Finite element analysis of tube drawing process with diameter expansion

This paper presents a tube drawing process with diameter expansion for producing a thin-walled tube effectively. In this proposed process, the tube was flared by a plug pushing into the tube, and then the tube was expanded by drawing the plug in the tube axial direction with chucking the flared tube edge. Optimum plug shape, such as the plug half angle and the corner radius, was investigated by a series of analyses using the finite element method (FEM) for improving the forming limit and the dimension accuracy. At first, a friction coefficient was determined to 0.3 by a comparison of the flaring limit between the analysis and the experiment of the tube flaring. As a result of the analyses in the drawing with the diameter expansion, the forming limit was high when the plug half angle was set to 18~30°. The thickness reduction ratio increased with an increase in the expansion ratio and the plug half angle. In addition, the overshoot, which is a difference between the plug diameter and the tube inner diameter after the drawing, was prevented by using the plug with the corner radius of 20 mm

Reaction-Diffusion Pattern in Shoot Apical Meristem of Plants

A fundamental question in developmental biology is how spatial patterns are self-organized from homogeneous structures. In 1952, Turing proposed the reaction-diffusion model in order to explain this issue. Experimental evidence of reaction-diffusion patterns in living organisms was first provided by the pigmentation pattern on the skin of fishes in 1995. However, whether or not this mechanism plays an essential role in developmental events of living organisms remains elusive. Here we show that a reaction-diffusion model can successfully explain the shoot apical meristem (SAM) development of plants. SAM of plants resides in the top of each shoot and consists of a central zone (CZ) and a surrounding peripheral zone (PZ). SAM contains stem cells and continuously produces new organs throughout the lifespan. Molecular genetic studies using Arabidopsis thaliana revealed that the formation and maintenance of the SAM are essentially regulated by the feedback interaction between WUSHCEL (WUS) and CLAVATA (CLV). We developed a mathematical model of the SAM based on a reaction-diffusion dynamics of the WUS-CLV interaction, incorporating cell division and the spatial restriction of the dynamics. Our model explains the various SAM patterns observed in plants, for example, homeostatic control of SAM size in the wild type, enlarged or fasciated SAM in clv mutants, and initiation of ectopic secondary meristems from an initial flattened SAM in wus mutant. In addition, the model is supported by comparing its prediction with the expression pattern of WUS in the wus mutant. Furthermore, the model can account for many experimental results including reorganization processes caused by the CZ ablation and by incision through the meristem center. We thus conclude that the reaction-diffusion dynamics is probably indispensable for the SAM development of plants

RNA-seq Transcriptional Profiling of an Arbuscular Mycorrhiza Provides Insights into Regulated and Coordinated Gene Expression in Lotus japonicus and Rhizophagus irregularis

Gene expression during arbuscular mycorrhizal development is highly orchestrated in both plants and arbuscular mycorrhizal fungi. To elucidate the gene expression profiles of the symbiotic association, we performed a digital gene expression analysis of Lotus japonicus and Rhizophagus irregularis using a HiSeq 2000 next-generation sequencer with a Cufflinks assembly and de novo transcriptome assembly. There were 3,641 genes differentially expressed during arbuscular mycorrhizal development in L. japonicus, approximately 80% of which were up-regulated. The up-regulated genes included secreted proteins, transporters, proteins involved in lipid and amino acid metabolism, ribosomes and histones. We also detected many genes that were differentially expressed in small-secreted peptides and transcription factors, which may be involved in signal transduction or transcription regulation during symbiosis. Coregulated genes between arbuscular mycorrhizal and root nodule symbiosis were not particularly abundant, but transcripts encoding for membrane traffic-related proteins, transporters and iron transport-related proteins were found to be highly co-up-regulated. In transcripts of arbuscular mycorrhizal fungi, expansion of cytochrome P450 was observed, which may contribute to various metabolic pathways required to accommodate roots and soil. The comprehensive gene expression data of both plants and arbuscular mycorrhizal fungi provide a powerful platform for investigating the functional and molecular mechanisms underlying arbuscular mycorrhizal symbiosis.ArticlePLANT AND CELL PHYSIOLOGY. 56(8):1490-1511 (2015)journal articl

