A Combined test using both cell sediment and supernatant cell-free DNA in pleural effusion shows increased sensitivity in detecting activating EGFR mutation in lung cancer patients.

INTRODUCTION: The aim of this study was to examine whether a combined test using both cell sediment and supernatant cytology cell-free DNA (ccfDNA) is more useful in detecting EGFR mutation than using cell sediment DNA or supernatant ccfDNA alone in pleural effusion of lung cancer patients. METHODS: A total of 74 lung adenocarcinoma patients with paired samples between primary tumour and corresponding metastatic tumour with both cell sediment and supernatant ccfDNA of pleural effusion cytology were enrolled in this study. Cell sediment and supernatant ccfDNA were analysed separately for EGFR mutations by polymerase chain reaction. RESULTS: Out of 45 patients with mutant EGFR in primary tumours, EGFR mutations were detected in 23 cell sediments of corresponding metastases (sensitivity; 51.1%) and 20 supernatant ccfDNA corresponding metastases (sensitivity; 44.4%). By contrast, the combined test detected EGFR mutations in 27 corresponding metastases (sensitivity; 60.0%), and had a higher sensitivity than the cell sediment or the supernatant ccfDNA alone (P < .05). Out of 45 patients with mutant EGFR, 24, three and 18 were cytologically diagnosed as positive, atypical or negative, respectively. The detection rate in the combined test was highest (95.8%) in the positive group, and mutant EGFR was also detected in four of 18 samples (22.2%) in the negative group. CONCLUSIONS: A combined test using both cell sediment DNA and supernatant ccfDNA samples increases the concordance rate of EGFR mutations between primary tumour and corresponding metastases. Our findings indicate that supernatant ccfDNA is useful even in cases where the cytological diagnosis is negative.

SurePath® LBC improves the diagnostic accuracy of intrahepatic and hilar cholangiocarcinoma.

INTRODUCTION: The current study aimed to compare cytology using SurePath® (SP)-LBC and biliary tissue histology (BTH) for the diagnosis of biliary disease. METHODS: Between January 2014 and December 2016, 57 patients underwent endoscopic retrograde cholangiopancreatography for the diagnosis of biliary disease. Biliary cytological samples were processed using SP-LBC and subsequently BTH was performed. A final diagnosis was confirmed by surgery (23 malignant cases) and clinical follow-up (34 benign and malignant cases): 18 extrahepatic cholangiocarcinoma; 17 intrahepatic/hilar cholangiocarcinoma (intra/H-CC); eight other malignant disease; and 14 benign biliary disease. The diagnoses made using SP-LBC and BTH were classified into four categories: (1) benign; (2) indeterminate; (3) suspicious for malignancy/malignant; and (4) inadequate. In addition, diagnostic accuracy was compared between SP-LBC and BTH. RESULTS: Although 23% (13/57) of BTH samples were classified as inadequate, all SP-LBC cases were classified as adequate. Among 43 malignant cases, 11 normal, four indeterminate and 28 suspicious for malignancy/malignant were found using SP-LBC (26%, 9% and 65%, respectively), in contrast to 10 inadequate, nine normal, 10 indeterminate and 14 suspicious for malignancy/malignant observed using BTH (23%, 21%, 23%, and 33%, respectively). The identification of malignant cells was strikingly different between SP-LBC and BTH. Furthermore, limited to intra/H-CC, accuracy was significantly higher using SP-LBC than using BTH (P < .001). CONCLUSIONS: SP-LBC of the biliary tract is a useful and reliable method for diagnosing biliary malignant disease and has an advantage over BTH for detecting malignant cells and accurately diagnosing intra/H-CC.

Multi-energy reconstructions, central electron temperature measurements, and early detection of the birth and growth of runaway electrons using a versatile soft x-ray pinhole camera at MST.

A multi-energy soft x-ray pinhole camera has been designed, built, and deployed at the Madison Symmetric Torus to aid the study of particle and thermal transport, as well as MHD stability physics. This novel imaging diagnostic technique employs a pixelated x-ray detector in which the lower energy threshold for photon detection can be adjusted independently on each pixel. The detector of choice is a PILATUS3 100 K with a 450 µm thick silicon sensor and nearly 100 000 pixels sensitive to photon energies between 1.6 and 30 keV. An ensemble of cubic spline smoothing functions has been applied to the line-integrated data for each time-frame and energy-range, obtaining a reduced standard-deviation when compared to that dominated by photon-noise. The multi-energy local emissivity profiles are obtained from a 1D matrix-based Abel-inversion procedure. Central values of Te can be obtained by modeling the slope of the continuum radiation from ratios of the inverted radial emissivity profiles over multiple energy ranges with no a priori assumptions of plasma profiles, magnetic field reconstruction constraints, high-density limitations, or need of shot-to-shot reproducibility. In tokamak plasmas, a novel application has recently been tested for early detection, 1D imaging, and study of the birth, exponential growth, and saturation of runaway electrons at energies comparable to 100 × Te,0; thus, early results are also presented.

