Novel Soybean Variety Lacking Raffinose Synthase 2 Activity.

Variation in the raffinose family oligosaccharide (RFO) content in soybean is advantageous for livestock farming and health science. In this study, a soybean variety (GmJMC172) with a significantly low stachyose content in its seeds was identified in the NARO Genebank core collection. The results of the single-nucleotide polymorphism (SNP) analysis suggested that this phenomenon was related to a single-base deletion, inducing a frameshift mutation in raffinose synthase 2 (RS2), rather than the polymorphisms in the RS3, RS4, and stachyose synthase (STS) sequences. Differences in the enzymatic properties between the native RS2 and truncated RS2 were examined by using a three-dimensional model predicted using Alphafold2. In addition to revealing the missing active pocket in truncated RS2, the modeled structure explained the catalytic role of W331* and suggested a sufficient space to bind both sucrose and raffinose in the ligand-binding pocket. The soybean line, with seeds available from the NARO Genebank, could serve as breeding materials for manipulating the RFO content.

Validation of the

The cluster structure of the neutron-rich isotope ^{10}Be has been probed via the (p,pα) reaction at 150 MeV/nucleon in inverse kinematics and in quasifree conditions. The populated states of ^{6}He residues were investigated through missing mass spectroscopy. The triple differential cross section for the ground-state transition was extracted for quasifree angle pairs (θ_{p},θ_{α}) and compared to distorted-wave impulse approximation reaction calculations performed in a microscopic framework using successively the Tohsaki-Horiuchi-Schuck-Röpke product wave function and the wave function deduced from antisymmetrized molecular dynamics calculations. The remarkable agreement between calculated and measured cross sections in both shape and magnitude validates the molecular structure description of the ^{10}Be ground-state, configured as an α-α core with two valence neutrons occupying π-type molecular orbitals.

Control of protein function through oxidation and reduction of persulfidated states.

Irreversible oxidation of Cys residues to sulfinic/sulfonic forms typically impairs protein function. We found that persulfidation (CysSSH) protects Cys from irreversible oxidative loss of function by the formation of CysSSO1-3H derivatives that can subsequently be reduced back to native thiols. Reductive reactivation of oxidized persulfides by the thioredoxin system was demonstrated in albumin, Prx2, and PTP1B. In cells, this mechanism protects and regulates key proteins of signaling pathways, including Prx2, PTEN, PTP1B, HSP90, and KEAP1. Using quantitative mass spectrometry, we show that (i) CysSSH and CysSSO3H species are abundant in mouse liver and enzymatically regulated by the glutathione and thioredoxin systems and (ii) deletion of the thioredoxin-related protein TRP14 in mice altered CysSSH levels on a subset of proteins, predicting a role for TRP14 in persulfide signaling. Furthermore, selenium supplementation, polysulfide treatment, or knockdown of TRP14 mediated cellular responses to EGF, suggesting a role for TrxR1/TRP14-regulated oxidative persulfidation in growth factor responsiveness.

Chemical curation to improve data accuracy: recent development of the 3DMET database.

We have developed a three-dimensional structure database of natural metabolites (3DMET). Early development of the 3DMET database relied on content auto-generated from 2D-structures of other chemical databases. From 2009, we began manual curation, obtaining new compounds from published works. In the process of curation, problems of digitizing 3D-structures from structure drawings of documents were accumulated. As the same as auto-generation, structure drawings should be also payed attention about stereochemistry. Our experiences in manual curation of 3DMET, as described herein, may be useful to others in this field of research and for the development of supporting systems of a chemical structure database. Manual curation is still necessary for proper database entry of the 3D-configurations of chiral atoms, a problem encountered frequently among natural products.

Current Challenges in Development of a Database of Three-Dimensional Chemical Structures.

We are developing a database named 3DMET, a three-dimensional structure database of natural metabolites. There are two major impediments to the creation of 3D chemical structures from a set of planar structure drawings: the limited accuracy of computer programs and insufficient human resources for manual curation. We have tested some 2D-3D converters to convert 2D structure files from external databases. These automatic conversion processes yielded an excessive number of improper conversions. To ascertain the quality of the conversions, we compared IUPAC Chemical Identifier and canonical SMILES notations before and after conversion. Structures whose notations correspond to each other were regarded as a correct conversion in our present work. We found that chiral inversion is the most serious factor during the improper conversion. In the current stage of our database construction, published books or articles have been resources for additions to our database. Chemicals are usually drawn as pictures on the paper. To save human resources, an optical structure reader was introduced. The program was quite useful but some particular errors were observed during our operation. We hope our trials for producing correct 3D structures will help other developers of chemical programs and curators of chemical databases.