On the Virulence of Escherichia coli Strain TK18-A Artificially Induced into the Air Sac of Day- Old Chicks

鶏呼吸器性マイコプラズマ病に関する基礎的検討の一環として,本病の一実験感染系確立の為に,さきに著者ら(1980)21)はM. gallisepticum (MG) 1RF株を用いて,今回と同様な実験を行った成績について報告した。今回は鶏大腸菌症の原因ならびにMG感染症の混合感染菌として重要な,E.coliの気嚢内人工感染による初生ヒナに対するvirulenceの程度を知る目的で本実験を行った。 E. coli TK18-A株(O群2)液体培養の10倍段階希釈(5.4×10^0-5/0.4ml)を,ブロイラー初生ヒナ(雄)の右後胸気嚢内に接種し,4週間観察した。各実験群20羽宛のヒナにつき,臨床症状,増体量,飼料要求率,病理変状,各臓器からのE. coli分離等の項目につき検討し,下記の成績が得られた。 1) 全群呼吸器症状はみられず,各群ヒナの死亡は主に感染後7日以内にみられ,例外的に10及び14日後1例ずつの死亡がみられた。菌接種0～10^5 CFU各群の死亡率は,それぞれ0,5,5,40,60,100,100%であった。供試菌のLD50は5.4×10^2.4 CFUであり,接種菌10CFU以下では死亡率は極めて低く,また接種菌数が減少するに従い,死亡時期がやや遅延する傾向がみられた。 2) 増体量・飼料要求率については,早期死亡例が多く,各群間の比較は困難であった。40%のヒナが生残した10^3 CFU菌接種群とそれ以下の菌接種群での成長曲線では,10^3 CFU菌接種群が最低を示した。 3) 肉眼病変出現率と接種菌数とはほぼ平行する傾向がみられ,LD50以上の菌接種群で病変陽性の傾向がみられた。気嚢病変は菌接種側で高率にみられた。10^3 CFU菌接種死亡例で顕著な病変が多数みられた。気嚢病変は,10^2-5 CFU菌接種群にみられた。 4) 以上の病変は,病理組織学的にも,全体的に滲出性の炎症病相が主体で,従来記載の大腸菌性敗血症病変とよく一致した。 5) E. coli分離率は接種菌数とほぼ平行し,10^4-5 CFU菌接種群死亡例では100%で大部分のヒナは典型的な急性敗血症死とみられた。 6) 本菌株のAID50とLD50の菌数はほぼ同一とみられた。The present experiments were performed to investigate the virulence of E. coli strain TK18－A40) (O-group 2) induced artificially into the air sac of day-old chicks. This was done with the further purpose to establish an experimental system using chicks for fundamental experiments on the efficacy in vivo of the antibacterial agents that control the avian colibacillosis and the avian respiratory mycoplasmosis complicated with E. coli. The inocula of 0.4 ml /chick of ten-fold serial dilutions of broth culture were artificially induced into the right posterior thoracic air sac of day-old male broiler chicks, and these chicks were observed during 4 weeks. The experimental groups were designed as groups of 20 chicks each, 6 groups included a non-infected control and 5 infected groups. The inoculum sizes of the organisms of the chicks ranged from 5.4×1 to 10^5 CFU* in 5 dilutions of the broth culture. The following results were obtained. 1) No clinical symptoms were observed except that some chicks died. Death of the chicks was mostly observed within 7 days post inoculation, exceptionally 2 chicks only died 10 and 14 days post inoculation. Mortalities in the group of chicks inoculated with 0－10^5 CFU were 0, 5, 5, 40, 60, 100 and 100%, respectively. The LD50 of the organisms was determined as being 5.4×10^2.4 CFU. A remarkably low mortality was observed in the chicks inoculated with organisms less than 10 CFU. A delaying tendency of the lethal period of the chicks was noticed in accordance to the decrease of the number of organisms inoculated. 2) As a large number of chicks died in the early stage of the experiments, it was difficult to determine a comparison of weight gain and feed conversion for the chicks of each group. In the groups of chicks inoculated with 10^3 or less CFU of the organisms, 40% or more survived ratios were observed. In comparison of the growth curve of these groups, the group inoculated with 10^3 CFU of the organisms showed the lowest curve. 3) An almost parallel tendency was observed between the gross lesion score and the inoculum size of the organisms. The positive tendency of the gross lesion score was investigated in the group of chicks inoculated with a number larger than LD50 of the organisms. A higher gross lesion score was observed on the inoculated side of the air sac of chicks. A large number of severe lesions was observed in chicks that had died after inoculation of 10^3 CFU of the organisms. Air sac lesions were observed in the group of chicks inoculated with 10^2-10^5 CFU of the organisms. 4) The major finding observed in the histopathological examination of chicks was generally an exudative inflammatory manifestation. These findings were similar to those of the septicemic avian colibacillosis described by many workers. 5) The recovery ratio of the organisms ran mostly parallel with the number of the inoculated organisms. One hundred percent ratios were observed in 2 groups of chicks inoculated with 10^4－10^5 CFU of the organisms. It was considered that most cases of the dead chicks were due to coli-septicemia. 6) The AID50 of the organisms in the present experimental conditions was estimated as almost the same number (CFU) of the organisms as that of the LD50.本研究の一部は，第87 回日本獣医学会(1979年4月)で口頭発表したものである

