Activation of Notch1 promotes development of human CD8 single positive T cells in humanized mice

Notch1 mutations are found in more than 50% of human T cell acute lymphoblastic leukemia (T-ALL) cells. However, the functions of Notch1 for human T cell development and leukemogenesis are not well understood. To examine the role of Notch1, human hematopoietic stem cells (HSCs), which had been transduced with a constitutively active form of Notch1 (ICN1), were transplanted into severely immunodeficient NOD/Shi-scid-IL2rγ^null (NOG) mice. We found that the great majority of the ICN1-expressing hematopoietic cells in the bone marrow expressed surface markers for T cells, such as CD3, CD4, and CD8, and that this T cell development was independent of the thymus. Accordingly, phenotypically mature CD8⁺ single positive (SP) T cells were observed in the spleen. Furthermore, T-ALL developed in one NOG recipient mouse out of 26 that had been secondary transferred with the T cells developed in the first NOG mice. These results indicate that Notch1 signaling in HSCs promotes CD8⁺ SP T cell development, and that T cell leukemogenesis may require additional oncogenic factors other than Notch1 activation.     

Laminin receptor mediates anti-inflammatory and anti-thrombogenic effects of pigment epithelium-derived factor in myeloma cells

Pigment epithelium-derived factor (PEDF) has anti-inflammatory and anti-thrombogenic properties both in cell culture and animal models. Although adipose triglyceride lipase (ATGL) and laminin receptor (LR) are two putative receptors for PEDF, which receptor mainly mediates the beneficial effects of PEDF is largely unknown. In this study, we addressed the issue. siRNA raised against LR (siLR) and siATGL transfection dramatically decreased LR and ATGL levels in human cultured myeloma cells, respectively. Ten nM PEDF significantly reduced vascular endothelial growth factor (VEGF), monocyte chemoattractant protein-1 (MCP-1), intercellular cell adhesion molecule-1 (ICAM-1) and plasminogen activator inhibitor-1 (PAI-1) mRNA levels in siCon- or siATGL-transfected myeloma cells, whereas PEDF increased rather than decreased these gene expressions in siLR-transfected cells. Neutralizing antibody directed against LR (LR-Ab) or LR antagonist actually bound to LR and reduced mRNA levels of VEGF, MCP-1, ICAM-1 and PAI-1 in myeloma cells. Further, pre-treatment of LR-Ab or LR antagonist suppressed the binding of PEDF to LR and resultantly blocked the effects of PEDF in myeloma cells. In addition, high concentration of LR agonist mimicked the actions of PEDF on these gene expressions in myeloma cells. This study indicates that PEDF causes anti-angiogenic, anti-inflammatory and anti-thrombogenic reactions in myeloma cells through the interaction with LR. Target domain of LR agonist and antagonist might be involved in the PEDF-signaling to gene suppression in myeloma cells.     

Genetic structure and putative selective sweep in the pioneer tree,Zanthoxylum ailanthoides

Zanthoxylum ailanthoides Siebold & Zucc. is one of the most frequently encountered pioneer trees in Japanese warm–temperate evergreen oak forests. Our previous study in one region of Japan suggested high levels of population differentiation and putative natural selection acting on one of the nuclear loci analyzed. Here, we extend our analysis to study the genetic structure of 10 populations of Z. ailanthoides across Japan using 9 simple sequence repeat (SSR) loci for a better understanding of its genetic structure. First, the southernmost population (Kagoshima) in the samples was found to have the highest genetic diversity, suggesting there was a glacial refugium at or near the location of the population. Second, relatively strong genetic differentiation was found among populations, and there was a positive correlation between genetic distances and geographic distances (Mantel test; P < 0.001). Based on this information, we analyzed nucleotide variation at the putatively selected locus homologous to the gene encoding the ADP-glucose pyrophosphorylase large subunit (agpL). Despite the strong genetic differentiation among populations suggested by the SSR loci, the agpL locus was monomorphic in almost all populations analyzed. The results of this study strongly supported the possibility of a selective sweep at or near the agpL locus.     

Discovery of imidazo[1,2-b]pyridazine derivatives as IKKβ inhibitors. Part 1: Hit-to-lead study and structure–activity relationship

Imidazo[1,2-b]pyridazine derivatives from high-throughput screening were developed as IKKβ inhibitors. By the optimization of the 3- and 6-position of imidazo[1,2-b]pyridazine scaffold, cell-free IKKβ inhibitory activity and TNFα inhibitory activity in THP-1 cell increased. Also, these compounds showed high kinase selectivity. The structure–activity relationship was revealed and the interaction model of imidazo[1,2-b]pyridazine compounds with IKKβ was constructed.     