Development of ion source with a washer gun for pulsed neutral beam injection.

A new type of economical neutral beam source has been developed by using a single washer gun, pulsed operation, and a simple electrode system. We replaced the conventional hot filaments for arc-discharge-type plasma formation with a single stainless-steel washer gun, eliminating the entire dc power supply for the filaments and the cooling system for the electrodes. Our initial experiments revealed successful beam extraction up to 10 kV and 8.6 A, based on spatial profile measurements of density and temperature in the plasma source. The system also shows the potential to control the beam profile by controlling the plasma parameters in the ion accumulation chamber.

A computational tool for simulation and design of tangential multi-energy soft x-ray pin-hole cameras for tokamak plasmas.

A new tool has been developed to calculate the spectral, spatial, and temporal responses of multi-energy soft x-ray (ME-SXR) pinhole cameras for arbitrary plasma densities (n e,D), temperature (T e), and impurity densities (n Z). ME-SXR imaging provides a unique opportunity for obtaining important plasma properties (e.g., T e, n Z, and Z eff) by measuring both continuum and line emission in multiple energy ranges. This technique employs a pixelated x-ray detector in which the lower energy threshold for photon detection can be adjusted independently. Simulations assuming a tangential geometry and DIII-D-like plasmas (e.g., n e,0 ≈ 8 × 1019 m-3 and T e,0 ≈ 2.8 keV) for various impurity (e.g., C, O, Ar, Ni, and Mo) density profiles have been performed. The computed brightnesses range from few 102 counts pixel-1 ms-1 depending on the cut-off energy thresholds, while the maximum allowable count rate is 104 counts pixel-1 ms-1. The typical spatial resolution in the mid-plane is ≈0.5 cm with a photon-energy resolution of 500 eV at a 500 Hz frame rate.

Simulation, design, and first test of a multi-energy soft x-ray (SXR) pinhole camera in the Madison Symmetric Torus (MST).

A multi-energy soft x-ray pinhole camera has been designed and built for the Madison Symmetric Torus reversed field pinch to aid the study of particle and thermal-transport, as well as MHD stability physics. This novel imaging diagnostic technique combines the best features from both pulse-height-analysis and multi-foil methods employing a PILATUS3 x-ray detector in which the lower energy threshold for photon detection can be adjusted independently on each pixel. Further improvements implemented on the new cooled systems allow a maximum count rate of 10 MHz per pixel and sensitivity to the strong Al and Ar emission between 1.5 and 4 keV. The local x-ray emissivity will be measured in multiple energy ranges simultaneously, from which it is possible to infer 1D and 2D simultaneous profile measurements of core electron temperature and impurity density profiles with no a priori assumptions of plasma profiles, magnetic field reconstruction constraints, high-density limitations, or need of shot-to-shot reproducibility. The expected time and space resolutions will be 2 ms and <1 cm, respectively.

The essential interaction between yeast mRNA capping enzyme subunits is not required for triphosphatase function in vivo.

The Saccharomyces cerevisiae mRNA capping enzyme consists of two subunits: an RNA 5'-triphosphatase (Cet1) and an mRNA guanylyltransferase (Ceg1). In yeast, the capping enzyme is recruited to the RNA polymerase II (Pol II) transcription complex via an interaction between Ceg1 and the phosphorylated carboxy-terminal domain of the Pol II largest subunit. Previous in vitro experiments showed that the Cet1 carboxy-terminal region (amino acids 265 to 549) carries RNA triphosphatase activity, while the region containing amino acids 205 to 265 of Cet1 has two functions: it mediates dimerization with Ceg1, but it also allosterically activates Ceg1 guanylyltransferase activity in the context of Pol II binding. Here we characterize several Cet1 mutants in vivo. Mutations or deletions of Cet1 that disrupt interaction with Ceg1 are lethal, showing that this interaction is essential for proper capping enzyme function in vivo. Remarkably, the interaction region of Ceg1 becomes completely dispensable when Ceg1 is substituted by the mouse guanylyltransferase, which does not require allosteric activation by Cet1. Although no interaction between Cet1 and mouse guanylyltransferase is detectable, both proteins are present at yeast promoters in vivo. These results strongly suggest that the primary physiological role of the Ceg1-Cet1 interaction is to allosterically activate Ceg1, rather than to recruit Cet1 to the Pol II complex.