Predictors of the response to treatment in acute lupus hemophagocytic syndrome.

OBJECTIVE: The objective of this paper is to identify predictors for the response to treatment of acute lupus hemophagocytic syndrome (ALHS). METHODS: We reviewed seven cases with ALHS admitted to our hospital and published ALHS cases identified in the 2001-2014 Medline database, and then conducted univariate and multivariate analyses to identify predictors for the response to treatment. RESULTS: Review of our cases showed a significant and negative correlation between serum ferritin and anti-DNA antibody (p = 0.0025). All three patients treated with cyclosporine A (CsA) were considered responders despite high serum ferritin and corticosteroid resistance. We also reviewed 93 patients with ALHS identified in 46 articles. Multiple logistic regression analysis identified C-reactive protein (CRP) (OR 0.83, p = 0.042) and hemoglobin (OR 1.53, p = 0.026) measured at diagnosis of ALHS as significant predictors of the response to corticosteroid monotherapy. Moreover, among 32 patients treated with CsA, serum ferritin was significantly higher in CsA responders (12163 ± 16864 µg/l, n = 22) than in non-responders (3456 ± 6267/µg/l, p = 0.020, n = 10). Leukocyte count was significantly lower in the CsA responders (1940.0 ± 972.3/µl) than in the non-responders (3253 ± 2198/µl, p = 0.034). CONCLUSION: Low CRP and high hemoglobin can predict a positive response to corticosteroid monotherapy while high serum ferritin and low leukocyte count can predict a positive response to CsA in patients with ALHS and therefore, when corticosteroid monotherapy is not effective in such cases, CsA could be the first choice of an additional immunosuppressive agent.

Pim-2 kinase is an important target of treatment for tumor progression and bone loss in myeloma.

Pim-2 kinase is overexpressed in multiple myeloma (MM) cells to enhance their growth and survival, and regarded as a novel therapeutic target in MM. However, the impact of Pim-2 inhibition on bone disease in MM remains unknown. We demonstrated here that Pim-2 expression was also upregulated in bone marrow stromal cells and MC3T3-E1 preosteoblastic cells in the presence of cytokines known as the inhibitors of osteoblastogenesis in MM, including interleukin-3 (IL-3), IL-7, tumor necrosis factor-α, transforming growth factor-ß (TGF-ß) and activin A, as well as MM cell conditioned media. The enforced expression of Pim-2 abrogated in vitro osteoblastogenesis by BMP-2, which suggested Pim-2 as a negative regulator for osteoblastogenesis. Treatment with Pim-2 short-interference RNA as well as the Pim inhibitor SMI-16a successfully restored osteoblastogenesis suppressed by all the above inhibitory factors and MM cells. The SMI-16a treatment potentiated BMP-2-mediated anabolic signaling while suppressing TGF-ß signaling. Furthermore, treatment with the newly synthesized thiazolidine-2,4-dione congener, 12a-OH, as well as its prototypic SMI-16a effectively prevented bone destruction while suppressing MM tumor growth in MM animal models. Thus, Pim-2 may have a pivotal role in tumor progression and bone loss in MM, and Pim-2 inhibition may become an important therapeutic strategy to target the MM cell-bone marrow interaction.

Three-dimensional morphology and bony range of movement in hip joints in patients with hip dysplasia.

To confirm whether developmental dysplasia of the hip has a risk of hip impingement, we analysed maximum ranges of movement to the point of bony impingement, and impingement location using three-dimensional (3D) surface models of the pelvis and femur in combination with 3D morphology of the hip joint using computer-assisted methods. Results of computed tomography were examined for 52 hip joints with DDH and 73 normal healthy hip joints. DDH shows larger maximum extension (p = 0.001) and internal rotation at 90° flexion (p < 0.001). Similar maximum flexion (p = 0.835) and external rotation (p = 0.713) were observed between groups, while high rates of extra-articular impingement were noticed in these directions in DDH (p < 0.001). Smaller cranial acetabular anteversion (p = 0.048), centre-edge angles (p < 0.001), a circumferentially shallower acetabulum, larger femoral neck anteversion (p < 0.001), and larger alpha angle were identified in DDH. Risk of anterior impingement in retroverted DDH hips is similar to that in retroverted normal hips in excessive adduction but minimal in less adduction. These findings might be borne in mind when considering the possibility of extra-articular posterior impingement in DDH being a source of pain, particularly for patients with a highly anteverted femoral neck.

