Porphyromonas gingivalis Lipopolysaccharides Promote Proliferation and Migration of Human Vascular Smooth Muscle Cells through the MAPK/TLR4 Pathway

Atherosclerosis is a major cause of mortality worldwide. The initial change in atherosclerosis is intimal thickening due to muscle cell proliferation and migration. A correlation has been observed between periodontal disease and atherosclerosis. Here, we investigated the proliferation and migration of human aortic smooth muscle cells (HASMCs) using Porphyromonas gingivalis-derived LPS (Pg-LPS). To elucidate intracellular signaling, toll-like receptor 4 (TLR4) and myeloid differentiation factor 88 (MyD88) of HASMCs were knocked down, and the role of these molecules in Pg-LPS-stimulated proliferation and migration was examined. The role of mitogen-activated protein kinase (MAPK) in HASMC proliferation and migration was further elucidated by MAPK inhibition. Pg-LPS stimulation increased the proliferation and migration of HASMCs and activated the TLR4/MyD88 pathway. TLR4 knockdown inhibited Pg-LPS stimulated HASMCs proliferation and migration. Pg-LPS stimulation led to the phosphorylation of P38 MAPK, JNK, and ERK, and MyD88 knockdown inhibited the phosphorylation of P38 MAPK and JNK but not ERK. P38 MAPK and SAPK/JNK inhibition did not suppress the proliferation of HASMCs upon Pg-LPS stimulation, but ERK inhibition significantly inhibited proliferation. SAPK/JNK and ERK inhibition suppressed Pg-LPS-stimulated migration of HASMCs. In conclusion, our findings suggest that Pg-LPS may promote atherosclerosis via the activation of MAPK through TLR4.     

Gluten Gel and Film Properties in the Presence of Cysteine and Sodium Alginate

ABSTRACT:  Wheat flour has an ability of forming dough by mixing with water, which exhibits a rheological property required for making bread. The major protein is gluten, which is a valuable protein material for food industry. In this study, gluten protein gels and films were formed with cysteine and sodium alginate. Adding cysteine improved gel and film properties (stress relaxation behavior, bending strength). The gel containing 0.01 M cysteine had a longer relaxation time and was more rigid than the gel without cysteine. Although adding sodium alginate to the gluten suspension containing cysteine improved the water-holding ability and homogeneity of the gel network, the film from this gel was more brittle than the gluten film with cysteine alone. Microstructural observations of the gels and films with scanning electron microscopy suggested that water evaporation was more heterogeneous from the gel containing sodium alginate than from the gel with cysteine alone. Fourier transform-infrared (FT-IR) analysis during film formation suggested that the presence of cysteine encourages interaction between gluten molecules and results in intermolecular β-sheet formation in earlier stages than in the no additive condition. FT-IR results also suggested that the combined effect of sodium alginate and cysteine on the protein secondary structure was remarkably different from that of cysteine alone. Our results suggest that addition of a suitable amount of cysteine (0.01 M) and heat treatment to 80 °C during gluten gel and film formation induces a homogenous network in the gel and film by regulating disulfide–sulfide interactions.     

Determination method for ractopamine in swine and cattle tissues using LC/MS

Simple and reliable methods using LC/MS have been developed for the determination of the beta-agonist ractopamine in swine and cattle tissues. Ractopamine was extracted with ethyl acetate from muscle and liver, and the ethyl acetate layer was evaporated to dryness. The residue was purified by partition with acetonitrile/n-hexane. In the case of fat, ractopamine was extracted and purified by partition with acetonitrile/n-hexane. The resulting acetonitrile solutions were evaporated to dryness. The residue was dissolved in methanol, and subjected to LC/MS. The LC separation was performed on a Wakosil-II 3C18HG column (150 x 3 mm i.d.) in isocratic mode with 0.05% trifluoroacetic acid-acetonitrile (80:20) as a mobile phase at a flow rate of 0.4 mL/min. The MS detection was performed in the selected ion recording (SIR) mode, with detection of the M + H+ ion of ractopamine (m/z 302) produced by electrospray ionization (ESI). The mean recoveries of the drug from swine muscle (0.01 microg fortified), fat (0.01 microg fortified) and liver (0.04 microg/g fortified) were 99.7%, 99.5% and 100.8%, and those from cattle samples were 108.3%, 97.0% and 109.4%, respectively. The relative standard deviations (RSDs) ranged from 0.1% to 9.5%. The limit of quantification (LOQ) of the drug was 1 ng/g.     

