An Expansion of Crystalline Sponge X-ray Analysis to Elucidate the Molecular Structure of Reactive Compounds via Ion Pair Formation.

The crystalline sponge (CS) method allows structural elucidation of a target compound (guest) in solution by single crystal X-ray diffraction through trapping the guest into the CS framework. In principle, the CS method is inapplicable to reactive compounds that break the CS framework, such as acidic, basic, or nucleophilic ones. Here, a solution to this problem is disclosed wherein an ion pair of the guest compound is formed during the guest-soaking step by adding a suitable reagent. The ion pair can be observed and does not damage the CS framework. Using the developed method, amino, guanidino, and amidino compounds have been successfully analyzed as ion pairs with sulfonic acids. Practical utility has been shown because the absolute configurations of optically resolved amine derivatives were revealed with only a few micrograms. This demonstrates that the ion-pair-soaking method is simple and expands the range of compounds applicable to the CS method.

Safe administration of rituximab for follicular lymphoma after obinutuzumab infusion-related reaction.

Obinutuzumab is a novel glycoengineered, type-II anti-CD20 monoclonal antibody that was recently developed to treat follicular lymphoma (FL), the most prevalent subtype of indolent B-cell lymphoma. Several intensely hypermetabolic lesions (SUVmax: 40) were identified in the post-mediastinal and paraaortic lymph nodes by 18F-FDG-PET maximum-intensity projection images of a 58-year-old man who presented with systemic lymphadenopathy. A biopsy at the time of laparotomy definitively diagnosed grade 1 FL. The patient was given the recommended standard premedication, comprising acetaminophen (1000 mg), diphenhydramine (50 mg), and dexamethasone (20 mg), and then started on six cycles of obinutuzumab plus cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP). However, the patient developed severe hypotension and dyspnea about 15 min after starting obinutuzumab. It was difficult to differentiate between a possible allergic reaction and infusion-related reaction. A pleural effusion was drained to reduce the tumor burden, after which a single course of CHOP was started. Rituximab (R) was added 10 days later without incident, and the patient completed six cycles of the R-CHOP therapy without adverse events. We conclude that R-CHOP was safe for administration to patients who react to infused obinutuzumab. Such patients should be carefully monitored during R infusion, given the risk of cross-reactivity.

Development of drug-loaded protein nanoparticles displaying enzymatically-conjugated DNA aptamers for cancer cell targeting.

Modification of protein-based drug carriers with tumor-targeting properties is an important area of research in the field of anticancer drug delivery. To this end, we developed nanoparticles comprised of elastin-like polypeptides (ELPs) with fused poly-aspartic acid chains (ELP-D) displaying DNA aptamers. DNA aptamers were enzymatically conjugated to the surface of the nanoparticles via genetic incorporation of Gene A* protein into the sequence of the ELP-D fusion protein. Gene A* protein, derived from bacteriophage ϕX174, can form covalent complexes with single-stranded DNA via the latter's recognition sequence. Gene A* protein-displaying nanoparticles exhibited the ability to deliver the anticancer drug paclitaxel (PTX), whilst retaining activity of the conjugated Gene A* protein. PTX-loaded protein nanoparticles displaying DNA aptamers known to bind to the MUC1 tumor marker resulted in increased cytotoxicity with MCF-7 breast cancer cells compared to PTX-loaded protein nanoparticles without the DNA aptamer modification.

Pulmonary Hyalinizing Granuloma Mimicking Primary Lung Cancer.

We herein report a case of pulmonary hyalinizing granuloma (PHG), which is a rare pulmonary mass. A 69-year-old man with no symptoms presented to our hospital because of the appearance of an abnormal shadow on chest X-ray. Computed tomography revealed a right middle-lobe mass with spicula and infiltration into the upper lobe. Since a bronchofiberscopic examination showed no malignant cells in the specimen, the patient underwent thoracoscopic surgery, which revealed PHG. Spiculation and interlobar infiltration, which comprise the characteristic features of primary lung cancer, are uncommon presentations of this rare entity.

