Subthreshold membrane potential dynamics of posterior parietal cortical neurons coupled with hippocampal ripples.

During behavioral states of immobility, sleep, and anesthesia, the hippocampus generates high-frequency oscillations called ripples. Ripples occur simultaneously with synchronous neuronal activity in the neocortex, known as slow waves, and contribute to memory consolidation. During these ripples, various neocortical regions exhibit modulations in spike rates and local field activity irrespective of whether they receive direct synaptic inputs from the hippocampus. However, little is known about the subthreshold dynamics of the membrane potentials of neocortical neurons during ripples. We patch-clamped layer 2/3 pyramidal cells in the posterior parietal cortex (PPC), a neocortical region that is involved in allocentric spatial representation of behavioral exploration and sequential series of relevant action potentials during ripples. We simultaneously monitored the membrane potentials of post hoc-identified PPC neurons and the local field potentials of the hippocampus in anesthetized mice. More than 50% of the recorded PPC neurons exhibited significant depolarizations and/or hyperpolarizations during ripples. Histological inspections of the recorded neurons revealed that the ripple-modulated PPC neurons were distributed in the PPC in a spatially non-biased fashion. These results suggest that hippocampal ripples are widely but selectively associated with the subthreshold dynamics of the membrane potentials of PPC neurons even though there is no monosynaptic connectivity between the hippocampus and the PPC.

COMPARATIVE STUDY ON PERFORMANCE OF VARIOUS ENVIRONMENTAL RADIATION MONITORS.

After the Fukushima Daiichi Nuclear Power Plant accident, the radiation dose for first responders was not evaluated accurately due to lack of the monitoring data. It has been important to evaluate a radiation dose for workers in emergency response at a nuclear accident. In this study, a new device which can evaluate both of external and internal exposure doses was developed and the performance of various environmental radiation monitors including commercially available monitors were tested and compared from the viewpoint of an environmental monitoring at emergency situation. Background counts of the monitors and the ambient dose equivalent rate were measured in Fukushima Prefecture. The detection limit for beta particles was evaluated by the method of ISO11929. The sensitivity for gamma-rays of the dust monitor using a ZnS(Ag) and a plastic scintillator was high, but that of the external exposure monitor using a silicon photodiode with CsI(Tl) crystal was relatively low. The detection limit ranged 190-280 Bq m-3 at 100 µSv h-1, exceeding the detection limit of 100 Bq m-3 in the minimum requirement by the National Regulation Authority in Japan. Use of the shielding with lead is necessary to achieve the minimum requirement. These results indicate that the dust monitor using a ZnS(Ag) scintillator and a plastic scintillator is suitable for the external exposure monitor and the developed internal exposure monitor is for the internal exposure monitor at emergency situation among the evaluated monitors. In the future study, the counting efficiency, the relative uncertainty and the performance of the detection for alpha particles will be evaluated, and it will be considered which type of a monitor is suitable after taking the portability into account.

Microscopic Investigation into the Electric Field Effect on Proximity-Induced Magnetism in Pt.

Electric field effects on magnetism in metals have attracted widespread attention, but the microscopic mechanism is still controversial. We experimentally show the relevancy between the electric field effect on magnetism and on the electronic structure in Pt in a ferromagnetic state using element-specific measurements: x-ray magnetic circular dichroism (XMCD) and x-ray absorption spectroscopy (XAS). Electric fields are applied to the surface of ultrathin metallic Pt, in which a magnetic moment is induced by the ferromagnetic proximity effect resulting from a Co underlayer. XMCD and XAS measurements performed under the application of electric fields reveal that both the spin and orbital magnetic moments of Pt atoms are electrically modulated, which can be explained not only by the electric-field-induced shift of the Fermi level but also by the change in the orbital hybridizations.

An introduction to optical super-resolution microscopy for the adventurous biologist.

