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Discordance between fractional flow reserve (FFR) and instantaneous wave-free ratio (iFR) occurs in approximately 20 % of cases. However, no studies have reported the discordance in patients with severe aortic stenosis (AS). We aimed to evaluate the diagnostic discordance between FFR and iFR in patients with severe AS.We examined 140 consecutive patients with severe AS (164 intermediate coronary artery stenosis vessels). FFR and iFR were calculated in four quadrants based on threshold FFR and iFR values of ≤0.8 and ≤0.89, respectively (Group 1: iFR0.89, FFR0.80; Group 2: iFR ≤0.89, FFR0.80; Group 3: iFR0.89, FFR ≤0.80; and Group 4: iFR ≤0.89, FFR ≤0.80). Concordant groups were Groups 1 and 4, and discordant groups were Groups 2 and 3. Positive and negative discordant groups were Groups 3 and 2, respectively.The median (Q1, Q3) FFR and iFR were 0.84 (0.76, 0.88) and 0.85 (0.76, 0.91), respectively. Discordance was observed in 48 vessels (29.3 %). In the discordant group, negative discordance (Group 2: iFR ≤0.89 and FFR0.80) was predominant (45 cases, 93.6 %). Multivariate analysis showed that the left anterior descending artery [odds ratio (OR), 3.88; 95 % confidence interval (CI): 1.54-9.79, p = 0.004] and peak velocity ≥5.0 m/s (OR, 3.21; 95%CI: 1.36-7.57, p = 0.008) were independently associated with negative discordance (FFR0.8 and iFR ≤0.89).In patients with severe AS, discordance between FFR and iFR was predominantly negative and observed in 29.3 % of vessels. The left anterior descending artery and peak velocity ≥5.0 m/s were independently associated with negative discordance.

A new strategy for the convergent synthesis of the ABCD ring system of indole terpene alkaloids has been developed based on a Sonogashira coupling of an o-iodoaniline (the A ring) with an alkyne bearing the D ring. After a tandem palladium-catalyzed cyclization, the tetracyclic ABCD ring structure found in the terpene indole alkaloids was obtained in good yield.

Sandwich freezing is a method of rapid freezing by sandwiching specimens between two metal disks and has been used for observing exquisite the close-to-native ultrastructure of living yeast and bacteria. Recently, this method has been found to be useful for preserving cell images of glutaraldehyde-fixed animal and human tissues. In the present study, this method was applied to observe the fine structure of mouse glomerular capillary loops. Morphometry was then performed, and the results were compared with the data obtained by an in vivo cryotechnique, which may provide the closest ultrastructure to the native state of living tissue. The results show that the ultrastructure of glomerular capillary loops obtained by sandwich freezing–freeze-substitution after glutaraldehyde fixation was close to that of the ultrastructure obtained by in vivo cryotechnique not only in the quality of cell image but also in quantitative morphometry. They indicate that the ultrastructure obtained by sandwich freezing–freeze-substitution after glutaraldehyde fixation may more closely reflect the living state of cells and tissues than conventional chemical fixation and dehydration at room temperature and conventional rapid freezing–freeze-substitution of excised tissues without glutaraldehyde fixation. Sandwich freezing–freeze-substitution techniques should be used routinely as a standard method for observing the close-to-native ultrastructure of biological specimens.

BACKGROUND/PURPOSE Data on the feasibility of coronary access (CA) through above or outside of the cylindrical shaped-transcatheter heart valve (THV) are very limited. The aims of the present study were to assess the feasibility of CA after transcatheter aortic valve replacement (TAVR) with the LOTUS using multi detector computed tomography (MDCT) and the reliability of algorithm detecting unfavorable CA. METHODS/MATERIALS Post-TAVR MDCT of 41 patients with 82 coronary arteries were evaluated. The relationship and distance between the THV flame and sinotubular junction (STJ) and coronary ostia were assessed. Unfavorable CA was defined as the valve-to-STJ distance < 2-mm or the valve-to-coronary ostia distance < 2-mm if the THV flame was above STJ or coronary ostia. RESULTS MDCT-identified unfavorable CA was observed in 29.3% for the left coronary artery and 41.5% for the right coronary artery. In total, 53.7% of patients had at least one unfavorable CA and 14.6% of those had unfavorable CA for both left and right coronary artery. While patients underwent coronary angiography after TAVR, the success rates of selective coronary cannulation were significantly lower in patients with MDCT-identified unfavorable CA in comparison to those with favorable CA for left (20.0% vs. 100%, P = 0.002) and right coronary artery (0% vs. 100%, P < 0.001). CONCLUSIONS Future CA through above or outside of a cylindrical shaped THV may be challenging with a significant probability. Our algorithm identifying unfavorable CA using post-MDCT seems to be useful for estimating the risk of unsuccessful selective cannulation.