Efficient synthesis of 6-<Emphasis Type="Italic">O</Emphasis>-palmitoyl-1,2-<Emphasis Type="Italic">O</Emphasis>-isopropylidene-α-<Emphasis Type="SmallCaps">d</Emphasis>-glucofuranose in an organic solvent system by lipase-catalyzed esterification

In order to synthesize a sugar ester at high concentration, 1,2-O-isopropylidene-α-d-glucofuranose (IpGlc), which is one of the sugar acetals and is more hydrophobic than unmodified glucose, was esterified with palmitic acid at 40°C using immobilized lipase from Candida antarctica in some organic solvents or their mixtures. Acetone + t-butyl alcohol (3:1 v/v) improved both the initial reaction rate and yield after 80 h: the product reached its maximum value (240 mmol/kg solvent; ca. 110 g/kg solvent) when 400 mmol IpGlc/kg solvent and 1,200 mmol palmitic acid/kg solvent were used in this solvent mixture.     

Nifedipine, a calcium channel blocker, inhibits inflammatory and fibrogenic gene expressions in advanced glycation end product (AGE)-exposed fibroblasts via mineralocorticoid receptor antagonistic activity

Advanced glycation end products (AGE) are involved in tissue damage and remodeling. This study investigated whether AGE could elicit inflammatory and fibrogenic reactions in fibroblast cell line MRC-5 cells via autocrine production of aldosterone and if nifedipine could block the AGE actions through mineralocorticoid receptor (MR) antagonistic activity. AGE significantly up-regulated monocyte chemoattractant protein-1 (MCP-1), transforming growth factor-β (TGF-β), type III collagen and receptor for AGE (RAGE) mRNA levels in MRC-5 cells, all of which were completely blocked by nifedipine or an MR antagonist spironolactone. Aldosterone also dose-dependently increased MCP-1, TGF-β and type III collagen mRNA levels in MRC-5 cells, which were suppressed by nifedipine, but not amlodipine, a control calcium channel blocker. Further, AGE significantly stimulated aldosterone generation in MRC-5 cells, which was partially blocked by nifedipine or spironolactone. In this study, we demonstrated for the first time that AGE could evoke inflammatory and fibrogenic reactions in MRC-5 cells via aldosterone production, which were blocked by the MR antagonistic activity of nifedipine. Our present study provides a unique beneficial aspect of nifedipine on tissue damage and remodeling; it could work as an anti-inflammatory and anti-fibrogenic agent against AGE via MR antagonistic activity.     

Nifedipine inhibits advanced glycation end products (AGEs) and their receptor (RAGE) interaction-mediated proximal tubular cell injury via peroxisome proliferator-activated receptor-gamma activation

There is a growing body of evidence that advanced glycation end products (AGEs) and their receptor (RAGE) interaction evokes oxidative stress generation and subsequently elicits inflammatory and fibrogenic reactions, thereby contributing to the development and progression of diabetic nephropathy. We have previously found that nifedipine, a calcium-channel blocker (CCB), inhibits the AGE-induced mesangial cell damage in vitro. However, effects of nifedipine on proximal tubular cell injury remain unknown. We examined here whether and how nifedipine blocked the AGE-induced tubular cell damage. Nifedipine, but not amlodipine, a control CCB, inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by the simultaneous treatment of GW9662, an inhibitor of peroxisome proliferator-activated receptor-γ (PPARγ). GW9662 treatment alone was found to increase RAGE mRNA levels in tubular cells. Further, nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-κB activation and increases in intercellular adhesion molecule-1 and transforming growth factor-beta gene expression in tubular cells, all of which were blocked by GW9662. Our present study provides a unique beneficial aspect of nifedipine on diabetic nephropathy; it could work as an anti-oxidative and anti-inflammatory agent against AGEs in tubular cells by suppressing RAGE expression via PPARγ activation.     

Clonal and spatial genetic structure within populations of a coastal plant, Carex kobomugi (Cyperaceae)

Clarification of clonal growth pattern is critical for understanding the population dynamics and reproductive system evolution of clonal plant species. The contribution of clonality to the spatial genetic structure (SGS) within populations is also an important issue. I examined the spatial distribution of genetic variability within two populations of the coastal plant Carex kobomugi using seven microsatellite loci. Genotyping of 226 and 140 ramets within 14 × 40 m and 14 × 34 m plots on two populations revealed 36 and 33 multilocus genotypes, respectively. To quantify the extent of intermingling among clones, for each genet, I calculated the dominance of ramets belonging to a particular genet within a spatial range of the genet. Furthermore, I analyzed spatial distribution of genotypes within 2 × 2 m and 1 × 2 m quadrats using second-order spatial statistics. These analyses indicated that clones are highly intermingled, suggesting a low level of spatial interaction among clones. Spatial autocorrelation analysis of kinship coefficient including all pairs of ramets showed significantly stronger SGS than analysis considering only pairs between different genets. I conclude that clonal propagation largely contributes to SGS at a fine scale.