) intake, bodyweight gain and DE expenditure in grazing activity. Five females were grazed for 17 h/day (LTG), 5 males were grazed for 7 h/day (STG) and they were fed differently. As a result, DE intake from pasture forage of LTG horses and STG horses was 27.3 and 12.7-13.9 Mcal/day, respectively. The average daily gain (ADG) of LTG and STG horses was 0.37 and 0.39-0.61 kg/day, respectively. The regression lines between DE intake and ADG of both groups were parallel and the difference in DE intake between them was 10 Mcal of DE. It was suggested that DE expenditure in grazing activity (entirely at night) of growing horses was 1 Mcal/h under the conditions of this study.

Oral vitamin E (alpha-tocopherol) supplementation has been reported to improve facial hyperpigmentation. The compound of alpha-tocopherol and ferulic acid, also an antioxidant connected with an ester bond, alpha-tocopheryl ferulate (alpha-TF) can absorb ultraviolet (UV) radiation and thus maintain tocopherol in a stable state. Our aim was to determine whether alpha-TF can be applied to improve and prevent facial hyperpigmentation induced by UV as a whitening agent as well as an antioxidant. In this study, the effects of alpha-TF on melanogenesis were examined using cultured human melanoma cells and normal human melanocytes in vitro. alpha-TF solubilized in 0.5% lecithin inhibited melanization significantly at the concentration of 30 micrograms/ml compared with arbutin (100 micrograms/ml), kojic acid (100 micrograms/ml), ascorbic acid (600 micrograms/ml), and tranexamic acid (600 micrograms/ml). alpha-TF had no effect on the protein amounts of tyrosinase, TRP (tyrosinase related protein)-1, and TRP-2 of human melanoma cells exposed to UV radiation, but inhibited tyrosine hydroxylase activity. alpha-TF neither directly inhibited tyrosinase activity of the large granule fraction extracted from melanoma cells, nor modulated glycosylation of tyrosinase. These results suggest that alpha-TF may be a candidate for whitening agent which suppresses melanogenesis, possibly by inhibiting tyrosine hydroxylase activity in an indirect manner. Further, alpha-TF decreased the amount of 8-hydroxydeoxyguanosine produced indirectly through active oxygen species (AOS) in guinea pig skin exposed to 2 times the minimal erythema dose of UVB radiation, but did not suppress the direct formation of cyclobutane pyrimidine dimers and (6-4) photoproducts. Thus alpha-TF may reduce AOS-induced DNA damage and thereby contribute at least in part to suppressing or retarding skin cancer development.

The oxidative breakdown of membrane polyunsaturated fatty acids is known to be accompanied by the formation of a complex mixture of lipid hydroperoxides and secondary products. These compounds are highly reactive and are capable of rapid reaction with cellular nucleophiles such as phospholipids and proteins; we found that these reaction products are candidates as important biomarkers to evaluate the antioxidative activity of dietary antioxidants. We have been involved in developing immuno-chemical detection methods for oxidative stress by the application of polyclonal and monoclonal antibodies. From our hypothesis that endogenous antioxidants in plants must play an important role in antioxidative defense systems against oxidative stress, an intensive search for novel natural antioxidants has been carried out on numerous plant materials, including those used as foods, and we have isolated and identified a number of lipid-soluble and water-soluble dietary antioxidants from crop seeds, sesame seeds and some spices. In this paper, we review the recent progress of research on functions of rice polyphenols.

It is known that the gaps between epithelium and the underlying connective tissue usually occur between the epithelial cells and the basement membrane, resulting in exposure of the cellular surface of the lamina densa. After dithiothreitol separation, the epithelia of oral mucosa, gingiva, and tongue were mechanically peeled off from the underlying connective tissues. This treatment severed the connections between the basement membrane and the underlying connective tissue and the anchoring fibrils were pulled off from the collagen layer. In contrast, connections between the epithelial cells and basement membrane were preserved, resulting in exposure of the interstitial surfaces of the laminae densae. Scanning electron-microscopic observations of those interstitial surfaces were possible using the specimens prepared as above. The basement membranes of these three oral epithelia were morphologically the same not only by transmission but also by scanning electron microscopy. Scanning electron-microscopic observations revealed that their laminae densae were composed of fine fibrils and demonstrated three-dimensionally the projection of the anchoring fibrils from the laminae densae to the interstitial side. These findings coincide with those for epidermal basement membrane, which had already been observed with the same method.