The fungal cell wall is the initial barrier for the fungi against diverse external stresses, such as osmolarity changes, harmful drugs, and mechanical injuries. This study explores the roles of osmoregulation and the cell wall integrity (CWI) pathway in response to high hydrostatic pressure in the yeastSaccharomyces cerevisiae. We demonstrate the roles of the transmembrane mechanosensor Wsc1 and aquaglyceroporin Fps1 in a general mechanism to maintain cell growth under high-pressure regimes. The promotion of water influx into cells at 25 MPa, as evident by an increase in cell volume and a loss of the plasma membrane eisosome structure, promotes the activation of Wsc1, an activator of the CWI pathway. The downstream mitogen-activated protein kinase Slt2 was hyperphosphorylated at 25 MPa. Glycerol efflux increases via Fps1 phosphorylation, which is initiated by downstream components of the CWI pathway and contributes to the reduction in intracellular osmolarity under high pressure. The elucidation of the mechanisms underlying adaption to high pressure through the well-established CWI pathway could potentially translate to mammalian cells and provide novel insights into cellular mechanosensation.

Background Difficulties are often encountered while controlling atrial fibrillation (AF), especially in hemodialysis (HD) patients. Previous data revealed that cryoballoon ablation (CBA) for treating paroxysmal atrial fibrillation (PAF) was not inferior to radiofrequency ablation (RFA); however, HD patients were excluded in this prior trial. Thus, the efficacy of CBA for HD patients is still unknown. Methods This retrospective study analyzed HD patients who underwent catheter ablation (CA) for AF from August 2011 to June 2019. Patients who received CBA (CBA group) and those who received RFA (RFA group) were compared. The primary endpoint was defined as freedom from a composite outcome (a documented recurrence of any atrial tachyarrhythmia or a prescription of antiarrhythmic drugs) at one year after CA. Results The RFA and CBA groups were composed of 21 and 23 patients, respectively. Freedom from a composite outcome was 58.4% in the RFA group and 68.2% in the CBA group (Log-rank: p = 0.571). Conclusion Our results suggest that patients on HD with AF who were treated with CBA tended to have better outcomes than patients treated with RFA. Therefore, CBA could be a suitable ablation method for HD patients.

We herein report the cytokine expression at different stages for three patients who developed cardiac complications after immune checkpoint inhibitor (ICI) therapy. Case 1 with biopsy-proven myocarditis showed increased levels of interleukin (IL)-8, monocyte chemotactic and activating factor, and granulocyte macrophage colony-stimulating factor (GM-CSF) when he developed Takotsubo cardiomyopathy. Case 2 with subclinical myocarditis showed predominant activation of IL-8 during the progressive clinical course. Case 3 with cytokine-releasing syndrome showed substantial activations of IL-6, IL-8, GM-CSF, and interferon-γ. Our data suggest the development of unique cytokine activation in individual patients with cardiac complications after ICI therapy.

We have been using sandwich freezing of living yeast and bacteria followed by freeze-substitution for observing close-to-native ultrastructure of cells. Recently, sandwich freezing of glutaraldehyde-fixed cultured cells and human tissues have been found to give excellent preservation of ultrastructure of cells and tissues. These studies, however, have been conducted using a handmade sandwich freezing device and have been limited in a few laboratories. To spread the use of this method to other laboratories, we fabricated and commercialized a new sandwich freezing device. The new device is inexpensive, portable and sterilizable. It can be used to rapid-freeze viruses, bacteria, yeast, cultured cells and animal and human tissues to a depth of 0.2 mm if tissues are prefixed with glutaraldehyde. The commercial availability of this device will expand application of rapid freezing to wide range of biological materials.

Many advances in PET/CT technology can potentially improve image quality and the ability to detect small lesions. A new digital TOF-PET/CT scanner based on silicon photomultipliers (SiPM) integrated with a Bayesian penalized likelihood (BPL) PET reconstruction algorithm (Q.Clear; GE Healthcare) has been introduced into clinical practice. The present study aimed to quantify the ability of a digital TOF-PET/CT scanner combined with BPL reconstruction to detect small lesions, and to determine the optimal penalization factor (β) in BPL to accurately detect such lesions. All PET data were acquired from a NEMA body phantom using a Discovery MI (DMI) PET/CT system (GE Healthcare). The phantom included six spheres with diameters of 4, 5, 6, 8, 10, and 13 mm, and contained a background activity level of 5.3 kBq/mL, with target-to-background ratios (TBR) of 4:1 and 8:1. Images were reconstructed using a baseline OSEM algorithm, with OSEM + PSF, OSEM + TOF, OSEM + PSF + TOF, and BPL + PSF + TOF (β: 50–400). The matrix size was 192 × 192 and 384 × 384. Data acquired in 100-min list mode were re-binned into acquisition times ranging from 2 to 100 min. The quantitative accuracy and detectability of small hot spheres were evaluated by physical assessment of a recovery coefficient (RC) and a detectability index (DI), as well as visual assessment of PET images at each acquisition time. The RC and DI of sub-centimeter spheres were improved, because the digital TOF-PET/CT scanner has a larger TOF performance gain due to better timing resolution. The RC and DI were higher with BPL in sub-centimeter spheres, than with other OSEM-based types of reconstruction. The BPL for an 8-mm sphere overestimated uptake due to edge artifact overshoot induced by PSF modeling. The variability of RC and DI for acquisition times and TBR differed considerably according to β values. The RC for ~ 8-mm spheres were > 1 at β values between 50 and 100, but were close to 1 at β value of 200. The visual scores for β = 200 in BPL were maximal, whereas those for spheres that were ≥ 6 mm exceeded the criterion of 3. The BPL in the digital TOF-PET/CT scanner improved the quantitation and detectability of sub-centimeter spheres compared with OSEM-based reconstruction. Optimization of the β value in BPL might allow the detection of lesions ≤ 6 mm, although detectability depended on the TBR of lesions. A β value of 200 seemed optimal for detecting sub-centimeter lesions.