Three-dimensional structure database of natural metabolites (3DMET): a novel database of curated 3D structures.

A database of 3D structures of natural metabolites has been developed called 3DMET. During the process of structure conversion from 2D to 3D, we found many structures were misconverted at chiral atoms and bonds. Several popular converters were tested in regard to their conversion accuracy. For verification, three canonical strings were also tested. No procedure could satisfactorily cover all the structures of the natural products. The misconverted structures had to be corrected manually. However, a nonnegligible number of mistakes were also observed even after manual curation, so a self-checking system was developed and introduced to our work flow. Thus, the 3D structures in our 3DMET database were evaluated in two steps: automatically and manually. The current version includes most of the natural products of the KEGG COMPOUND collection [ http://www.genome.jp/kegg/compound/ ] and is searchable by string, value range, and substructure. 3DMET can be accessed via http://www.3dmet.dna.affrc.go.jp/ , which also has detailed manuals.

Small molecule antibody targeting HLA class I inhibits myeloma cancer stem cells by repressing pluripotency-associated transcription factors.

Cancer stem cells have been proposed to be responsible for tumorigenesis and recurrence in various neoplastic diseases, including multiple myeloma (MM). We have previously reported that MM cells specifically express HLA class I at high levels and that single-chain Fv diabody against this molecule markedly induces MM cell death. Here we investigated the effect of a new diabody (C3B3) on cancer stem cell-like side population (SP) cells. SP fraction of MM cells highly expressed ABCG2 and exhibited resistance to chemotherapeutic agents; however, C3B3 induced cytotoxicity in both SP cells and main population (MP) cells to a similar extent. Moreover, C3B3 suppressed colony formation and tumorigenesis of SP cells in vitro and in vivo. Crosslinking of HLA class I by C3B3 mediated disruption of lipid rafts and actin aggregation, which led to inhibition of gene expression of ß-catenin and pluripotency-associated transcription factors such as Sox2, Oct3/4 and Nanog. Conversely, knockdown of Sox2 and Oct3/4 mRNA reduced the proportion of SP cells, suggesting that these factors are essential in maintenance of SP fraction in MM cells. Thus, our findings reveal that immunotherapeutic approach by engineered antibodies can overcome drug resistance, and provide a new basis for development of cancer stem cell-targeted therapy.

The serine/threonine kinase Pim-2 is a novel anti-apoptotic mediator in myeloma cells.

Bone marrow stromal cells (BMSCs) and osteoclasts (OCs) confer multiple myeloma (MM) cell survival through elaborating factors. We demonstrate herein that IL-6 and TNF family cytokines, TNFα, BAFF and APRIL, but not IGF-1 cooperatively enhance the expression of the serine/threonine kinase Pim-2 in MM cells. BMSCs and OCs upregulate Pim-2 expression in MM cells largely via the IL-6/STAT3 and NF-κB pathway, respectively. Pim-2 short interfering RNA reduces MM cell viability in cocultures with BMSCs or OCs. Thus, upregulation of Pim-2 appears to be a novel anti-apoptotic mechanism for MM cell survival. Interestingly, the mammalian target of rapamycin inhibitor rapamycin further suppresses the MM cell viability in combination with the Pim-2 silencing. The Pim inhibitor (Z)-5-(4-propoxybenzylidene) thiazolidine-2, 4-dione and the PI3K inhibitor LY294002 cooperatively enhance MM cell death. The Pim inhibitor suppresses 4E-BP1 phosphorylation along with the reduction of Mcl-1 and c-Myc. Pim-2 may therefore become a new target for MM treatment.

Oligomeric peroxiredoxin-I is an essential intermediate for p53 to activate MST1 kinase and apoptosis.

