Diagnostic evaluation of 2', 5'-oligoadenylate synthetase activities and antibodies against Epstein-Barr virus and Coxiella burnetii in patients with chronic fatigue syndrome in Japan

To investigate the association of viral infections with chronic fatigue syndrome (CFS), we assayed 2', 5'-oligoadenylate synthetase (2-5AS) activities in peripheral blood mononuclear cells from CFS patients in Japan. These patients were diagnosed in two hospitals, H1 and H2, located in different areas of the country. The activities were detected in 19 (86%) and 7 (32%) of each of the 22 patients in H1 and H2, respectively, while they were detected in only four (11%) out of the 38 healthy controls. IFN-alpha was similarly detected in a few CFS patients and healthy controls. We also assayed the antibody titers against Epstein-Barr virus (EBV) and Coxiella burnetii in these patients. The EBV anti-EA-IgG antibodies were detected in two (9%) and seven (32%) of each of the 22 patients in H1 and H2, respectively. Anti-C. burnetii IgG antibodies were detected in six (27%) out of 22 patients in H1 but not in 22 patients in H2, while they were detected in one (11%) of the nine healthy controls. Some CFS patients may be associated with EBV or C. burnetii infection. There were some statistical correlations between the 2-5AS activities and antibody titers of EA-IgG (P < 0.05, Student's t-test) but not to the antibody titers of C. burnetii. The up-regulation of 2-5AS activities suggests immunological dysfunctions with some virus infections in the CFS patients. Our results indicate that 2-5AS activities are useful for a diagnostic marker of CFS and for exploring the complicated pathogenesis of CFS.     

Synthesis of an unnatural N-glycan-linked dolichyl pyrophosphate precursor

An unnatural alpha-D-mannopyranose-linked chitobiosyl dolichyl pyrophosphate, a stereoisomer of the N-glycan biosynthesis intermediate, was synthesized. The protected trisaccharide, alpha-D-Man-(1-->4)-beta-D-GlcNAc-(1-->4)-D-GlcNAc, carrying a 4-methylbenzoyl group was prepared for the convenience of a TLC analysis. 1-O-Phosphorylation, condensation with dolichyl phosphate, and subsequent deprotection afforded the title compound.     

Organogenesis of heart-vascular system derived from mouse 2 cell stage embryos and from early embryonic stem cells in vitro

Regenerative medical treatment with embryonic stem cells (an ES cell) is a goal for organ transplantation. Structures that are tubular in nature (i.e. blood capillaries) were induced from early embryonic stem (EES) cells in vitro using embryotrophic factor (ETFs). In addition, cardiac muscle cells could be identified as well. However, differentiation of EES cells into a complete cardiovascular system was difficult because 3 germ layer primordial organs are directed embryologically in various ways and it is not possible to guide only cardiovascular organs. Thus, we introduced ETFs after the formation of an embryoid body and were successful in cloning cell clusters that beat, thus deriving only cardiovascular organs. The application of this to the treatment of various cardiovascular diseases is promising.     

Properties of a novel extracellular cell-free ice nuclei from ice-nucleating Pseudomonas antarctica IN-74

Some ice-nucleating bacterial strains, including Pantoea ananatis (Erwinia uredovora), Pseudomonas fluorescens, and Pseudomonas syringae isolates, were examined for the ability to shed ice nuclei into the growth medium. A novel ice-nucleating bacterium, Pseudomonas antarctica IN-74, was isolated from Ross Island, Antarctica. Cell-free ice nuclei from P. antarctica IN-74 were different from the conventional cell-free ice nuclei and showed a unique characterization. Cell-free ice nuclei were purified by centrifugation, filtration (0.45 microm), ultrafiltration, and gel filtration. In an ice-nucleating medium in 1 liter of cell culture, maximum growth was obtained with the production of 1.9 mg of cell-free ice nuclei. Ice nucleation activity in these cell-free ice nuclei preparations was extremely sensitive to pH. It was demonstrated that the components of cell-free ice nuclei were protein (33%), saccharide (12%), and lipid (55%), indicating that cell-free ice nuclei were lipoglycoproteins. Also, carbohydrate and lipid stains showed that cell-free ice nuclei contained both carbohydrate and lipid moieties.     

Stimulatory effect of a dietary casein phosphopeptide preparation on the mucosal IgA response of mice to orally ingested lipopolysaccharide from Salmonella typhimurium

The effect on immunoglobulin production of a commercially available casein phosphopeptide preparation (CPP-III) consisting mainly of bovine alpha s2-casein (1-32) and beta-casein (1-28) in mice that had orally ingested lipopolysaccharide (LPS) from Salmonella typhimurium was investigated. No significant difference in body weight gain was observed between the mice fed on the CPP-III-added diet and those fed on the control diet. The mice fed on the CPP-III-added diet exhibited similar serum and intestinal IgG, IgM, and IgE responses towards LPS to those fed on the control diet. In contrast, fecal and intestinal anti-LPS IgA and total IgA in mice fed on the CPP-III-added diet were significantly higher than in those fed on the control diet. Spleen cells from mice fed on the CPP-III-added diet produced larger amounts of IgA, IL-5, and IL-6 than cells from mice fed on the control diet. These results suggest that dietary casein phosphopeptide may protect a host from invasion of the intestinal mucosa by food-born pathogenic microorganisms.     