The uptake of cholesterol from the host is closely linked to the proliferation of pathogenic fungi and protozoa during infection. For some pathogenic fungi

This retrospective study was aimed (i) at elucidating the correlation between fatty liver diagnoses based on the plain computed tomography (CT) value and those based on the attenuation coefficient (AC) value determined with the ultrasound-guided attenuation parameter (UGAP) and (ii) at evaluating the diagnostic power of AC values. We included 125 patients who underwent blood tests, abdominal ultrasonography and abdominal CT at our department between April 2020 and March 2021. Hepatic fat infiltration was categorized as S0 (5%), S1 (≥5 and 30%), S2 (≥30 and50%) or S3 (≥50%). The diagnostic ability of UGAP-determined AC was evaluated using receiver operating characteristic (ROC) curve analysis, and the correlation between AC value and fatty liver grade by CT value. The coefficient of correlation (r) between the AC value and plain CT value was -0.6188, indicating a moderate relationship. For diagnosing grade ≥S1 (n = 44), the area under the ROC curve (AUROC) was 0.8541, sensitivity 84.1%, specificity 81.5% and cutoff value 0.676 dB/cm/MHz. In diagnosing grade ≥S2 (n = 35), the AUROC was 0.8603, sensitivity 88.6%, specificity 81.1% and cutoff value 0.694 dB/cm/MHz. In diagnosing grade = S3 (n = 18), the AUROC was 0.9016, sensitivity 94.5%, specificity 81.9% and cutoff value, 0.704 dB/cm/MHz. The AC value is useful in diagnosing fatty liver.

Background Giant cell myocarditis (GCM) is a rare cause of fulminant heart failure (HF). The most common presentation is progressive hemodynamic deterioration, and a few cases present with idiopathic complete atrioventricular block (cAVB). The prognosis of GCM is poor, and GCM patients usually die of HF and ventricular arrhythmia unless cardiac transplantation is performed. Few reports have described the effects of treatments such as immunosuppression and detailed reverse remodelling in GCM patients. Case summary A 69-year-old female presented with cAVB. Transvenous pacemaker was implanted via the left subclavian vein. One and a half months later, she exhibited left ventricular dyssynchrony and lower left ventricular ejection fraction (LVEF), resulting in hospitalization for HF. She received cardiac resynchronization therapy; however, this had no apparently positive effects on her cardiac function. To investigate the cause of the lower LVEF, an endomyocardial biopsy was taken from the right ventricular septum. She was diagnosed with GCM and immediately received immunosuppression therapy with prednisolone and ciclosporin. This resulted in the functional recovery of the right ventricle; on the other hand, the left ventricle had still not recovered based on transthoracic echocardiography. Fortunately, she successfully recovered from severe HF without recurrence. Discussion This is a case of fulminant HF due to GCM which initially presented as cAVB. Moreover, this case demonstrates the quite difference of the functional recovery between the left ventricle and the right ventricle with immunosuppression therapy.

The neutron-emission spectrum from a 197Au target excited with a 16.6 ±0.2 MeV, monoenergetic, linearly polarized photon beam was measured using the time-of-flight method at the NewSUBARU-BL01. Two...

Chemical fixation has been used for observing the ultrastructure of cells and tissues. However, this method does not adequately preserve the ultrastructure of cells; artifacts and extraction of cell contents are usually observed. Rapid freezing is a better alternative for the preservation of cell structure. Sandwich freezing of living yeast or bacteria followed by freeze-substitution has been used for observing the exquisite natural ultrastructure of cells. Recently, sandwich freezing of glutaraldehyde-fixed cultured cells or human tissues has also been used to reveal the ultrastructure of cells and tissues. These studies have thus far been carried out with a handmade sandwich freezing device, and applications to studies in other laboratories have been limited. A new sandwich freezing device has recently been fabricated and is now commercially available. The present paper shows how to use the sandwich freezing device for rapid freezing of biological specimens, including bacteria, yeast, cultured cells, isolated cells, animal and human tissues, and viruses. Also shown is the preparation of specimens for ultrathin sectioning after rapid freezing and procedures for freeze-substitution, resin embedding, trimming of blocks, cutting of ultrathin sections, recovering of sections, staining, and covering of grids with support films.

We present the case of a 56-year-old woman diagnosed with primary biliary cholangitis (PBC). She has continuously taken 600 mg/day of ursodeoxycholic acid. Edema of the lower limbs manifested on July 20, 20XX; after 2 weeks, she manifested rapid weight gain and nettle rash on the limbs and trunk. She was admitted to our hospital on August 22. She had marked eosinophilia, hypoalbuminemia, anemia, non-pitting lower limbs edema, and nettle rash of the limbs and the trunk. We ruled out other diseases that may have caused the edema and suspected her with episodic angioedema with eosinophilia (EAE). The peripheral blood eosinophil count rapidly decreased after the administration of 30 mg prednisolone. The edema and nettle rash improved on the 7th day of admission, and the hypoalbuminemia and anemia improved on the 14th day. Prednisolone was tapered and discontinued, and there was no relapse of edema. We revised our diagnosis to non-recurrent EAE. She was diagnosed with asymptomatic PBC; therefore, anemia and hypoalbuminemia were considered not PBC but chronic inflammation and decrease in appetite. In this case, elevation of serum IgG4 was observed at onset and at remission. This suggests that IgG4 may be involved in the development of EAE in patients with chronic liver disease.

