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Dynamic covalent bonds are useful tools in a wide range of applications. Although various reversible chemical reactions have been studied for this purpose, the requirement for harsh conditions, such as high temperature and low or high pH, to activate generally stable covalent bonds limits their potential applications involving biomolecules or household utilization. Here, we report the design, synthesis, characterization, and dynamic covalent bonding properties of 1,2-disubstituted 1,2-dihydro-1,2,4,5-tetrazine-3,6-dione (TETRAD). Hetero-Diels-Alder reactions of TETRAD with furan derivatives and their retro-reactions proceeded rapidly at room temperature under neutral conditions, enabling a chemically induced sol-gel transition system.

Cancer cells communicate with each other via exosomes in the tumor microenvironment. However, measuring trace amounts of proteins in exosomes is difficult, and thus the cancer stemness-promoting mechanisms of exosomal proteins have not been elucidated. In the present study, we attempted to quantify trace amounts of 78-kDa glucose-regulated protein (GRP78), which is involved in cancer progression, in exosomes released from cultured gastric cancer cells using an ultrasensitive ELISA combined with thio-NAD cycling. We also evaluated the cancer stemness-promoting effects by the application of high-GRP78-containing exosomes to cultured gastric cancer cells. The ultrasensitive ELISA enabled the detection of GRP78 at a limit of detection of 0.16 pg/mL. The stemness of cancer cultured cells incubated with high-GRP78-containing exosomes obtained from GRP78-overexpressed cells was increased on the basis of both an MTT assay and a wound healing assay. Our results demonstrated that the ultrasensitive ELISA has strong potential to measure trace amounts of proteins in exosomes. Further, exosomes with a high concentration of GRP78 promote the cancer stemness of surrounding cells. The technique for quantifying proteins in exosomes described here will advance our understanding of cancer stemness progression via exosomes.

In Saccharomyces cerevisiae, Avt4 exports neutral and basic amino acids from vacuoles. Previous studies have suggested that the GATA transcription factors, Gln3 and Gat1, which are key regulators that adapt cells in response to changes in amino acid status, are involved in the AVT4 transcription. Here, we show that mutations in the putative GATA-binding sites of the AVT4 promoter reduced AVT4 expression. Consistently, a chromatin immunoprecipitation (ChIP) assay revealed that Gat1-Myc13 binds to the AVT4 promoter. Previous microarray results were confirmed that gln3∆gat1∆ cells showed a decrease in expression of AVT1 and AVT7, which also encode vacuolar amino acid transporters. Additionally, ChIP analysis revealed that the AVT6 encoding vacuolar acidic amino acid exporter represents a new direct target of the GATA transcription factor. The broad effect of the GATA transcription factors on the expression of AVT transporters suggests that vacuolar amino acid transport is integrated into cellular amino acid homeostasis.

SummaryThe efficiency of in vitro embryo production technologies would be improved by the development of suitable non-invasive biomarkers that allow the selection of good quality cumulus–oocyte complexes (COCs). The present study used whole, single oocyte culture to investigate whether the expression levels of follicle-stimulating hormone receptor (FSHR), insulin-like factor 1 receptor (IGF1R) and three steroidogenesis-related enzymes (CYP11al, CYP19al and HSD3β) in cumulus cells reflected the developmental competence of COCs. Cumulus cells were collected from single COCs before maturation culture and relative mRNA levels were assessed using real-time PCR. The analysis indicated that mRNAs for FSHR, IGF1R, CYP11al and HSD3β were present at higher levels in cumulus cells from COCs that failed to form blastocysts compared with cumulus cells from COCs that formed blastocysts. Moreover, FSHR and IGF1R mRNA levels were positively correlated with those of genes for steroidogenesis-related enzymes. In conclusion, poor developmental competence of COCs was related to higher expression of FSHR, IGF1R, CYP11al and HSD3β in cumulus cells, which may indicate the advanced differentiation of cumulus cells into granulosa cells.

In this study, we aimed to elucidate the utility of transrectal ultrasonography (TRUS) and blood lactate concentration (bLac) measurement to diagnose cows with severe uterine torsion. We investigated the association of TRUS and bLac measurement with macroscopic findings on laparotomy for severe uterine torsion in nine cows. We found that an increased ultrasonographic cross-sectional thickness (15-25 mm) and multiple hypoechogenic areas corresponded to macroscopic vascular compromise in the uterus on laparotomy. In addition, bLac was elevated (≥5.0 mmol/l) in cows showing uterine necrosis on laparotomy. A combined diagnostic approach with TRUS and bLac measurement enables assessment of the uterine vascular status and has utility for selecting the treatment option (including laparotomy) and predicting the outcomes.