The effects of ERN1 on gene expression during early rhizobial infection in Lotus japonicus

Legumes develop root nodules in association with compatible rhizobia to overcome nitrogen deficiency. Rhizobia enter the host legume, mainly through infection threads, and induce nodule primordium formation in the root cortex. Multiple transcription factors have been identified to be involved in the regulation of the establishment of root nodule symbiosis, including ERF Required for Nodulation1 (ERN1). ERN1 is involved in a transcription network with CYCLOPS and NODULE INCEPTION (NIN). Mutation of ERN1 often results in misshapen root hair tips, deficient infection thread formation, and immature root nodules. ERN1 directly activates the expression of ENOD11 in Medicago truncatula to assist cell wall remodeling and Epr3 in Lotus japonicus to distinguish rhizobial exopolysaccharide signals. However, aside from these two genes, it remains unclear which genes are regulated by LjERN1 or what role LjERN1 plays during root nodule symbiosis. Thus, we conducted RNA sequencing to compare the gene expression profiles of wild-type L. japonicus and Ljern1-6 mutants. In total, 234 differentially expressed genes were identified as candidate LjERN1 target genes. These genes were found to be associated with cell wall remodeling, signal transduction, phytohormone metabolism, and transcription regulation, suggesting that LjERN1 is involved in multiple processes during the early stages of the establishment of root nodule symbiosis. Many of these candidate genes including RINRK1 showed decreased expression levels in Ljnin-2 mutants based on a search of a public database, suggesting that LjERN1 and LjNIN coordinately regulate gene expression. Our data extend the current understanding of the pleiotropic role of LjERN1 in root nodule symbiosis

Isolation and Phenotypic Characterization of Lotus japonicus Mutants Specifically Defective in Arbuscular Mycorrhizal Formation

