Prevalence and Genetic Characterization of Pertactin-Deficient Bordetella pertussis in Japan

The adhesin pertactin (Prn) is one of the major virulence factors of Bordetella pertussis, the etiological agent of whooping cough. However, a significant prevalence of Prn-deficient (Prn−) B. pertussis was observed in Japan. The Prn− isolate was first discovered in 1997, and 33 (27%) Prn− isolates were identified among 121 B. pertussis isolates collected from 1990 to 2009. Sequence analysis revealed that all the Prn− isolates harbor exclusively the vaccine-type prn1 allele and that loss of Prn expression is caused by 2 different mutations: an 84-bp deletion of the prn signal sequence (prn1ΔSS, n = 24) and an IS481 insertion in prn1 (prn1::IS481, n = 9). The frequency of Prn− isolates, notably those harboring prn1ΔSS, significantly increased since the early 2000s, and Prn− isolates were subsequently found nationwide. Multilocus variable-number tandem repeat analysis (MLVA) revealed that 24 (73%) of 33 Prn− isolates belong to MLVA-186, and 6 and 3 Prn− isolates belong to MLVA-194 and MLVA-226, respectively. The 3 MLVA types are phylogenetically closely related, suggesting that the 2 Prn− clinical strains (harboring prn1ΔSS and prn1::IS481) have clonally expanded in Japan. Growth competition assays in vitro also demonstrated that Prn− isolates have a higher growth potential than the Prn+ back-mutants from which they were derived. Our observations suggested that human host factors (genetic factors and immune status) that select for Prn− strains have arisen and that Prn expression is not essential for fitness under these conditions

Cartilage-Specific Over-Expression of CCN Family Member 2/Connective Tissue Growth Factor (CCN2/CTGF) Stimulates Insulin-Like Growth Factor Expression and Bone Growth

Previously we showed that CCN family member 2/connective tissue growth factor (CCN2) promotes the proliferation, differentiation, and maturation of growth cartilage cells in vitro. To elucidate the specific role and molecular mechanism of CCN2 in cartilage development in vivo, in the present study we generated transgenic mice overexpressing CCN2 and analyzed them with respect to cartilage and bone development. Transgenic mice were generated expressing a ccn2/lacZ fusion gene in cartilage under the control of the 6 kb-Col2a1-enhancer/promoter. Changes in cartilage and bone development were analyzed histologically and immunohistologically and also by micro CT. Primary chondrocytes as well as limb bud mesenchymal cells were cultured and analyzed for changes in expression of cartilage-related genes, and non-transgenic chondrocytes were treated in culture with recombinant CCN2. Newborn transgenic mice showed extended length of their long bones, increased content of proteoglycans and collagen II accumulation. Micro-CT analysis of transgenic bones indicated increases in bone thickness and mineral density. Chondrocyte proliferation was enhanced in the transgenic cartilage. In in vitro short-term cultures of transgenic chondrocytes, the expression of col2a1, aggrecan and ccn2 genes was substantially enhanced; and in long-term cultures the expression levels of these genes were further enhanced. Also, in vitro chondrogenesis was strongly enhanced. IGF-I and IGF-II mRNA levels were elevated in transgenic chondrocytes, and treatment of non-transgenic chondrocytes with recombinant CCN2 stimulated the expression of these mRNA. The addition of CCN2 to non-transgenic chondrocytes induced the phosphorylation of IGFR, and ccn2-overexpressing chondrocytes showed enhanced phosphorylation of IGFR. Our data indicates that the observed effects of CCN2 may be mediated in part by CCN2-induced overexpression of IGF-I and IGF-II. These findings indicate that CCN2-overexpression in transgenic mice accelerated the endochondral ossification processes, resulting in increased length of their long bones. Our results also indicate the possible involvement of locally enhanced IGF-I or IGF-II in this extended bone growth

CCN Family Member 2/Connective Tissue Growth Factor (CCN2/CTGF) Has Anti-Aging Effects That Protect Articular Cartilage from Age-Related Degenerative Changes

