Seven sesquiterpene lactones from Inula britannica var. chinensis

Four known sesquiterpene lactones, tomentosin, ivalin, 4-epi-isoinuviscolide and gaillardin, together with three new lactones, inuchinenolides A, B and C, were identified in the whole plant of Inula britannica var. chinensis.     

Carbon cycling and sequestration in a Japanese red pine (Pinus densiflora) forest on lava flow of Mt. Fuji

Biometric-based carbon flux measurements were conducted in a pine forest on lava flow of Mt. Fuji, Japan, in order to estimate carbon cycling and sequestration. The forest consists mainly of Japanese red pine (Pinus densiflora) in a canopy layer and Japanese holly (Ilex pedunculosa) in a subtree layer. The lava remains exposed on the ground surface, and the soil on the lava flow is still immature with no mineral soil layer. The results showed that the net primary production (NPP) of the forest was 7.3 ± 0.7 t C ha^?1 year^?1, of which 1.4 ± 0.4 t C ha^?1 year^?1 was partitioned to biomass increment, 3.2 ± 0.5 t C ha^?1 year^?1 to above-ground fine litter production, 1.9 t C ha^?1 year^?1 to fine root production, and 0.8 ± 0.2 t C ha^?1 year^?1 to coarse woody debris. The total amount of annual soil surface CO₂ efflux was estimated as 6.1 ± 2.9 t C ha^?1 year^?1, using a closed chamber method. The estimated decomposition rate of soil organic matter, which subtracted annual root respiration from soil respiration, was 4.2 ± 3.1 t C ha^?1 year^?1. Biometric-based net ecosystem production (NEP) in the pine forest was estimated at 2.9 ± 3.2 t C ha^?1 year^?1, with high uncertainty due mainly to the model estimation error of annual soil respiration and root respiration. The sequestered carbon being allocated in roughly equal amounts to living biomass (1.4 t C ha^?1 year^?1) and the non-living C pool (1.5 t C ha^?1 year^?1). Our estimate of biometric-based NEP was 25 % lower than the eddy covariance-based NEP in this pine forest, due partly to the underestimation of NPP and difficulty of estimation of soil and root respiration in the pine forest on lava flows that have large heterogeneity of soil depth. However, our results indicate that the mature pine forest acted as a significant carbon sink even when established on lava flow with low nutrient content in immature soils, and that sequestration strength, both in biomass and in soil organic matter, is large.     

Tumor cells enhance their own CD44 cleavage and motility by generating hyaluronan fragments

Hyaluronan (HA) is an extracellular matrix glycosaminoglycan that interacts with cell-surface receptors, including CD44. Although HA usually exists as a high molecular mass polymer, HA of a much lower molecular mass that shows a variety of biological activities can be detected under certain pathological conditions, particularly in tumors. We previously reported that low molecular weight HAs (LMW-HAs) of a certain size range induce the proteolytic cleavage of CD44 from the surface of tumor cells and promote tumor cell migration in a CD44-dependent manner. Here, we show that MIA PaCa-2, a human pancreatic carcinoma cell line, secreted hyaluronidases abundantly and generated readily detectable levels of LMW-HAs ranging from approximately 10- to 40-mers. This occurred in the absence of any exogenous stimulation. The tumor-derived HA oligosaccharides were able to enhance CD44 cleavage and tumor cell motility. Inhibition of the CD44-HA interaction resulted in the complete abrogation of these cellular events. These results are consistent with the concept that tumor cells generate HA oligosaccha-rides that bind to tumor cell CD44 through the expression of their own constitutive hyaluronidases. This enhances their own CD44 cleavage and cell motility, which would subsequently promote tumor progression. Such an autocrine/paracrine-like process may represent a novel activation mechanism that would facilitate and promote the malignant potential of tumor cells.     

