Lysophosphatidic acid,a growth factor-like lipid,in the saliva

Lysophosphatidic acid is a multifunctional phospholipid mediator and elicits a variety of biological responses in vitro and in vivo. Evidence is accumulating that lysophosphatidic acid plays important physiological roles in diverse mammalian tissues and cells. In the present study, we first examined whether lysophosphatidic acid is present in human saliva. We found that a significant amount of lysophosphatidic acid is present in the saliva (0.785 nmol/ml). The predominant fatty acyl moiety of lysophosphatidic acid was 18:1n-9 + n-7 followed by 18:0 and 16:0. A small amount of lysoplasmanic acid, an alkyl ether-linked analog of lysophosphatidic acid, was also detected in the saliva (0.104 nmol/ml). We found that physiologically relevant concentrations of lysophosphatidic acid induced accelerated growth of cells of mouth, pharynx, and esophagus origin in vitro. Lysophosphatidic acid also induced rapid increases in the intracellular free Ca2+ concentrations in these cells. We obtained evidence that lysophosphatidic acid receptor mRNAs are actually present in these cells. These results strongly suggest that lysophosphatidic acid is involved in wound healing in the upper digestive organs such as the mouth, pharynx, and esophagus.     

Inhibition of hyaluronan synthesis in Streptococcus equi FM100 by 4-methylumbelliferone.

As observed previously in cultured human skin fibroblasts, a decrease of hyaluronan production was also observed in group C Streptococcus equi FM100 cells treated with 4-methylumbelliferone (MU), although there was no effect on their growth. In this study, the inhibition mechanism of hyaluronan synthesis by MU was examined using Streptococcus equi FM100, as a model. When MU was added to a reaction mixture containing the two sugar nucleotide donors and a membrane-rich fraction as an enzyme source in a cell-free hyaluronan synthesis experiment, there was no change in the production of hyaluronan. On the contrary, when MU was added to the culture medium of FM100 cells, hyaluronan production in the isolated membranes was decreased in a dose-dependent manner. However, when the effect of MU on the expression level of hyaluronan synthase was examined, MU did not decrease either the mRNA level of the has operon containing the hyaluronan synthase gene or the protein level of hyaluronan synthase. Solubilization of the enzyme from membranes of MU-treated cells and addition of the exogenous phospholipid, cardiolipin, rescued hyaluronan synthase activity. In the mass spectrometric analysis of the membrane phospholipids from FM100 cells treated with MU, changes were observed in the distribution of only cardiolipin species but not of the other major phospholipid, PtdGro. These results suggest that MU treatment may cause a decrease in hyaluronan synthase activity by altering the lipid environment of membranes, especially the distribution of different cardiolipin species, surrounding hyaluronan synthase.     

Studies on the functional site on staphylococcal enterotoxin A responsible for production of murine gamma interferon

To identify the functional region(s) associated with induction of gamma interferon on the staphylococcal enterotoxin A molecule, native staphylococcal enterotoxin A molecules and 12 various synthetic peptides corresponding to different regions of entire staphylococcal enterotoxin A were compared to induce gamma interferon production in murine spleen cells. The native staphylococcal enterotoxin A molecule induced gamma interferon production, whereas all of the 12 synthetic peptides did not. Pre-treatment of the murine spleen cells with synthetic peptide A-9 (corresponding to amino acid residues 161-180) significantly inhibited the staphylococcal enterotoxin A-induced gamma interferon production, whereas those with other synthetic peptides did not. When native staphylococcal enterotoxin A was pre-treated with either anti-staphylococcal enterotoxin A serum or anti-peptide sera, anti-staphylococcal enterotoxin A serum and antisera to peptides A-1 (1-20), A-7 (121-140), A-8 (141-160), A-9 (161-180) and A-10 (181-200) inhibited the staphylococcal enterotoxin A-induced gamma interferon production. From these findings, the amino acid residues 161-180 on the staphylococcal enterotoxin A molecule may be an essential region for murine gamma interferon production. Furthermore, the neutralizing epitopes may be also located on regions of amino acid residues 1-20, 121-140, 141-160 and 181-200 on the staphylococcal enterotoxin A molecule.     

Evidence that 2-arachidonoylglycerol but not N-palmitoylethanolamine or anandamide is the physiological ligand for the cannabinoid CB2 receptor. Comparison of the agonistic activities of various cannabinoid receptor ligands in HL-60 cells

We examined the effect of 2-arachidonoylglycerol, an endogenous cannabinoid receptor ligand, on the intracellular free Ca(2+) concentrations in HL-60 cells that express the cannabinoid CB2 receptor. We found that 2-arachidonoylglycerol induces a rapid transient increase in intracellular free Ca(2+) concentrations in HL-60 cells. The response was affected by neither cyclooxygenase inhibitors nor lipoxygenase inhibitors, suggesting that arachidonic acid metabolites are not involved. Consistent with this notion, free arachidonic acid was devoid of any agonistic activity. Importantly, the Ca(2+) transient induced by 2-arachidonoylglycerol was blocked by pretreatment of the cells with SR144528, a CB2 receptor-specific antagonist, but not with SR141716A, a CB1 receptor-specific antagonist, indicating the involvement of the CB2 receptor but not the CB1 receptor in this cellular response. G(i) or G(o) is also assumed to be involved, because pertussis toxin treatment of the cells abolished the response. We further examined the structure-activity relationship. We found that 2-arachidonoylglycerol is the most potent compound among a number of naturally occurring cannabimimetic molecules. Interestingly, anandamide and N-palmitoylethanolamine, other putative endogenous ligands, were found to be a weak partial agonist and an inactive ligand, respectively. These results strongly suggest that the CB2 receptor is originally a 2-arachidonoylglycerol receptor, and 2-arachidonoylglycerol is the intrinsic natural ligand for the CB2 receptor that is abundant in the immune system.     

