Search

From the viewpoint of evaluating the instrumental performance of high-resolution electron microscopy (HREM), the Scherzer condition was investigated using information theory. As a result, the optimum defocus amount Δf can be expressed based on [Formula: see text] , and the formula [Formula: see text] is obtained. Furthermore, a procedure for measuring point resolution using the through-focus technique is developed, and a new method for determining the spherical aberration coefficient using the variance of Δf is introduced in the procedure., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Mitochondrial cloning is a promising approach to achieve homoplasmic mitochondrial DNA (mtDNA) mutations. We previously developed a microfluidic device that performs single mitochondrion transfer from a mtDNA-intact cell to a mtDNA-less (ρ 0 ) cell by promoting cytoplasmic connection through a microtunnel between them. In the present study, we described a method for generating transmitochondrial cybrids using the microfluidic device. After achieving mitochondrial transfer between HeLa cells and thymidine kinase-deficient ρ 0 143B cells using the microfluidic device, selective culture was carried out using a pyruvate and uridine (PU)-absent and 5-bromo-2'-deoxyuridine-supplemented culture medium. The resulting cells contained HeLa mtDNA and 143B nuclei, but both 143B mtDNA and HeLa nuclei were absent in these cells. Additionally, these cells showed lower lactate production than parent ρ 0 143B cells and disappearance of PU auxotrophy for cell growth. These results suggest successful generation of transmitochondrial cybrids using the microfluidic device. Furthermore, we succeeded in selective harvest of generated transmitochondrial cybrids under a PU-supplemented condition by removing unfused ρ 0 cells with puromycin-based selection in the microfluidic device., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

To develop a ready-to-drink (RTD) milk coffee retaining the original coffee flavor, the effects of processing conditions during manufacture on retronasal-arma (RA) compounds from the milk coffee were investigated by gas chromatography-mass spectrometry using an RA simulator (RAS). Thirteen of 46 detected compounds in the RAS effluent (RAS compounds) decreased significantly following pH adjustment of coffee (from pH 5.1 to 6.8) and 5 compounds increased. RAS compounds from coffee tended to decrease through the pH adjustment and subsequent sterilization. Significantly higher amounts of 13 RAS compounds were released from the milk coffee produced using a blending-after-sterilization (BAS) process without the pH adjustment than from that using a blending-before-sterilization (BBS) process with the pH adjustment. In BAS-processed milk coffee, significantly lower amounts of 8 high-volatility compounds and 1H-pyrrole were released from coffee containing infusion-sterilized (INF) milk than from coffee containing plate-sterilized (PLT) milk, whereas 3 low-volatility compounds were released significantly more from coffee using PLT milk. Principal component analysis revealed that the effect of the manufacturing process (BAS, BBS, or homemade (blending unsterilized coffee without pH adjustment with sterilized milk)) on milk coffee volatiles was larger than that of the sterilization method (INF or PLT) for milk, and that the sterilization method could result in different RAS volatile characteristics in BAS and homemade processes. In conclusion, a BAS process was found to be superior to a BBS process for the manufacture of an RTD milk coffee that retains volatile characteristics similar to that of a homemade milk coffee., Practical Application: Ready-to-drink (RTD) milk coffee manufactured using the conventional blending-before-sterilization process does not retain its original coffee flavor due to pH adjustment of the coffee during the process. The new blending-after-sterilization (BAS) process enabled the production of RTD milk coffee whose volatiles are closer to that of homemade milk coffee, as demonstrated by the results of RAS-GC-MS analysis. The BAS process has already been applied to the manufacture of RTD milk coffees in Japan., (© 2018 Institute of Food Technologists®.)

Detection of foodborne pathogens at very low levels is still a challenge. A custom-built multichannel surface plasmon resonance (SPR) biosensor and simultaneous enrichment broth (SEB) were used to develop a simultaneous detection method for 3 important foodborne pathogens, Escherichia coli O157:H7 (O157:H7), Salmonella enteritidis, and Listeria monocytogenes, at a very low level. These 3 foodborne pathogens at a very low level (14, 6, and 28 CFU/25 g (mL) for O157:H7, S. enteritidis, and L. monocytogenes, respectively) were inoculated in SEB and incubated at 37 ˚C for 24 h. Sample prepared from the simultaneous enrichment culture was analyzed using the multichannel SPR biosensor and sensor chip immobilized with polyclonal antibodies specific to each of the target pathogens. O157:H7, S. enteritidis, and L. monocytogenes in chicken were detected simultaneously at an inoculum dose of 14, 6, and 28 CFU/25 g, respectively. Our method using a custom-built multichannel SPR biosensor and enrichment in SEB is expected as a rapid and simultaneous detection method for low levels of O157:H7, S. enteritidis, and L. monocytogenes in food., Practical Application: Our method is expected as a rapid and simultaneous detection method for pathogens at very low levels. It has great potential for safety control of food and microbiological detection applications., (© 2017 Institute of Food Technologists®.)

An abrupt onset of the new tongue-shaped deformation of magnetic surface in magnetized plasmas, which was conjectured in since the 1960s but has not been observed, is experimentally identified just before an abrupt onset of a large-scale collapse event. Two novel properties of the event are identified. First, the transition of symmetry of perturbation (rather than a growth of linearly unstable MHD modes) was found to be a key for the onset of abrupt collapse, i.e., the transition of symmetry gives a new route to the collapse from stable state. Second, as a phase-space response of ions, the distortion from Maxwell-Boltzmann distribution of epithermal ions was observed for the first time.