Distribution of deoxynivalenol and nivalenol in milling fractions from fusarium-infected Japanese wheat cultivars

The fate of the Fusarium mycotoxins deoxynivalenol and nivalenol during the milling of Japanese wheat cultivars artificially infected with Fusarium was investigated. Grain samples with different mycotoxin concentrations were milled using a laboratory-scale test mill to produce eight fractions: three breaking flours (1B, 2B, and 3B), three reduction flours (1M, 2M, and 3M), wheat bran, and wheat shorts. Patent flour for human consumption was made from the 1B, 2B, 1M, and 2M flours, and low-grade flour was made from 3B and 3M flours. The four resulting samples (patent flour, low-grade flour, bran, and shorts) were analyzed for deoxynivalenol and/or nivalenol with an in-house validated analytical method using high-performance liquid chromatography with UV absorbance detection. In samples with different mycotoxin concentrations, the distribution of those toxins differed among the milling fractions. Grains with a lower level of contamination produced bran and shorts samples with a high relative concentration of nivalenol. A high percentage of nivalenol was found in patent flour, followed by bran. Contrary to the less-contaminated sample, the concentration of nivalenol in moderately contaminated grain was high only in the shorts sample. The highest percentage of deoxynivalenol and nivalenol was observed in the patent flour. The results of this study indicate that the distribution of deoxynivalenol and nivalenol in milled Japanese wheat could be influenced by the contamination level of the original grain, and the milling process is not always effective for removal of toxins from wheat grains.     

Overexpression of NUDT7, a candidate quantitative trait locus for pork color,downregulates heme biosynthesis in L6 myoblasts

While testing a quantitative trait locus (QTL) for pork color in a cross population of pigs from the mating of Large White dams to a Japanese wild boar, our laboratory discovered a candidate gene (NUDT7) that might affect heme biosynthesis in porcine muscle. Therefore, this experiment was designed to test the effect of NUDT7 on heme biosynthesis in cultured myoblasts. Rat L6 myoblasts were transfected with a mammalian expression vector for pig NUDT7 immediately after the induction of cell differentiation, and samples were harvested at 2, 4, 6, and 8 days. Expression of exogenous NUDT7 mRNA was highest on day 4, when the heme content was substantially lower (P < 0.01) than that of the control (14.2 vs. 63.9 pmol/10⁵ cells). These results suggest that overexpression of pig NUDT7 may be associated with heme biosynthesis downregulation in skeletal muscle, which may partially explain differences in meat color among breeds of livestock.     

Formation of damascenone derived from glycosidically bound precursors in green tea infusions

Damascenone is well-known for its potent flavour with an extremely low odour threshold. Several glycosidically bound precursors of damascenone have been isolated from several plants, but little is known about their occurrences in green tea infusions. In this work, three major glycosidic precursors of damascenone, 9-O-β-d-glucopyranosyl-megastigma-6,7-dien-3,5,9-triol (1a), 9-O-β-d-glucopyranosyl-3-hydroxy-7,8-didehydro-β-ionol (2a), and 3-O-β-d-glucopyranosyl-3-hydroxy-7,8-didehydro-β-ionol (2b) were isolated and identified in green tea infusions, and the stereochemistries at C-3 and C-9 positions of aglycone parts of the three glycosidic precursors were determined as (3S, 9R)-1a, (3R, 9R)-2a, and (3R, 9R)-2b, respectively. Compounds 1a, 2a, and 2b as well as 3-O-β-d-glucopyranosyl-megastigma-6,7-dien-3,5,9-triol (1b) were hydrolysed to form damascenone in a model system with strong acidic conditions (pH 2.0) and at high temperature (90 °C). In contrast to hydrolysis of 2a and 2b, more damascenone was transformed from 1a and 1b. Furthermore, the β-d-glucosyl moiety at the C-3 position gave a higher dehydration rate from megastigma-6,7-dien-3,5,9-triol to 3-hydroxy-7,8-didehydro-β-ionol than compound 1a carrying the sugar residue at C-9 position. Interestingly, the four glycosidic precursors of damascenone were not hydrolysed to give damascenone under slightly acidic conditions (pH 5.4 and 120 °C for 10 min), but they could be transformed to damascenone in the presence of green tea infusions even under the equal conditions.     

Synthesis of polypyridine-graft-PEG copolymer for protein repellent and stable interface

Polypyridine grafted with poly(ethylene glycol) (Py-g-PEG) have been synthesized. Radical copolymerization of methyl-terminated PEG macromonomer with 4-pyridylmethyl methacrylate homogeneously proceeded and the obtained copolymer spontaneously adsorbs from aqueous solution onto gold surfaces, where the pyridine parts act as the multipoint anchor to the surface and the PEG parts provide the strong steric repulsion between the chains. As a result, the highly protein repellent and stable surface was constructed through multipoint pyridine attachment as compared with singlepoint pyridine attachment. Py-g-PEGs thus synthesized are promising material to functionalize metal and semiconductor material and to self-assemble into micelle in biotechnological and biomedical field.     

Intestinal responses to hyperosmolar glucose infusion in dogs

Physiological and morphological changes of small intestine after hyperosmolar glucose infusions into canine jejunum were studied using in vivo perfusion model. Infusions of 40 ml of 50% and 20% glucose solution into the jejunal loops induced biphasic osmolar degression in the lumen. Osmolarity of jejunal venous blood was rapidly increased and maintained the maximal level (approximately 320 mOM/L). Blood flow to the jejunal loop was significantly increased after 50% glucose infusion compared to 5% glucose infusion. Most characteristic electron microscopic change of jejunal epithelial cell was pseudo-pod like process projected into the jejunal lumen, which was very similar to that of cholera.