Several symbiotic mutants of legume plants defective in nodulation have also been shown to be mutants related to arbuscular mycorrhizal (AM) symbiosis. The origin of the AM symbiosis can be traced back to the early land plants. It has therefore been postulated that the older system of AM symbiosis was partially incorporated into the newer system of legume-rhizobium symbiosis. To unravel the genetic basis of the establishment of AM symbiosis, we screened about 34,000 plants derived from ethyl methanesulfonate (EMS)-mutagenized Lotus japonicus seeds by microscopic observation. As a result, three lines (ME778, ME966 and ME2329) were isolated as AM-specific mutants that exhibit clear AM-defective phenotypes but form normal effective root nodules with rhizobial infection. In the ME2329 mutant, AM fungi spread their hyphae into the intercellular space of the cortex and formed trunk hyphae in the cortical cells, but the development of fine branches in the arbuscules was arrested. The ME2329 mutant carried a nonsense mutation in the STR-homolog gene, implying that the line may be an str mutant in L. japonicus. On the ME778 and ME966 mutant roots, the entry of AM fungal hyphae was blocked between two adjacent epidermal cells. Occasionally, hyphal colonization accompanied by arbuscules was observed in the two mutants. The genes responsible for the ME778 and ME966 mutants were independently located on chromosome 2. These results suggest that the ME778 and ME966 lines are symbiotic mutants involved in the early stage of AM formation in L. japonicus.ArticlePLANT AND CELL PHYSIOLOGY. 55(5):928-941 (2014)journal articl

Expression and function of inducible co-stimulator in patients with systemic lupus erythematosus: possible involvement in excessive interferon-γ and anti-double-stranded DNA antibody production

Inducible co-stimulator (ICOS) is the third member of the CD28/cytotoxic T-lymphocyte associated antigen-4 family and is involved in the proliferation and activation of T cells. A detailed functional analysis of ICOS on peripheral blood T cells from patients with systemic lupus erythematosus (SLE) has not yet been reported. In the present study we developed a fully human anti-human ICOS mAb (JTA009) with high avidity and investigated the immunopathological roles of ICOS in SLE. JTA009 exhibited higher avidity for ICOS than a previously reported mAb, namely SA12. Using JTA009, ICOS was detected in a substantial proportion of unstimulated peripheral blood T cells from both normal control individuals and patients with SLE. In CD4(+)CD45RO(+ )T cells from peripheral blood, the percentage of ICOS(+ )cells and mean fluorescence intensity with JTA009 were significantly higher in active SLE than in inactive SLE or in normal control individuals. JTA009 co-stimulated peripheral blood T cells in the presence of suboptimal concentrations of anti-CD3 mAb. Median values of [(3)H]thymidine incorporation were higher in SLE T cells with ICOS co-stimulation than in normal T cells, and the difference between inactive SLE patients and normal control individuals achieved statistical significance. ICOS co-stimulation significantly increased the production of IFN-γ, IL-4 and IL-10 in both SLE and normal T cells. IFN-γ in the culture supernatants of both active and inactive SLE T cells with ICOS co-stimulation was significantly higher than in normal control T cells. Finally, SLE T cells with ICOS co-stimulation selectively and significantly enhanced the production of IgG anti-double-stranded DNA antibodies by autologous B cells. These findings suggest that ICOS is involved in abnormal T cell activation in SLE, and that blockade of the interaction between ICOS and its receptor may have therapeutic value in the treatment of this intractable disease

A Set of Lotus japonicus Gifu × Lotus burttii Recombinant Inbred Lines Facilitates Map-based Cloning and QTL Mapping

Model legumes such as Lotus japonicus have contributed significantly to the understanding of symbiotic nitrogen fixation. This insight is mainly a result of forward genetic screens followed by map-based cloning to identify causal alleles. The L. japonicus ecotype ‘Gifu’ was used as a common parent for inter-accession crosses to produce F2 mapping populations either with other L. japonicus ecotypes, MG-20 and Funakura, or with the related species L. filicaulis. These populations have all been used for genetic studies but segregation distortion, suppression of recombination, low polymorphism levels, and poor viability have also been observed. More recently, the diploid species L. burttii has been identified as a fertile crossing partner of L. japonicus. To assess its qualities in genetic linkage analysis and to enable quantitative trait locus (QTL) mapping for a wider range of traits in Lotus species, we have generated and genotyped a set of 163 Gifu × L. burttii recombinant inbred lines (RILs). By direct comparisons of RIL and F2 population data, we show that L. burttii is a valid alternative to MG-20 as a Gifu mapping partner. In addition, we demonstrate the utility of the Gifu × L. burttii RILs in QTL mapping by identifying an Nfr1-linked QTL for Sinorhizobium fredii nodulation