Influence of combustion-originated dioxins in atmospheric deposition on water quality of an urban river in Japan.

Bulk (wet and dry) deposition samples were collected in Saitama Prefecture, Japan throughout a year (February 8, 2012 to February 7, 2013) to estimate the influence of dioxins emitting from waste incinerators on river water quality. The annual deposition flux of dioxins was 3.3ng-toxic equivalent (TEQ)/m2/year. Source identification using indicative congeners estimated that 82% of dioxin TEQ in the bulk deposition (2.7ng-TEQ/m2/year) was combustion-originated, indicating that most of the dioxins in the deposition were derived from waste incinerators. In Saitama prefecture the annual flux of combustion-originated dioxins in depositions was apparently consistent with that of dioxin emission into the air from waste incinerators. The TEQ of combustion-originated dioxins in the deposition per rainfall was 2.4pg-TEQ/L on annual average, exceeding the environmental quality standard (EQS) for water in Japan of 1pg-TEQ/L. This suggests there is a possibility that dioxins in atmospheric deposition have a significant influence on the water quality of urban rivers which rainwater directly flows into because of many paved areas in the basins. The influence of combustion-originated dioxin in the deposition on the water quality of Ayase River, an urban river heavily polluted with dioxins, was estimated at 0.29pg-TEQ/L on annual average in 2015. It seems that dioxins in atmospheric deposition from waste incinerators have a significant influence on water quality of some urban rivers via rainwater though the dioxins in the ambient air have achieved the EQS for atmosphere at all monitoring sites in Japan.

A magnetic synapse: multilevel spin-torque memristor with perpendicular anisotropy.

Memristors are non-volatile nano-resistors which resistance can be tuned by applied currents or voltages and set to a large number of levels. Thanks to these properties, memristors are ideal building blocks for a number of applications such as multilevel non-volatile memories and artificial nano-synapses, which are the focus of this work. A key point towards the development of large scale memristive neuromorphic hardware is to build these neural networks with a memristor technology compatible with the best candidates for the future mainstream non-volatile memories. Here we show the first experimental achievement of a multilevel memristor compatible with spin-torque magnetic random access memories. The resistive switching in our spin-torque memristor is linked to the displacement of a magnetic domain wall by spin-torques in a perpendicularly magnetized magnetic tunnel junction. We demonstrate that our magnetic synapse has a large number of intermediate resistance states, sufficient for neural computation. Moreover, we show that engineering the device geometry allows leveraging the most efficient spin torque to displace the magnetic domain wall at low current densities and thus to minimize the energy cost of our memristor. Our results pave the way for spin-torque based analog magnetic neural computation.

Targeting of EGF-displayed protein nanoparticles with anticancer drugs.

The development of protein-based carriers for drug delivery has been well studied. We previously constructed a protein-based nanoparticle consisting of genetically engineered elastin-like polypeptides (ELPs) with a fused poly-aspartic acid tail (ELPD ). The size of the self-assembled ELPD nanoparticles was regulated by charged repulsion of the poly-aspartic acid chains. In the present study, epidermal growth factor (EGF) was genetically fused to the C-terminus of ELPD to impart an active targeting ability to the ELPD nanoparticles. We examined the nanoparticle formation with EGF as well as its targeting ability. ELPD with fused EGF was found to form nanoparticles that displayed multivalent EGFs on their surface. EGF-displayed nanoparticles loaded with the anti-cancer drug paclitaxel were internalized into cells overexpressing the EGF receptor, and induced cell death.

Molecular characterization and subcellular localization of Arabidopsis class VIII myosin, ATM1.

