Discovery of 4-[4-({(3R)-1-butyl-3-[(R)-cyclohexyl(hydroxy)methyl]-2,5-dioxo-1,4,9-triazaspiro[5.5]undec-9-yl}methyl)phenoxy]benzoic acid hydrochloride: a highly potent orally available CCR5 selective antagonist

Based on the original spirodiketopiperazine design framework, further optimization of an orally available CCR5 antagonist was undertaken. Structural hybridization of the hydroxylated analog 4 derived from one of the oxidative metabolites and the new orally available non-hydroxylated benzoic acid analog 5 resulted in another potent orally available CCR5 antagonist 6a as a clinical candidate. Full details of a structure-activity relationship (SAR) study and ADME properties are presented.     

Lack of association between orexin receptor gene polymorphisms and obstructive sleep apnea syndrome in Japanese

Sleep and Biological Rhythms - The pathogenesis of obstructive sleep apnea (OSA) is not fully understood, particularly at the genetic level. The orexins have pleiotropic effects on several aspects...     

Inhibitory effect of porphyran, prepared from dried "Nori", on contact hypersensitivity in mice

Porphyran is a major component of the red algae, Porphyra tenera and P. yezoensis, which are processed into a sheet type of dried food, "Nori". Porphyran has been reported to activate murine macrophages by in vitro and i.p. injection studies. The contact hypersensitivity (CHS) reaction in mice is commonly used as a model to evaluate the anti-allergic activity of food and food components. We therefore studied the effect of porphyran on the CHS reaction in Balb/c mice to evaluate anti-allergic activity of porphyran. We found that an oral administration of porphyran (2% in drinking water) suppressed the CHS reaction (ear edema) induced by 2,4,6-trinitrochlorobenzene. We also found that porphyran suppressed the serum level of IgE and the production of interferon-gamma (IFN-gamma) in the challenged ear lobe. We conclude from these results that the CHS reaction was suppressed by oral porphyran due to the decreased serum level of IgE and the production of IFN-gamma in the challenged ear lobe.     

Arum- and Paris-type arbuscular mycorrhizas in a mixed pine forest on sand dune soil in Niigata Prefecture, central Honshu, Japan

Arbuscular mycorrhizas (AM) are the most widespread mycorrhiza in nature and form two morphologies, Arum- and Paris-type. The determining factors defining the two different morphologies are not well understood. In this study, the distribution of Arum- and Paris-type AM was determined in a mixed pine forest. A total of 35 plant species belonging to 20 families and 32 genera were identified and examined for AM colonization and morphological types. AM morphological types in 14 families were confirmed as follows: Arum-type in Rosaceae, Oleaceae, Lauraceae, Vitaceae and Compositae, Paris-type in Aquifoliaceae, Ulmaceae, Araliaceae, Theaceae, Magnoliaceae, Rubiaceae and Dioscoraceae, and both and/or intermediate types in Caprifoliaceae and Gramineae. Plant families whose AM morphological status was previously unknown were clarified as follows: Polygonaceae and Commelinaceae showed Arum-type morphology; Celastraceae, Menispermaceae and Elaeagnaceae had typical Paris-type morphology. The proportion of Arum-type to Paris-type species decreased in the following order: annuals > perennials > deciduous species > evergreen species, and pioneer group > early successional group > late successional group. Evergreen plants had a higher tendency to form Paris-type AM than annuals, perennials and deciduous plants. The results indicate that environmental changes, such as shade during plant succession, control the distribution of plant growth forms in mixed pine forest and may also play a part in the distribution of Arum- and Paris-type morphology. The identity of the plant seems to strongly influence AM morphology, though control by the fungal genome cannot be ruled out.     

Transgenic rats overexpressing the human MrgX3 gene show cataracts and an abnormal skin phenotype

The human MrgX3 gene, belonging to the mrgs/SNSRs (mas related genes/sensory neuron specific receptors) family, was overexpressed in transgenic rats using the actin promoter. Two animal lines showed cataracts with liquification/degeneration and swelling of the lens fiber cells. The transient epidermal desquamation was observed in line with higher gene expression. Histopathology of the transgenic rats showed acanthosis and focal parakeratosis. In the epidermis, there was an increase in cellular keratin 14, keratin 10, and loricrin, as well as PGP 9.5 in innervating nerve fibers. These phenotypes accompanied an increase in the number of proliferating cells. These results suggest that overexpression of the human MrgX3 gene causes a disturbance of the normal cell-differentiation process.     

