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Summary Platelet transfusions are critical for severe thrombocytopenia but depend on blood donors. The shortage of donors and the potential of universal HLA-null platelet products have stimulated research on the ex vivo differentiation of human pluripotent stem cells (hPSCs) to platelets. We recently established expandable immortalized megakaryocyte cell lines (imMKCLs) from hPSCs by transducing MYC, BMI1, and BCL-XL (MBX). imMKCLs can act as cryopreservable master cells to supply platelet concentrates. However, the proliferation rates of the imMKCLs vary with the starting hPSC clone. In this study, we reveal from the gene expression profiles of several MKCL clones that the proliferation arrest is correlated with the expression levels of specific cyclin-dependent kinase inhibitors. Silencing CDKN1A and p53 with the overexpression of MBX was effective at stably inducing imMKCLs that generate functional platelets irrespective of the hPSC clone. Collectively, this improvement in generating imMKCLs should contribute to platelet industrialization and platelet biology., Graphical abstract, Highlights • Expression of CDKIs is negatively correlated with MKCL proliferation potential • Silencing CDKN1A and p53 improved the induction efficiency of imMKCLs, imMKCLs derived from human iPSCs potentially overcome the many issues depending on blood donations for platelet transfusions. However, the rate of stable proliferation depends on the starting PSC clone. In this article, Takayama and colleagues show that silencing CDKN1A and p53 simultaneously with the overexpression of MYC/BMI1/BCL-XL dramatically increased the success rate of imMKCL establishment irrespective of the clone.

Pyrvinium pamoate (PP), a classical anthelminthic, potently inhibited proliferation and STAT3 Tyr705 phosphorylation of human myeloma (U266B1 and PCM6)/erythroleukemia (HEL 92.1.7) cells. PCM6 cell proliferation was markedly impaired by STAT3 siRNA knockdown. PP inhibited ATP production/O(2) consumption in those three cells and mitochondrial respiratory complex (I+III, but not II+III) activity in mouse kidney mitochondrial fractions. PP inhibition of ATP production, STAT3 Tyr705 phosphorylation, and proliferation was absent in mitochondrial DNA-deficient HEL 92.1.7-ρ(0) cells. Moreover, PP acted synergistically with dexamethasone to inhibit PCM6 cell proliferation. In conclusion, we identified PP as a potential anticancer drug directed against mitochondrial respiratory complex I/STAT3.

The lagena of pigeons is a unique organ and it is concluded that it is a key element in the magnetic sensor system of pigeons and migrating birds. The lagenal otolith in pigeons contains more iron than saccular and utricular otoliths. The function of the lagena of pigeons was clarified because the homing ability of pigeons was largely disrupted after unilateral lagenal nerve section and attachment of magnetic balls with a magnetic field strength under 5 Gauss. The lagena of pigeons may have a navigational function as a geomagnetic sensor.Otoliths of many kinds of fishes and birds were analyzed.The otoliths of fish and birds were analyzed using synchrotron X-ray fluorescence analysis. Behavioral experiments concerning homing ability of pigeons were done by sectioning their lagenal nerves or interfering with the function of the lagena using a magnet. Twenty-one birds were treated in this way and 30 birds from the same loft of racing pigeons were used as controls.By comparing the compositions of the three different kinds of otoliths among several species of sea fish and birds, it was found that the saccular and utricular otoliths contain scarcely detectable levels of iron but that iron is present in significant quantities in the lagenal otoliths of the birds and sea fish. The results of homing tests clearly revealed a magnetic influence on the function of the lagena in terms of navigation ability of pigeons. The treated pigeons were either lost or significantly delayed while the controls returned within 30 min of release.

To examine the measuring accuracy for river discharge with floats, flood discharges obtained with float and ADCP measurements have been compared. Since the ADCP can measure vertical velocity distributions and water depth simultaneously, it may be one of accurate discharge measurements under various flow conditions except high flow conditions (e. g., u>3m/s). We also try to improve the accuracy of the float measurement with special attention to evaluations of lateral and vertical velocity distributions. The observed results indicate that the measured discharges obtained with floats are relatively larger than those with the ADCPs. The modifications of the lateral and vertical velocity profiles presented here may improve appreciably the measuring accuracy for flood discharge with floats.