Direct measurement of density oscillation induced by a radio-frequency wave.

An O-mode reflectometer at a frequency of 25.85 GHz was applied to plasmas heated by the high harmonic fast wave (21 MHz) in the TST-2 spherical tokamak. An oscillation in the phase of the reflected microwave in the rf range was observed directly for the first time. In TST-2, the rf (250 kW) induced density oscillation depends mainly on the poloidal rf electric field, which is estimated to be about 0.2 kV/m rms by the reflectometer measurement. Sideband peaks separated in frequency by ion cyclotron harmonics from 21 MHz, and peaks at ion cyclotron harmonics which are suggested to be quasimodes generated by parametric decay, were detected.

Measurement of thickness of film deposited on the plasma-facing wall in the QUEST tokamak by colorimetry.

After several experimental campaigns in the Kyushu University Experiment with Steady-state Spherical Tokamak (QUEST), the originally stainless steel plasma-facing wall (PFW) becomes completely covered with a deposited film composed of mixture materials, such as iron, chromium, carbon, and tungsten. In this work, an innovative colorimetry-based method was developed to measure the thickness of the deposited film on the actual QUEST wall. Because the optical constants of the deposited film on the PFW were position-dependent and the extinction coefficient k1 was about 1.0-2.0, which made the probing light not penetrate through some thick deposited films, the colorimetry method developed can only provide a rough value range of thickness of the metal-containing film deposited on the actual PFW in QUEST. However, the use of colorimetry is of great benefit to large-area inspections and to radioactive materials in future fusion devices that will be strictly prohibited from being taken out of the limited area.

Validations of calibration-free measurements of electron temperature using double-pass Thomson scattering diagnostics from theoretical and experimental aspects.

This paper evaluates the accuracy of electron temperature measurements and relative transmissivities of double-pass Thomson scattering diagnostics. The electron temperature (Te) is obtained from the ratio of signals from a double-pass scattering system, then relative transmissivities are calculated from the measured Te and intensity of the signals. How accurate the values are depends on the electron temperature (Te) and scattering angle (θ), and therefore the accuracy of the values was evaluated experimentally using the Large Helical Device (LHD) and the Tokyo spherical tokamak-2 (TST-2). Analyzing the data from the TST-2 indicates that a high Te and a large scattering angle (θ) yield accurate values. Indeed, the errors for scattering angle θ = 135° are approximately half of those for θ = 115°. The method of determining the Te in a wide Te range spanning over two orders of magnitude (0.01-1.5 keV) was validated using the experimental results of the LHD and TST-2. A simple method to provide relative transmissivities, which include inputs from collection optics, vacuum window, optical fibers, and polychromators, is also presented. The relative errors were less than approximately 10%. Numerical simulations also indicate that the Te measurements are valid under harsh radiation conditions. This method to obtain Te can be considered for the design of Thomson scattering systems where there is high-performance plasma that generates harsh radiation environments.

A model of plasma current through a hole of Rogowski probe including sheath effects.

In TST-2 Ohmic discharges, local current is measured using a Rogowski probe by changing the angle between the local magnetic field and the direction of the hole of the Rogowski probe. The angular dependence shows a peak when the direction of the hole is almost parallel to the local magnetic field. The obtained width of the peak was broader than that of the theoretical curve expected from the probe geometry. In order to explain this disagreement, we consider the effect of sheath in the vicinity of the Rogowski probe. A sheath model was constructed and electron orbits were numerically calculated. From the calculation, it was found that the electron orbit is affected by E × B drift due to the sheath electric field. Such orbit causes the broadening of the peak in the angular dependence and the dependence agrees with the experimental results. The dependence of the broadening on various plasma parameters was studied numerically and explained qualitatively by a simplified analytical model.

Cyclic GMP phosphodiesterase inhibitors. 1. The discovery of a novel potent inhibitor, 4-((3,4-(methylenedioxy)benzyl)amino)-6,7,8-trimethoxyquinazoline.