The coupling between the transport and magnetic topology is an important issue because the structure of magnetic islands, embedded in a toroidal equilibrium field, depends on the nature of the transport at the edge of the islands. Measurements of modulated heat pulse propagation in the DIII-D tokamak have revealed the existence of self-regulated oscillations in the radial energy transport into magnetic islands that are indicative of bifurcations in the island structure and transport near the q = 2 surface. Large amplitude heat pulses are seen in one state followed by small amplitude pulses later in the discharge resulting in a repeating cycle of island states. These two states are interpreted as a bifurcation of magnetic island with high and low heat pulse accessibility. This report describes the discovery of a bifurcation in the coupled dynamics between the transport and topology of magnetic islands in tokamak plasmas.

A Phased Array Antenna (PAA) was considered as launching and receiving antennae in reflectometry to attain good directivity in its applied microwave range. A well-focused beam was obtained in a launching antenna application, and differential-phase evolution was properly measured by using a metal reflector plate in the proof-of-principle experiment at low power test facilities. Differential-phase evolution was also evaluated by using the PAA in the Q-shu University Experiment with Steady State Spherical Tokamak (QUEST). A beam-forming technique was applied in receiving phased-array antenna measurements. In the QUEST device that should be considered as a large oversized cavity, standing wave effect was significantly observed with perturbed phase evolution. A new approach using derivative of measured field on propagating wavenumber was proposed to eliminate the standing wave effect.

The radial variations in the velocity of longitudinal waves propagating through Japanese cedar and Japanese cypress were experimentally investigated. In addition, the tracheid length (TL), microfibril angle (MFA), air-dried density (AD), and moisture content (MC) were measured in order to determine the effect of wood properties on velocity variations within the wood trunk. For both species, the longitudinal wave velocities measured in the longitudinal direction (V(L)) exhibited minimum values near the pith. For Japanese cedar, V(L) increased from 3600m/s toward the bark and soon attained a constant value (=4500m/s). On the other hand, for Japanese cypress, V(L) kept increasing from 4000m/s near the pith to 4800m/s at the bark. These radial variations in V(L) coincided with those in the tracheid length. V(L) exhibited strong correlations with TL and MFA with a significant level of (p<0.01). These findings suggest that the TL and MFA greatly affect the radial variation in the ultrasonic wave velocity in softwood., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Unlabelled: In this study, we focused on the antihypertensive effect of newly fermented salt-free soy sauce (SFS) in spontaneously hypertensive rats (SHRs). A 13-wk oral administration of SFS was performed daily in 8- to 21-wk-old SHRs with a dose of 200 mg/kg of body weight/d. The systolic (SBP) and diastolic (DBP) blood pressures were measured for each week during the experimental period. As a result, a significant (P < 0.01) BP lowering effect was observed from the 5th d to the end of the experiment in the SFS group compared with the control group (DeltaSBP(21-wk): 27 mmHg, DeltaDBP(21-wk): 20 mmHg). In contrast to such substantial effect by SFS intake, angiotensin I-converting enzyme activities in blood and local organs as well as pharmaceutical parameters such as serum Na(+) or K(+) level did not show any significant difference between in both SFS and control groups. Vasoconstriction experiment using thoracic aorta rings from 21-wk SHRs provided an interesting result that the rings from the SFS group evoked a >2-fold higher increase in the angiotensin II-stimulated constrictive response compared with the rings from the control group (P = 0.012), which suggested that the SFS-intake would be effective in possessing a higher vessel tone., Practical Application: In this study, we investigated the antihypertensive effect of newly fermented SFS in SHRs. As a result of 13-wk SFS-administration, we found a significant antihypertensive effect of the SFS. This finding strongly demonstrates that the developed SFS would be greatly beneficial for health and useful for health-related industries.

Amphiregulin, an EGF receptor (EGFR) ligand, is essential for epithelial development in various organs. A recent report suggested that amphiregulin acts as a protective factor in a liver injury model. Little is known about the roles of amphiregulin in lung injury and pulmonary fibrosis. The purpose of the present study was to investigate the role of amphiregulin in an experimental model of bleomycin-induced pneumopathy in mice. C57BL/6 mice were administered a bleomycin hydrochloride solution intratracheally. Recombinant human amphiregulin was injected intraperitoneally at 6, 8, 10, and 12 days after the bleomycin instillation. The grades of inflammation and fibrosis were assessed histologically and biochemically, and the numbers of apoptotic cells were counted after TdT-mediated dUTP nick end labeling (TUNEL) staining in the lung tissues. We also examined downstream survival signals of EGFR, namely phosphorylated Akt and phosphorylated Erk, in lung tissues by Western blotting analysis and immunohistochemistry. Expression of intrinsic amphiregulin was increased in murine lung tissues after bleomycin instillation. Administration of recombinant amphiregulin improved the survival rate and suppressed the degrees of inflammation and fibrosis and the number of TUNEL-positive cells in lung tissues. Amphiregulin treatment enhanced the activation of Akt and Erk in lung epithelial cells. Amphiregulin may play a protective role in bleomycin-induced pneumopathy in mice, probably through the activation of survival signals. Administration of amphiregulin may be a novel therapeutic strategy against lung injury and fibrosis.