Land plants possess myosin classes VIII and XI. Although some information is available on the molecular properties of class XI myosins, class VIII myosins are not characterized. Here, we report the first analysis of the enzymatic properties of class VIII myosin. The motor domain of Arabidopsis class VIII myosin, ATM1 (ATM1-MD), and the motor domain plus one IQ motif (ATM1-1IQ) were expressed in a baculovirus system and characterized. ATM1-MD and ATM1-1IQ had low actin-activated Mg(2+)-ATPase activity (Vmax = 4 s(-1)), although their affinities for actin were high (Kactin = 4 µM). The actin-sliding velocities of ATM1-MD and ATM1-1IQ were 0.02 and 0.089 µm/s, respectively, from which the value for full-length ATM1 is calculated to be ∼0.2 µm/s. The results of actin co-sedimentation assay showed that the duty ratio of ATM1 was ∼90%. ADP dissociation from the actin·ATM1 complex (acto-ATM1) was extremely slow, which accounts for the low actin-sliding velocity, low actin-activated ATPase activity, and high duty ratio. The rate of ADP dissociation from acto-ATM1 was markedly biphasic with fast and slow phase rates (5.1 and 0.41 s(-1), respectively). Physiological concentrations of free Mg(2+) modulated actin-sliding velocity and actin-activated ATPase activity by changing the rate of ADP dissociation from acto-ATM1. GFP-fused full-length ATM1 expressed in Arabidopsis was localized to plasmodesmata, plastids, newly formed cell walls, and actin filaments at the cell cortex. Our results suggest that ATM1 functions as a tension sensor/generator at the cell cortex and other structures in Arabidopsis.

High domain wall velocities via spin transfer torque using vertical current injection.

Domain walls, nanoscale transition regions separating oppositely oriented ferromagnetic domains, have significant promise for use in spintronic devices for data storage and memristive applications. The state of these devices is related to the wall position and thus rapid operation will require a controllable onset of domain wall motion and high speed wall displacement. These processes are traditionally driven by spin transfer torque due to lateral injection of spin polarized current through a ferromagnetic nanostrip. However, this geometry is often hampered by low maximum wall velocities and/or a need for prohibitively high current densities. Here, using time-resolved magnetotransport measurements, we show that vertical injection of spin currents through a magnetic tunnel junction can drive domain walls over hundreds of nanometers at ~500 m/s using current densities on the order of 6 MA/cm(2). Moreover, these measurements provide information about the stochastic and deterministic aspects of current driven domain wall mediated switching.

[Clinical evaluation of calculating carboplatin doses using modification of diet in renal disease (MDRD) estimate and adverse events].

A modified diet in renal disease (MDRD), a formula to estimate glomerular filtration rate (GFR), was proposed by Levey in 2006. In this study, we compared the dosage of carboplatin (CBDCA) calculated using MDRD with that calculated by conservative creatinine clearance (Ccr), and investigated the actual dosage given and the incidence of its adverse effects. In the 101 patients undergoing chemotherapy including CBDCA, the dosage calculated from the estimated GFR was 16% lower than that based on the estimated Ccr. This difference was greater in those under 65 years, women and those with a body mass index (BMI) > or =25. The most prominent incidence of adverse effects was thrombocytopenia in patients with lung cancer. In men, a serum creatinine level of > or =0. 6 mg/dL, GFR of <50 mL/min/1. 73 m(2) and a combined use of gemcitabine were cited as the factors responsible for the development of thrombocytopenia. It was concluded that the MDRD formula is an effective tool for evaluating patients with impaired renal function. It was suggested, on the other hand, that the dosage for medication should be decided by giving due consideration to factors other than renal functions.

Bacterial population dynamics in a reverse-osmosis water purification system determined by fluorescent staining and PCR-denaturing gradient gel electrophoresis.