Advances in biotreatment of acid mine drainage and biorecovery of metals: 1. Metal precipitation for recovery and recycle

Acid mine drainage (AMD), an acidic metal-bearingwastewater, poses a severe pollution problem attributedto post mining activities. The metals usuallyencountered in AMD and considered of concern for riskassessment are arsenic, cadmium, iron, lead, manganese,zinc, copper and sulfate. The pollution generated byabandoned mining activities in the area of Butte, Montanahas resulted in the designation of the Silver Bow Creek–ButteArea as the largest Superfund (National Priorities List) sitein the U.S. This paper reports the results of bench-scalestudies conducted to develop a resource recovery basedremediation process for the clean up of the Berkeley Pit.The process utilizes selective, sequential precipitation (SSP)of metals as hydroxides and sulfides, such as copper, zinc,aluminum, iron and manganese, from the Berkeley Pit AMDfor their removal from the water in a form suitable foradditional processing into marketable precipitates and pigments.The metal biorecovery and recycle process is based on completeseparation of the biological sulfate reduction step and themetal precipitation step. Hydrogen sulfide produced in the SRBbioreactor systems is used in the precipitation step to forminsoluble metal sulfides. The average metal recoveries usingthe SSP process were as follows: aluminum (as hydroxide) 99.8%,cadmium (as sulfide) 99.7%, cobalt (as sulfide) 99.1% copper(as sulfide) 99.8%, ferrous iron (sulfide) 97.1%, manganese(as sulfide) 87.4%, nickel (as sulfide) 47.8%, and zinc (as sulfide)100%. The average precipitate purity for metals, copper sulfide,ferric hydroxide, zinc sulfide, aluminum hydroxide and manganesesulfide were: 92.4, 81.5, 97.8, 95.6 , 92.1 and 75.0%, respectively.The final produced water contained only calcium and magnesiumand both sulfate and sulfide concentrations were below usablewater limits. Water quality of this agriculturally usable watermet the EPA's gold standard criterion.     

A rapamycin-sensitive pathway down-regulates insulin signaling via phosphorylation and proteasomal degradation of insulin receptor substrate-1

Insulin receptor substrate-1 (IRS-1) is a major substrate of the insulin receptor and acts as a docking protein for Src homology 2 domain containing signaling molecules that mediate many of the pleiotropic actions of insulin. Insulin stimulation elicits serine/threonine phosphorylation of IRS-1, which produces a mobility shift on SDS-PAGE, followed by degradation of IRS-1 after prolonged stimulation. We investigated the molecular mechanisms and the functional consequences of these phenomena in 3T3-L1 adipocytes. PI 3-kinase inhibitors or rapamycin, but not the MEK inhibitor, blocked both the insulin-induced electrophoretic mobility shift and degradation of IRS-1. Adenovirus-mediated expression of a membrane-targeted form of the p110 subunit of phosphatidylinositol (PI) 3-kinase (p110CAAX) induced a mobility shift and degradation of IRS-1, both of which were inhibited by rapamycin. Lactacystin, a specific proteasome inhibitor, inhibited insulin-induced degradation of IRS-1 without any effect on its electrophoretic mobility. Inhibition of the mobility shift did not significantly affect tyrosine phosphorylation of IRS-1 or downstream insulin signaling. In contrast, blockade of IRS-1 degradation resulted in sustained activation of Akt, p70 S6 kinase, and mitogen-activated protein (MAP) kinase during prolonged insulin treatment. These results indicate that insulin-induced serine/threonine phosphorylation and degradation of IRS-1 are mediated by a rapamycin-sensitive pathway, which is downstream of PI 3-kinase and independent of ras/MAP kinase. The pathway leads to degradation of IRS-1 by the proteasome, which plays a major role in down-regulation of certain insulin actions during prolonged stimulation.     

Glial cells contribute more to iron and aluminum accumulation but are more resistant to oxidative stress than neuronal cells

Iron (Fe) and aluminum (Al) have been implicated in the pathogenesis of Alzheimer's disease (AD). In this study, we examined neuronal and glial cells to clarify which contributes most to metal accumulation after internalization through the transferrin-independent iron uptake (Tf-IU) systems in primary neuronal and glial predominant (NP and GP) cells from rat cerebral cortex, which affect the accumulation of transition metals in a variety of cultured cells. Al more significantly upregulated the Tf-IU activity in GP cells than in NP cells. GP cells were more resistant to Fe and Al exposure than NP cells. However, a chemiluminescence analysis specific for reactive oxygen species (ROS) showed that ROS levels in Fe- or Al-loaded NP cells were twice as high as in Fe- or Al-loaded GP cells. Northern blot analysis and gel retardation assay showed that the Al and Fe exposure taken up by the cells suppress Tf receptor mRNA expression to a greater extent in GP than NP cells, indicating that Al and Fe more markedly accumulate in glial than in neuronal cells. These results suggest that glial cells rather than neuronal cells contribute to the metal accumulation and are more resistant to oxidative stress caused by metals than neuronal cells. The present study may help to explain the pathogenesis of neurodegeneration in AD disorders caused by metal-generated oxidative stress.     

Silefrin, a sodefrin-like pheromone in the abdominal gland of the sword-tailed newt, Cynops ensicauda

Sodefrin-like female-attracting pheromone was purified from the abdominal glands of male sword-tailed newts, Cynops ensicauda, by gel-filtration chromatography and reversed-phase high-performance liquid chromatography. The final product comprises 10 amino acid residues with the sequence SILSKDAQLK which coincided with the sequence deduced from its precursor cDNA. This peptide was designated silefrin. The sequence of silefrin was different from that of sodefrin by two amino acid residues, with substitutions Leu for Pro and Gln for Leu at positions 3 and 8, respectively. Both native and synthetic silefrin exerted an equipotent activity in attracting conspecific females.