The baroreflex system is the fastest mechanism in the body to regulate arterial pressure. Because the neural system (i.e., autonomic nervous system) mediates the baroreflex and the system operates under the closed-loop condition, the quantitative dynamic characteristics of the baroreflex system remained unknown until recently despite the fact that a countless number of observational and qualitative studies had been conducted. In order to develop the artificial baroreflex system, i.e., the bionic baroreflex system, we first anatomically isolated the carotid sinuses to open the baroreflex loop and identified the open-loop transfer function of the baroreflex system using white noise pressure perturbations. We found that the baroreflex system is basically a lowpass filter and remarkably linear. As an actuator to implement the bionic baroreflex system, we then stimulated the sympathetic efferent nerves at various parts of the baroreflex loop and identified the transfer functions from the stimulation sites to systemic arterial pressure. We found that the actuator responses can be described remarkably well with linear transfer functions. Since transfer functions of the native baroreflex and of the actuator were identified, the controller that is required to reproduce the native baroreflex transfer function can be easily derived from those transfer functions. To examine the performance of bionic baroreflex system, we implemented it animal models of baroreflex failure. The bionic baroreflex system restored normal arterial pressure regulation against orthostatic stresses that is indistinguishable from the native baroreflex system.

The pressure-volume relationship of the heart was first reported more than a century ago. It was not widely accepted, however, until the mid-1970s. The pressure-volume diagram became a central theme of cardiac mechanics once it was shown to be a good representation of ventricular mechanics. Early in 1980s, the introduction of the ventricular interaction with afterload using effective arterial elastance made it possible to translate ventricular mechanical properties represented by the pressure-volume relationship to the pumping ability of the heart. Furthermore incorporating the framework of ventricular arterial interaction into the classic Guyton's circulatory equilibrium early in 2000s enabled us to express quantitatively how mechanical properties of the ventricles and vascular systems determine the circulatory equilibrium. Successful quantitative descriptions of circulatory equilibrium using the pressure-volume concept would promote basic cardiovascular physiology and accelerate its clinical applications.

The inwardly rectifying properties and molecular basis of ATP-sensitive K(+) channels (K(ATP) channels) have now been established for several cell types. However, these aspects of nonvascular smooth muscle K(ATP) channels still remain to be defined. In this study, we investigated the molecular basis of the pore of K(ATP) channels of pig urethral smooth muscle cells through a comparative study of the inwardly rectifying properties, conductance, and regulation by PKC of native and homo- and heteroconcatemeric recombinant Kir6.x channels coexpressed with sulfonylurea receptor subunit SUR2B in human embryonic kidney (HEK) 293 cells by the patch-clamp technique (conventional whole-cell and cell-attached modes). In conventional whole-cell clamp recordings, levcromakalim (> or = 1 microM) caused a concentration-dependent increase in current that demonstrated strong inward rectification at positive membrane potentials. In cell-attached mode, the unitary amplitude of levcromakalim-induced native and recombinant heteroconcatemeric Kir6.1-Kir6.2 K(ATP) channels also showed strong inward rectification at positive membrane potentials. Phorbol 12,13-dibutyrate, but not the inactive phorbol ester, 4alpha-phorbol 12,13-didecanoate, enhanced the activity of native and heteroconcatemeric K(ATP) channels at -50 mV. The conductance of the native channels at approximately 43 pS was consistent with that of heteroconcatemeric channels with a pore-forming subunit composition of (Kir6.1)(3)-(Kir6.2). RT-PCR analysis revealed the expression of Kir6.1 and Kir6.2 transcripts in pig urethral myocytes. Our findings provide the first evidence that the predominant K(ATP) channel expressed in pig urethral smooth muscle possesses a unique, heteromeric pore structure that differs from the homomeric Kir6.1 channels of vascular myocytes and is responsible for the differences in inward rectification, conductance, and PKC regulation exhibited by the channels in these smooth muscle cell types.

Adipose differentiation-related protein (ADRP) is highly expressed in macrophages and human atherosclerotic lesions. We demonstrated that Toll-like receptor (TLR) 4-mediated signals, which are involved in atherosclerosis formation, enhanced the expression of ADRP in macrophages. Lipopolysaccharide (LPS) enhanced the ADRP expression in RAW264.7 cells or peritoneal macrophages from wild-type mice, but not in macrophages from TLR4-deficient mice. Actinomycin D almost completely abolished the LPS effect, whereas cycloheximide decreased the expression at 12 h, indicating that the LPS-induced ADRP expression was stimulated at the transcriptional level and was also mediated by new protein synthesis. LPS enhanced the ADRP promoter activity, in part, by stimulating activator protein (AP)-1 binding to the Ets/AP-1 element. In addition, preceding the increase of the ADRP mRNA, LPS induced the expression of interleukin (IL)-6, IL-1alpha, and interferon-beta mRNAs, all of which stimulated the ADRP expression. Antibodies against these cytokines or inhibitors of c-Jun NH(2)-terminal kinase and nuclear factor (NF)-kappaB suppressed the ADRP mRNA level. Thus TLR4 signals stimulate the ADRP expression both in direct and indirect manners. Pycnogenol (PYC), an extract of French maritime pine, suppressed the expression of ADRP and the above-mentioned cytokines. PYC suppressed the ADRP promoter activity and enhancer activity of AP-1 and NF-kappaB, whereas it did not affect the LPS-induced DNA binding of these factors. In conclusion, TLR4-mediated signals stimulate the ADRP expression in macrophages while PYC antagonizes this process. PYC, a widely used dietary supplement, might be useful for prevention of atherosclerosis.