A new method that optimizes a control map of hydrocarbon (HC) addition to diesel exhaust gas for HC type Selective Catalytic Reduction DeNO x catalyst system has been developed. This method comprises a numerical model and a new optimization technique. The numerical model is capable of predicting performance of HC DeNO x with diesel fuel as a supplemental reductant. The control map of HC addition was optimized with the evolutionary programming based on the evolution of living things. As a result, NO x conversion with the optimized control map was found to be greater by 21% than that with the conventional control on Japanese 10–15 mode.

The ultrastructural differences between the inner ears of Japanese red-bellied newts (Cynops pyrrhogaster) and black-spotted pond frogs (Rana nigromaculata) were investigated. Scanning electron microscopic observations showed apparent morphological differences in the shape of the ampulla cristae and the localization of the striola in the saccular macula. There were differences in the length of the kinocilia of the sensory hairs in each sensory region. In addition, the diameters of the bundles of stereocilia differed between the two species: the bundles of stereocilia in the semicircular cristae were thicker in frogs than in newts, while those of the utricular and lagenal maculae were thicker in newts than in frogs.

Maternal sound stress (800 Hz; 77 dB, every other minute for 15 min/day, from day 10 to 18 of gestation), combined with forced swimming stress (15 min/day), was found to cause potentiation of sound-induced loss of locomotor activity, referred to as emotional behavior, of male offspring, but not that of female offspring, at 4 weeks of age. Maternal stress also caused an increase in the total number of errors by male, but not female offspring in the water-maze test at 6 weeks of age. These effects of stress on emotional behavior and learning behavior were abolished when dams were pretreated with buspirone (30 min before the stress, from day 8 to 18 of gestation). Thus, prenatal stress might have sex-dependent effects on emotional behavior and learning ability of neonatal rats.

The mystery of the homing ability of pigeons has been the subject of much interest and it is widely believed that information from the earth's magnetic field may be involved. However, no specific magnetic sensory organ has yet been identified. The recent finding of magnetic materials in the lagenal otolith of fish and birds raises the possibility that these structures may be key elements in the elusive magnetic sensory system. For the elemental analysis of materials X-ray fluorescence using synchrotron radiation is one of the most powerful techniques available and was used in this study for analysis of the otoliths. By comparing the compositions of the three different kinds of otoliths among several species of sea fish and birds, we found that the saccular and utricular otoliths rarely contain detectable levels of iron but that iron is present in significant quantities in the lagenal otoliths of the birds. The lagenal otolith comprises tiny magnetic particles of low inertia that are displaced by imposed magnetic fields, providing the animal with geomagnetic sensory input, from which the brain would infer navigational information.

Three ganglioside molecular species, HPG-8, HPG-3, and HPG-1, have been obtained from the polar fraction of the chloroform/methanol extract of the sea cucumber Holothuriapervicax. The structures of these gangliosides have been determined on the basis of chemical and spectroscopic evidence. They represent new ganglioside molecular species. HPG-8 is a sulfated monosialo-ganglioside, while HPG-3 and HPG-1 are disialo-gangliosides possessing 24-linked tandem-type disialosyl moieties. These three gangliosides qualitative neuritogenic activity toward the rat pheochromocytoma cell line, PC-12 cells.

In vivo electrophysiological and patch-clamp studies were performed to determine whether 20-hydroxyecdysone (20-HE), a neurosteroid, influenced neuronal activities of the medial vestibular nucleus (MVN) using chloral hydrate-anesthetized rats and dissociated MVN neurons, respectively. Single neuronal activities of MVN were extracellularly recorded with a glass-insulated silver wire microelectrode attached along a seven-barreled micropipette. Each micropipette was filled with 20-HE, glutamate, bicuculline or 2 M NaCl. These chemicals were applied microiontophoretically to the immediate vicinity of the target neurons. Microiontophoretically applied 20-HE (20-80 nA) dose-dependently decreased rotation-induced firings of both type I and II neurons, which were identified according to their responses to horizontal sinusoidal rotations. Microiontophoretically applied bicuculline, a GABAA receptor antagonist, inhibited 20-HE-induced decreases in neuronal firing of MVN. These findings suggest that 20-HE potentiates the action of GABA, probably by acting directly on the GABAA receptor of MVN neurons. In addition, microiontophoretically applied 20-HE decreased firings induced by glutamate in both type I and II neurons. This decrease by 20-HE was also antagonized with bicuculline. Furthermore, the effects of 20-HE on GABA-induced currents in acutely dissociated MVN neurons were investigated using the whole-cell patch-clamp technique. Under voltage-clamp conditions, GABA (10 microM)-induced currents were potentiated in the presence of 20-HE (100 microM). These findings suggest that 20-HE inhibits MVN neurons by acting on the modulatory site on GABA receptor-ion channel complexes to potentiate GABA inhibition.

