Leukocyte activation by (1→3)-β-D glucans

We studied the activities of several kinds of β-glucans, including sonifilan, grifolan, Sclerotinia sclerotiorum glucan, laminarin and zymosan, on macrophages. Preculture of macrophages with inactive β-glucans rendered the cells unresponsive to subsequent stimulation with grifolan, suggesting a specific pathway in the β-glucan structure. The importance of protein C and phosphorylation of mitogen-activated protein kinase was demonstrated in the activation with grifolan or zymosan. Immunoprecipitation of complement receptor (CR3), coprecipitated other proteins carrying phosphotyrosine residues in stimulation with grifolan. These data suggest that protein kinase C and tyrosine kinases are essential for signal transduction, and that CR3 might participate in the activation through interaction with other intracellular proteins

Lipase-Catalyzed Synthesis of Unsaturated Acyl L-Ascorbates and Their Ability to Suppress the Autoxidation of Polyunsaturated Fatty Acids

ABSTRACT: L-Ascorbic acid and various polyunsaturated fatty acids (PUFA) were condensed at 55°C by the immobilized lipase Chirazyme L-2 in dry acetone to produce the unsaturated acyl ascorbates. The PUFA moieties of the products were much more resistant to autoxidation at 65°C and nearly 0% relative humidity than the corresponding unmodified PUFA. The effects of the molar ratio of ascorbic acid or linoleoyl ascorbate to linoleic acid on the autoxidation of linoleic acid were examined. The autoxidation of linoleic acid was effectively suppressed at molar ratios greater than or equal to 0.2 when either ascorbic acid or linoleoyl ascorbate was mixed with linoleic acid. The addition of lauroyl ascorbate, synthesized through the enzyme-catalyzed condensation of ascorbic acid and lauric acid in acetone, to docosahexaenoic acid also significantly suppressed the autoxidation of docosahexaenoic acid at molar ratios of ≥0.2. Paper no. J9826 in JAOCS 78, 823-826 (August 2001). KEY WORDS: Acyl ascorbate, L-ascorbic acid, autoxidation, condensation, immobilized lipase, polyunsaturated fatty acid. Much attention has been paid to the use of polyunsaturated fatty acids (PUFA) as components in foods (1). However, PUFA are susceptible to autoxidation (2,3), and the autoxidation causes deterioration of the foods. L-Ascorbic acid is a hydrophilic antioxidant with a strong reducing ability. The lipase-catalyzed synthesis of acyl ascorbate in a solid-phase system (4) or in an organic solvent (5-9) has been reported. However, its ability to suppress lipid autoxidation has not been reported. In a previous paper (10), we reported the synthesis of 6-O-eicosapentaenoyl L-ascorbate by the lipase-catalyzed condensation of eicosapentaenoic acid and L-ascorbic acid in acetone and compared its autoxidation process to that of the unmodified eicosapentaenoic acid. In the work described in this paper, some PUFA L-ascorbates were synthesized using an immobilized lipase from Candida antarctica, Chirazyme ® L-2, and their autoxidation processes were then observed. The PUFA used were linoleic, α-linolenic, γ-linolenic, arachidonic, and docosahexaenoic acids. The effect of the molar ratio of unmodified L-ascorbic acid or linoleoyl ascorbate to linoleic acid on the suppression of the autoxidation of linoleic acid was examined. We previously reported the lipase-catalyzed condensation of ascorbic acid with various medium-chain fatty acids having carbon numbers of 6, 8, 10, and 12 in acetonitrile (11). Therefore, the ability of lauroyl ascorbate to suppress the autoxidation of docosahexaenoic acid was also evaluated in the present work. EXPERIMENTAL PROCEDURES Materials. γ-Linolenic and docosahexaenoic acids were supplied by the Maruha Corporation (Tokyo, Japan), and their purities were both greater than 95% based on gas chromatographic (GC) analysis. L-Ascorbic acid, linoleic acid, acetone, and hexane were purchased from Nacalai Tesque (Kyoto, Japan). α-Linolenic, arachidonic, and lauric acids were purchased from Sigma Chemical (St. Louis, MO). Immobilized lipase from C. antarctica, Chirazyme ® L-2 c.-f. C2, was obtained from Roche Molecular Biochemicals (Mannheim, Germany). The enzyme is the same as Novozym ® 435 according to the manufacturer. Soybean oil was purchased from Wako Pure Chemical Industries (Osaka, Japan). Condensation reaction. Acetone was first dehydrated by adding 5 Å molecular sieves. The water content of the acetone was about 0.01% (vol/vol), and was determined for each experiment by a Karl-Fischer titration. L-Ascorbic acid (0.125 mmol) and a PUFA [linoleic acid (0.577 mmol)], γ-linolenic acid (0.600 mmol), arachidonic acid, (0.638 mmol), α-linolenic acid (0.611 mmol), and docosahexaenoic acid (0.648 mmol)] were weighed into an amber glass vial with a screw cap, and 200 mg of Chirazyme L-2 and 2.5 mL of dehydrated acetone were added to the vial. The headspace of the vial was filled with nitrogen, and the vial was tightly sealed. The vial was then immersed in a waterbath at 55°C with vigorous shaking to commence the condensation reaction. At appropriate intervals, 10 µL of the reaction mixture was taken and mixed with 50 µL of a 50 mM solution of toluene in high-performance liquid chromatography (HPLC) eluent [acetonitrile/tetrahydrofuran/0.1% (vol/vol) phosphoric acid (50:22:28 by vol) as the internal standard for the HPLC analysis and then with 40 µL of HPLC eluent. The analysis was carried out by HPLC with a YMC-Pack C8 column (4.6 mm × 250 mm; YMC Inc., Kyoto, Japan) and an ultraviolet (UV) detector (245 nm). The mixture (20 µL) was applied to the column and eluted with the eluent at 1.5 mL/min. The retention times o