We analyzed low molecular weight organic acids and aldehydes in throughfall under pine forest, and organic acids in dew on chemically inert surfaces and pine needle surfaces at urban- and mountain-facing sites of pine forest in western Japan. Low molecular weight organic acids and aldehydes accounted for less than 5% of the dissolved organic carbon in throughfall at both sites. Formaldehyde at both sites and formate at the mountain-facing site were found at significantly lower concentrations in throughfall than in rainfall, which may be explained by the degradation and/or retention of these components by the pine canopy as the incident precipitation passed through it. The oxalate concentration in throughfall was significantly higher than those in rainfall at both sites, suggesting that oxalate was derived from leaching from the pine foliage. At both sites, organic acid concentrations were higher in dew on the pine needles than in throughfall or dew on chemically inert surfaces. This could be due to the long contact time of dew on pine needles, during which leached substances from pine needles and dry deposits accumulated on their surfaces can dissolve into the small volume of dew. The role of enhanced concentrations of oxalate in an aqueous phase on the plant surfaces (e.g., dew) is discussed in relation to hydroxyl radical formation via the photo-Fenton reaction.

Recent reports have demonstrated that erythroid progenitor cells contain and secrete various angiogenic cytokines. Here, the impact of erythroid colony-forming cell (ECFC) implantation on therapeutic angiogenesis was investigated in murine models of hindlimb ischemia. During the in vitro differentiation, vascular endothelial growth factor (VEGF) secretion by ECFCs was observed from day 3 (burst-forming unit erythroid cells) to day 10 (erythroblasts). ECFCs from day 5 to day 7 (colony-forming unit erythroid cells) showed the highest VEGF productivity, and day 6 ECFCs were used for the experiments. ECFCs contained larger amounts of VEGF and fibroblast growth factor-2 (FGF-2) than peripheral blood mononuclear cells (PBMNCs). In tubule formation assays with human umbilical vein endothelial cells, ECFCs stimulated 1.5-fold more capillary growth than PBMNCs, and this effect was suppressed by antibodies against VEGF and FGF-2. Using an immunodeficient hindlimb ischemia model and laser-Doppler imaging, we evaluated the limb salvage rate and blood perfusion after intramuscular implantation of ECFCs. ECFC implantation increased both the salvage rate (38% vs. 0%, P < 0.05) and the blood perfusion (82.8% vs. 65.6%, P < 0.01). In addition, ECFCs implantation also significantly increased capillaries with recruitment of vascular smooth muscle cells and the capillary density was 1.6-fold higher than in the control group. Continuous production of human VEGF from ECFCs in the skeletal muscle was confirmed at least 7 days after the implantation. Implantation of ECFCs promoted angiogenesis in ischemic limbs by supplying angiogenic cytokines (VEGF and FGF-2), suggesting a possible novel strategy for therapeutic angiogenesis.

Thrombin and other proteinases exert vascular effects by activating the proteinase-activated receptors (PARs). The expression of PARs has been shown to be upregulated after balloon injury and in human arteriosclerosis. However, the relationship between the receptor upregulation and the alteration of vasomotor function remains to be elucidated. We herein demonstrated that the contractile responses to the PAR-1 and PAR-2 agonist were markedly enhanced in the rabbit femoral arteries after balloon injury. Neointimal thickening was established 4 wk after the injury. No histological change was observed in the sham operation, where the saphenous artery was ligated without any balloon injury. The contractile response to K(+) depolarization was significantly attenuated 1 wk after the injury and then partly recovered after 4 wk. Thrombin, PAR-1-activating peptide, trypsin, and PAR-2-activating peptide induced no significant contraction in the control. All these stimulants induced enhanced responses 1 wk after balloon injury. Such enhanced responses were seen 4 wk after the injury, except for thrombin. There was no change in the Ca(2+) sensitivity of the contractile apparatus as evaluated in the permeabilized preparations. PAR-1-activating peptide (100 mumol/l), but no other stimulants, induced an enhanced contraction in the sham operation. The expression of PAR-1 and PAR-2 slightly increased after the sham operation, whereas it markedly and significantly increased after balloon injury. Our observations suggest that balloon injury induced the receptor upregulation, thereby enhancing the contractile response before the establishment of vascular lesions. The local inflammation associated with the sham operation may also contribute to the receptor upregulation.

Acoustic pulse echoes generated by femtosecond laser irradiation were detected using time-resolved x-ray triple-crystal diffractometry. The determined time-dependent longitudinal strain component for pulse echoes in silicon and gallium arsenide plates showed that the polarity of the strain pulse was dependent on the optically induced initial stress, and that the bipolar pulse waveform was gradually deformed and broadened in the course of propagation. The three-dimensional wave front distortion of pulse echoes was shown simply as the pulse duration broadening, which was consistent with a boundary roughness for an unpolished plate.