The efficiency of in vitro embryo production technologies would be improved by the development of suitable non-invasive biomarkers that allow the selection of good quality cumulus-oocyte complexes (COCs). The present study used whole, single oocyte culture to investigate whether the expression levels of follicle-stimulating hormone receptor (FSHR), insulin-like factor 1 receptor (IGF1R) and three steroidogenesis-related enzymes (CYP11al, CYP19al and HSD3β) in cumulus cells reflected the developmental competence of COCs. Cumulus cells were collected from single COCs before maturation culture and relative mRNA levels were assessed using real-time PCR. The analysis indicated that mRNAs for FSHR, IGF1R, CYP11al and HSD3β were present at higher levels in cumulus cells from COCs that failed to form blastocysts compared with cumulus cells from COCs that formed blastocysts. Moreover, FSHR and IGF1R mRNA levels were positively correlated with those of genes for steroidogenesis-related enzymes. In conclusion, poor developmental competence of COCs was related to higher expression of FSHR, IGF1R, CYP11al and HSD3β in cumulus cells, which may indicate the advanced differentiation of cumulus cells into granulosa cells.

The aim of this study was to investigate how intraduodenal infusions of fatty acids (FA) affect appetite-related gut peptides such as glucagon-like peptide-1 (GLP-1) and ghrelin in sheep. We hypothesized that these peptides can be highly reactive to unsaturated long-chain FA, because they are well known to decrease dry matter intake (DMI). Four ewes were fitted with a duodenal cannula and a jugular vein catheter for a 6-h duodenal infusion of the 9 FA (C8:0, C10:0, C12:0, C14:0, C16:0, C18:0, C18:1, C18:2, and C18:3) and water (control). The concentration of each FA was 1.6 g per metabolic body weight (BW), approximately corresponding to the amount of supplemented fat in a standard dairy cow diet. Each infusion was separated by at least 2 d. During the infusion period, blood samples were collected periodically to determine changes in plasma GLP-1, ghrelin, and metabolite concentrations. Duodenal infusions of C18:1, C18:2, and C18:3 led to higher plasma GLP-1 (P < 0.05) and lower glucose (P < 0.05) than control. Plasma ghrelin concentrations were greater in C18:1 and C18:3 infusions than control (P < 0.05). Plasma ketone bodies were higher in C8:0 and C10:0 infusions (P < 0.05), but plasma triglyceride concentrations were lower in C8:0, C10:0, C12:0, and C16:0 infusions (P < 0.05) than control. Fatty acid infusions except for C18:3 led to higher plasma NEFA concentrations than control (P < 0.05). These results confirmed that the hypophagic effect of dietary unsaturated long-chain FA is mediated by GLP-1 (an anorexigenic effect) secretion. However, we also observed higher plasma ghrelin (an orexigenic effect) partially by unsaturated long-chain FA. Thus, the gut peptide secretions when ruminant animals ingest FA supplements would complexly affect satiety and further studies are needed to determine their each impact on DMI.

In order to obtain more information on the development of bovine and ovine fetal mammary glands, a series of mammary glands from fetuses of different ages were analyzed. A total of 16 bovine fetuses with curved crown rump lengths ranging from 12 cm (80 days) to 75 cm (240 days) and 15 ovine fetuses ranging from 55 days to 131 days were examined. We used hematoxylin and eosin stain and Oil-Red-O stain to analyze the developmental and morphogenetic processes of mammary glands. In addition, we used immunohistochemical staining to determine the pattern of expression of cytokeratin 18 (CK18) during luminal epithelial differentiation, α-smooth-muscle actin (α-SMA) for myoepithelial differentiation, Ki-67 for cell proliferation, and estrogen receptor α (ERα). Our analyzes showed: (a) The primary mammary duct begin to proliferate in a lengthwise within the teat at 90 days in bovine fetuses and 63 days in ovine fetus; (b) luminal epithelial cells and myoepithelial cells appeared from 90 days in bovine fetuses and 63 days in ovine fetus; (c) proliferation of epithelial cells appeared to coincide with the development of the primary and secondary ducts; and (d) ERα was not found in the fetal mammary gland, but adipocytes showed the presence of ERα. Overall, these results indicate that the sequence of events in the prenatal development of the mammary gland of sheep is similar to that of cattle.