The bacterial population dynamics in an industrial scale reverse-osmosis (RO) water purification system were analyzed by fluorescent staining methods and denaturing gradient gel electrophoresis (DGGE). Bacterial numbers increased with storage in a tank, and bacterial diversity changed during the water purification process. A DNA sequence-based analysis of the major bands on the DGGE gel revealed that Simonsiella sp. (Betaproteobacteria) was abundant in the source water (activated sludge-treated waste effluent), while Bosea sp. and Rhizobium sp. (Alphaproteobacteria), which usually exist in an oligotrophic environment, became abundant during the water purification process. These results suggest the importance of microbiological monitoring by culture-independent methods for quality control in RO water purification systems. These methods could provide an early warning of impending problems and clarify critical steps in controlling specific bacteria contributing to the contamination of RO water systems.

Histamine H4 receptor agonists have more activities than H4 agonism in antigen-specific human T-cell responses.

Histamine not only mediates immediate allergic reactions, it also regulates cellular immune responses. H4R is the most recently identified histamine receptor (HR). In the present study, we examined the in vitro effect of histamine and H4R agonists on the responses of human T cells to purified protein derivative from Mycobacterium tuberculosis (PPD) and to Cry j1, the major allergen of Cryptomeria japonica pollen. Dimaprit, clobenpropit and clozapine, which are H4R agonists, dose-dependently blocked both PPD-induced interferon-gamma and Cry j1-induced interleukin-5 production by both peripheral blood mononuclear cells (PBMCs) and antigen-specific T-cell lines. However, the addition of thioperamide, an H3R/H4R antagonist, as well as a mixture of d-chlropheniramine, famotidine and thioperamide, did not reverse the inhibition. Pretreatment of PBMCs with SQ22536 and 8-bromoadenosine-3',5'-cyclic monophosphorothioate, Rp-isomer, had varying abilities to reverse the inhibitory effects of H4R agonists, except for clobenpropit. Moreover, the addition of H4R agonists induced annexin-V expression on PBMCs, especially in CD19(+) and CD4(+) cells. cDNA microarray analysis revealed that, among 16,600 genes tested, increased expression following treatment with clozapine was seen in 0 x 8% of the genes, whereas decreased expression was seen in 3 x 0% of the genes. These results suggest that H4R agonists inhibit antigen-specific human T-cell responses, although H4R does not appear to be important for this effect. In addition, the present study indicated that there may be orphan receptors or HR subtypes which can bind dimaprit, clobenpropit and clozapine, and that can exert an inhibitory effect on antigen-specific cellular responses via a cAMP/cAMP-dependent protein kinase-dependent, apoptotic pathway.

Expression of IL-12 and T helper cell 1 cytokines in the fluid of paranasal sinus mucoceles.

OBJECTIVE: The objective of this study was to assess the expression of regulatory cytokines and T helper cell (Th)1/Th2 cytokines in paranasal sinus mucoceles. MATERIALS AND METHODS: Fluid samples of 12 paranasal sinus mucoceles were assessed by enzyme-linked immunosorbent assay for concentrations of regulatory cytokines (interleukin [IL]-10 and IL-12), Th1 cytokines (IL-2 and interferon gamma), and Th2 cytokines (IL-4 and IL-5). RESULTS: IL-12 was detected in all samples, whereas IL-10 was detected in only one case. The concentration of IL-12 tended to correlate with that of interferon gamma and was significantly and positively correlated with that of IL-2. CONCLUSIONS: Th1 cytokines and the Th1 regulatory cytokine IL-12, but not IL-10, potentially play a key role in the pathogenesis of paranasal sinus mucoceles. Together with our recent report showing that lipopolysaccharide is highly detected in mucocele fluid, the data from this study suggest that the Th1 response induced by lipopolysaccharide may affect the immunological inflammation in the epithelium of paranasal sinus mucoceles.

TH1/TH2 and regulatory cytokines in adults with otitis media with effusion.