To examine the role of connective tissue growth factor CCN2/CTGF (CCN2) in the maintenance of the articular cartilaginous phenotype, we analyzed knee joints from aging transgenic mice (TG) overexpressing CCN2 driven by the Col2a1 promoter. Knee joints from 3-, 14-, 40-, and 60-day-old and 5-, 12-, 18-, 21-, and 24-month-old littermates were analyzed. Ccn2-LacZ transgene expression in articular cartilage was followed by X-gal staining until 5 months of age. Overexpression of CCN2 protein was confirmed through all ages in TG articular cartilage and in growth plates. Radiographic analysis of knee joints showed a narrowing joint space and other features of osteoarthritis in 50% of WT, but not in any of the TG mice. Transgenic articular cartilage showed enhanced toluidine blue and safranin-O staining as well as chondrocyte proliferation but reduced staining for type X and I collagen and MMP-13 as compared with those parameters for WT cartilage. Staining for aggrecan neoepitope, a marker of aggrecan degradation in WT articular cartilage, increased at 5 and 12 months, but disappeared at 24 months due to loss of cartilage; whereas it was reduced in TG articular cartilage after 12 months. Expression of cartilage genes and MMPs under cyclic tension stress (CTS) was measured by using primary cultures of chondrocytes obtained from wild-type (WT) rib cartilage and TG or WT epiphyseal cartilage. CTS applied to primary cultures of mock-transfected rib chondrocytes from WT cartilage and WT epiphyseal cartilage induced expression of Col1a1, ColXa1, Mmp-13, and Mmp-9 mRNAs; however, their levels were not affected in CCN2-overexpressing chondrocytes and TG epiphyseal cartilage. In conclusion, cartilage-specific overexpression of CCN2 during the developmental and growth periods reduced age-related changes in articular cartilage. Thus CCN2 may play a role as an anti-aging factor by stabilizing articular cartilage

沖縄における縄文時代の木材利用について―木材利用解明の可能性―

publisher奈良「縄文時代の木材利用」とは、縄文時代の人々と森林の密接なつながりを示すものである。それについては、日本各地の遺跡から得られた情報によって日本列島全体に関わる利用形態の動きが解明されつつある。それらは出土遺物のなかで木製品を中心として花粉やプラント・オパールなどの植物遺体の分析結果から、木材を含めた植物の利用形態が浮き彫りにされた。特に鳥浜貝塚や三内丸山遺跡においては顕著で、縄文時代には木材の持つ性質によって意図的に樹種を選択していることや特定の樹種に限って栽培・管理を行ったことなどがわかっている

Gas-Selective Catalytic Regulation by a Newly Identified Globin-Coupled Sensor Phosphodiesterase Containing an HD-GYP Domain from the Human Pathogen <i>Vibrio fluvialis</i>

Globin-coupled sensors constitute an important family of heme-based gas sensors, an emerging class of heme proteins. In this study, we have identified and characterized a globin-coupled sensor phosphodiesterase containing an HD-GYP domain (GCS-HD-GYP) from the human pathogen Vibrio fluvialis, which is an emerging foodborne pathogen of increasing public health concern. The amino acid sequence encoded by the AL536_01530 gene from V. fluvialis indicated the presence of an N-terminal globin domain and a C-terminal HD-GYP domain, with HD-GYP domains shown previously to display phosphodiesterase activity toward bis(3′,5′)-cyclic dimeric guanosine monophosphate (c-di-GMP), a bacterial second messenger that regulates numerous important physiological functions in bacteria, including in bacterial pathogens. Optical absorption spectral properties of GCS-HD-GYP were found to be similar to those of myoglobin and hemoglobin and of other bacterial globin-coupled sensors. The binding of O2 to the Fe(II) heme iron complex of GCS-HD-GYP promoted the catalysis of the hydrolysis of c-di-GMP to its linearized product, 5′-phosphoguanylyl-(3′,5′)-guanosine (pGpG), whereas CO and NO binding did not enhance the catalysis, indicating a strict discrimination of these gaseous ligands. These results shed new light on the molecular mechanism of gas-selective catalytic regulation by globin-coupled sensors, with these advances apt to lead to a better understanding of the family of globin-coupled sensors, a still growing family of heme-based gas sensors. In addition, given the importance of c-di-GMP in infection and virulence, our results suggested that GCS-HD-GYP could play an important role in the ability of V. fluvialis to sense O2 and NO in the context of host–pathogen interactions

Menaquinone-4 Suppresses Lipopolysaccharide-Induced Inflammation in MG6 Mouse Microglia-Derived Cells by Inhibiting the NF-κB Signaling Pathway