The KIF3 motor transports N-cadherin and organizes the developing neuroepithelium

In the developing brain, the organization of the neuroepithelium is maintained by a critical balance between proliferation and cell-cell adhesion of neural progenitor cells. The molecular mechanisms that underlie this are still largely unknown. Here, through analysis of a conditional knockout mouse for the Kap3 gene, we show that post-Golgi transport of N-cadherin by the KIF3 molecular motor complex is crucial for maintaining this balance. N-cadherin and beta-catenin associate with the KIF3 complex by co-immunoprecipitation, and colocalize with KIF3 in cells. Furthermore, in KAP3-deficient cells, the subcellular localization of N-cadherin was disrupted. Taken together, these results suggest a potential tumour-suppressing activity for this molecular motor.     

Characterization of histopathology and gene-expression profiles of synovitis in early rheumatoid arthritis using targeted biopsy specimens

The disease category of early rheumatoid arthritis (RA) has been limited with respect to clinical criteria. Pathological manifestations of synovitis in patients whose disease is clinically classified as early RA seem to be heterogeneous, with regular variations. To clarify the relation between the molecular and histopathological features of the synovitis, we analyzed gene-expression profiles in the synovial lining tissues to correlate them with histopathological features. Synovial tissues were obtained from knee joints of 12 patients with early RA by targeted biopsy under arthroscopy. Surgical specimens of long-standing RA (from four patients) were examined as positive controls. Each histopathological parameter characteristic of rheumatoid synovitis in synovial tissues was scored under light microscopy. Total RNAs from synovial lining tissues were obtained from the specimens selected by laser capture microdissection and the mRNAs were amplified by bacteriophage T7 RNA polymerase. Their cDNAs were analyzed in a cDNA microarray with 23,040 cDNAs, and the levels of gene expression in multilayered lining tissues, compared with those of normal-like lining tissues in specimens from the same person, were determined to estimate gene-expression profiles characteristic of the synovial proliferative lesions in each case. Based on cluster analysis of all cases, gene-expression profiles in the lesions in early RA fell into two groups. The groups had different expression levels of genes critical for proliferative inflammation, including those encoding cytokines, adhesion molecules, and extracellular matrices. One group resembled synovitis in long-standing RA and had high scores for some histopathological features – involving accumulations of lymphocytes and plasma cells – but not for other features. Possible differences in the histopathogenesis and prognosis of synovitis between the two groups are discussed in relation to the candidate genes and histopathology.     

Combined effects of ATP on the therapeutic efficacy of antimicrobial drug regimens against Mycobacterium avium complex infection in mice and roles of cytosolic phospholipase A2-dependent mechanisms in the ATP-mediated potentiation of antimycobacterial host resistance

ATP, which serves as a mediator of intramacrophage signaling pathways through purinoceptors, is known to potentiate macrophage antimycobacterial activity. In this study we examined the effects of ATP in potentiating host resistance to Mycobacterium avium complex (MAC) infection in mice undergoing treatment with a drug regimen using clarithromycin and rifamycin and obtained the following findings. First, the administration of ATP in combination with the clarithromycin and rifamycin regimen accelerated bacterial elimination in MAC-infected mice without causing changes in the histopathological features or the mRNA expression of pro- or anti-inflammatory cytokines from those in the mice not given ATP. Second, ATP potentiated the anti-MAC bactericidal activity of macrophages cultivated in the presence of clarithromycin and rifamycin. This effect of ATP was closely related to intracellular Ca2+ mobilization and was specifically blocked by a cytosolic phospholipase A2 (cPLA2) inhibitor, arachidonyl trifluoromethylketone. Third, intramacrophage translocation of membranous arachidonic acid molecules to MAC-containing phagosomes was also specifically blocked by arachidonyl trifluoromethylketone. In the confocal microscopic observation of MAC-infected macrophages, ATP enhanced the intracellular translocation of cPLA2 into MAC-containing phagosomes. These findings suggest that ATP increases the host anti-MAC resistance by potentiating the antimycobacterial activity of host macrophages and that the cPLA2-dependent generation of arachidonic acid from the phagosomal membrane is essential for such a phenomenon.