Ever since the inception of light microscopy, the laws of physics have seemingly thwarted every attempt to visualize the processes of life at its most fundamental, sub-cellular, level. The diffraction limit has restricted our view to length scales well above 250 nm and in doing so, severely compromised our ability to gain true insights into many biological systems. Fortunately, continuous advancements in optics, electronics and mathematics have since provided the means to once again make physics work to our advantage. Even though some of the fundamental concepts enabling super-resolution light microscopy have been known for quite some time, practically feasible implementations have long remained elusive. It should therefore not come as a surprise that the 2014 Nobel Prize in Chemistry was awarded to the scientists who, each in their own way, contributed to transforming super-resolution microscopy from a technological tour de force to a staple of the biologist's toolkit. By overcoming the diffraction barrier, light microscopy could once again be established as an indispensable tool in an age where the importance of understanding life at the molecular level cannot be overstated. This review strives to provide the aspiring life science researcher with an introduction to optical microscopy, starting from the fundamental concepts governing compound and fluorescent confocal microscopy to the current state-of-the-art of super-resolution microscopy techniques and their applications.

Angiogenic conditioning of peripheral blood mononuclear cells promotes fracture healing.

OBJECTIVES: The objective of this study was to investigate the therapeutic effect of peripheral blood mononuclear cells (PBMNCs) treated with quality and quantity control culture (QQ-culture) to expand and fortify angiogenic cells on the acceleration of fracture healing. METHODS: Human PBMNCs were cultured for seven days with the QQ-culture method using a serum-free medium containing five specific cytokines and growth factors. The QQ-cultured PBMNCs (QQMNCs) obtained were counted and characterised by flow cytometry and real-time polymerase chain reaction (RT-PCR). Angiogenic and osteo-inductive potentials were evaluated using tube formation assays and co-culture with mesenchymal stem cells with osteo-inductive medium in vitro. In order to evaluate the therapeutic potential of QQMNCs, cells were transplanted into an immunodeficient rat femur nonunion model. The rats were randomised into three groups: control; PBMNCs; and QQMNCs. The fracture healing was evaluated radiographically and histologically. RESULTS: The total number of PBMNCs was decreased after QQ-culture, however, the number of CD34+ and CD206+ cells were found to have increased as assessed by flow cytometry analysis. In addition, gene expression of angiogenic factors was upregulated in QQMNCs. In the animal model, the rate of bone union was higher in the QQMNC group than in the other groups. Radiographic scores and bone volume were significantly associated with the enhancement of angiogenesis in the QQMNC group. CONCLUSION: We have demonstrated that QQMNCs have superior potential to accelerate fracture healing compared with PBMNCs. The QQMNCs could be a promising option for fracture nonunion.Cite this article: K. Mifuji, M. Ishikawa, N. Kamei, R. Tanaka, K. Arita, H. Mizuno, T. Asahara, N. Adachi, M. Ochi. Angiogenic conditioning of peripheral blood mononuclear cells promotes fracture healing. Bone Joint Res 2017;6: 489-498. DOI: 10.1302/2046-3758.68.BJR-2016-0338.R1.

Following the stability of amphiphilic nanoaggregates by using intermolecular energy transfer.

An intermolecular energy transfer system is developed for studying the stability of nanoaggregate(s) (NAs) in complex solution and cell culture by one- and two-photon fluorescence microscopy and optical imaging. The system allows facile addition of one or more tumor targeting molecules, one of which is exemplified here. NAs functionalized with an MRI and optical probe, with and without folic acid, remain stable in fetal bovine serum for at least 4 hours. HeLa cell cultures showed a clear difference between NAs non-targeted and targeted to folate receptors, with both NAs appearing to be taken up by the cells through different mechanisms. An MRI relaxivity, r1, of 9 mM-1 s-1 at 310 K and 1.4 T was measured associated with the increased rotational correlation time of the NAs. These NAs may have application in the targeted drug delivery of hydrophobic drugs such as doxorubicin (DOX).

The AURKA/TPX2 axis drives colon tumorigenesis cooperatively with MYC.