OBJECTIVE: Otitis media with effusion is one of the most common and intractable ear diseases. However, the role of Th1, Th2, and immunoregulatory cytokines on the pathogenesis of the disease in adult patients remains to be determined. The aim of this study is to disclose the cytokine expression in middle ear effusions (MEEs) in adults and to compare the profile on the basis of the presence of allergic rhinitis and the type of effusions. STUDY DESIGN: A prospective controlled clinical study. PATIENTS: MEEs were collected from 80 adult subjects. The concentration of interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12, and interferon (IFN)-gamma in MEEs were determined by using enzyme-linked immunosorbent assay. RESULTS: IL-2, IL-4, IL-5, IL-10, IL-12, and IFN-gamma in MEEs were detected in 60 (75.0%), 33 (41.3%), 42 (52.5%), 14 (17.5%), 80 (100%), and 66 (82.5%) samples, respectively. Among these cytokines, only the concentration of IL-4 in the allergic rhinitis-positive group was significantly higher than that in the allergic rhinitis-negative group. On the other hand, IL-2, IL-12, and IFN-gamma were detected, regardless of the presence of allergic rhinitis, and the concentration of these cytokines correlated with each other. The correlation between the concentration of IL-4 and IL-5 was also detected. In addition, both the incidence rate and the concentration of IL-10 in MEEs were significantly higher in the mucoid type compared with those in the serous type effusions. CONCLUSION: Regardless of allergic status, IL-12 may play a critical role in the pathogenesis of otitis media with effusion by affecting the production of IL-2 and IFN-gamma. In addition, IL-4 may have some impact on the immunologic condition in adults with allergic rhinitis. IL-10 potentially affects the viscosity of MEEs.

E prostanoid 2 (EP2)/EP4-mediated suppression of antigen-specific human T-cell responses by prostaglandin E2.

Prostaglandin E2 (PGE2) is a lipid mediator that displays important immunomodulatory properties, such as polarization of cytokine production by T cells. Recent investigations have revealed that the effect of PGE2 on cytokine production is greatly influenced by external stimuli; however, it is unclear whether PGE2 plays a significant role in major histocompatibility complex-mediated antigen-specific T-cell responses via binding to one of four subtypes of E prostanoid (EP) receptor alone or in combination. In the present study, we sought to determine the effect of PGE2 on antigen-specific CD4+ T-cell responses in humans, especially in terms of receptor specificity. We used purified protein derivative (PPD) and Cry j 1 as T helper type 1 (Th1) and Th2-inducing antigens, respectively. We generated several different Cry j 1- and PPD-specific T-cell lines (TCLs). PGE2 significantly and dose-dependently inhibited the proliferation and subsequent production of interleukin-4 by Cry j 1-specific TCLs and of interferon-gamma by PPD-specific TCLs upon antigen stimulation. Administration of EP2 receptor agonist and EP4 receptor agonist suppressed these responses in an adenylate cyclase-dependent manner, while EP1 and EP3 receptor agonists did not. Messenger RNA for EP2, EP3 and EP4, but not EP1, receptors were detected in Cry j 1- and PPD-specific TCLs, and no differences in EP receptor expression were observed between them. Furthermore, PGE2 and EP2 receptor agonist significantly inhibited interleukin-5 and interferon-gamma production by peripheral blood mononuclear cells in response to Cry j 1 and PPD stimulation, respectively. These results suggest that PGE2 suppresses both Th1- and Th2-polarized antigen-specific human T-cell responses via a cAMP-dependent EP2/EP4-mediated pathway.

Presence and characterization of prostaglandin D2-related molecules in nasal mucosa of patients with allergic rhinitis.