A newly synthesized compound, 4-((3,4-(methylenedioxy)benzyl)amino)-6,7,8-trimethoxyquinazoline (6), had a potent (IC50 = 0.36 microM) inhibitory action on cyclic GMP phosphodiesterase (cGMP-PDE) isolated from porcine aorta; its inhibitory activities toward other PDE isozymes were at least 10-fold weaker. In addition, 6 relaxed porcine coronary arteries precontracted with PGF2 alpha (EC50 = 1.96 +/- 0.58 microM). At the concentration of 30 microM, 6 caused elevation of the intracellular cGMP level in porcine coronary arteries without any change in cAMP level. Various other 4-substituted 6,7,8-trimethoxyquinazolines were also synthesized and evaluated for cGMP-PDE inhibitory activity. From their structure-activity relationships, we concluded that the 4-((3,4-(methylenedioxy)benzyl)-amino) group is essential for potent inhibition of cGMP-PDE.

Cyclic GMP phosphodiesterase inhibitors. 2. Requirement of 6-substitution of quinazoline derivatives for potent and selective inhibitory activity.

We synthesized various 4-[[3,4-(methylenedioxy)benzyl]amino]quinazolines substituted at the 5- to 8-positions and evaluated their inhibitory activities toward cyclic GMP phosphodiesterase (cGMP-PDE) from porcine aorta. Monosubstitution at the 6-position was essential for the inhibitory activity, and the preferred substituents were compact and hydrophobic: methoxy (3b, IC50 = 0.23 microM), methyl (3c, 0.10 microM), chloro (3d, 0.019 microM), thiomethyl (3f, 0.031 microM), and cyano (3p, 0.090 microM) groups. Compounds 3b-d,f,p lacked inhibitory activity toward other PDE isozymes (all IC50 values > 100 microM), and their relaxing activities in porcine coronary arteries were well correlated with the inhibitory activities toward cGMP-PDE (r = 0.88, p < 0.05). One of these compounds, 3b, elevated the intracellular cGMP level in isolated porcine coronary arteries without causing any change in the cAMP level. We consider that this series of compounds dilates coronary arteries via potent and specific inhibition of cGMP-PDE.

4-Benzylamino-1-chloro-6-substituted phthalazines: synthesis and inhibitory activity toward phosphodiesterase 5.

We synthesized various 4-benzylamino-1-chloro-6-substituted phthalazines (15) and 4-benzylamino-1-chloro-7-substituted phthalazines (16) and evaluated their inhibitory activity toward phosphodiesterase 5 (PDE5) purified from porcine platelets. The PDE5-inhibitory activities of 15 were greater than those of the isomers (16). The preferred substituent at the 4-position of phthalazine was a (3-chloro-4-methoxybenzyl)amino group, and those at the 6-position were cyano, nitro, and trifluoromethyl groups. Compounds 15a (IC50 = 4.8 nM), 15f (3.5 nM), and 15i (5.3 nM) were more potent inhibitors than E4021 (8.6 nM). Compounds 15a and 15f also showed vasorelaxant activity in isolated porcine coronary arteries precontracted with prostaglandin F2alpha (10(-5) M). The EC50 values for vasorelaxant action of 15a, 15f, and E4021 were 150, 160, and 980 nM, respectively. These results show that novel PDE5 inhibitors possessing a potent vasorelaxant effect may exist among phthalazine derivatives.

4-(3-Chloro-4-methoxybenzyl)aminophthalazines: synthesis and inhibitory activity toward phosphodiesterase 5.

We synthesized various 4-(3-chloro-4-methoxybenzyl)aminophthalazines substituted at the 1- and 6-positions and evaluated their inhibitory activity toward phosphodiesterase 5 (PDE5) and their vasorelaxant activity in isolated porcine coronary arteries precontracted with prostaglandin F2alpha (10(-5) M). The preferred substituents at the 1-position of the phthalazine were 4-hydroxypiperidino, 4-hydroxymethylpiperidino, 4-(2-hydroxyethyl)piperidino, and 4-oxopiperidino. Among these compounds, [4-(3-chloro-4-methoxybenzyl)amino-1-(4-hydroxy)piperidino]-6-phthala zinecarbonitrile monohydrochloride (13) exhibited potent PDE5 inhibitory activity (IC(50) = 0.56 nM) with >1700-fold high selectivity over other PDE isozymes (PDE1-4). Compound 13 exhibited the most potent vasorelaxant action (EC(50) = 13 nM) in this series of compounds. Compound 13 reduced mean pulmonary arterial pressure by 29.9 +/- 3.1% when administered intravenously at 30 microg/kg to the chronically hypoxic rats and had an apparent oral bioavailability of about 19.5% in rats and was selected for further biological evaluation.