The maturation rate of canine oocytes during in vitro maturation (IVM) needs to be improved. The present study was designed to evaluate the effects of insulin-like growth factor-1 (IGF-1) on the IVM of canine oocytes. Ovaries were obtained by ovariohysterectomy and were sliced to release cumulus-oocyte complexes (COCs). In Experiment 1, the effects of different concentrations of IGF-1 on the nuclear maturation of oocytes was investigated. The COCs were cultured in a modified medium (mTCM199) with IGF-1 (0, 0.5, 5, 10, and 50 µg/ml). At the end of the 48 h culture, oocytes were fixed and stained to evaluate their nuclear stage. Supplementation with 50 µg/ml IGF-1 induced a significantly higher metaphase II (MII) rate (P < 0.05) compared to the 0 and 0.5 μg/ml IGF-1 groups. In Experiment 2, the expression levels of insulin receptor (INSR), IGF-1 receptor (IGF-1R), and IGF-2 receptor (IGF-2R) genes, localized to canine oocytes and cumulus cells, were investigated before and after IVM. The expression level of IGF-1R in cumulus cells after IVM was higher than that before IVM (P < 0.05). In Experiment 3, it was investigated whether an inhibitor of PTEN (phosphatase and tensin homolog), bpV, affects the nuclear maturation of oocytes. Regardless of bpV supplementation at a concentration of 0.2 to 200 µmol/l, there was no significant difference in the proportion of oocytes that reached the MII stage. These results indicated that IGF-1 has a favorable effect on the IVM of canine oocytes, possibly through the stimulation of the Ras/MAPK pathway via IGF-1R expressed in cumulus cells.

In order to determine blood lactate concentrations (bLac) and their validity as a diagnostic marker in bovine uterine torsion, blood samples were taken from 54 Holstein cows with uterine torsion before the correction of torsion. bLac in a group of cows with and without uterine necrosis were 15.0 and 3.0 mmol/l, respectively (P5.0 and >6.5 mmol/l, respectively. These findings suggest that in dairy cows with uterine torsion, an increase in bLac is a diagnostic predictor of uterine necrosis as well as poor prognosis in dams.

Rare-earth-free phosphors based on vanadate compounds were investigated, where the vanadates included chloride vanadates (MII2VO4Cl), pyrovanadates (MII2V2O7), orthovanadates (MII3(VO4)2) with divalent cations MII of Mg, Sr, Ba, and Zn, and oxofluorovanadates (AIVOF4) with an alkali metal AI. A chloride pyrolysis method and a liquid phase precipitation method were proposed for preparing the chloride vanadates and pyro- and orthovanadates, respectively. These vanadate compounds showed self-activated photoluminescence (PL) based on the VO4 clusters against the ultraviolet (UV) light irradiation. The colors of PL covered almost the whole visible-light region from blue to yellow as Sr2VO4Cl (deep blue), Ca2VO4Cl (sky blue), Ba2V2O7 (green), Sr2V2O7 (yellowish green), Zn3(VO4)2 (yellow), and Mg3(VO4)2 (yellow). A correlation was suggested from these compounds between the luminescent colors and the structural feature as the longer V–O distances in the VO4 tetrahedra in the crystal structures led to the longer wavelength in PL. This seemed to be also applicable for the oxofluorovanadates AIVOF4 (AI = K and Cs) which contain the VOF4 polyhedra with one O2− ion and four F- ions as the ligands, as they exhibited the reddish PL.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder. We screened 751 familial ALS patient whole-exome sequences and identified six mutations including p.D40G in the ANXA11 gene in 13 individuals. The p.D40G mutation was absent from 70,000 control whole-exome sequences. This mutation segregated with disease in two kindreds and was present in another two unrelated cases (P = 0.0102), and all mutation carriers shared a common founder haplotype. Annexin A11-positive protein aggregates were abundant in spinal cord motor neurons and hippocampal neuronal axons in an ALS patient carrying the p.D40G mutation. Transfected human embryonic kidney cells expressing ANXA11 with the p.D40G mutation and other N-terminal mutations showed altered binding to calcyclin, and the p.R235Q mutant protein formed insoluble aggregates. We conclude that mutations in ANXA11 are associated with ALS and implicate defective intracellular protein trafficking in disease pathogenesis.