The objectives of this study were to estimate the structure of the lipid hydroperoxide-modified lysine residue and to prove the presence of the adducts in vivo. The reaction of lipid hydroperoxide toward the lysine moiety was investigated employing N-benzoyl-glycyl-L-lysine (Bz-Gly-Lys) as a model compound of Lys residues in protein and 13-hydroperoxyoctadecadienoic acid (13-HPODE) as a model of the lipid hydroperoxides. One of the products, compound X, was isolated from the reaction mixture of 13-HPODE and Bz-Gly-Lys and was then identified as N-benzoyl-glycyl-Nepsilon-(hexanonyl)lysine. To prove the formation of Nepsilon-(hexanonyl)lysine, named HEL, in protein exposed to the lipid hydroperoxide, the antibody to the synthetic hexanonyl protein was prepared and then characterized in detail. Using the anti-HEL antibody, the presence of HEL in the lipid hydroperoxide-modified proteins and oxidized LDL was confirmed. Furthermore, the positive staining by anti-HEL antibody was observed in human atherosclerotic lesions using an immunohistochemical technique. The amide-type adduct may be a useful marker for the lipid hydroperoxide-derived modification of biomolecules.

Methylglyoxal (MG), an endogenous metabolite that increases in diabetes and is a common intermediate in the Maillard reaction (glycation), reacts with proteins and forms advanced glycation end products. In the present study, we identify a novel MG-arginine adduct and also characterize the structure of a major fluorescent adduct. In addition, we describe the immunochemical study on the MG-arginine adducts using monoclonal antibody directed to MG-modified protein. Upon incubation of Nalpha-acetyl-L-arginine with MG at 37 degrees C, two nonfluorescent products and one fluorescent product were detected as the major products. The nonfluorescent products were identified as the Ndelta-(5-hydro-5-methyl-4-imidazolon-2-yl)-L-ornithine derivatives (5-hydro-5-methylimidazolone) and a novel MG-arginine adduct having a tetrahydropyrimidine moiety (Ndelta-(4-carboxy-4,6-dimethyl-5, 6-dihydroxy-1,4,5,6-tetrahydropyrimidine-2-yl)-L-ornithine). On the basis of the following chemical and spectroscopic evidence, the major fluorescent product, putatively identified as Ndelta-(5-methylimidazolon-2-yl)-L-ornithine (5-methylimidazolone), was found to be identical to Ndelta-(5-hydroxy-4, 6-dimethylpyrimidine-2-yl)-L-ornithine (argpyrimidine): (i) the low and high resolution fast atom bombardment-mass spectrometry gave a molecular ion peak at m/z of 297 (M+H) and a molecular formula of C10H25O6N4, respectively, which coincided with argpyrimidine; (ii) the 1H NMR spectrum of this product in d6-Me2SO showed a singlet at 2.10 ppm corresponding to six protons; (iii) the peak corresponding to the 5-methylimidazolone derivative was not detected by the liquid chromatography-mass spectrometry with the mode of selected ion monitoring; (iv) incubation of 5-hydro-5-methylimidazolone, a putative precursor of 5-methylimidazolone, at 37 degrees C for 14 days scarcely generated 5-methylimidazolone. On the other hand, as an immunochemical approach to the detection of these MG adducts, we raised the monoclonal antibodies (mAb3C and mAb6B) directed to the MG-modified protein and found that they specifically recognized the major fluorescent product, argpyrimidine, as the dominant epitope. The immunohistochemical analysis of the kidneys from diabetic patients revealed the localization of argpyrimidine in intima and media of small artery walls. Furthermore, the accumulation of argpyrimidine was also observed in some arterial walls of the rat brain after middle cerebral artery occlusion followed by reperfusion. These results suggest that argpyrimidine may contribute to the progression of not only long term diabetic complications, such as nephropathy and atherosclerosis, but also the tissue injury caused by ischemia/reperfusion.

Three cases of bifid ribs were found in two cadavers during routine dissections at the Iwate Medical University School of Dentistry. All of the cases were found in the third or the fourth rib. The distal parts of the osseous rib bifurcated with an angle of 60 degrees and both of the branches had their own costal cartilage. The costal cartilage fused again to form the trunk which was connected to the sternum. The space between the two branches was filled with presumably normal intercostal muscles. Blood supply was maintained by a small branch from the interthoracic artery to the upper branches. However, the intercostal nerves did not branch toward the upper branch but only ran along the lower margins of the lower branches of the bifid ribs.

Various immobilized lectins can be successfully used for fractionation and for structural studies of asparagine-linked sugar chains of glycoproteins (see Note 9). This method needs10 ng of a radiolabeled oligosaccharide prepared from a glycoprotein by hydrazinolysis or by digestion with endo-beta-N-acetylglucosaminidases. The fractionation and the structural assessment through the use of immobilized lectins make the subsequent structural studies much easier.