We previously reported that Candida albicans responded to mild heat stress in a range of temperature elevations simulating fever, and concluded that mild heat stress increases susceptibility to antifungal drugs. In this study, we show that mild heat stress causes a morphological change in hyphae during the process of biofilm formation. We found that mild heat stress extended the period of hyphal stage maintenance in C. albicans biofilm. Although the rate of hyphal change from yeast form to hyphal form reached the maximum within 3 hr, later, almost every cell quickly reverted to the yeast growth phase within 6 hr at 37°C but not at 39°C, or under mild heat stress. Electron microscopy using a smart specimen preparation technique revealed that mild heat stress significantly increased the thickness of the inner cell wall accompanied by a decrease in density of the outer cell wall in the hyphae of C. albicans biofilm. To identify the gene responsible for the morphological changes associated with mild heat stress, we performed microarray gene expression analysis. Eleven genes were upregulated and 17 genes were downregulated under mild heat stress in biofilm cells. The increased PHR1 gene expression in response to mild heat stress was confirmed in quantitative RT-PCR analysis. The mutant upregulated PHR1 expression showed the same sensitivity against antifungal drug micafungin as dependent on mild heat stress. Our findings point to possible therapeutic effects of hyperthermia as well as to the effect of fever during infections.

We analyse the performance of a gas time projection chamber (TPC) as a high-performance gamma-ray telescope and polarimeter in the e$^+$e$^-$ pair creation regime. We use data collected at a gamma-ray beam of known polarisation. The TPC provides two orthogonal projections $(x,z)$ and $(y,z)$ of the tracks induced by each conversion in the gas volume. We use a simple vertex finder in which vertices and pseudo-tracks exiting from them are identified. We study the various contributions to the single-photon angular resolution using Monte Carlo simulations and compare them with the experimental data and find that they are in excellent agreement. The distribution of the azimutal angle of pair conversions shows a bias due to the non-cylindrical-symmetric structure of the detector. This bias would average out for a long duration exposure on a space mission, but for this pencil-beam characterisation we have ensured its accurate simulation by a double systematics control scheme, data taking with the detector rotated at several angles with respect to the beam polarisation direction and systematics control with a non-polarised beam. We measure, for the first time, the polarisation asymmetry of a linearly polarised gamma-ray beam in the low energy pair creation regime. This sub-GeV energy range is critical for cosmic sources as their spectra are power laws which fall quickly as a function of increasing energy. This work could pave the way to extending polarised gamma-ray astronomy beyond the MeV energy regime., Comment: Submitted to Astroparticle Physics

The N-to-Mo ratio x in epitaxial cubic molybdenum nitride γ -MoN x (001) layers was varied from x = 0.69–1.25 by adjusting the substrate temperature T s = 1000–600 °C during reactive magnetron sputter deposition. X-ray diffraction, reflection, and reciprocal space mapping measurements in combination with Rutherford backscattering spectroscopy indicate that the layers exhibit negligible strain and that increasing x = 0.69–1.25 leads to an increasing lattice constant a = 4.16–4.26 A and a decreasing density ρ = 8.7–6.8 g/cm 3 which is primarily attributed to a decrease in the cation site occupation from 0.89 to 0.70. Time resolved pump-probe reflectivity measurements of the sound velocity parallel to the [001] growth direction indicate that increasing the nitrogen concentration leads to a velocity reduction, corresponding to a decrease in C 11 from 502 ± 31 to 355 ± 30 GPa for x = 0.69–1.25. Similarly, the transverse surface acoustic wave velocity measured by surface Brillouin scattering spectroscopy decreases from 4.00–3.52 km/s, providing values for C 44 from 100 ± 7 to 73 ± 3 GPa. The softening with increasing N concentration is primarily attributed to the decreasing Mo site occupancy, which causes a reduction in the valence electron concentration leading to a weakening of N Mo bonds associated with hybridized N 2 p - Mo 4 d e g orbitals.

In planarization processes of sapphire, lapping process takes a long time because sapphire is a hard material. In contrast, superfinishing, which involves fixed abrasive machining, can substitute for lapping, and it would be possible to shorten the amount of processing time. In this work, vitrified-bonded diamond superabrasive stones with different grain diameters are developed. Then, multistage superfinishing is investigated by combining these stones. Results indicate that the multistage process is capable of producing a 2 nmRa surface, equivalent of a lapped surface in less than 10 min. To improve the process of multistage superfinishing, a removal amount estimation method is developed based on the real contact pressure calculation. The working area ratio of the stone was calculated by considering elastic deformation during superfinishing. The contact ratio of sapphire is calculated considering the roughness of the pre-finished surface and grain depth of cut. Accordingly, the real contact pressure is calculated to estimate the removal amount during superfinishing and finished surface roughness was expected.