A castrated male border collie 23 months of age weighing 19.4 kg was referred to the Animal Medical Center of Nihon University with complaints of visual disturbance and behavioral abnormality, hyperacusis and morbid fear. The MRI examination revealed the slight dilated cerebral sulci and cerebellar fissures and left ventricular enlargement. This is the first report of MRI findings of canine neuronal ceroid lipofuscinosis.

Heat capacities of crystalline charge-transfer (CT) complexes between trans-stilbene (STB) and tetracyanoquinodimethane (TCNQ) or (TCNQ)F4 were precisely measured below room temperature by adiabatic calorimetry. Although orientational disorder is seemingly absent, a glass transition due to freezing of the crankshaft motion is observed in STB-(TCNQ)F4 around 240 K as in the case of highly disordered STB-TCNQ around 250 K. Assessment of the degree of CT by IR and structural methods indicates that STB-(TCNQ)F4 is not a fully but weakly (only partially) ionic complex with a similar degree of CT of 0.1-0.2 to that in STB-TCNQ, despite the large difference in acceptor ability.

Heat capacity of the halogen-bridged mixed-valence complex Pt2(dta)4I (dta = CH3CS2-) has been measured in the temperature region between 6 and 386 K. The complex exhibited a phase transition of or...

Anomalies in the heat capacity due to spin reorientation transitions have been successfully detected for two orthoferrites by chopped-light ac calorimetry on single-crystals. Onset and completion of the spin reorientation were clearly detected as jumps in heat capacity at 82 and 89 K for TmFeO3 and at 53 and 59 K for HoFeO3. The magnitude of the anomalies is less than 1% with respect to the total heat capacity at the transition temperatures. The shapes of the anomalies are compared with available theoretical models.

Heat capacity of a quasi-one-dimensional organic conductor (DIMET) 2 I 3 has been measured by chopped-light ac calorimetry below room temperature. An anomaly due to a metal-insulator transition has been clearly detected around 40 K. The magnitude of the anomaly is analyzed through comparison with other Q1D conductors. A possible SDW mechanism is suggested.

The crystal of \ensuremath{\lambda}-${(\mathrm{BETS})}_{2}{(\mathrm{F}\mathrm{e}}_{x}{\mathrm{Ga}}_{1\ensuremath{-}x}{)\mathrm{C}\mathrm{l}}_{4}$ with $x\ensuremath{\approx}0.45$ undergoes successive superconducting and superconductor-to-insulator transitions around liquid helium temperature. The large diamagnetic susceptibility (\ensuremath{\approx}-35 emu/mol at 4 K) observed in the superconducting phase of the system with $x\ensuremath{\approx}0.47,$ which exhibits a superconducting transition at 4.3 K and superconductor-to-insulator transition at 3.7 K, can be regarded as evidence for the bulk nature of the superconductor-to-insulator transition. The anisotropy of the susceptibility and spin-flop behavior indicate that the system takes a \ensuremath{\pi}-d coupled antiferromagnetic insulating ground state, which is essentially the same as that of \ensuremath{\lambda}-${(\mathrm{BETS})}_{2}{\mathrm{FeCl}}_{4}$ exhibiting a coupled metal-insulator and antiferromagnetic transition at 8.5 K. The easy axis is parallel to the c direction. Unlike the Ga-rich system $(xl0.35)$ with superconducting ground state, the Fe-rich system such as \ensuremath{\lambda}-${(\mathrm{BETS})}_{2}{(\mathrm{F}\mathrm{e}}_{0.7}{\mathrm{Ga}}_{0.3}{)\mathrm{C}\mathrm{l}}_{4}$ shows a metal-insulator transition at ambient pressure. At high pressure, however, the \ensuremath{\pi} and d electrons tend to be independent of each other and the system takes a superconducting ground state.