Brain-Derived Neurotrophic Factor Stimulates Interactions of Shp2 with Phosphatidylinositol 3-Kinase and Grb2 in Cultured Cerebral Cortical Neurons

Shp2, a protein tyrosine phosphatase possessing SH2 domains, is utilized in the intracellular signaling of various growth factors. Shp2 is highly expressed in the CNS. Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, which also shows high levels of expression in the CNS, exerts neurotrophic and neuromodulatory effects in CNS neurons. We examined how BDNF utilizes Shp2 in its signaling pathway in cultured cerebral cortical neurons. We found that BDNF stimulated coprecipitation of several tyrosine-phosphorylated proteins with anti-Shp2 antibody and that Grb2 and phosphatidylinositol 3-kinase (PI3-K) were coprecipitated with anti-Shp2 antibody in response to BDNF. In addition, both anti-Grb2 and anti-PI3-K antibodies coprecipitated Shp2 in response to BDNF. The BDNF-stimulated coprecipitation of the tyrosine-phosphorylated proteins, Grb2, and PI3-K with anti-Shp2 antibody was completely inhibited by K252a, an inhibitor of TrkB receptor tyrosine kinase. This BDNF-stimulated Shp2 signaling was markedly sustained as well as BDNF-induced phosphorylation of TrkB and mitogen-activated protein kinases. In PC12 cells stably expressing TrkB, both BDNF and nerve growth factor stimulated Shp2 signaling similarly to that by BDNF in cultured cortical neurons. These results indicated that Shp2 shows cross-talk with various signaling molecules including Grb2 and PI3-K in BDNF-induced signaling and that Shp2 may be involved in the regulation of various actions of BDNF in CNS neurons.     

Structural aspects of nucleic acid-sensing Toll-like receptors

Invading pathogens elicit potent immune responses in cells through interactions between structurally conserved molecules derived from the pathogens and specialized innate immune receptors such as the Toll-like receptors (TLRs). Nucleic acid is one of the principal TLR ligands. Nucleic acid-sensing TLRs recognize an array of nucleic acids, including double-stranded RNA, single-stranded RNA, and DNAs with specific sequence motifs. Although ligand-induced dimerization is commonly observed followed by TLR activation, both the specific recognition mechanisms and the ligand–receptor interactions vary among different TLRs. In this review, we highlight our current understanding of how these receptors recognize their cognate ligands based on the recent advances in structural biology.     

Inhibitory CD161 receptor identified in glioma-infiltrating T cells by single-cell analysis

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Mutations affecting early distribution of primordial germ cells in Medaka (Oryzias latipes) embryo

The development of germ cells has been intensively studied in Medaka (Oryzias latipes). We have undertaken a large-scale screen to identify mutations affecting the development of primordial germ cells (PGCs) in Medaka. Embryos derived from mutagenized founder fish were screened for an abnormal distribution or number of PGCs at embryonic stage 27 by RNA in situ hybridization for the Medaka vasa homologue (olvas). At this stage, PGCs coalesce into two bilateral vasa-expressing foci in the ventrolateral regions of the trunk after their migration and group organization. Nineteen mutations were identified from a screen corresponding to 450 mutagenized haploid genomes. Eleven of the mutations caused altered PGC distribution. Most of these alterations were associated with morphological abnormalities and could be grouped into four phenotypic classes: Class 1, PGCs dispersed into bilateral lines; Class 2, PGCs dispersed in a region more medial than that in Class 1; Class 3, PGCs scattered laterally and over the yolk sac area; and Class 4, PGCs clustered in a single median focus. Eight mutations caused a decrease in the number of PGCs. This decrease was observed in the offspring of heterozygous mothers, indicating the contribution of a maternal factor in determining PGC abundance. Taken together, these mutations should prove useful in identifying molecular mechanisms underlying the early PGC development and migration.     