Mammalian Ste20-like kinase-1 (MST1) kinase mediates H2O2-induced cell death by anticancer drugs such as cisplatin in a p53-dependent manner. However, the mechanism underlying MST1 activation by H2O2 remains unknown. Here we show that peroxiredoxin-I (PRX-I) is an essential intermediate in H2O2-induced MST1 activation and cisplatin-induced cell death through p53. Cell stimulation with H2O2 resulted in PRX-I oxidation to form homo-oligomers and interaction with MST1, leading to MST1 autophosphorylation and augmentation of kinase activity. In addition, RNA interference knockdown experiments indicated that endogenous PRX-I is required for H2O2-induced MST1 activation. Live-cell imaging showed H2O2 generation by cisplatin treatment, which likewise caused PRX-I oligomer formation, MST1 activation and cell death. Cisplatin-induced PRX-I oligomer formation was not observed in embryonic fibroblasts obtained from p53-knockout mice, confirming the importance of p53. Indeed, ectopic expression of p53 induced PRX-I oligomer formation and cell death, both of which were cancelled by the antioxidant NAC. Moreover, we succeeded in reconstituting H2O2-induced MST1 activation in vitro, using purified PRX-I and MST1 proteins. Collectively, our results show a novel PRX-I function to cause cell death in response to high levels of oxidative stress by activating MST1, which underlies the p53-dependent cytotoxicity caused by anticancer agents.

Grafting with hydroxyapatite granules for defects of acetabular bone at revision total hip replacement: a minimum ten-year follow-up.

The long-term results of grafting with hydroxyapatite granules for acetabular deficiency in revision total hip replacement are not well known. We have evaluated the results of revision using a modular cup with hydroxyapatite grafting for Paprosky type 2 and 3 acetabular defects at a minimum of ten years' follow-up. We retrospectively reviewed 49 acetabular revisions at a mean of 135 months (120 to 178). There was one type 2B, ten 2C, 28 3A and ten 3B hips. With loosening as the endpoint, the survival rate was 74.2% (95% confidence interval 58.3 to 90.1). Radiologically, four of the type 3A hips (14%) and six of the type 3B hips (60%) showed aseptic loosening with collapse of the hydroxyapatite layer, whereas no loosening occurred in type 2 hips. There was consolidation of the hydroxyapatite layer in 33 hips (66%). Loosening was detected in nine of 29 hips (31%) without cement and in one of 20 hips (5%) with cement (p = 0.03, Fisher's exact probability test). The linear wear and annual wear rate did not correlate with loosening. These results suggest that the long-term results of hydroxyapatite grafting with cement for type 2 and 3A hips are encouraging.

A modular femoral neck and head system works well in cementless total hip replacement for patients with developmental dysplasia of the hip.

We compared a modular neck system with a non-modular system in a cementless anatomical total hip replacement (THR). Each group consisted of 74 hips with developmental hip dysplasia. Both groups had the same cementless acetabular component and the same articulation, which consisted of a conventional polyethylene liner and a 28 mm alumina head. The mean follow-up was 14.5 years (13 to 15), at which point there were significant differences in the mean total Harris hip score (modular/non-modular: 98.6 (64 to 100)/93.8 (68 to 100)), the mean range of abduction (32 degrees (15 degrees to 40 degrees )/28 (0 degrees to 40 degrees )), use of a 10 degrees elevated liner (31%/100%), the incidence of osteolysis (27%/79.7%) and the incidence of equal leg lengths (> or = 6 mm, 92%/61%). There was no disassociation or fracture of the modular neck. The modular system reduces the need for an elevated liner, thereby reducing the incidence of osteolysis. It gives a better range of movement and allows the surgeon to make an accurate adjustment of leg length.

Choledochal cyst associated with duodenal atresia: case report and review of the literature.

We report a rare case of choledochal cyst (CC) associated with congenital duodenal atresia (DA) and annular pancreas (AP). A girl was born at 37 weeks of gestation weighing 2,974 g with a prenatal diagnosis of DA. She underwent a duodenoduodenostomy for type III DA with an AP 1 day after birth. At 4 years of age, she was admitted for evaluation of cholangitis and pancreatitis. Radiological studies demonstrated a fusiform-type CC with pancreaticobiliary maljunction (PBMJ). Excision of the CC and hepaticojejunostomy were performed. The patient was discharged without complications. Despite the fact that CC, DA, and AP are embryologically closely related entities, to the best of our knowledge, only eight such cases have been documented. We must be aware of the possible combination of CC in the follow-up of the patients with DA associated with AP.