BACKGROUND: The MYC oncogene has long been established as a central driver in many types of human cancers including colorectal cancer. However, the realization of MYC-targeting therapies remains elusive; as a result, synthetic lethal therapeutic approaches are alternatively being explored. A synthetic lethal therapeutic approach aims to kill MYC-driven tumors by targeting a certain co-regulator on the MYC pathway. PATIENTS AND METHODS: We analyzed copy number and expression profiles from 130 colorectal cancer tumors together with publicly available datasets to identify co-regulators on the MYC pathway. Candidates were functionally tested by in vitro assays using colorectal cancer and normal fibroblast cell lines. Additionally, survival analyses were carried out on another 159 colorectal cancer patients and public datasets. RESULTS: Our in silico screening identified two MYC co-regulator candidates, AURKA and TPX2, which are interacting mitotic regulators located on chromosome 20q. We found the two candidates showed frequent co-amplification with the MYC locus while expression levels of MYC and the two genes were positively correlated with those of MYC downstream target genes across multiple cancer types. In vitro, the aberrant expression of MYC, AURKA and TPX2 resulted in more aggressive anchorage-independent growth in normal fibroblast cells. Furthermore, knockdown of AURKA or TPX2, or treatment with an AURKA-specific inhibitor effectively suppressed the proliferation of MYC-expressing colorectal cancer cells. Additionally, combined high expression of MYC, AURKA and TPX2 proved to be a poor prognostic indicator of colorectal cancer patient survival. CONCLUSIONS: Through bioinformatic analyses and experiments, we proposed TPX2 and AURKA as novel co-regulators on the MYC pathway. Inhibiting the AURKA/TPX2 axis would be a novel synthetic lethal therapeutic approach for MYC-driven cancers.

Molecular and clinical characterization of glucokinase maturity-onset diabetes of the young (GCK-MODY) in Japanese patients.

AIMS: To investigate the molecular and clinical characteristics of the largest series of Japanese patients with glucokinase maturity-onset diabetes of the young (GCK-MODY), and to find any features specific to Asian people. METHODS: We enrolled 78 Japanese patients with GCK-MODY from 41 families (55 probands diagnosed at the age of 0-14 years and their 23 adult family members). Mutations were identified by direct sequencing or multiplex ligation-dependent probe amplification of all exons of the GCK gene. Detailed clinical and laboratory data were collected on the probands using questionnaires, which were sent to the treating physicians.　Data on current clinical status and HbA1c levels were also collected from adult patients. RESULTS: A total of 35 different mutations were identified, of which seven were novel. Fasting blood glucose and HbA1c levels of the probands were ≤9.3 mmol/l and ≤56 mmol/mol (7.3%), respectively, and there was considerable variation in their BMI percentiles (0.4-96.2). In total, 25% of the probands had elevated homeostatic assessment of insulin resistance values, and 58.3% of these had evidence of concomitant Type 2 diabetes in their family. The HbA1c levels for adults were slightly higher, up to 61 mmol/mol (7.8%). The incidence of microvascular complications was low. Out of these 78 people with GCK-MODY and 40 additional family members with hyperglycaemia whose genetic status was unknown, only one had diabetic nephropathy. CONCLUSIONS: The molecular and clinical features of GCK-MODY in Japanese people are similar to those of other ethnic populations; however, making a diagnosis of GCK-MODY was more challenging in patients with signs of insulin resistance.

Altered cerebellum development and dopamine distribution in a rat genetic model with congenital hypothyroidism.

Thyroid hormones play crucial roles in the development and functional maintenance of the central nervous system. Despite extensive studies of the neural function of thyroid hormones, little is known about the effects of hypothyroidism on behavioural traits and the mechanisms underlying such effects. In the present study, we report an investigation of congenitally hypothyroid mutant rdw rats, revealing a novel function of thyroid hormones in the central nervous system. The rdw rats were subjected to behavioural analyses such as the rotarod test, open field test and circadian activity measurement. To determine the cause of behavioural disorders, cerebellar morphogenesis was examined by immunohistochemical analysis, and the axonal transport of dopamine in the nigrostriatal pathway was analysed by high-performance liquid chromatography and western blotting. The effects of thyroxine administration to the rdw rats were examined by behavioural analysis. The rdw rats showed severe impairment of motor coordination and balance. This could be explained by the fact that the rats showed severe retardation of cerebellar morphogenesis, which correlates with the small somata and poor dendritic arborisation of Purkinje cells and retarded migration of granule cells particularly during the first two postnatal weeks. Moreover, the rdw rats showed hypoactivity, characterised by decreased circadian locomotor activity. After weaning, thyroxine administration improved the dwarfism in rdw rats but had no effect on cerebellar function. In addition, the rdw rats showed anxiety and depression intrinsically to novel surroundings. Interestingly, the rdw rats showed high levels of dopamine in the substantia nigra and low levels in the striatum, an important centre for the coordination of behaviour. Furthermore, low levels of tubulin in the striatum were detected, indicating the aberrant axonal transport of dopamine in the nigrostriatal pathway as a result of the reduced delivery of microtubules. These findings indicate an important function of thyroid hormones in cerebellar formation and in the regulation of axonal transport of dopamine. Moreover, rdw rats will be useful for studies of brain function and behavioural disorders in congenital hypothyroidism.