Pyridoxine 5'-phosphate oxidase is a candidate gene responsible for hypertension in Dahl-S rats

To identify candidate genes responsible for hypertension in Dahl salt-sensitive rats (Dahl-S), an oligonucleotide microarray analysis was performed to find differentially expressed genes in kidneys of Dahl-S and Lewis rats. We obtained 101 F2 male rats from Dahl-S and Lewis rats and performed precise measurements of blood pressure (BP) and heart rate by telemetric monitoring at 14 weeks of age after 9 weeks of salt-loading. The correlation analysis between genotypes of differentially expressed genes and BP in F2 rats indicated that pyridoxine 5'-phosphate oxidase (Pnpo) and catecholamine-O-methyltransferease (Comt) showed a highly significant association with BP. However, in the case of Comt, the Dahl-S genotype correlated with low BP. Short/branched chain acyl-CoA dehydrogenase and Sah also showed a significant association with systolic blood pressure. The present study provided evidence that Pnpo is a candidate gene responsible for hypertension in Dahl-S rats.     

Rat NAD+-dependent 3alpha-hydroxysteroid dehydrogenase (AKR1C17): a member of the aldo-keto reductase family highly expressed in kidney cytosol

Mammalian 3α-hydroxysteroid dehydrogenases (3α-HSDs) have been divided into two types: Cytosolic NADP(H)-dependent 3α-HSDs belonging to the aldo-keto reductase family, and mitochondrial and microsomal NAD⁺-dependent 3α-HSDs belonging to the short-chain dehydrogenase/reductase family. In this study, we characterized a rat aldo-keto reductase (AKR1C17), whose functions are unknown. The recombinant AKR1C17 efficiently oxidized 3α-hydroxysteroids and bile acids using NAD⁺ as the preferred coenzyme at an optimal pH of 7.4-9.5, and was inhibited by ketamine and organic anions. The mRNA for AKR1C17 was detected specifically in rat kidney, where the enzyme was more highly expressed as a cytosolic protein than NADP(H)-dependent 3α-HSD (AKR1C9). Thus, AKR1C17 represents a novel NAD⁺-dependent type of cytosolic 3α-HSD with unique inhibitor sensitivity and tissue distribution. In addition, the replacement of Gln270 and Glu276 of AKR1C17 with the corresponding residues of NADP(H)-dependent 3α-HSD resulted in a switch in favor of NADP⁺ specificity, suggesting their key roles in coenzyme specificity.     

Suppression of matrix metalloproteinase production in nasal fibroblasts by tranilast, an antiallergic agent, in vitro

Allergic rhinitis is an inflammatory disease characterized by nasal wall remodeling with intense infiltration of eosinophils and mast cells/basophils. Matrix metalloproteinases (MMPs), MMP-2 and MMP-9, are the major proteolytic enzymes that induce airway remodeling. These enzymes are also important in the migration of inflammatory cells through basement membrane components. We evaluated whether tranilast (TR) could inhibit MMP production from nasal fibroblasts in response to tumor necrosis factor-alpha (TNF-alpha) stimulation in vitro. Nasal fibroblasts (NF) were established from nasal polyp tissues taken from patients with allergic rhinitis. NF (2 x 10(5) cells/mL) were stimulated with TNF-alpha in the presence of various concentrations of TR. After 24 hours, the culture supernatants were obtained and assayed for MMP-2, MMP-9, TIMP-1, and TIMP-2 levels by ELISA. The influence of TR on mRNA expression of MMPs and TIMPs in cells cultured for 12 hours was also evaluated by RT-PCR. TR at more than 5 x 10(-5) M inhibited the production of MMP-2 and MMP-9 from NF in response to TNF-alpha stimulation, whereas TIMP-1 and TIMP-2 production was scarcely affected. TR also inhibited MMP mRNA expression in NF after TNF-alpha stimulation. The present data suggest that the attenuating effect of TR on MMP-2 and MMP-9 production from NF induced by inflammatory stimulation may underlie the therapeutic mode of action of the agent in patients with allergic diseases, including allergic rhinitis.