The overactivation of microglia is known to trigger inflammatory reactions in the central nervous system, which ultimately induce neuroinflammatory disorders including Alzheimer&#8217;s disease. However, increasing evidence has shown that menaquinone-4 (MK-4), a subtype of vitamin K2, can attenuate inflammation in the peripheral system. Whereas it was also observed at high levels within the brain, its function in this organ has not been well characterized. Therefore, we investigated the effect of MK-4 on microglial activation and clarified the underlying mechanism. Mouse microglia-derived MG6 cells were exposed to lipopolysaccharide (LPS) either with or without MK-4 pretreatment. Cell responses with respect to inflammatory cytokines (Il-1&#946;, Tnf-&#945;, and Il-6) were measured by qRT-PCR. We further analyzed the phosphorylation of TAK1, IKK&#945;/&#946;, and p65 of the NF-&#954;B subunit by Western blotting. We observed that in LPS-induced MG6 cells, MK-4 dose-dependently suppressed the upregulation of inflammatory cytokines at the mRNA level. It also significantly decreased the phosphorylation of p65, but did not affect that TAK1 and IKK&#945;/&#946;. Furthermore, the nuclear translocation of NF-&#954;B in LPS-induced MG6 cells was inhibited by MK-4. These results indicate that MK-4 attenuates microglial inflammation by inhibiting NF-&#954;B signaling

Top-Down Approach toward Building Ubiquitous Sensor Network Applications

Tiny networked sensor devices will be disseminated over our physical life space, and take a significant role in realizing a ubiquitous computing environment. As sensor network technologies and its applications are still at its early stages, the architecture of the sensor network heavily relies on the applications. We believe it quite essential to come up with various kinds of applications, and obtain sufficient knowledge of sensor network architecture and its peripheral technologies through demonstrations. In this paper, we introduce our sensor network applications and present the way sensor network is used in them. 1

Sequence Finishing and Gene Mapping for Candida albicans Chromosome 7 and Syntenic Analysis Against the Saccharomyces cerevisiae Genome

The size of the genome in the opportunistic fungus Candida albicans is 15.6 Mb. Whole-genome shotgun sequencing was carried out at Stanford University where the sequences were assembled into 412 contigs. C. albicans is a diploid basically, and analysis of the sequence is complicated due to repeated sequences and to sequence polymorphism between homologous chromosomes. Chromosome 7 is 1 Mb in size and the best characterized of the 8 chromosomes in C. albicans. We assigned 16 of the contigs, ranging in length from 7309 to 267,590 bp, to chromosome 7 and determined sequences of 16 regions. These regions included four gaps, a misassembled sequence, and two major repeat sequences (MRS) of >16 kb. The length of the continuous sequence attained was 949,626 bp and provided complete coverage of chromosome 7 except for telomeric regions. Sequence analysis was carried out and predicted 404 genes, 11 of which included at least one intron. A 7-kb indel, which might be caused by a retrotransposon, was identified as the largest difference between the homologous chromosomes. Synteny analysis revealed that the degree of synteny between C. albicans and Saccharomyces cerevisiae is too weak to use for completion of the genomic sequence in C. albicans

Fucoxanthin Ameliorates Atopic Dermatitis Symptoms by Regulating Keratinocytes and Regulatory Innate Lymphoid Cells

Fucoxanthin (FX) is a xanthophyll that is contained abundantly in marine plants. The biological action of FX includes its antioxidant and anti-lipogenic activities, while the precise action of its mechanisms on skin cells has not yet been clarified. The current study examined the effect of FX in comparison with tacrolimus (TAC) on NC/Nga mice, which are an atopic dermatitis (AD) model. FX topical treatment dramatically ameliorated itching behavior over the TAC treatment, which was insufficient for improvement of AD symptoms. In Nc/Nga mice, FX or TAC applied to the skin inhibited eosinophil infiltration with decreased expression of Il-33. FX also stimulated Il-2, Il-5, Il-13, Il-10, and TGF-&beta; expression levels, and Sca1+Il-10+TGF-&beta;+ regulatory innate lymphoid cells (ILCreg) were dominantly observed in FX treated skin epidermal keratinocytes and dermal layers. This combined evidence demonstrated that FX exerts anti-inflammatory effects on keratinocytes and ameliorates AD symptoms by regulating ILCreg to normalize immune responses in an atopic dermatitis model