Mechanical responses and stress fluctuations of a supercooled liquid in a sheared non-equilibrium state.

A steady shear flow can drive supercooled liquids into a non-equilibrium state. Using molecular dynamics simulations under steady shear flow superimposed with oscillatory shear strain for a probe, non-equilibrium mechanical responses are studied for a model supercooled liquid composed of binary soft spheres. We found that even in the strongly sheared situation, the supercooled liquid exhibits surprisingly isotropic responses to oscillating shear strains applied in three different components of the strain tensor. Based on this isotropic feature, we successfully constructed a simple two-mode Maxwell model that can capture the key features of the storage and loss moduli, even for highly non-equilibrium state. Furthermore, we examined the correlation functions of the shear stress fluctuations, which also exhibit isotropic relaxation behaviors in the sheared non-equilibrium situation. In contrast to the isotropic features, the supercooled liquid additionally demonstrates anisotropies in both its responses and its correlations to the shear stress fluctuations. Using the constitutive equation (a two-mode Maxwell model), we demonstrated that the anisotropic responses are caused by the coupling between the oscillating strain and the driving shear flow. Due to these anisotropic responses and fluctuations, the violation of the fluctuation-dissipation theorem (FDT) is distinct for different components. We measured the magnitude of this violation in terms of the effective temperature. It was demonstrated that the effective temperature is notably different between different components, which indicates that a simple scalar mapping, such as the concept of an effective temperature, oversimplifies the true nature of supercooled liquids under shear flow. An understanding of the mechanism of isotropies and anisotropies in the responses and fluctuations will lead to a better appreciation of these violations of the FDT, as well as certain consequent modifications to the concept of an effective temperature.

Mast cells promote the growth of Hodgkin's lymphoma cell tumor by modifying the tumor microenvironment that can be perturbed by bortezomib.

Hodgkin's lymphoma is frequently associated with mast cell infiltration that correlates directly with disease severity, but the mechanisms underlying this relationship remain unclear. Here, we report that mast cells promote the growth of Hodgkin's tumor by modifying the tumor microenvironment. A transplantation assay shows that primary murine mast cells accelerate tumor growth by established Hodgkin's cell lines, and promote marked neovascularization and fibrosis. Both mast cells and Hodgkin's cells were sensitive to bortezomib, but mast cells were more resistant to bortezomib. However, bortezomib inhibited degranulation, PGE(2)-induced rapid release of CCL2, and continuous release of vascular endothelial growth factor-A from mast cells even at the concentration that did not induce cell death. Bortezomib-treated mast cells lost the ability to induce neovasculization and fibrosis, and did not promote the growth of Hodgkin tumor in vivo. These results provide further evidence supporting causal relationships between inflammation and tumor growth, and demonstrate that bortezomib can target the tumor microenvironment.

Glucocorticoid attenuates brain-derived neurotrophic factor-dependent upregulation of glutamate receptors via the suppression of microRNA-132 expression.