Lipoprotein peroxidation, especially the modification of apolipoprotein B-100, has been implicated to play an important role in the pathogenesis of atherosclerosis. However, there have been few detailed insights into the chemical mechanism of derivatization of apolipoproteins during oxidation. In the present study, we provide evidence that the formation of the toxic pollutant acrolein (CH2=CH-CHO) and its conjugate with lysine residues is involved in the oxidative modification of human low density lipoprotein (LDL). Upon incubation with LDL, acrolein preferentially reacted with lysine residues. To determine the structure of acrolein-lysine adduct in protein, the reaction of acrolein with a lysine derivative was carried out. Employing Nalpha-acetyllysine, we detected a single product, which was identified to be a novel acrolein-lysine adduct, Nalpha-acetyl-Nepsilon-(3-formyl-3,4-dehydropiperidino )lysine. The acid hydrolysis of the adduct led to the derivative that was detectable with amino acid analysis. It was revealed that, upon in vitro incubation of LDL with acrolein, the lysine residues that had disappeared were partially recovered by Nepsilon-(3-formyl-3, 4-dehydropiperidino)lysine. In addition, we found that the same derivative was detected in the oxidatively modified LDL with Cu2+ and that the adduct formation was correlated with LDL peroxidation assessed by the consumption of alpha-tocopherol and cholesteryl ester and the concomitant formation of cholesteryl ester hydroperoxide. Enzyme-linked immunosorbent assay that measures free acrolein revealed that a considerable amount of acrolein was released from the Cu2+-oxidized LDL. Furthermore, metal-catalyzed oxidation of arachidonate was associated with the formation of acrolein, indicating that polyunsaturated fatty acids including arachidonate represent potential sources of acrolein generated during the peroxidation of LDL. These results indicate that acrolein is not just a pollutant but also a lipid peroxidation product that could be ubiquitously generated in biological systems.

Volatile components formed from the reaction of monosaccharides or disaccharides with beta-alanine were investigated in a dry condition as a model system of cookie processing. Maltol is a common compound formed in the Maillard reaction, but it was very difficult to detect it in previous experiments using actual cookie materials. In this work, we investigated the principal compounds and maltol formation from the reaction of monosaccharides or disaccharides with beta-alanine at 150 degrees C for 10 min. Neither the reaction of monosaccharides nor the disaccharides with beta-alanine resulted in the formation of maltol. 2,3-Dihydro-3,5-dihydroxy-6-methyl-4(H)-pyran-4-one (DDMP) was detected as a principal product from the reaction of monosaccharides with beta-alanine. 5-Hydroxymethyl-2-furfural was also confirmed as being a major product in both reactions.

The reservoir and transit capacity of the post-gastrectomy jejunal pouch was evaluated, using a radioisotopic method, to examine the relationship of the gastric emptying to postprandial symptoms and to the food intake status.Thirty-seven patients who had undergone total gastrectomy for cancer (Roux-Y reconstruction, 8; Hunt- Lawrence pouch and Roux-Y, 15; pouch interposition, 5; modified pouch interposition, 9) were retrospectively studied. Based on the percent retention in the gastric substitute, the emptying curves were classed as showing delayed, intermediate, and rapid emptying types.All of the patients with pouch reconstruction showed either delayed or intermediate emptying. High frequency of the sensation of epigastric fullness, nausea, or vomiting was demonstrated in the patients with delayed emptying (p0.01). The patients with delayed emptying showed poor food intake compared to those with intermediate emptying (p0.05). Given the X-ray video film and endoscopic findings, the delayed emptying was thought to be due to poor drainage of the efferent loop resulting from the post-operative adhesions.The present study revealed that delayed emptying is associated with postprandial symptoms and with poor food intake. The examination of gastric emptying is useful in evaluating or predicting the postoperative status.

The DNA base-modified product 8-hydroxy-2'-deoxyguanosine (8-OHdG) is one of the most commonly used markers for the evaluation of oxidative DNA damage. A monoclonal antibody specific for 8-OHdG (N45.1) was characterized and applied in quantitative immunohistochemistry. N45.1 recognized both the modified base and deoxyribose structure of 8-OHdG and required a concentration two orders higher of 8-hydroxyguanosine as a competitor in the ELISA. In addition, N45.1 did not cross-react with the original four deoxyribonucleosides, other DNA base-modified products such as 8-hydroxy-2'-deoxyadenosine and O6-methyl-2'-deoxyguanosine, or urine components such as uric acid, creatine, and creatinine. A ferric nitrilotriacetate-induced rat renal carcinogenesis model was used for the evaluation of quantitative immunohistochemistry. The 8-OHdG index of quantitative immunohistochemistry, as analyzed by NIH image freeware, correlated reasonably well with the 8-OHdG amount determined by high-performance liquid chromatography with an electrochemical detector-except for a difference in peak time, which could be attributed to the selection of target location. The present method has advantages over the high-performance liquid chromatography/electrochemical detector, gas chromatography/mass spectrometry, and 32P-postlabeling methods in that it allows localization of 8-OHdG to be specified without the risk of artifactual production of 8-OHdG during the DNA extraction and hydrolytic processes.