Pectic polysaccharides of rice endosperm cell walls

Two pectic polysaccharide fractions were purified from rice endosperm cell walls. Methylation analysis including carboxyl-reduction and also selective     

Comprehensive analysis of classical and newly described staphylococcal superantigenic toxin genes in Staphylococcus aureus isolates

We describe a comprehensive detection system for 18 kinds of classical and newly described staphylococcal superantigenic toxin genes using four sets of multiplex PCR. Superantigenic toxin genotyping of Staphylococcus aureus for 69 food poisoning isolates and 97 healthy human nasal swab isolates revealed 32 superantigenic toxin genotypes and showed that many S. aureus isolates harbored multiple toxin genes. Analysis of the relationship between toxin genotypes and toxin genes encoding profiles of mobile genetic elements suggests its possible role in determining superantigenic toxin genotypes in S. aureus as combinations of toxin gene-encoding mobile genetic elements.     

TH1 and TH2 cytokine inhibition by 3,5-bis(trifluoromethyl)pyrazoles,a novel class of immunomodulators

In order to discover novel immunomodulators for application in treating autoimmune diseases, a stable Jurkat transfectant was constructed in which luciferase reporter gene is driven by a full-length IL-2 promotor. A chemical library was screened to identify compounds that inhibited luciferase expression in Jurkat transfectants stimulated with PMA and ionomycin. A class of compounds (bis-trifluoromethyl pyrazole, BTPs) was identified from this screen. BTPs were shown to inhibit anti-CD3 and anti-CD28 antibody-induced IL-2 secretion, mixed lymphocyte reaction, and Con A-induced T cell proliferation in normal human peripheral blood T cells. In addition, mRNA levels of IL-4, IL-5, IL-9, IL-10, IL-13, IL-15, and IFN-gamma were markedly inhibited by BTPs in peripheral blood mononuclear cells stimulated by Con A as determined by multi-probe RNA protection assay. Furthermore, IL-2, IL-4, IL-5, and IFN-gamma secretion by Hut 78 cells or CD3(+) T cells stimulated with PMA plus ionomycin or anti-CD3 antibody plus PMA were inhibited in a concentration-dependent manner by BTPs. Therefore, BTPs inhibit a wide spectrum of cytokine production including TH1 and TH2 type cytokines. Taken together, these compounds may be useful for treating autoimmune diseases and organ transplant rejection.     

Food habits of fishes on an exposed sandy beach at Fukiagehama,South-West Kyushu Island,Japan

To clarify the feeding habits and major food sources of sandy beach fishes, the gut contents of 55 fish species collected on a sandy beach at Fukiagehama, South-West Kyushu Island, Japan, were examined. Ontogenetic changes in food preference were recognized in nine species (Hypoatherina valenciennei, Lateolabrax japonicus, Trachurus japonicus, Sillago japonica, Sphyraena japonica, Paralichthys olivaceus, Heteromycteris japonica, Paraplagusia japonica, and Takifugu niphobles). A cluster analysis based on dietary overlaps showed that the sandy beach fish assemblage comprised six trophic groups (mysid, amphipod, zooplankton, juvenile fish, terrestrial insect, and mollusk feeders). Of these, the first three groups were the most abundantly represented, whereas the last two were represented by only a single species. These results indicated that epibenthic macrofauna, such as mysids and gammaridean amphipods, and zooplankton, were important food resources for the fish assemblage at the study site, but infaunal macrobenthos, such as polychaetes and bivalves, being relatively unimportant.     

A novel comprehensive analysis method for Staphylococcus aureus pathogenicity islands

Staphylococcus aureus pathogenicity islands (SaPIs) form a growing family of mobile genetic elements (MGEs) in Staphylococci. Horizontal genetic transfer by MGEs plays an important role in the evolution of S. aureus. Several SaPIs carry staphylococcal enterotoxin and SE‐like toxin genes. To comprehensively investigate the diversity of SaPIs, a series of primers corresponding to sequences flanking six SaPI insertion sites in S. aureus genome were designed and a long and accurate (LA)‐PCR analysis method established. LA‐PCR products of 13–17 kbp were observed in strains with seb, selk or selq genes. Restriction fragment length polymorphism (RFLP) analysis showed that the products have different RFLP characteristics than do previously described SaPIs; they were therefore predicted to include new SaPIs. Nucleotide sequencing analysis revealed seven novel SaPIs: seb‐harboring SaPIivm10, SaPishikawa11, SaPIivm60, SaPIno10 and SaPIhirosaki4, selk and selq‐harboring SaPIj11 and non‐superantigen‐harboring SaPIhhms2. These SaPIs have mosaic structures containing components of known SaPIs and other unknown genes. Strains carrying different SaPIs were found to have significantly different production of superantigen toxins. The present results show that the LA‐PCR approach can comprehensively identify SaPI diversity and is useful for investigating the evolution of S. aureus pathogenicity.