λ-(BETS)2GaBrxCl4-x [BETS = bis(ethylenedithio)tetraselenafulvalene; 0 ≤ x ≤ 2] is a molecular superconductor with strongly correlated conduction electrons. The electrical transport properties of λ-(BETS)2GaBrxCl4-x are drastically changed by varying the bromine content x or by applying pressure. At ambient pressure, the superconducting transition could be observed for x < 0.75. The pressure and x dependencies of Tc were examined. The M−H curve (M = magnetization; H = magnetic field) at 2 K indicated the almost perfect Meissner state of the superconducting phase of λ-(BETS)2GaCl4. The Hc1 is ∼8 Oe for H⊥ and 12 Oe for H∥, where H⊥ and H∥ are the magnetic fields perpendicular and parallel to the c axis, respectively. The magnetic susceptibility of λ-(BETS)2GaBrxCl4-x increases with decreasing temperature to ∼60 K, below which the susceptibility becomes x-dependent and tends to be suppressed with increasing x. The isotropic decrease of the static susceptibility at lower temperature observed in the insulatin...

Organic conductors based on the unsymmetrical π donor molecule EDT-TTF (ethylenedithiotetrathiafulvalene) or its selenium-substituted analogs (EDST, EDTS) and tetrahedral anions GaCl 4 – were prepared. The crystal structure determinations and the extended Huckel tight-binding band calculations indicate these systems to be quasi-one-dimensional conductors similar to TMTTF or TMTSF systems (TM systems). Electrical resistivity and magnetic susceptibility measurements and low-temperature X-ray diffraction experiments suggested a spin-Peierls ground state for (EDT-TTF) 2 GaCl 4 . (EDST) 2 GaCl 4 and (EDTS) 2 GaCl 4 exhibit metallic behavior down to ca. 40 K. The electric and magnetic properties of (EDST) 2 GaCl 4 suggested a semimetallic state at low temperature. In spite of the similarity in the crystal and electronic band structures between TM and EDT systems, these two series of quasi-one-dimensional conductors do not share the same ‘generalized phase diagram’. The electron–lattice interaction seems to be important in EDT conductors. The electric and magnetic properties of the isostructural systems with magnetic FeCl 4 – anions were also examined. The magnetic interaction between the high-spin Fe 3+ ions was found to be very weak.

β-[(CH3)4N][Pd(dmit)2]2 is isomorphic to [(CH3)4N][Ni(dmit)2]2, which is the first pure π acceptor molecular conductor exhibiting a superconducting transition. The crystal structure at low temperature, pressure dependence of the resistivity and the magnetic susceptibility of β-[(CH3)4N][Pd(dmit)2]2 were examined. The general features of the phase diagram of β-[(CH3)4N][Pd(dmit)2]2 resemble those of the organic superconductors. At ambient pressure, the resistivity increases gradually with lowering temperature. A superconducting transition was observed at 6—9 kbar. The highest Tc was 6.5 K. Between 5 and 6.3 kbar, a characteristic rapid resistivity decrease was observed at the small temperature range above the offset temperature of the superconducting transition. Although the resistivity is not zero, this temperature region can be regarded as a “pre-superconducting region.” Above 9 kbar the metallic state is stabilized down to low temperature. Above 5 kbar, most of the crystals exhibited resistivity anomali...

Single crystals of [Me3HN][Ni(dmise)2]2, [Me2H2N][Ni(dmise)2]2, [MeH3N][Ni(dmise)2]2, and Cs[Pd(dmise)2]2 were grown electrochemically. The crystal structures of [Me3HN][Ni(dmise)2]2, [Me2H2N][Ni(d...

The ambient-pressure superconductor a-(EDT-TTF)[Ni(dmit)2], which is a charge transfer salt formed between EDT-TTF and Ni(dmit)2, exhibits a superconducting transition at 1.3 K and a phase transition around 20 K featuring a characteristic resistance hump. We estimated that the room-temperature electrical conductivity was 140–370 and 14–36 S cm−1 for the current parallel and perpendicular to the conducting plane respectively. We measured the upper critical magnetic field (Hc2) in the superconducting phase. The temperature dependence of Hc2 was estimated |d Hc2/dT| T = Te = −0.27 T K−1 (B||c-axis); |dHc2/d T| T = Tc = −3.5 T K−1 (B||ab-plane). The anisotropy of Hc2 within the ab-plane was found to be quite small, indicating a two-dimensionally isotrope character within the ab-plane in the superconducting state of the salt.