Brain-specific microRNAs (miRs) may be involved in synaptic plasticity through the control of target mRNA translation. Brain-derived neurotrophic factor (BDNF) also contributes to the regulation of synaptic function. However, the possible involvement of miRs in BDNF-regulated synaptic function is poorly understood. Importantly, an increase in glucocorticoid levels and the downregulation of BDNF are supposed to be involved in the pathophysiology of depressive disorders. Previously, we reported that glucocorticoid exposure inhibited BDNF-regulated synaptic function via weakening mitogen-activated protein kinase/extracellular signal-regulated kinase1/2 (MAPK/ERK) and/or phospholipase C-gamma (PLC-gamma) intracellular signaling in cultured neurons [Kumamaru et al (2008) Mol Endocrinol 22:546-558; Numakawa et al (2009) Proc Natl Acad Sci U S A 106:647-652]. Therefore, in this study, we investigate the possible influence of glucocorticoid on BDNF/miRs-stimulated biological responses in cultured cortical neurons. Significant upregulation of miR-132 was caused by BDNF, although miR-9, -124, -128a, -128b, -134, -138, and -16 were intact. Transfection of exogenous ds-miR-132 induced marked upregulation of glutamate receptors (NR2A, NR2B, and GluR1), suggesting that miR-132 has a positive effect on the increase in postsynaptic proteins levels. Consistently, transfection of antisense RNA to inhibit miR-132 function decreased the BDNF-dependent increase in the expression of postsynaptic proteins. U0126, an inhibitor of the MAPK/ERK pathway, suppressed the BDNF-increased miR-132, suggesting that BDNF upregulates miR-132 via the MAPK/ERK1/2 pathway. Interestingly, pretreatment with glucocorticoid (dexamethasone, DEX) reduced BDNF-increased ERK1/2 activation, miR-132 expression, and postsynaptic proteins. We demonstrate that the exposure of neurons to an excess glucocorticoid results in a decrease in the BDNF-dependent neuronal function via suppressing miR-132 expression.

Carbon-ion radiotherapy: clinical aspects and related dosimetry.

The features of relativistic carbon-ion beams are attractive from the viewpoint of radiotherapy. They exhibit not only a superior physical dose distribution but also an increase in biological efficiency with depth, because energy loss of the beams increases as they penetrate the body. This paper reviews clinical aspects of carbon-beam radiotherapy using the experience at the National Institute of Radiological Sciences. The paper also outlines the dosimetry related to carbon-beam radiotherapy, including absolute dosimetry of the carbon beam, neutron measurements and radiation protection measurements.

Characterization of internal structure of the nucleolar organizing region in rice (Oryza sativa L.).

The genomic sequence of the nucleolar organizing region (NOR) in rice has not been characterized fully because of the difficulty in assembling repetitive sequences in silico. Here, we used a cytogenetic approach to elucidate the internal structure of the NOR. We detected one locus of the 18S rRNA genes on 'Nipponbare' chromosome. High-resolution fiber-fluorescence in situ hybridization made it possible to visualize each rRNA gene unit in the array of rRNA genes. Signals of pairs of alternating 18S and 25S rRNA genes were detected uniformly along the DNA fiber. Intergenic spacers were shorter than the transcribed region. The rRNA genes were infrequently interrupted. These and previous results based on the sequencing of genome fragments, PCR analysis and Southern blot hybridization suggest that the internal region of the NOR is filled with a uniform array of canonical rRNA genes separated by spacers carrying three 254-bp sub-repeats.

Oxydemeton-methyl resistance, mechanisms, and associated fitness cost in green peach aphids (Hemiptera: Aphididae).

Susceptibility to oxydemeton-methyl and imidacloprid, and the inhibitory effects of oxydemeton-methyl and some organophosphate compounds on acetylcholinesterase (AChE) and carboxylesterase activity were studied in two populations (Karaj and Rasht) of green peach aphids, Myzus persicae (Sulzer). Results show that the Karaj population was resistant to oxydemeton-methyl but susceptible to imidacloprid. The esterase activity of the resistant and susceptible populations suggests that one of the resistance mechanisms to oxydemeton-methyl was esterase-based. The inhibition assay shows that the AChE of the Karaj population is less sensitive to oxydemeton-methyl and paraoxon derivatives. Regarding the paraoxon derivatives, the smaller paraoxon side chain is more potent against the modified AChE than against the AChE from the susceptible strain. Fertility life table parameters of green peach aphid populations resistant and susceptible to oxydemeton-methyl also were studied under laboratory conditions. The standard errors of the population growth parameters were calculated using the Jackknife method. Results showed that susceptible strain exhibits a significantly higher r(m) than the resistant strain, probably because the resistant strain had a higher generation time than the susceptible strain. These results suggested that the resistant Karaj strain may be less fit than the susceptible strain.

Monitoring the bacterial community during fermentation of sunki, an unsalted, fermented vegetable traditional to the Kiso area of Japan.