A shortage of organic substances (COD) may cause problems for biological nutrient removal, that is, lower influent COD concentration leads to lower nutrient removal rates. Biological phosphorus removal and denitrification are reactions in which COD is indispensable. As for biological simultaneous nitrogen and phosphorus removal systems, a competition problem of COD utilisation between polyphosphate accumulating organisms (PAOs) and non-polyphosphate-accumulating denitrifiers is not avoided. From the viewpoint of effective utilisation of limited influent COD, denitrifying phosphorus-removing organisms (DN-PAOs) can be effective. In this study, DN-PAOs activities in modified UCT (pre-denitrification process) and DEPHANOX (post-denitrification process) wastewater treatments were compared. In conclusion, the post-denitrification systems can use influent COD more effectively and have higher nutrient removal efficiencies than the conventional pre-denitrification systems.

The study of optical illusion is an important method to elucidate the mechanism of visual perception. However, many details about the cause of optical illusions are still unclear. In this research, based on the characteristic of the physiological structure of the retina, we proposed an on-center receptive field model of the retina. Using this model, we simulated the distributions of the inducing field of some visual stimulus. Comparing to the past studies' results, the validity of the proposed model was proofed. Furthermore, we simulated the distributions of the inducing field of some typical illusions. The simulation results can explain these illusion phenomenon rationally. Therefore, it suggested that some of illusions are probably engendered by the distributions of the inducing field in the retina which generated by the illusions stimuli. The practicality of the proposed model was also verified.

Gene transfer to vein grafts has therapeutic potential to prevent late graft failure; however, certain issues, including efficacy and safety, have hindered the clinical application of this treatment modality. Here, we report the successful and efficient gene transfer of plasmid DNA via ex vivo electroporation into veins as well as into vein grafts. Two approaches were used: one involved transluminal in situ gene transfer using a T-shaped electrode (the "Lu" method), and the other was an adventitial ex vivo approach using an electroporation cuvette followed by vein grafting (the "Ad" method). The Lu method was carried out at 10 V, with optimal gene transfer efficiency in the in situ jugular veins of rabbits, and transgene expression was observed primarily in endothelial cells. However, when these veins were grafted into the arterial circulation, no luciferase activity was detected; this effect was probably due to the elimination of the gene-transferred cells as a result of endothelial denudation. In contrast, optimal and satisfactory gene transfer was obtained with the vein grafts subjected to the Ad method at 30 V, and transgene expression was seen primarily in adventitial fibroblasts. Gene transfer of endothelial nitric oxide synthase cDNA to the vein graft via the Ad method successfully limited the extent of intimal hyperplasia, even under hyperlipidemic conditions, at 4 wk after grafting. We thus propose that the Ad method via ex vivo electroporation may provide a novel, safe, and clinically available technique for nonviral gene transfer to sufficiently prevent late graft failure.

Reduced nitric oxide (NO) in the brain might contribute to enhanced sympathetic drive in heart failure (HF). The aim of this study was to determine whether increased NO production induced by local overexpression of endothelial NO synthase (eNOS) in the nucleus tractus solitarius (NTS) of the brain stem reduces the enhanced sympathetic drive in mice with HF. Myocardial infarction (MI) was induced in mice by ligating the left coronary artery. MI mice exhibited left ventricular dilatation and a reduced left ventricular ejection fraction. Urinary norepinephrine excretion in MI mice was greater than that in sham-operated mice, indicating that sympathetic drive was enhanced in this model. Thus this model has features that are typical of HF. Western blot analysis and immunohistochemical staining for neuronal NOS (nNOS) indicated that nNOS protein expression was significantly reduced in the brain stem of MI mice. MI mice had a significantly smaller increase in blood pressure evoked by intracisternal injection of N(G)-monomethyl-L-arginine than sham-operated mice. Adenoviral vectors encoding either eNOS (AdeNOS) or beta-galactosidase (Adbeta gal) were transfected into the NTS to examine the effect of increased NO production in the NTS on the enhanced sympathetic drive in HF. After the gene transfer, urinary norepinephrine excretion was reduced in AdeNOS-transfected MI mice but not in Adbeta gal-transfected MI mice. These results indicate that nNOS expression in the brain stem, especially in the NTS, is reduced in the MI mouse model of HF, and increased NO production induced by overexpression of eNOS in the NTS attenuates the enhanced sympathetic drive in this model.

Crohn's disease is an idiopathic inflammatory condition. However, little is known about the changes that occur in the muscularis externa, despite the fact that this tissue contributes to motility changes and stricture formation. We characterized immune activity in the muscularis externa from intestinal segments of Crohn's disease patients and evaluated the role of IL-4 and -13 as well as signal transducer and activator of transcription (STAT)6 in the contractility of the cultured human intestinal smooth muscle cells. CD3+ve cells (P < 0.01) and IL-4 protein (P < 0.01) were significantly increased in the muscularis externa of Crohn's disease patients compared with noninflamed controls. Preincubation of human cultured smooth muscle cells with IL-4 (P < 0.001) or IL-13 (P < 0.05) significantly enhanced carbachol-induced contraction, and this was significantly inhibited by the STAT6 inhibitor leflunomide (P < 0.0001). A similar profile was observed in muscle cells isolated from Crohn's disease patients. Both IL-4 and IL-13 increased specific STAT6-DNA binding in control cells, and this was inhibited by anti-STAT6 Ab (P < 0.05) or leflunomide (P < 0.05). IL-4 and IL-13 mediate the hypercontractility of intestinal muscle via a STAT6 pathway at the level of the smooth muscle cell. The STAT6 pathway may contribute to the hypercontractility of intestinal muscle in Crohn's disease.