This paper describes the high speed air-jet-assisted machining of nickel-base superalloy with a SiC whisker reinforced alumina insert. This machining method showed already good performance in high speed machining of Inconel 718. In this paper, the influence of the application direction of coolant on the tool wear and tool life was investigated for obtaining much better cutting performance of the ceramic tool. The coolant was applied from the side of flank face with three different directions: two oblique directions from the end and side flank faces and the perpendicular direction to the cutting edges. In contrast, the application direction of the air jet was always perpendicular to the cutting edges. The experimental results showed that the best performance in the air-jet-assisted machining was obtained by applying coolant from the side of side flank at a cutting speed of 420 m/min and from the side of end flank at a cutting speed of 780 m/min. The necessary condition found for the best performance was that the size of notch wear was a little larger than that of flank wear.

We constructed deletion mutants of Cryptococcus neoformans var neoformans (serotype D) genes encoding late ergosterol biosynthetic pathway enzymes and found that the mutations enhanced susceptibility to various drugs including micafungin, one of the echinocandins, to which wild-type Cryptococcus strains show no susceptibility. Furthermore, through isolation of a mutant resistant to micafungin from a micafungin-sensitive erg mutant and genetic analysis of it, we found that the responsible mutation occurred in the hotspot 2 of FKS1 encoding β-1, 3-glucan synthase, indicating that micafungin inhibited the growth of the erg mutant via inhibiting Fks1 activity. Addition of ergosterol to the culture of the erg mutants recovered the resistance to micafungin, suggesting that the presence of ergosterol in membrane inhibits the accession of micafungin to its target. We found that a loss of one of genes encoding subunits of v-ATPase, VPH1, made Cryptococcus cells sensitive to micafungin. Our observation that the erg2 vph1 double mutant was more sensitive to micafungin than either single mutant suggests that these two genes act differently in becoming resistant to micafungin. The erg mutants allowed us to study the physiological significance of β-1, 3-glucan synthesis in C. neoformans; the inhibition of β-1, 3-glucan synthesis induced cell death and changes in cellular morphology. By observing the erg mutant cells recovering from the growth inhibition imposed by micafungin, we recognized β-1, 3-glucan synthesis would suppress filamentous growth in C. neoformans.

Current {\gamma}-ray telescopes based on photon conversions to electron-positron pairs, such as Fermi, use tungsten converters. They suffer of limited angular resolution at low energies, and their sensitivity drops below 1 GeV. The low multiple scattering in a gaseous detector gives access to higher angular resolution in the MeV-GeV range, and to the linear polarisation of the photons through the azimuthal angle of the electron-positron pair. HARPO is an R&D program to characterise the operation of a TPC (Time Projection Chamber) as a high angular-resolution and sensitivity telescope and polarimeter for {\gamma} rays from cosmic sources. It represents a first step towards a future space instrument. A 30 cm cubic TPC demonstrator was built, and filled with 2 bar argon-based gas. It was put in a polarised {\gamma}-ray beam at the NewSUBARU accelerator in Japan in November 2014. Data were taken at different photon energies from 1.7 MeV to 74 MeV, and with different polarisation configurations. The electronics setup is described, with an emphasis on the trigger system. The event reconstruction algorithm is quickly described, and preliminary measurements of the polarisation of 11 MeVphotons are shown., Comment: Proceedings VCI2016

Most cancer cells rely on glycolysis to generate ATP, even when oxygen is available. However, merely inhibiting the glycolysis is insufficient for the eradication of cancer cells. One main reason for this is that cancer cells have the potential to adapt their metabolism to their environmental conditions. In this study, we investigated how cancer cells modify their intracellular metabolism when glycolysis is suppressed, using PANC-1 pancreatic cancer cells and two other solid tumor cell lines, A549 and HeLa. Our study revealed that glycolytically suppressed cells upregulated mitochondrial function and relied on oxidative phosphorylation (OXPHOS) to obtain the ATP necessary for their survival. Dynamic changes in intracellular metabolic profiles were also observed, reflected by the reduced levels of TCA cycle intermediates and elevated levels of most amino acids. Glutamine and glutamate were important for this metabolic reprogramming, as these were largely consumed by influx into the TCA cycle when the glycolytic pathway was suppressed. During the reprogramming process, activated autophagy was involved in modulating mitochondrial function. We conclude that upon glycolytic suppression in multiple types of tumor cells, intracellular energy metabolism is reprogrammed toward mitochondrial OXPHOS in an autophagy-dependent manner to ensure cellular survival.