Structure-activity relationship of N-[2-(dimethylamino)-6-[3-(5-methyl-4-phenyl-1H-imidazol-1-yl)propoxy] phenyl]-N'-pentylurea and analogues. Novel potent inhibitors of acyl-CoA:cholesterol O-acyltransferase with antiatherosclerotic activity.

We have discovered N-butyl-N'-[2-(dimethylamino)-6-[3-(4-phenyl-1H- imidazol-1-yl)propoxy]phenyl]urea (4), a novel, potent, and systemically bioavailable inhibitor of ACAT (acylCoA:cholesterol O-acyltransferase). The structure-activity relationships (SARs) of this lead compound 4 were investigated by systematic modification of four regions in the molecule. The compounds prepared in this study were tested for in vitro inhibitory activity toward both aortic and intestinal ACATs, and selected compounds were further tested for in vivo hypocholesterolemic activity. The studies not only resulted in the discovery of N-[2-(dimethylamino)-6-[3-(5-methyl-4-phenyl-1H-imidazol-1-yl) propoxy]phenyl]-N'-pentylurea (24), with potent activity and moderate plasma level after oral administration, but also revealed the SAR in each modified region. Four compounds (4, 13, 14, 24) were further selected for testing of in vivo antiatherosclerotic activity; 4, 13, and 24 reduced atherosclerotic plaque development to 38-45% of the control value in terms of area, while 14 did not have a significant antiatherosclerotic effect.

Changes in enzyme activities in skin fibroblasts derived from persons of various ages.

We examined antioxidant enzyme activities (catalase, glutathione peroxidase, and superoxide dismutase) in cultured skin fibroblasts (passage number 2-3) derived from 30 persons of various ages. With increasing ages, catalase activity decreased, glutathione peroxidase activity increased slightly, and superoxide dismutase activity was unchanged. After UVA irradiation (4.8 joule/cm2) of the fibroblasts, only catalase activity decreased by 70%. This suggests that catalase may play an important role in the aging of human skin fibroblasts.

Eight years of experience with transjugular retrograde obliteration for gastric varices with gastrorenal shunts.

BACKGROUND AND OBJECTIVES: There is no standard treatment for gastric varices. Transjugular retrograde obliteration (TJO) is one way of obliterating gastric varices with gastrorenal shunts, in which blood flow is abundant. Our aim was to examine our experience with TJO during an 8-year period and to determine the long-term effects of this treatment. METHODS: We performed TJO procedures in 52 patients to obliterate gastric varices. All the patients had liver cirrhosis. Sixteen had hepatocellular carcinoma (HCC) without vascular invasion. We inserted an angiographic catheter with an occlusive balloon through the right internal jugular vein into the gastrorenal shunt or the gastric varices. After controlling the other blood-draining routes with a microcoil or absolute ethanol, or both, we injected 5% ethanolamine oleate with iopamidol into the gastric varices under fluoroscopy. RESULTS: The gastric varices were successfully obliterated by TJO in all cases. The complications were all minor and transient. The mortality rate for TJO was 0%. There was no recurrence and no bleeding of gastric varices at all after TJO. Patient survival differed depending on the presence or absence of HCC (P <.05). The development of HCC in the cirrhotic liver was the most common cause of late death. Gastrointestinal bleeding was not a cause of death. The occurrence rate of esophageal varices after TJO was high, but these varices could be treated easily by endoscopic injection sclerotherapy before they bled. CONCLUSIONS: Portal blood flow through the gastrorenal shunt is diverted to the porto-azygos venous system after the gastrorenal shunt is obliterated by TJO. TJO is a safe option that we recommend for treating gastric varices with gastrorenal shunts, provided that the TJO is followed by endoscopic injection sclerotherapy.

Injection sclerotherapy of esophageal varices for patients undergoing emergency and elective surgery.

From October 1977 to September 1981, 68 patients with esophageal varices (30 emergency cases of bleeding and 38 elective cases) were treated by injecting 5% ethanolamine oleate into varices, using an esophagofiberscope. Esophageal bleeding was successfully controlled in 29 of 30 patients who had emergency surgery. None of the 38 patients who underwent elective operation had bleeding after treatment. When recurrence occurred 1 or 2 years after treatment, the same procedure was repeated. Pleuritis occurred in one of the patients who had emergency surgery, and bleeding (300 to 400 ml) from the esophagocardial junction occurred in two patients who underwent elective operation. These patients were treatment conservatively.