BACKGROUND: Prostaglandin D2 (PGD2) is the major prostanoid produced in the acute phase of allergic reactions. However, its pathophysiological role in addition to the pathway of production in allergic rhinitis remains unclear. We sought to determine the expression of synthases and receptors for PGD2 in human nasal mucosa. These expressions were compared between allergic and nonallergic patients. METHODS: The expression and localization of hematopoietic-type (h)-PGD2 synthase (PGDS) and lipocalin-type (l)-PGDS were detected by immunohistochemistry. The expression of D prostanoid (DP) receptor and chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) was determined by quantitative real-time PCR. RESULTS: The h-PGDS but not l-PGDS was clearly expressed in nasal mucosa. The expression of h-PGDS in allergic patients was significantly higher than in control patients without mucosal hypertrophy. A variety of infiltrating cells including mast cells, eosinophils, macrophages, and lymphocytes as well as constitutive cells such as epithelial cells and fibroblasts expressed h-PGDS. The expression of both DP and CRTH2 was confirmed also. Although either the amount of DP or the amount of CRTH2 was not correlated with serum levels of IgE, the amount of CRTH2 but not DP was highly and significantly correlated with the number of eosinophils infiltrating into nasal musosa. CONCLUSION: These results suggest that PGD2 is released via the action of h-PGDS from various cells, and the expression of h-PGDS may be associated with the hypertrophic inflammation in the nose. In addition, ligation of PGD2 to CRTH2 appears to be selectively involved in eosinophil recruitment into the nose regardless of atopic status.

Molecular characterization of a novel translocation t(5;14)(q21;q32) in a patient with congenital abnormalities.

Chromosomal translocations are frequently found to be associated with various malignant disorders as well as congenital abnormalities. We report the characterization of a novel reciprocal translocation t(5;14)(q21;q32) in a patient with congenital abnormalities manifested by severe mental retardation, athetotic tetraplegia, microcephaly, peculiar facies (upward slanting of palpebral fissures), clinodactyly of the fifth fingers, and overlapping toes. Using a JHGP24 lymphoblast cell line derived from this patient, metaphase fluorescence in situ hybridization with bacterial artificial chromosome and cosmid probes and subsequent molecular analysis mapped the translocation breakpoint to the nucleotide level. Sequence analysis of the breakpoint junctions revealed the presence of a homologous sequence, GTGGC, along with a single nucleotide substitution and an insertion in der(14), and a single nucleotide deletion in the der(5) chromosome. We also attempted to identify and characterize the transcripts near the breakpoint by 5' and 3' rapid amplification of cDNA ends. Although we found several transcripts near the breakpoint of chromosome 14, the lack of significant ORFs within these transcripts suggests they are likely to be non-coding RNAs. These transcripts may have an important role in the neurogenesis or differentiation.

A series of maturity onset diabetes of the young, type 2 (MODY2) mouse models generated by a large-scale ENU mutagenesis program.

Mutant mouse models are indispensable tools for clarifying the functions of genes and for elucidating the underlying pathogenic mechanisms of human diseases. Currently, several large-scale mutagenesis projects that employ the chemical mutagen N-ethyl-N-nitrosourea (ENU) are underway worldwide. One specific aim of our ENU mutagenesis project is to generate diabetic mouse models. We screened 9375 animals for dominant traits using a clinical biochemical test and thereby identified 11 mutations in the glucokinase (Gk) gene that were associated with hyperglycemia. GK is a key regulator of insulin secretion in the pancreatic beta-cell. Approximately 190 heterozygous mutations in the human GK gene have been reported to cause maturity onset diabetes of the young, type 2 (MODY2). In addition, five mutations have been reported to cause permanent neonatal diabetes mellitus (PNDM) when present on both alleles. The mutations in our 11 hyperglycemic mutants are located at different positions in Gk. Four have also been found in human MODY2 patients, and another mutant bears its mutation at the same location that is mutated in a PNDM patient. Thus, ENU mutagenesis is effective for developing mouse models for various human genetic diseases, including diabetes mellitus. Some of our Gk mutant lines displayed impaired glucose-responsive insulin secretion and the mutations had different effects on Gk mRNA levels and/or the stability of the GK protein. This collection of Gk mutants will be valuable for understanding GK gene function, for dissecting the function of the enzyme and as models of human MODY2 and PNDM.