Embryonic rather than extraembryonic tissues have more impact on the development of placental hyperplasia in cloned mice.

Somatic cell cloning by nuclear transfer (NT) in mice is associated with hyperplastic placentas at term. To dissect the effects of embryonic and extraembryonic tissues on this clone-associated phenotype, we constructed diploid (2n) fused with (<-->) tetraploid (4n) chimeras from NT- and fertilization-derived (FD) embryos. Generally, the 4n cells contributed efficiently to all the extraembryonic tissues but not to the embryo itself. Embryos constructed by 2n NT<-->4n FD aggregation developed hyperplastic placentas (0.33+/-0.22 g) with a predominant contribution by NT-derived cells. Even when the population of FD-derived cells in placentas was increased using multiple FD embryos (up to four) for aggregation, most placentas remained hyperplastic (0.36+/-0.13 g). By contrast, placentas of the reciprocal combination, 2n FD<-->4n NT, were less hyperplastic (0.15+/-0.02 g). These nearly normal-looking placentas had a large proportion of NT-derived cells. Thus, embryonic rather than extraembryonic tissues had more impact on the onset of placental hyperplasia, and that the abnormal placentation in clones occurs in a noncell-autonomous manner. These findings suggest that for improvement of cloning efficiency we should understand the mechanisms regulating placentation, especially those of embryonic origin that might control the proliferation of trophoblastic lineage cells.

Topographic analysis of the inferior parietal lobule in high-resolution 3D MR imaging.

BACKGROUND AND PURPOSE: Recent functional MR imaging studies have revealed that the inferior parietal lobule (IPL) supports numerous functions; however, the precise organization of the IPL remains unclear. It has been previously reported that the individual components of the IPL show variable morphologic features; thus, any investigations of IPL function must take into account the variable topography of the IPL. However, it is difficult to understand the topography of the IPL only in transaxial and/or sagittal MR images. We generated volume rendering (VR) images of the brain from high-resolution 3D MR imaging data and analyzed the topography of the IPL. MATERIALS AND METHODS: Forty healthy volunteers were examined with 1.5T MR imaging, and VR images were generated. Sulcal continuity, sulcal connections, and the gyral pattern of the IPL were investigated. RESULTS: The main sulci and gyri were able to be identified excellently on VR images of all volunteers. One long, continuous postcentral sulcus (postCS) was seen in 47.5% and 40% in the right and left hemisphere, respectively. The frequency with which the postCS connected with the anterior end of the IPS was 72.5% and 75% in the right and left, respectively. The gyral pattern of the IPL showed variable morphology. CONCLUSIONS: VR images can depict the sulcal continuity and connections of the IPL accurately. VR imaging is a very useful noninvasive technique to observe the topography of the IPL and should contribute to the investigation of functional localization.

Development of new indices to evaluate protein-protein interfaces: assembling space volume, assembling space distance, and global shape descriptor.

Protein-protein interaction is fundamental to initiate the cellular functions of proteins, and thus structural analyses of protein interfaces are the first step to understand their functions at the molecular level. Although shape complementarities have been used to evaluate the fitness of interfaces, the conventional method did not distinguish between two main components of complementarity, the well-fitting of the surface shapes and the size of the gap regions between the pair of molecules, and could not evaluate the global shape of the interfaces. Therefore, we now propose three new indices to describe protein interfaces: assembling space volume (ASV), assembling space distance (AS-distance), and global shape descriptor (GS). The ASV is calculated using Delaunay tessellation, and the AS-distance is calculated as the ratio of ASV to delta-ASA (accessible surface area). The GS is calculated as the ratio of the volume of Delaunay tetrahedra with a long edge to that of all tetrahedra in the assembling space. To evaluate the feasibility of the three indices, we applied our method to homo-dimer proteins, and performed systematic comparisons with SURFNET by Laskowski and shape complementarity by Lawrence and Colman. As a result, our indices behave differently from the existing ones, and shed light on new features of protein-protein interfaces, as general trends of AS-distances for all protein interfaces.