Two types of BETS conductors are reported. Needle crystals of λ-BETS2GaXxY4-x gave a series of new superconductors: λ-BETS2GaCl4 (Tc=7 K), λ-BETS2GaBrCl3 (8 K), λ-BETS2GaCl3F (3.5 K), λ-BETS2GaBr0.5Cl3.5 (6.5 K). λ-BETS2GaBr1.5Cl2.5, which is a semiconductor at 1 bar, exhibited a superconducting transition at 8 K and ca. 1 kbar. Pressure and temperature dependencies of the resistivities of λ-BETS2GaXxY4-x closely resemble those of κ-type BEDT-TTF superconductors. Many BETS metals with magnetic ions (Fe3+, Mn2+, Co2+, Cu2+) were prepared. Among them, κ-type MX4 (M=Fe, Co; X=Cl, Br) salts and θ-type Cu2Cl6 salts are metallic down to at least 4 K. The static susceptibility of λ-BETS2FeCl4 shows a characteristic magnetic field dependence below its metal-insulator transition temperature.

The band energy scheme of [(CH3)2(C2H5)2N][Pd(dmit)2]2 at high pressure was discussed and it might be possible that the insulating phase above 7 kbar is related to the 1D metal instability. The low-temperature crystal structures of 2D M(dmit)2(M=Ni, Pd) compounds, α-[(CH3)2(C2H5)2N]-[Ni(dmit)2]2 and (C7H16N)[Ni(dmit)2]2 were determined by imaging plate(IP) system equipped with a closed-cycle herium refrigirator. The X-ray satellite reflections of γ-(EDT-TTF)[Pd(dmit)2]2 indicating CDW formation below 80 K were also observed by IP system. [(CH3)3HN] [Ni(dmise)2]2 has a nearly three-dimensional Fermi surface owing to a strong intersheet interaction between their selone groups.

Electrical resistivity under a high pressure and the upper critical magnetic field ( H c2 ) in the superconducting state were investigated on the ambient-pressure superconductor α-(EDT-TTF)[Ni(dmit) 2 ], which exhibits a phase transition around 20 K and the superconducting transition at 1.3 K under ambient pressure. Above 9.9 kbar, the superconducting transition was not observed down to 0.5 K, while the former transition survived up to 13.7 kbar. The temperature dependence of H c2 was estimated: |d H c2 /d T | T = T c ≈-0.27 T/K ( B ∥ c * -axis); |d H c2 /d T | T = T c ≈-3.5 T/K ( B ∥ a b -plane). The anisotropy of H c2 within the a b -plane was found to be quite small indicating the two-dimensionally isotropic character within the a b -plane in the superconducting state of the salt.

α-(EDT–TTF)[Ni(dmit)2] comprises crossing columns of EDT–TTF and Ni(dmit)2 and exhibits unique metallic behaviour with a characteristic resistivity peak at around 14 K. The possibility of the development of the superstructure was examined down to 9 K by using a low-temperature imaging plate X-ray system, but no extra X-ray reflections could be found below 14 K. The 14 K anomaly is, therefore, thought not to originate from charge density wave (CDW) instability. New phases of γ-(EDT–TTF)[Pd(dmit)2] and γ-(EDT–TTF)[Ni(dmit)2] were found. γ-(EDT–TTF)[Pd(dmit)2] has parallel columns of EDT–TTF and Pd(dmit)2 and shows a resistivity anomaly around 80 K. Below 80 K, X-ray satellite reflections indicating CDW formation were observed. γ-(EDT–TTF)[Ni(dmit)2] undergoes a metal–insulator transition at around 100 K.

Wood creosote is a medicine that has been listed in the Japanese Pharmacopoeia (JP) since the first edition published in 1886. Medicines containing wood creosote and other natural ingredients have been very popular in Japan and Southeast Asian countries. In Japan, one such medicine, named Seirogan, has been used for more than 100 years. In this paper, we report the results of our examination on the historical aspects of wood creosote. One finding was that creosote, called "kereosote" at that time, was imported to Japan for the first time to Nagasaki by Johann Erdewin Niemann, who was the Director of the Dutch Mercantile House, and prescribed by Johannes Lijdius Catharinus Pompe van Meerdervoort and Anthonius Franciscus Bauduin. From our findings, we concluded that wood creosote was one of the essential medicines for the successful introduction and progression of Western medicine in Japan. Furthermore, we found that Dutch physicians introduced wood creosote to Japanese physicians, including Taizen Sato, Dokai Hayashi, and Jun Matsumoto, and that wood creosote was subsequently popularized by Rintaro (Ogai) Mori during the Russo-Japanese war. In addition, we examined the original plant for wood creosote, and consequently confirmed that the 15th edition of the JP, Supplement Two, clarifying the original plant for wood creosote, matches the pharmaceutical and historical facts. We also provide drug information relating to distinguishing between wood creosote and the creosote bush.