We previously reported that Candida albicans responded to mild heat stress in a range of temperature elevations simulating fever, and concluded that mild heat stress increases susceptibility to antifungal drugs. In this study, we show that mild heat stress causes a morphological change in hyphae during the process of biofilm formation. We found that mild heat stress extended the period of hyphal stage maintenance in C. albicans biofilm. Although the rate of hyphal change from yeast form to hyphal form reached the maximum within 3 hr, later, almost every cell quickly reverted to the yeast growth phase within 6 hr at 37°C but not at 39°C, or under mild heat stress. Electron microscopy using a smart specimen preparation technique revealed that mild heat stress significantly increased the thickness of the inner cell wall accompanied by a decrease in density of the outer cell wall in the hyphae of C. albicans biofilm. To identify the gene responsible for the morphological changes associated with mild heat stress, we performed microarray gene expression analysis. Eleven genes were upregulated and 17 genes were downregulated under mild heat stress in biofilm cells. The increased PHR1 gene expression in response to mild heat stress was confirmed in quantitative RT-PCR analysis. The mutant upregulated PHR1 expression showed the same sensitivity against antifungal drug micafungin as dependent on mild heat stress. Our findings point to possible therapeutic effects of hyperthermia as well as to the effect of fever during infections.

Backgrounds: Recently, several studies have demonstrated the usefulness of cold polypectomy (CP), a safe and simple method for the removal of small polyps. We investigated the safety and efficacy o...

99m Tc with a 6 hour half-life is one of the most important medical isotopes used for the Single-Photon Emission Computed Tomography (SPECT) inspection in hospitals of US, Canada, Europe and Japan. 99m Tc isotopes are extracted by the milking method from parent 99Mo isotopes with a 66 hour half-life. The supply of 99Mo isotopes now encounters a serious crisis. Hospitals may not suitably receive 99Mo medical isotopes in near future, due to difficulties in production by research nuclear reactors. Many countries are now looking for alternative ways to generate 99Mo isotopes other than those with research reactors. We discuss a sustained availability of 99m Tc isotopes via the nat Mo(γ, n) photonuclear reaction, and discuss to solve technical problems for extracting pure 99m Tc isotopes from other output materials of photonuclear reactions.

Urinary hippuric acid (HA) and phenylacetylglycine (PAG) are biomarker candidates for drug-induced phospholipidosis (PLD). To confirm their utility in preclinical and clinical settings, it is essential to develop and validate their quantification method in advance. In this study, we have applied liquid chromatography-tandem mass spectrometry (LC/MS/MS) for simultaneous quantification of HA and PAG in rat urine, and matrix based ion suppression was assessed by post-column infusion assay. Effective sample dilution reduced matrix effect of urine to be negligible level and calibration curves showed good correlation between those in urine diluent and buffer alone. Reliability of this assay was confirmed by the assessments for intra- and inter-day precisions and accuracies of quality control samples. The method was applied to rat urine after multiple oral administrations of PLD-inducing drugs, and the changes in HA and PAG concentrations and their ratio were successfully detected as rat plasma in previous report. This is the first report to quantify HA and PAG easily and accurately as potential biomarkers to monitor PLD status. This assay would be useful tool for monitoring PLD in toxicological studies by non-invasive sampling.

Metastin/kisspeptin is an endogenous ligand of KISS1 Receptor (KISS1R). Metastin and KISS1R are suggested to play crucial roles in regulating the secretion of gonadotropin-releasing hormone (GnRH), and continuous administration of metastin derivatives attenuated the plasma testosterone levels in male rats. Our optimization studies of metastin derivatives led to the discovery of 1 (Ac-d-Tyr-d-Trp-Asn-Thr-Phe-azaGly-Leu-Arg(Me)-Trp-NH2, TAK-683), which suppressed plasma testosterone in rats at lower doses than those of leuprolide. Although 1 possessed extremely potent pharmacological activity, 20 mg/mL aqueous solution of 1 has a gel formation property. In order to improve this physicochemical property, we substituted d-Trp at position 47 with a variety of amino acids; we identified that substitution with cyclic amino acids, which could change peptide conformation, retained its potency. Especially, analogue 24 (TAK-448) with trans-4-hydroxyproline (Hyp) at position 47 showed not only superior pharmacologica...

Structome analysis is a useful tool for identification of unknown microorganisms that cannot be cultured. In 2012, we discovered a unique deep-sea microorganism with a cell structure intermediate between those of prokaryotes and eukaryotes and described its features using freeze-substitution electron microscopy and structome analysis (quantitative and three-dimensional structural analysis of a whole cell at the electron microscopic level). We named it Myojin parakaryote Here we describe, using serial ultrathin sectioning and high-voltage electron microscopy tomography of freeze-substituted specimens, the structome analysis and 3D reconstruction of another unique spiral bacteria, found in the deep sea off the coast of Japan. The bacteria, which is named as 'Myojin spiral bacteria' after the discovery location and their morphology, had a total length of 1.768 ± 0.478 µm and a total diameter of 0.445 ± 0.050 µm, and showed either clockwise or counter-clockwise spiral. The cells had a cell surface membrane, thick fibrous layer, ribosomes and inner fibrous structures (most likely DNA). They had no flagella. The bacteria had 322 ± 119 ribosomes per cell. This ribosome number is only 1.2% of that of Escherichia coli and 19.3% of Mycobacterium tuberculosis and may reflect a very slow growth rate of this organism in the deep sea.

Rapid freeze-freeze substitution after glutaraldehyde fixation (CF-FS method) obtained the natural and fine structures of macrophages and engulfed yeast cells. Culturing macrophages on single hole molybdenum grids placed in culture dishes made possible the rapid freezing of cells by the 'open sandwich method'. This method may be convenient when rapid-freezing cannot be performed immediately, or when a rapid-freezing device is not available in the lab.

