Dopamine D1-like receptor antagonist attenuates Th17-mediated immune response and ovalbumin antigen-induced neutrophilic airway inflammation

Allergic airway inflammation is generally considered a Th2-type immune response. Recent studies, however, demonstrated that Th17-type immune responses also play important roles in this process, especially in the pathogenesis of neutrophilic airway inflammation, a hallmark of severe asthma. We previously reported that dendritic cells release dopamine to naive CD4(+) T cells in Ag-specific cell-cell interaction, in turn inducing Th17 differentiation through dopamine D1-like receptor (D1-like-R). D1-like-R antagonist attenuates Th17-mediated diseases such as experimental autoimmune encephalomyelitis and autoimmune diabetes. However, the effect of antagonizing D1-like-R on Th17-mediated airway inflammation has yet to be studied. In this study, we examined whether D1-like-R antagonist suppresses OVA-induced neutrophilic airway inflammation in OVA TCR-transgenic DO11.10 mice and then elucidated the mechanism of action. DO11.10 mice were nebulized with OVA or PBS, and some mice received D1-like-R antagonist orally before OVA nebulization. D1-like-R antagonist significantly suppressed OVA-induced neutrophilic airway inflammation in DO11.10 mice. It also inhibited the production of IL-17 and infiltration of Th17 cells in the lung. Further, D1-like-R antagonist suppressed the production of IL-23 by lung CD11c(+) APCs. In contrast, D1-like-R antagonist did not increase Foxp3(+) regulatory T cells in the lung. D1-like-R antagonist neither suppressed nonspecific LPS-induced neutrophilic airway inflammation nor OVA-induced eosinophilic airway inflammation. These results indicate that D1-like-R antagonist could suppress Th17-mediated neutrophilic airway inflammation, raising the possibility that antagonizing D1-like-R serves as a promising new strategy for treating neutrophil-dominant severe asthma.     

2-aminomethylphenylamine as a novel scaffold for factor Xa inhibitor

We have been researching orally active factor Xa inhibitor for a long time. We explored the new diamine linker using effective ligands to obtain a new attractive original scaffold 2-aminomethylphenylamine derivative. Compound 1D showed very strong in vitro and in vivo factor Xa inhibitory activity, as well as favorable PK profiles in po administration to monkeys.     

Inhibitory effect of chitinases isolated from Semillon grapes (Vitis vinifera) on growth of grapevine pathogens

Characteristics and antifungal activity of chitinases in Semillon grapes were investigated. Chitinases were isolated from the juice of Semillon grapes by chitin affinity chromatography. Native and SDS-PAGE analyses of the fraction showing chitin affinity (active fraction) demonstrated only the presence of protein bands of chitinases. Three types of class IV chitinases (chi-1a, chi-1b and chi-2) were purified from the active fraction. These chitinases actively hydrolyzed chitin under acidic conditions (pH 4.0–4.5). The isoelectric points and the molecular weights of chi-1a, chi-1b and chi-2 were 4.73, 4.60, and 7.87, and 32.1 kDa, 31.6 kDa, and 29.0 kDa, respectively. The active fraction was found to inhibit Botrytis cinerea mycelial growth and the inhibitory effect was due to the activity of chitinases. The active fraction inhibited twenty strains of B. cinerea collected from the experimental vineyard. The effect of chitinases was enhanced in media containing more than 20% sugar. When the active fraction was tested on Glomerella cingulata, the growth inhibitory effect observed was markedly less than that seen on B. cinerea.     

Perenniporiol derivatives six triterpenoids from the cultured mycelia of Perenniporia ochroleuca

Six new triterpenoids, (26S)-26-O-methylperenniporiol, (26S)-3-acetyl-26-O-methylperenniporiol, perenniporiol, 3-acetylperenniporiol, (26S-15-deacetoxy-7,11-dihydro-26-O-methylperenniporiol and 15-deacetoxy-7,11-dihydroperenniporiol were isolated from the cultured mycelia of Perenniporia ochroleuca. The structures of the first two compounds were determined using spectroscopic and X-ray analyses, and the structures of the other compounds were elucidated by spectroscopic data.     