牛ふん堆肥の腐熟過程における特徴を調べるために,牛ふんにパーライトまたはオガクズを混合し,2通りの方法で堆肥化実験を行った.ひとつは強制通気を行う機械撹拌堆肥化,もうひとつは強制通気を行わない堆積堆肥化である.堆肥化のための副資材として,生化学的反応に不活性な無機物のパーライトを用いることによって,牛ふんそれ自身の腐熟特性を調べることができた.堆肥化過程において,有機物,全炭素,ヘミセルロース,セルロースの含有量は減少し,一方灰分,全窒素,リグニンの含有量は相対的に増加し,その結果C/N比は減少した.このような変化は腐熟の指標として役立っものであろう.牛ふんーオガクズ混合物の堆積堆肥化では,通気性が良くなかったので,腐熟が効率的に進行しなかった.通気条件の改善については,パーライトのほうがオガクズよりも効果的とみなされた.

In order to clarify the mechanism of otoconia formation, the pathway of calcium transport in the utricular supporting cells was investigated. Potassium pyroantimonate (PA) precipitation, which indicates the localization of calcium ions, was seen not only in otoconia but also in supporting cell cytoplasm. In the latter, deposits of PA were detected in the secretory granules and endoplasmic reticulum (ER). These deposits were not present in cells pretreated with ethylene glycol-O,O,-bis (2-aminoethyl)-N,N,N',N'-tetra-acetic acid (EGTA). Exposure of the supporting cells to streptomycin sulfate (SM) increased the number of lysosomes. These lysosomes contained many small deposits of PA. The remaining granules and ER in cytoplasm also contained small deposits of PA. The findings suggest that otoconia are formed by the vestibular supporting cells in which calcium ions might be transported via ER-secretory granule-lysosome-cytoplasmic protrusion.

Formation and fate of abnormal (giant) otoconia of the guinea pig following streptomycin intoxication were investigated using scanning electron microscopy. The giant otoconia formed as multifaceted morphology in their early developmental period. They grew up the the transitional type and finally to the cylindrical type. It has been suggested that the giant otoconia found following streptomycin intoxication may be formed mainly by dissolution of normal otoconia due to the loss of environmental calcium, followed by recrystallization as giant crystals. These phenomena seemed to be closely related to the otoconial dynamics which may regulate calcium ion homeostasis of the endolymph.

The calcium ion turnover into otoconia of adult guinea pigs was investigated by the use of different fluorochromes, i.e. tetracycline (TC), calcein (Cal) and alizarin complexone (AC). The administration of all fluorochromes induced yellow (TC), green (Cal) and red (AC) fluorescence on the outer surface of otoconia, respectively. Sequential polychromatic labeling with AC followed by TC induced only red fluorescence after 1 week administration of TC, combined fluorescence after 2 weeks administration of TC and only yellow fluorescence after more than 3 weeks of administration of TC. The otoconia labeled with both AC and TC showed red fluorescence on the side faces of otoconia and yellow fluorescence on the terminal faces. These results may indicate the existence of a dynamic exchange of calcium ions in the otoconia and this exchange is mainly restricted to the terminal faces. The sequential polychromatic labeling may thus be of great use for the further investigation of the calcium dynamics of otoconia.