Neuroendocrine Carcinoma of the Stomach: A Case Study

Gastric neuroendocrine carcinomas are rare and have a poor prognosis, and the diagnostic criteria for this disease have recently changed. We herein report a case of sporadic gastric neuroendocrine carcinoma. A 75-year-old man was referred to our hospital with epigastric pain. Endoscopic examination revealed a localized ulcerative lesion (diameter, 4 cm) at the upper stomach. The diagnosis on biopsy was neuroendocrine carcinoma. Total gastrectomy with D2 lymphadenectomy, splenectomy, and cholecystectomy was performed. Pathologically, the tumor infiltrated the subserosal layer, and 6/49 lymph nodes were involved. The tumor was uniform in shape and arranged in a rosette-like structure to form solid nests, with medium-sized, round-to-cuboid-shaped tumor cells and intense mitosis 46/10 HPF. It was positive for synaptophysin and chromogranin A, and the Ki-67 labeling index was 70–80%. The diagnosis of neuroendocrine carcinoma was made according to the WHO 2010 criteria. The patient was followed up for three years without recurrence

School grade and sex differences in domain-specific sedentary behaviors among Japanese elementary school children: a cross-sectional study

Background: It is vital to reduce the proportion of sedentary behavior in children. Understanding the duration and behavioral context is needed. The present study examined school-grade and sex differences in domain-specificsedentary times and concurrence with screen-time guidelines among Japanese elementary school children.Methods: A total of 625 children (330 boys) were surveyed in 2010 and 2014. Using a questionnaire, data regarding participants’ grade (first through third grades: lower grades; fourth through six grades: higher grades), sex, weight, and height were collected in addition to the time spent per day engaging in each specific sedentary behavior separately:(1) reading or listening to music, (2) TV or video viewing, (3) TV game use, (4) internet use excluding class, (5) homework, and (6) car travel. Two-way analysis of covariance and logistic regression analyses, adjusted for BMI and moderate to vigorous physical activity, were used to examine school-grade and sex differences in sedentary behaviors and the independent risk of exceeding recommended total daily screen time (< 2 h).Results:On 625 children, mean minutes (SD) of sedentary behavior per week in (1) – (6) were 90.3 (123.4), 535.0 (356.6),167.3 (222.1), 23.9 (70.9), 264.9 (185.3), and 33.4 (61.2) in weekdays and 42.1 (70.0), 323.9 (232.0), 123.0 (96.4),15.8 (49.9), 74.4 (96.4), and 71.3 (84.9) in weekends, respectively. There were differences in the minutes of sedentary behavior between participants of 2010 and 2014; e.g., TV game use and homework in weekdays and weekdays and car travel in weekends. Boys spent more time in TV game use, and girls spent more time reading, listening to music, doing homework, and car travel. Higher-grade students spent more time reading or listening to music, using a computer, and doing homework. Higher-grade students were 2.09 times (95% CI: 1.32 − 3.30) in whole week, 2.08 times (95% CI: 1.45 − 3.00) in weekday, and 1.88 times (95% CI: 1.29 − 2.74) in weekend more likely to spend ≥2 h per day in domains (2) − (4) (screen-time) than lower-grade students.Conclusions: Time spent engaging in each domain-specific sedentary behavior differed according to sex and school grade. Higher-grade students were less likely to meet screen-time guidelines. These findings highlight the need fordomain-focused strategies to decrease sedentary behavior in Japanese school-age children

Protective role of vascular endothelial growth factor in endotoxin-induced acute lung injury in mice