Monocyte chemoattractant protein-1 (MCP-1) is a proinflammatory chemokine and may play an important role in the development of pulmonary fibrosis. We examined a new therapeutic strategy that comprises the transfection of the mutant MCP-1 gene into skeletal muscles as a biofactory for anti-MCP-1 therapy against bleomycin-induced pulmonary fibrosis in mice. Overexpression of the mutant MCP-1 gene at 10-14 days after intratracheal instillation of bleomycin resulted in decreased DNA damage, apoptosis, and pulmonary fibrosis at 14 days. However, overexpression of the mutant MCP-1 at 0-4 days after bleomycin instillation did not result in decreased pathological grade, DNA damage, or apoptosis at 7 and 14 days. Because, in this model, inflammatory cell infiltration begins at 3 days and is followed by interstitial fibrosis, it is likely that MCP-1 has an important role to play in the development of fibrogenesis but not in the development of early lung inflammation. This method does not require the use of viral vector or neutralizing antibody, and, as such, it is possible to avoid problems regarding the pathogenicity of the viral vector or immunocomplex. This new strategy may be a beneficial method of treating pulmonary fibrosis from the viewpoint of clinical application.

Alveolar epithelial cells are known to be present at the primary site of lung damage in pulmonary fibrosis. Apoptosis has been implicated as being involved in epithelial cell damage and pulmonary fibrosis. Because the cyclin-dependent kinase inhibitor p21 induces G1 arrest and DNA repair and because it also prevents apoptosis in some cells, we hypothesized that p21 gene transfer may attenuate bleomycin-induced pulmonary fibrosis in mice, the pathogenesis of which likely involves epithelial cell apoptosis. Human p21 protein was expressed in mouse alveolar epithelial cells at 1-7 days in vitro and was detected predominantly in lung epithelial cells at 1-7 days in vivo after adenoviral transfer of the human p21 gene. Inflammatory cell infiltration and fibrosis had already begun at 7 days in this model. Adenoviral transfer of the human p21 gene at 7 days after intratracheal instillation of bleomycin led to a decrease in the number of apoptotic cells, lung inflammation, and fibrosis at 14 days. Therefore, the forced expression of p21 exerted both anti-apoptotic and anti-fibrotic effects, which would facilitate the ultimate goal of treatment for pulmonary fibrosis.

We have recently demonstrated the direct involvement of the death receptor-mediated apoptotic pathways in cisplatin-induced renal tubular cell (RTC) death. Reactive oxygen species are thought to be a major cause of cellular damage in such injury. The aim of this study was to examine the mechanism through which antioxidants ameliorate cisplatin-induced RTC death, with special emphasis on death receptor-mediated apoptotic pathways. Cisplatin was added to cultures of normal rat kidney (NRK52E) cells or injected in rats. NRK52E cells and rats were also treated with dimethylthiourea (DMTU), a hydroxyl radical scavenger. We then examined the mRNA levels of death ligands and receptors, caspase-8 activity, cell viability, cell death, renal function, and histological alterations. RT-PCR indicated cisplatin-induced upregulation of Fas, Fas ligand, and TNF-alpha mRNAs and complete inhibition by DMTU in vitro and in vivo. Cisplatin increased caspase-8 activity of NRK52E cells, and DMTU prevented such activation. Exposure to cisplatin reduced viability of NRK52E cells, examined by WST-1 assay, and increased apoptosis and necrosis of the cells, examined by terminal deoxynucleotidyl transferase dUTP nick-end labeling assay and fluorescence-activated cell sorter analysis. DMTU abrogated cisplatin-induced changes in cell viability and apoptosis and/or necrosis. Cisplatin-induced renal dysfunction and histological damage were also prevented by DMTU. DMTU did not hinder cisplatin incorporation into RTCs. Our results suggest that antioxidants can ameliorate cisplatin-induced acute renal failure through inactivation of the death receptor-mediated apoptotic pathways.

The effects of various fractions of copper (Cu) and zinc (Zn) on soil bacteria were evaluated by the heavy metal tolerance level of the bacterial community (IC50) in soil samples collected near a mine. The IC50 values had no relationship with the total concentrations of Zn and Cu in the soils, but were weakly correlated with the 0.05 M CaCl2-extractable form of each metal in the soils (Cu: R2 = 0.670, p < 0.01; Zn: R2 = 0.453, p < 0.05). It was found that the IC50 correlated strongly with the total concentration of each metal in the extracts from water-saturated soil samples, described below as "soil solution" (Cu: R2 = 0.789, p < 0.01; Zn: R2 = 0.617, p < 0.01). The speciation of these metals in the soil solutions was estimated using an equilibrium thermodynamic computer model, SOILCHEM. Simulated free Cu ion ranged from 18 to 98% of total Cu, and organic complexes of Cu ranged from < 1 to 56%. In all samples, Zn existing as the free ion was estimated to be more than 80% of total Zn in the soil solutions. The IC50 values were also correlated with the estimated free metal ion activities, but with slightly lower correlation coefficients than found for total concentration in the soil solutions (Cu: R2 = 0.735, p < 0.01; Zn: R2 = 0.610, p < 0.01). The results suggest that not only high metal ion activities, but also total dissolved metal concentrations in soil solutions may affect the bacterial community.