The effects of streptomycin on the otoconial layer of the adult guinea pig were investigated using scanning electron microscopy. Administration of streptomycin induced a reduction of otoconia with formation of giant otoconia. These phenomena reached their maximum 4 weeks after the cessation of streptomycin intoxication in the utricle and between 2 and 6 weeks after in the saccule. Otoconia began to recover in the utricle 4 weeks and in the saccule 6 weeks after the cessation of streptomycin intoxication with an increasing number of small otoconia. Eight or 10 weeks after the cessation of streptomycin intoxication, the otoconial layer has completely recovered in both the utricle and saccule.

New antimicrobial substances against Streptomyces scabies, rosmic acid and rosmanol-related compounds, were isolated from leaves of rosemary (Rosmarinus officinalis L.). The active compounds were separated in pure form by a combination of gel permeation chromatography and reverse-phase HPLC. Their structures were determined by two-dimensional NMR experiments and HR-MS analyses.

We have proposed a method of product safety assessment by using hazard based FTA in our previous study. In the study, a risk level for a product was calculated by using FTA, and introduced into a manufacturer management system which set criteria for a design and shipment stage in manufacturing. A risk level for a product has a certain distribution in the process of manufacturing due to components quality and their reliability distribution. It is useful to find the probability distribution form of a risk level for a product so that a risk level can be used as criteria of design quality target and shipment condition in a manufacturing management system. In this paper we focus on a failure rate for a component of a product which is the dominant cause of unsafe incidents of the product. The NITE database of incident reports was analyzed for an electric fan to calculate a component failure rate. The failure rate is composed of an accidental and wear-out failure modes. Assuming that the probability distribution is identical at the crossing point of two modes, we estimate a failure rate at the crossing point. We compare the distribution with the normal, Weibull, and log-normal distribution by correlation coefficient and choose the distribution with the maximum correlation. Furthermore we apply the probability distribution to the failure rate calculated by IEC/TR62380 and find that the probability of failure rate for a standard environment is 50% and severe environment failure rate becomes 70% of probability of the distribution that is obtained by the incident database analysis. The knowledge of probability distribution can help to set a risk level standard as criteria for a management system according to an environment of product usage.

The effects of streptomycin on the calcium ion turnover into otoconia of adult guinea-pigs investigated by the use of tetracycline. The oral administration of tetracycline induced the deposition of tetracycline (fluorescence) on the outer surface of otoconia indicating the existence of dynamic exchange and/or uptake of calcium ions in the otoconia. The significant finding is that streptomycin specifically interfered with calcium uptake into the otoconia which indicated that the decrease in calcium uptake caused by streptomycin may be closely related to the loss of otoconia as well as to a decrease in the calcium contents of otoconia. The decrease in calcium incorporation into otoconia caused by streptomycin was recovered within 6 weeks after the last injection of streptomycin. The number of otoconia with fluorescence in the central portion as well as their outer surface was increased. It is therefore suggested that the recovery of calcium uptake as well as new otoconial regeneration may play an important role for the recovery from loss of otoconia.

The canine nasal mucosa was studied by in vitro bioassay. Non-adrenergic, non-cholinergic (NANC) vasodilator response to transmural electrical stimulation was observed in the presence of guanethidine and atropine. This vasodilator response was abolished by NG-nitro- l -arginine ( l -NA) which is an inhibitor of nitric oxide (NO) formation but not by NG-nitro- d -arginine. The inhibitory effect of l -NA was partially reversed by treatment with l -arginine but not with d -arginine. The vasodilator response was significantly suppressed by tetrodotoxin. The present results indicate that NO may mediate neurogenic vasodilation in the canine nasal mucosa.

The cytoskeletal organization of the guinea pig vestibular sensory cells was investigated employing the saponin perfusion method using scanning electron microscopy as well as immunohistochemical technique. The intermediate filaments were demonstrated in the sensory cells. They were usually seen surrounding the nucleus and extending through the cytoplasm which connected the nuclear membrane to the plasma membrane, cuticular plate or other cytoorganelles. These findings may suggest that the intermediate filaments provide mechanical support to the cell and its nucleus. Microtubules were found mainly in the supranuclear portion of the cells and run parallel to the main axis of the cell body, which is directly connected to the cuticular plate. These findings may suggest that the microtubules provide mechanical support to the cell and may be closely related to the sensory cell transduction system. The actin filaments were densely packed in the site of the cuticular plate. They also distributed throughout the cytoplasm, which appeared as a dense network in the periphery region. These findings support the idea that the actin filaments could be responsible for sensory cell transduction as well as to give mechanical strength to the surface of the cell and enable the cell to change its shape and move.