<p>Abstract</p> <p>Background</p> <p>Vascular endothelial growth factor (VEGF), a substance that stimulates new blood vessel formation, is an important survival factor for endothelial cells. Although overexpressed VEGF in the lung induces pulmonary edema with increased lung vascular permeability, the role of VEGF in the development of acute lung injury remains to be determined.</p> <p>Methods</p> <p>To evaluate the role of VEGF in the pathogenesis of acute lung injury, we first evaluated the effects of exogenous VEGF and VEGF blockade using monoclonal antibody on LPS-induced lung injury in mice. Using the lung specimens, we performed TUNEL staining to detect apoptotic cells and immunostaining to evaluate the expression of apoptosis-associated molecules, including caspase-3, Bax, apoptosis inducing factor (AIF), and cytochrome C. As a parameter of endothelial permeability, we measured the albumin transferred across human pulmonary artery endothelial cell (HPAEC) monolayers cultured on porous filters with various concentrations of VEGF. The effect of VEGF on apoptosis HPAECs was also examined by TUNEL staining and active caspase-3 immunoassay.</p> <p>Results</p> <p>Exogenous VEGF significantly decreased LPS-induced extravascular albumin leakage and edema formation. Treatment with anti-VEGF antibody significantly enhanced lung edema formation and neutrophil emigration after intratracheal LPS administration, whereas extravascular albumin leakage was not significantly changed by VEGF blockade. In lung pathology, pretreatment with VEGF significantly decreased the numbers of TUNEL positive cells and those with positive immunostaining of the pro-apoptotic molecules examined. VEGF attenuated the increases in the permeability of the HPAEC monolayer and the apoptosis of HPAECs induced by TNF-α and LPS. In addition, VEGF significantly reduced the levels of TNF-α- and LPS-induced active caspase-3 in HPAEC lysates.</p> <p>Conclusion</p> <p>These results suggest that VEGF suppresses the apoptosis induced by inflammatory stimuli and functions as a protective factor against acute lung injury.</p

Lipopolysaccharide Interaction with Cell Surface Toll-like Receptor 4-MD-2: Higher Affinity than That with MD-2 or CD14

Toll-like receptors (TLRs) are innate recognition molecules for microbial products, but their direct interactions with corresponding ligands remain unclarified. LPS, a membrane constituent of gram-negative bacteria, is the best-studied TLR ligand and is recognized by TLR4 and MD-2, a molecule associated with the extracellular domain of TLR4. Although TLR4-MD-2 recognizes LPS, little is known about the physical interaction between LPS and TLR4-MD-2. Here, we demonstrate cell surface LPS–TLR4-MD-2 complexes. CD14 greatly enhances the formation of LPS–TLR4-MD-2 complexes, but is not coprecipitated with LPS–TLR4-MD-2 complexes, suggesting a role for CD14 in LPS loading onto TLR4-MD-2 but not in the interaction itself between LPS and TLR4-MD-2. A tentative dissociation constant (Kd) for LPS–TLR4-MD-2 complexes was ∼3 nM, which is ∼10–20 times lower than the reported Kd for LPS–MD-2 or LPS–CD14. The presence of detergent disrupts LPS interaction with CD14 but not with TLR4-MD-2. E5531, a lipid A antagonist developed for therapeutic intervention of endotoxin shock, blocks LPS interaction with TLR4-MD-2 at a concentration 100 times lower than that required for blocking LPS interaction with CD14. These results reveal direct LPS interaction with cell surface TLR4-MD-2 that is distinct from that with MD-2 or CD14

MR perfusion imaging by alternate slab width inversion recovery arterial spin labeling (AIRASL): a technique with higher signal-to-noise ratio at 3.0 T

Object: To propose a new arterial spin labeling (ASL) perfusion-imaging method (alternate slab width inversion recovery ASL: AIRASL) that takes advantage of the qualities of 3.0 T. Materials and methods: AIRASL utilizes alternate slab width IR pulses for labeling blood to obtain a higher signal-to-noise ratio (SNR). Numerical simulations were used to evaluate perfusion signals. In vivo studies were performed to show the feasibility of AIRASL on five healthy subjects. We performed a statistical analysis of the differences in perfusion SNR measurements between flow-sensitive alternating inversion recovery (FAIR) and AIRASL. Results: In signal simulation, the signal obtained by AIRASL at 3.0 and 1.5 T was 1.14 and 0.85%, respectively, whereas the signal obtained by FAIR at 3.0 and 1.5 T was 0.57 and 0.47%, respectively. In an in vivo study, the SNR of FAIR (3.0 T) and FAIR (1.5 T) were 1.73 ± 0.49 and 1.02 ± 0.20, respectively, whereas the SNRs of AIRASL (3.0 T) and AIRASL (1.5 T) were 3.93 ± 1.65 and 1.34 ± 0.31, respectively. SNR in AIRASL at 3.0 T was significantly greater than that in FAIR at 3.0 T. Conclusion: The most significant potential advantage of AIRASL is its high SNR, which takes advantage of the qualities of 3.0 T. This sequence can be easily applied in the clinical setting and will enable ASL to become more relevant for clinical application. © 2012 ESMRMB.Article in Press エンバーゴ設定 Thesis of Fujiwara, Yasuhiro / 藤原 康博 博士学位論文(金沢大学 / 大学院医薬保健学総合研究科