AIMS: To investigate the microbial community in sunki, an indigenous, unsalted, fermented vegetable, made from the leaves of red beet. METHODS AND RESULTS: Fermenting samples were collected at 1- to 2-day intervals from four houses and investigated by culture-dependent and culture-independent techniques. PCR-Denaturing-Gradient-Gel-Electrophoresis profiles indicated that the bacterial community was stable and Lactobacillus delbrueckii, Lact. fermentum and Lact. plantarum were dominant during the fermentation. This result agreed well with that obtained by the culturing technique. Moulds, yeasts or bacteria other than lactic acid bacteria (LAB) were not detected. CONCLUSIONS: The bacterial community was stable throughout the fermentation, and Lact. delbrueckii, Lact. fermentum and Lact. plantarum were dominant. The acidic pH and lactic acid produced by LAB probably preserve the sunki from spoilage. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the use of both culture-dependent and culture-independent techniques to study the bacterial community in sunki. A combination of culture-dependent and culture-independent techniques is necessary for the analysis of complex microbial communities.

Clinical features of chromosome 16q22.1 linked autosomal dominant cerebellar ataxia in Japanese.

Chromosome 16q22.1-linked autosomal dominant cerebellar ataxia (16q-ADCA) is strongly associated with a substitution in the puratrophin-1 gene. This locus overlaps with spinocerebellar ataxia type 4 (SCA4) which shows ataxia with prominent sensory axonal neuropathy. We found that 16q-ADCA is a common ADCA subtype in the Tohoku District of Japan. The clinical feature of Japanese 16q-ADCA is characterized as late-onset pure cerebellar ataxia.

Calreticulin as a new histone binding protein in mitotic chromosomes.

Calreticulin (CRT) is a multifunctional Ca(2+)-binding protein that mainly functions in the endoplasmic reticulum as a molecular chaperone for newly synthesized proteins. Recently we reported the protein composition of human metaphase chromosomes (Uchiyama et al., 2004), which included CRT. Here we describe new characteristics of CRT in vitro as well as its localization on the surface of metaphase chromosomes in vivo. CRT was detected in the chromosomal fraction by Western blotting and its binding partners were identified as core and linker histones by ligand overlay assay. Surface plasmon resonance sensor analyses revealed that CRT is bound to chromatin fibers. Moreover, we found that CRT has both supercoiling activity, which assists core histone assembly into chromatin fibers, and binding ability to histone H2A/H2B dimers and histone H3/H4 tetramers. Unlike the chromosome scaffold proteins, indirect immunofluorescent staining revealed that CRT is located on the surface of metaphase chromosomes. These results suggest that CRT plays a role which involves chromatin dynamics on the surface of mitotic chromosomes.

Serial changes in plasma brain natriuretic peptide concentration at the infarct and non-infarct sites in patients with left ventricular remodelling after myocardial infarction.

OBJECTIVES: To clarify the role of infarct and non-infarct sites on left ventricular (LV) remodelling after myocardial infarction by measuring brain natriuretic peptide (BNP) from each site. METHODS AND RESULTS: BNP from the aorta and the anterior interventricular vein (AIV) was measured in 45 patients with first anterior myocardial infarction at one, six, and 18 months. The LV was significantly dilated (> 10 ml/m(2) of end diastolic volume from one to 18 months) in 20 patients (remodelling (R) group) but not in 25 others (non-remodelling (NR) group). Patient characteristics and LV functions did not differ significantly at one month but plasma BNP concentration was higher in group R than in group NR (336 (288) v 116 (106) pg/ml, p < 0.01), predicting the degree of LV dilatation. The difference in BNP concentration between the aortic root and AIV (DeltaBNP), reflecting BNP secreted from the infarct site, did not differ at one month. In both groups BNP and DeltaBNP significantly decreased from one to six months (p < 0.05) and decreased from six months to 18 months, but the change was not significant. BNP and DeltaBNP were significantly higher in group R than in group NR after six months, when LV dilatation was not evident in both groups. CONCLUSION: Enhanced BNP secretion at one month in the non-infarct and infarct ventricular sites predicts subsequent LV dilatation (that is, remodelling). The slower process of LV remodelling decreased BNP secretion at both sites. Thus, BNP concentration should be useful for monitoring ventricular remodelling after infarction.