A hi-speed system to execute certification path validation of the X.509 Certificate was developed. The algorithm used not only certificates and certificate revocation information but also certification path information as cached information in order to speed up transactions. The effect of it was confirmed by the performance experiment of the certification path validation system in test and real environment.

Lawsonia intracellularis is an obligate intracellular pathogenic bacterium that causes proliferative enteropathy in domestic and experimental animals. In this study, we improved the in vitro cultivation method of L. intracellularis to increase the passage efficiency and showed that L. intracellularis isolated from a rabbit and a pig have different antigenic properties. Bacteria should be recovered from infected cells before cell death due to infection to obtain higher bacterial passage efficiency, and measurement of LDH activity in the cell culture medium was useful for determining the timing of bacterial passage. L. intracellularis isolated from the rabbit and pig showed different band patterns in immunoblotting. Our results should be helpful in the development of serological diagnosis and epidemiological investigation methods.

After thoroughly studying the chronology of the therapeutic use of wood creosote, we obtained novel findings on its botanical origin. Furthermore, we could demonstrate the importance of differentiating between wood creosote and coal tar creosote, which is clearly stipulated by Japanese Pharmacopoeia.

We have examined time-averaged charges of the molecular donors around the phase boundary between the insulating, superconducting, and metal phases for ${\ensuremath{\beta}}^{\ensuremath{''}}$-type ET [$\mathrm{ET}=\text{bis}$-ethelenedithio-tetrathiafulvalene] salts with one (two) hole(s) per two (three) molecules by using vibrational spectroscopy. Around the phase boundary, the site charges are neither those expected for a well-developed charge-ordered (CO) state nor a homogeneous distribution. The molecular charges exhibit an inhomogeneous distribution just above the insulator-superconductor transition temperature. We have analyzed the distribution of the site charges from the viewpoint of the alternation of intersite Coulomb interactions along the stacking direction. The closeness in the energy of several CO patterns mostly contributes to the inhomogeneous distribution, whereas a large difference (no difference) in the energy levels contributes to the CO (homogeneous) state. Our observations indicate that the instability due to the closeness in the energy of several distributions correlates with the insulator-superconductor transition in nondimerized or weakly dimerized molecular conductors.

The electronic structure of ultrathin Ag films with 1/3 monolayer (ML) of Pb alloyed at the surface was investigated by angle-resolved photoemission spectroscopy. Compared to clean ultrathin Ag films, the energy positions of the quantum-well states (QWSs) moved closer to the Fermi level due to the change of the potential barrier at the film/vacuum interface. We found that the parabolic band dispersion of the QWSs become disturbed where they cross the surface-state bands, and furthermore, they were shown to follow the periodicity that is only present at the surface. Our results suggest that it is possible to tune the properties of the QWSs whose thickness is more than 10 ML just by submonolayer deposition of heteroelements at the surface.