The interaction of the endolymphatic flow between the anterior semicircular canal (asc) and the posterior semicircular canal (psc) was studied by three-dimensions in intact membranous labyrinths isolated from bull frogs. The asc and the psc nerves were cut 3mm from the ampullae to prevent neural interaction between the two semicircular canals. Two experiments were performed.Experiment 1: Changes of psc nerve potentials caused by stimulation of asc. The crus of the asc was cut beside the common osseous crus. A polyethylene tube connected with amicrosyringe was inserted into the crus of the asc. The syringe was driven with a micromanipulator for mechanical induction of endolymphatic flow into the asc. The psc and the asc nerve activities were recorded simultaneously with glass suction electrodes.Experiment 2: Changes of asc nerve potentials caused by stimulation of psc. The crus of the psc was cut beside the common osseous crus, a polyethylene tube was inserted, and the psc was stimulated in the same manner as was the asc in experiment 1.In experiment 1, psc nerve activity decreased when the asc was given an excitatory stimulus and increased when the asc was given an inhibitory stimulus. In experiment 2, asc nerve activity decreased when the psc was given an excitatory stimulus and increased when the psc was given an inhibitory stimulus. These results indicate that there is endolymphatic interaction between the asc and the psc.The volume of endolymphatic fluid flowing between the asc and the psc was estimated to be about 3% of the mechanical endolymphatic flow produced as a stimulus.The endolymphatic interaction probably enhances the sensitivity of the vestibular organs to various degrees of stimulation.

This paper presents a management system for the pursuit of product safety using hazard-based assessment during the product life cycle. A safety level index is initially calculated by hazard levels and safeguards reliability value using FTA diagram. Reliability of components are calculated by using IEC TR 62380. We also discuss an improvement scheme for product safety estimation. The management system can control manufacturers internal activities commonly through its functions of manufacturers department relating its product life cycle. For the manufacturers, the introduction of the management system to a company can contribute to set the common standardized level of product safety. Manufacturers are required to maintain to the same level of product safety over various categories of products.

Pyrvinium pamoate (PP) is an FDA-approved classical anthelmintic, but is now attracting particular attention as an anti-cancer drug after recent findings of its potent cytotoxicity against various cancer cell lines only during glucose starvation, as well as its anti-tumor activity against hypovascular pancreatic cancer cells transplanted in mice. The molecular mechanisms by which PP promotes such preferential toxicity against cancer cells are currently under extensive investigation. PP suppressed the NADH-fumarate reductase system that mediates a reverse reaction of the mitochondrial electron-transport chain complex II in anaerobic organisms such as parasitic helminthes or mammalian cells under tumor microenvironment-mimicking hypoglycemic/hypoxic conditions, thereby inhibiting efficient ATP production. PP also inhibited the unfolded protein response induced by glucose starvation, thereby inhibiting the proliferation of pancreatic cancer cells. Even under normoglycemic/normoxic conditions, PP suppressed the mitochondrial electron-transport chain complex I and thereby STAT3, inhibiting the proliferation of myeloma/erythroleukemia cells. Here, we review accumulating knowledge on its working mechanisms and evaluate PP as a novel anti-cancer drug that targets mitochondrial respiration.

Vestibular sensory cells were isolated from the utricular macula or cristae ampullares of the guinea pig by enzymatic and mechanical dissociation. The isolated cells were classified into three types: flask-shaped type I sensory cells, rod-shaped type II sensory cells and round-shaped supporting cells. The cilia of type I sensory cells in the crista ampullaris were longer than those in the corresponding cell type in the utricular macula, while no morphological differences of the cell bodies were noted between crista ampullaris and utricular macula. Isolated living vestibular cells have a motile capacity. After exposure to a hypo-osmotic medium, the type I sensory cells showed tilting of the hair bundle. This change in shape may be closely related to the active mechanical transduction control.