A new control method of nonlinear dynamic systems is proposed based on the impulse responses of universal learning networks (ULNs), ULNs form a superset of neural networks. They consist of a number of interconnected nodes where the nodes may have any continuously differentiable nonlinear functions in them and each pair of nodes can be connected by multiple branches with arbitrary time delays. A generalized learning algorithm is derived for the ULNs, in which both the first order derivatives (gradients) and the higher order derivatives are incorporated. One of the distinguished features of the proposed control method is that the impulse response of the systems is considered as an extended part of the criterion function and it can be calculated by using the higher order derivatives of ULNs. By using the impulse response as the criterion function, nonlinear dynamics with not only quick response but also quick damping and small steady state error can be more easily obtained than the conventional nonlinear control systems with quadratic form criterion functions of state and control variables.

Universal learning networks (ULNs) and robust control system design are discussed, ULNs provide a generalized framework to model and control complex systems. They consist of a number of interconnected nodes where the nodes may have any continuously differentiable nonlinear functions in them and each pair of nodes can be connected by multiple branches with arbitrary time delays. Therefore, physical systems which can be described by differential or difference equations and also their controllers can be modeled in a unified way. So, ULNs constitute a superset of neural networks or fuzzy neural networks. In order to optimize the systems, a generalized learning algorithm is derived for the ULNs, in which both the first order derivatives (gradients) and the higher order derivatives are incorporated. The derivatives are calculated by using forward or backward propagation schemes. These algorithms for calculating the derivatives are extended versions of back propagation through time (BPTT) and real time recurrent learning (RTRL) by Williams in the sense that generalized nonlinear functions and higher order derivatives are dealt with. As an application of ULNs, the higher order derivative, one of the distinguished features of ULNs, is applied to realizing a robust control system in this paper. In addition, it is shown that the higher order derivatives are effective tools to realize sophisticated control of nonlinear systems. Other features of ULNs such as multiple branches with arbitrary time delays and using a priori information will be discussed in other papers.

There have been many applications of the Hilbert curve, such as image processing, image compression, computer hologram, etc. The Hilbert curve is a one-to-one mapping between N-dimensional space and one-dimensional (l-D) space which preserves point neighborhoods as much as possible. There are several algorithms for N-dimensional Hilbert scanning, such as the Butz algorithm and the Quinqueton algorithm. The Butz algorithm is a mapping function using several bit operations such as shifting, exclusive OR, etc. On the other hand, the Quinqueton algorithm computes all addresses of this curve using recursive functions, but takes time to compute a one to-one mapping correspondence. Both algorithms are complex to compute and both are difficult to implement in hardware. In this paper, we propose a new, simple, nonrecursive algorithm for N-dimensional Hilbert scanning using look-up tables. The merit of our algorithm is that the computation is fast and the implementation is much easier than previous ones.

According to recent knowledge of brain science it is suggested that there exists functions distribution, which means that specific parts exist in the brain for realizing specific functions. This paper introduces a new brain-like model called Learning Petri Network (LPN) that has the capability of functions distribution and learning. The idea is to use Petri net to realize the functions distribution and to incorporate the learning and representing ability of neural network into the Petri net. The obtained LPN can be used in the same way as a neural network to model and control dynamic systems, while it is distinctive to a neural network in that it has the capability of functions distribution. An application of the LPN to nonlinear crane control systems is discussed. It is shown via numerical simulations that the proposed LPN controller has superior performance to the commonly-used neural network one.

Choriocarcinoma developed from a complete hydatidiform mole has an unique genetic feature that involves monoallelic contribution from the paternal genome. To determine the chromosome carrying putative tumor suppressor gene(s), microcell-hybrids were isolated following fusion of choriocarcinoma cells with microcells from mouse A9 cells containing a single human chromosome (1, 2, 6, 7, 9 or 11). Microcell-hybrids with the introduction of chromosome 7 were suppressed or modulated for tumorigenicity and exhibited altered in vitro growth properties. Introduction of chromosomes 1, 2, 6, 9 or 11 had no effect. Tumorigenic revertants isolated from microcell-hybrids with the introduced chromosome 7 contains reduced numbers of chromosome 7. These findings suggest that chromosome 7 contains a putative tumor suppressor gene(s) for choriocarcinoma. Alterations in tumorigenic phenotypes seen in microcell-hybrids were not associated with the presence of either ERV3 or H-plk locus located on the introduced chromosome 7, indicating the putative tumor suppressor gene(s) is outside of ERV3 and H-plk gene loci. Furthermore, we obtained evidence to define a critical region on chromosome 7 (7p12-7q11.23) that was frequently lost in surgically removed choriocarcinoma tissues and cell lines. Using a panel of microsatellite markers, biallelic deletions were observed, which strongly suggests the presence of a tumor suppressor gene(s) within this critical region.

We examined argyrophil nuclear organizer regions (AgNOR) and proliferating cell nuclear antigen (PCNA) in 41 patients with surgically-treated thymic disease. AgNOR count and PCNA labeling index (LI) in thymic carcinoma were significantly higher than those in thymoma and thymic hyperplasia. A positive correlation was observed between the PCNA LI and argyrophilic nucleolar organizer region (AgNOR) counts within all thymic disease (r=0.31, P=0.002). The PCNA LI in invasive thymoma was lower than that in non-invasive thymoma. In survival analysis, the cut-off values for the PCNA LI and AgNOR count were chosen to produce two categories with equal numbers of 26 thymoma patients. There were no significant difference in the survival rate between the lower and higher group patients in relation with AgNOR count and PCNA LI. We conclude that combining AgNOR and PCNA may discriminate the biological activity of thymic disease. These staining methods can be performed with ease and, applied in a clinical laboratory on a routine basis to help predict cytological malignancy of thymic disease.