Catalytically inactive cyclooxygenase 2 and absence of prostaglandin E2 biosynthesis in murine peritoneal macrophages following in vivo phagocytosis of heat-killed Mycobacterium bovis bacillus Calmette-Guérin

Over 25 years ago, it was observed that peritoneal macrophages (Mphi) isolated from mice given heat-killed Mycobacterium bovis bacillus Calmette-Guérin (HK-BCG) i.p. did not release PGE(2). However, when peritoneal Mphi from untreated mice are treated with HK-BCG in vitro, cyclooxygenase 2 (COX-2), a rate-limiting enzyme for PGE(2) biosynthesis, is expressed and the release of PGE(2) is increased. The present study of peritoneal Mphi obtained from C57BL/6 mice and treated either in vitro or in vivo with HK-BCG was undertaken to further characterize the cellular responses that result in suppression of PGE(2) release. The results indicate that Mphi treated with HK-BCG in vivo express constitutive COX-1 and inducible COX-2 that are catalytically inactive, are localized subcellularly in the cytoplasm, and are not associated with the nuclear envelope (NE). In contrast, Mphi treated in vitro express catalytically active COX-1 and COX-2 that are localized in the NE and diffusely in the cytoplasm. Thus, for local Mphi activated in vivo by HK-BCG, the results indicate that COX-1 and COX-2 dissociated from the NE are catalytically inactive, which accounts for the lack of PGE(2) production by local Mphi activated in vivo with HK-BCG. Our studies further indicate that the formation of catalytically inactive COX-2 is associated with in vivo phagocytosis of HK-BCG, and is not dependent on extracellular mediators produced by in vivo HK-BCG treatment. This attenuation of PGE(2) production may enhance Mphi-mediated innate and Th1-acquired immune responses against intracellular infections which are suppressed by PGE(2).     

Theanine, r-glutamylethylamide, increases neurotransmission concentrations and neurotrophin mRNA levels in the brain during lactation

Theanine (r-glutamylethylamide) is one of the major amino acid components in green tea. Recent studies suggest that theanine affects neurotransmission, especially inhibitory neurotransmission. In this study, we investigated whether theanine affects brain development in infant rats, because inhibitory neurotransmission is required for mature brain function. Mother rats were fed theanine ad libitum after confinement. The body weight gain rate of infants was not different from control infants. We detected theanine in the infant serum and measured neurotransmitter concentration and nerve growth factor (NGF) mRNA level in the infant rat brain. Some neurotransmitters, including dopamine, serotonin, glycine and GABA concentration, increased in the infant brain and NGF mRNA level increased in the cerebral cortex and hippocampus. However, these differences were lost by the end of nerve maturity. These results suggest that theanine enhanced synthesis of nerve growth factor and neurotransmitters during a nerve maturing period and promoted central nerve system maturation (CNS). Thus, theanine accelerated maturation. In conclusion, theanine may assist in healthy brain function development.     

Involvement of up-regulated Necl-5/Tage4/PVR/CD155 in the loss of contact inhibition in transformed NIH3T3 cells

Normal cells show contact inhibition of cell movement and proliferation, but this is lost following transformation. We found that Necl-5, originally identified as a poliovirus receptor and up-regulated in many cancer cells, enhances growth factor-induced cell movement and proliferation. We showed that when cells contact other cells, Necl-5 interacts in trans with nectin-3 and is removed by endocytosis from the cell surface, resulting in a reduction of cell movement and proliferation. We show here that up-regulation of the gene encoding Necl-5 by the oncogene V12-Ki-Ras causes enhanced cell movement and proliferation. Upon cell-cell contact, de novo synthesis of Necl-5 exceeds the rate of Necl-5 endocytosis, eventually resulting in a net increase in the amount of Necl-5 at the cell surface. In addition, expression of the gene encoding nectin-3 is markedly reduced in transformed cells. Thus, up-regulation of Necl-5 following transformation contributes to the loss of contact inhibition in transformed cells.     

Proteomic study on X-irradiation-responsive proteins and ageing: search for responsible proteins for radiation adaptive response

We investigated high- or low-dose irradiation-responsive proteins using proteomics on two-dimensional (2D) PAGE, and the effects of ageing on cell responses to radiation in variously aged rat astrocytes. After 5 Gy irradiation, the relative abundance of peroxiredoxin 2, an antioxidant enzyme, and latexin, an inhibitor of carboxypeptidase, increased. The induction of these proteins was suppressed by ageing, suggesting that the response to high-dose radiation decreased with ageing. The relative abundance of elongation factor 2 (EF-2) fragment increased 3 h and reduced 24 h after 0.1 Gy irradiation. Temporal enhancement of the EF-2 fragment due to low-dose irradiation was suppressed by ageing. Since radiation adaptive response in cultured astrocytes was observed 3 h but not 24 h after 0.1 Gy irradiation and suppressed by ageing as previously reported, alteration of the EF-2 fragment corresponded to the radiation adaptive response. We also examined phospho-protein profiles, resulting in the relative abundance of phospho-EF-1beta and phospho-beta-actin being altered by 0.1 Gy irradiation; however, ageing did not affect the alteration of phospho-EF-1beta and phospho-beta-actin, unlike the EF-2 fragment. The results suggested that the EF-2 fragment was a possible candidate for the protein responsible for the radiation adaptive response in cultured astrocytes.