Lysine-specific demethylases 1 and 2 (LSD1 and LSD2) are flavoenzyme demethylases, and their inhibitors are considered as potential chemical tools and anticancer agents. Here we report polyamine-based inhibitors of LSD1 and LSD2. In the initial screening, partially constrained polyamine 2 which contains three trans-cyclopentane units with a total of six stereogenic centers, showed the most potent LSD1-inhibitory activity. We then prepared a set of optical isomers of 2 and evaluated their inhibitory activities toward LSD1, LSD2, monoamine oxidases A and B (MAO-A and MAO-B). Optical isomers of 2 showed LSD1-inhibitory activity with Ki values of 2.2 to 6.4 μM, and LSD2-inhibitory activity with Ki values of 4.4 to 39 μM; there was a general preference for LSD1 to LSD2. All of them showed weak to negligible inhibition of MAO-A and MAO-B. This selectivity seemed to reflect the differences in the size and shape of the catalytic cavity of target enzymes, and our strategy of employing a set of optical isomers appears to be an effective approach for exploring the structural features of this family of enzymes. Polyamine 9 showed most potent LSD1-inhibitory activity (Ki = 2.2 μM in vitro), and it also inhibited the proliferation of HL-60 cells (IC50 = 49 μM). On the other hand, 12 was the most potent inhibitors of LSD2 with in vitro Ki values of 4.4 μM.

A series of structome analyses, that is, quantitative and three-dimensional structural analysis of a whole cell at the electron microscopic level, have already been achieved individually in Exophiala dermatitidis, Saccharomyces cerevisiae, Mycobacterium tuberculosis, Myojin spiral bacteria, and Escherichia coli. In these analyses, sample cells were processed through cryo-fixation and rapid freeze-substitution, resulting in the exquisite preservation of ultrastructures on the serial ultrathin sections examined by transmission electron microscopy. In this paper, structome analysis of non pathogenic Mycolicibacterium smegmatis, basonym Mycobacterium smegmatis, was performed. As M. smegmatis has often been used in molecular biological experiments and experimental tuberculosis as a substitute of highly pathogenic M. tuberculosis, it has been a task to compare two species in the same genus, Mycobacterium, by structome analysis. Seven M. smegmatis cells cut into serial ultrathin sections, and, totally, 220 serial ultrathin sections were examined by transmission electron microscopy. Cell profiles were measured, including cell length, diameter of cell and cytoplasm, surface area of outer membrane and plasma membrane, volume of whole cell, periplasm, and cytoplasm, and total ribosome number and density per 0.1 fl cytoplasm. These data are based on direct measurement and enumeration of exquisitely preserved single cell structures in the transmission electron microscopy images, and are not based on the calculation or assumptions from biochemical or molecular biological indirect data. All measurements in M. smegmatis, except cell length, are significantly higher than those of M. tuberculosis. In addition, these data may explain the more rapid growth of M. smegmatis than M. tuberculosis and contribute to the understanding of their structural properties, which are substantially different from M. tuberculosis, relating to the expression of antigenicity, acid-fastness, and the mechanism of drug resistance in relation to the ratio of the targets to the corresponding drugs. In addition, data obtained from cryo-transmission electron microscopy examination were used to support the validity of structome analysis. Finally, our data strongly support the most recent establishment of the novel genus Mycolicibacterium, into which basonym Mycobacterium smegmatis has been classified.

The prevalence of nonencapsulated Streptococcus pneumoniae (NESp) has increased with the introduction of pneumococcal conjugate vaccines in children; however, the bacteriological characteristics of NESp have not been sufficiently clarified. In this study, NESp strains isolated from the nasopharyngeal carriage of children from four nursery schools in Japan were analyzed for molecular type, antibiotic susceptibility, and biofilm productivity. A total of 152 putative S. pneumoniae strains were identified by optochin-susceptibility analysis, of which 21 were not serotypeable by slide agglutination, quellung reaction, or multiplex PCR. Among these 21 strains, three were lytA-negative and, therefore, not S. pneumoniae. The remaining 18 strains were positive for lytA, ply, pspK, and bile solubility and were confirmed as NESp. Therefore, the isolation rate of NESp in the S. pneumoniae strains in this study was 12.0% (18/149). Molecular-typing analyses classified five strains as two existing sequence types (STs; ST7502 and ST7786), and 13 strains formed four novel STs. Horizontal spread was suspected, because strains with the same ST were often isolated from the same nursery school. The NESp isolates were generally susceptible to most antimicrobials, with the exception of macrolides; however, all isolates possessed more than one abnormal penicillin-binding protein gene. Furthermore, NESp strains were more effective than encapsulated counterparts at forming biofilms, which showed obvious differences in morphology. These data indicated that NESp strains should be continuously monitored as emerging respiratory pathogens.