Although in vitro maturation (IVM) of oocytes has been successfully established for many species, the efficiency of IVM in canine oocytes is still very low. As growth factors have been shown to promote oocyte maturation in some species, we investigated whether use of transforming growth factor α (TGF-a) and insulin-like growth factor 1 (IGF-1) might overcome the difficulties of achieving meiotic maturation in cultured canine cumulus-oocyte complexes (COC). Ovaries were obtained from bitches at 6 months to 7 years of age by ovariohysterectomy and were sliced repeatedly to release COC. In the first experiment, the COC were cultured at 38.8°C for 48 h in 5% CO2 in air in medium 199 supplemented with either TGF-a (0, 1, 10, or 100 ng mL–1) or IGF-1 (0, 0.5, 5, 10, or 50 µg mL–1). In the second experiment, the synergistic effect of TGF-a and IGF-1 was investigated by culturing COC in medium 199 supplemented with both TGF-a (0, 1, 10, or 100 ng mL–1) and IGF-1 (0, 0.5, 5, 10, or 50 µg mL–1). At the end of the culture period, the oocytes were denuded of cumulus cells by pipetting with a fine bore glass pipette; the denuded oocytes were then fixed in Carnoy's solution and stained with Hoechst 33342. The nuclear configuration and chromatin morphology of the oocytes were evaluated under confocal laser scanning microscopy. The cells were assigned to 1 of the following meiotic stages: germinal vesicle (GV), germinal vesicle breakdown (GVBD), metaphase I (MI), or metaphase II (MII). Data were analysed by ANOVA with Fisher's PLSD test. In experiment 1, no significant difference were observed in the rates of cells maturing to the MI and MII stages, but that in the 10 ng mL–1 of TGF-a group (56.3%) were larger than in the other treatment groups (38.8–51.0%). The frequencies of MII stage cells in the 5, 10, and 50 µg mL–1 of IGF-1 treatment groups (9.8, 13.3, and 12.2%, respectively) were significantly higher than in the 0.5 µg mL–1 of IGF-1 group and the control group (5.3 and 2.2%, respectively). In experiment 2, the frequency of MI and MII cells in the control, 1 ng mL–1 of TGF-a plus 0.5 µg mL–1 of IGF-1, 10 ng mL–1 of TGF-a plus 5 µg mL–1 of IGF-1, 10 ng mL–1 of TGF-a plus 10 µg mL–1 of IGF-1, and 100 ng mL–1 of TGF-a plus 50 µg mL–1 of IGF-1 group were 44.1, 36.1, 63.5, 70.8, and 50.8%, respectively. The frequency of MII cells in the control group and the same treatment groups were 2.8, 7.2, 10.4, 15.3, and 10.8%, respectively. Both frequencies in the 10 ng mL–1 of TGF-a plus 10 µg mL–1 of IGF-1 group were significantly higher than in the control group. The TGF-a may act in a paracrine fashion on the surrounding granulosa cells, and IGF-1 may play multiple roles in cellular metabolism, proliferation, growth, and differentiation in canine oocyte maturation, as has been reported for many other species. In conclusion, these results demonstrate that a synergistic effect between TGF-a and IGF-1 produces an increased rate of in vitro maturation to the MI and MII stages in canine oocytes.

We report the anisotropy of the upper critical magnetic field and the pressure-temperature phase diagram for a molecular superconductor, α-EDT-TTF[Ni (dmit) 2], which is a charge-transfer salt formed between EDT-TTF and Ni (dmit) 2. The critical field was found to be almost isotropic within the conductive ab-plane. The pressure-temperature (P-T) phase diagram shows decreasing of a superconducting transition temperature by increasing an applied pressure. These results are discussed in connection with the electrical-anisotropy studies reported for the normal-conducting state of this salts.

Heat capacity of halogen-bridged one-dimensional binuclear metal complex (so-called MMX chain) having four n-pentyl groups, Pt2(n-PenCS2)4I, was measured by adiabatic calorimetry. A first-order phase transition was observed at 207.4 K when measurement was made after cooling from room temperature. The enthalpy and entropy of transition were determined to be 10.19 kJ mol(-1) and 49.1 J K(-1) mol(-1), respectively. A monotropic phase transition was observed at 324 K on heating, and the entropy of transition was essentially null. The sample once heated above 324 K never returned to the initial phase at room temperature and underwent a higher-order phase transition at 173 K and a first-order phase transition at 220.5 K. The enthalpy and entropy of the first-order phase transition were estimated to be 11.6 kJ mol(-1) and 52.4 J K(-1) mol(-1), respectively. The magnitude of the entropy gain at the phase transition from the initial room-temperature phase to the high-temperature phase at 324 K shows that in Pt2(n-PenCS2)4I a large amount of entropy reserved in alkyl chain is transferred to dithiocarboxylato groups upon the phase transition, as in the cases of Pt2(n-PrCS2)4I and Pt2(n-BuCS2)4I.

Meissner and diamagnetic shielding effects were investigated for an organic superconductor, α-EDT-TTF[Ni(dmit) 2 ] (dmit = 4,5-dimercapto-1,3-dithiole-2-thione, EDT-TTF=ethylenedithio-tetrathiafulvalene), under ambient pressure. The onset of the diamagnetic transition occurred at 1.39 K. From the measurements of the Meissner effect, the volume fraction of the superconducting phase was estimated to be more than 62%. From the measurements of the diamagnetic shielding effect, this fraction was estimated to be about 93%. This high level of volume fraction shows that the superconducting phase is the main phase in α-EDT-TTF[Ni(dmit) 2 ].