The life-cycle system of Ulotrichales, a major order of Ulvophyceae, remains controversial because it is unclear whether the Codiolum phase, a characteristic unicellular diploid generation in ulotrichalean algae, is a zygote or a sporophyte. This controversy inhibits the understanding of the diversified life cycles in Ulvophyceae. To distinguish between zygotes and sporophytes, we have to examine not only whether diploid generations function as sporophytes, but also whether mitosis occurs before meiosis in diploid generations. However, the nuclear behavior in the Codiolum phases is largely unknown, probably because no suitable methods are available. Using fluorescent microscopy with ethidium bromide and transmission electron microscopy of cell-wall-dissected specimens, we report the nuclear behavior in the Codiolum phases of an ulotrichalean alga with a representative life cycle, Monostroma angicava. Each vegetative Codiolum phase had a single polyploid nucleus due to endoreduplication, a type of mitosis without nuclear division. During zoosporogenesis, the nucleus had a structure that would be a meiosis-specific complex. We quantitatively showed that Codiolum phases grew extremely large and produced numerous zoospores. Our results suggest that an event comparable to mitosis occurs before meiosis in the Codiolum phase of M. angicava. This nuclear behavior and the functions (growth and zoospore production abilities) correspond to those of sporophytes. Therefore, the life-cycle system of M. angicava is a heteromorphic haplo-diplontic cycle. This system appears to be widely adopted among other ulotrichalean algae.

The present paper describes the applicability of the active flow control device, mini electromagnetic flap actuators attached on the leading edge of an airfoil, for the flow separation under both the steady and the unsteady flow conditions in the low Reynolds number region. At first, lift and drag have been measured for a wide variety of the wind speed Reynolds numbers and the angles of attack for the steady flow condition. Then, effects of some simple feedback flow controls, where the time-dependent signal of the lift-drag ratio have been used to detect the stall and served as a trigger to start the actuation, have been explored under the unsteady flow condition for evading the stall. In every low Reynolds number ranging from 30 000 to 80 000, the present actuators worked quite well to delay the stall, increasing in the lift and delaying the stall angle of attack. These aerodynamic modifications by the flap actuators obtained from the steady flow were found to be available even if the manipulation of the actuators started after the stall. Activation threshold of the lift-drag ratio as the input for the feedback control was determined from a stall classification map obtained under the steady flow experiment. Effectiveness of this feedback control was then demonstrated under the condition of the wind speed decrease (Reynolds number from 80 000 to 40 000) keeping the angle of attack constant at 11°, at which the stall occurs without the active control. Immediately after the sudden velocity decrease, the decrease in the lift-drag ratio were detected and the dynamic actuations were successfully started, resulting in evading the stall and keeping high and stable lift. An additional operation of the feedback, in which the running actuation is turned off when the lift-drag ratio shows lower than the second threshold value after operation, was revealed to be effective to keep the high lift force under the condition combined with the wind speed increase and decrease within the low Reynolds number range treated in this study.

Our basic cell biology research was aimed at investigating the effect on eukaryotic cells of the sudden loss of the F-actin cytoskeleton. Cells treated with latrunculin A (LA) in yeast extract peptone dextrose (YEPD) medium were examined using phase-contrast and fluorescent microscopy, freeze-substitution, transmission and scanning electron microscopy, counted using a Bürker chamber and their absorbance measured. The cells responded to the presence of LA, an F-actin inhibitor, with the disappearance of actin patches, actin cables and actin rings. This resulted in the formation of larger spherical cells with irregular morphology in the cell walls and ultrastructural disorder of the cell organelles and secretory vesicles. Instead of buds, LA-inhibited cells formed only 'table-mountain-like' wide flattened swellings without apical growth with a thinner glucan cell-wall layer containing β-1,3-glucan microfibrils. The LA-inhibited cells lysed. Actin cables and patches were required for bud formation and bud growth. In addition, actin patches were required for the formation of β-1,3-glucan microfibrils in the bud cell wall. LA has fungistatic, fungicidal and fungilytic effects on the budding yeast Saccharomyces cerevisiae.

This paper describes the notch and flank wear specific to a SiC whisker reinforced alumina tool in air jet assisted (AJA) turning of nickel-base superalloy Inconel 718 at high cutting speeds. An AJA machining experiment has revealed that the air jet applied to the tool tip in addition to coolant dramatically reduces the depth-of-cut notch wear. As a result, the width of flank wear, but not the size of notch wear, determined the life of a ceramic tool in AJA machining of Inconel 718. This is a reason for the large extension and small variation of the tool life when high speed AJA machining is adopted. The maximum tool life length reached 2160 m at a cutting speed of 660 m/min under the given cutting conditions. Finally, the mechanisms of the notch and flank wear of a SiC whisker reinforced alumina tool in AJA machining are discussed from the viewpoints of tribochemical reactions and tool wear anisotropy.

Tolvaptan (TLV) is a new vasopressin type 2 receptor antagonist effective in patients with heart failure (HF). We herein describe the case of an 84-year-old woman who developed acute renal injury induced by hypersensitivity to TLV. The patient had received an implanted pacemaker and was diagnosed with exacerbation of chronic HF due to atrial fibrillation, mitral regurgitation, tricuspid regurgitation and left ventricular dyssynchrony. Treatment with tolvaptan increased the urine volume, improved the dyspnea and decreased the edema. However, the patient's renal function and hyperkalemia worsened, and the blood eosinophil count increased without signs of dehydration or hypotension. Positive findings on a drug-induced lymphocyte stimulation test for TLV were consistent with this diagnosis.