Dukes' classification for colorectal cancer is easy to remember and accurate in estimating survival. The Japanese definition of early gastric cancer have some problems, since it includes node-positive cases showing a worse prognosis. The aim of this study was to clarify whether or not Dukes' A tumor can be used as new criteria for early gastric cancer. Using 217 patients who underwent radical gastrectomy and lymph node dissection for primary gastric adenocarcinoma, long-term results and cumulative survival rates after operation were examined. Patients consisted of three groups: Group 1, 20 patients with node-negative tumor invading down to the muscularis propria, Group 2, 175 patients with node-negative tumor limited to the mucosa or submucosa, and Group 3, 22 patients with node-positive tumor limited to the mucosa or submucosa. The 10-year survival rates for Group 1, Group 2, and Group 3 patients were 94.7%, 96.9%, and 85.1%, respectively. The survival rate of Group 3 patients was significantly lower than that of Group 1 and Group 2 patients (p<0.05). The prognosis of patients with Dukes' A tumor (Group 1 plus Group 2) was excellent, showing 10-year survival rate of 96.6%. Mucosal or submucosal cancer brought a poor prognosis when there was lymph node metastasis. Early gastric cancer would better be defined as a Dukes' A tumor which includes node-negative tumor not beyond the muscularis propria.

Integrin alpha 6 is believed to be involved in malignant biological development. High tumor/normal (T/N) ratio of integrin alpha 6 was observed in intestinal type tumors in gastric carcinoma. In an immunohistochemical study, an intense and continuous staining was seen at the basement layer in the intestinal type tumors. Concerning the variant, there was a significant correlation between the expression ratio of variant B/variant A and the depth of tumor invasion. In conclusion, i) integrin alpha 6 expression was intimately correlated with histologic differentiation, and ii) the overexpression of variant B may be correlated with tumor invasion of gastric carcinoma.

p21(WAF1/CIP1) is a potent inhibitor of various cyclin-dependent kinases, the expression of which is transcriptionally regulated by tumor suppressor gene product p53. We immunohistochemically examined the expression of p53 and p21(WAF1/CIP1) in 61 esophageal squamous cell carcinomas. p53 protein was expressed in 37 (61%) of 61 carcinomas. p21(WAF1/CIP1) was consistently expressed in the normal stratified esophageal mucosa. In the carcinomas, the expression of p21(WAF1/CIP1) protein was markedly reduced or not expressed in 33 (54%) cases. Clinicopathologic analyses revealed that no significant correlation exists either between p53-positive and -negative cases or between p21(WAF1)/(CIP1)-positive and -negative cases. Twenty-four cases were p53-positive/p21(WAF1/CIP1) negative, 15 were p53-negative/p21(WAF1/CIP1)-positive, 13 were positive for both and 9 were negative for both, and these findings thus showed an inverse correlation of the positivity between p53 and p21(WAF1/CIP1) (p<0.05). Furthermore, of the 13 cases with positive staining for both, the distribution of the expression was mutually exclusive in 6 cases and coincidental in the remaining 7 cases. These findings showed the p53-dependent expression of p21(WAF1/CIP1) was observed in esophageal squamous cell carcinomas, while the lack of an absolute correlation between abnormal p53 expression and p21(WAF1/CIP1) expression suggested that the p53-independent expression of p21(WAF1/CIP1) might also occur in some portions of the esophageal squamous carcinomas, suggesting that the molecular mechanisms of esophageal carcinogenesis appear to be complicated.

Aggressive angiomyxoma is a distinctive and very rare soft tissue tumor occurring almost exclusively in women. Only seven cases occurring in men have been previously reported. We herein report a case of a 74-year-old man who was admitted to our hospital presenting with a huge right inguinal mass that had grown over a 14-year period, in order to undergo surgical treatment for gastric cancer. The inguinal tumor was well-defined and demonstrated a glistening appearance on the cut surface. Histologically, the tumor was composed of bland-looking spindle and stellate cells with delicate cytoplasmic processes, which sparsely populated the fibromyxoid matrix. A prominent vascular component was also present. Immunohistochemically, the stromal cells stained consistently for vimentin and variably for muscle-specific actin, but not for alpha-smooth muscle actin, desmin, and S-100 protein. The gastric cancer was microscopically diagnosed to be papillary adenocarcinoma with serosal invasion and showing immunoreactivity for p53 protein, but not for aggressive angiomyxoma.

Cytochrome p4501A1 gene (CYP1A1) and glutathione S-transferase mu gene (GSTM1) are involved in the metabolic activation or detoxification of environmental carcinogens including benzo[a]pyrene in tobacco smoke. Individuals with both Val/Val and C type of CYP1A1 (CYP1A1; Val/Val and CYP1A1; C) or homozygous null (-/-) genotype of GSTM1 gene (GSTM1; -/-) show increased susceptibility to lung cancer. The incidence of p53 gene mutations are related to the smoking index of the lung cancer patients. Therefore we determined genotypes of these enzymes and screened p53 gene mutations in 123 non-small cell lung cancer (NSCLC) patients. p53 gene mutations were found in 35% (43/123) of the patients. The incidence of p53 gene mutation CYP1A1; Val/Val (60.0%), CYP1A1; C (50.0%) tended to be higher than those of CYPIAI; Ile/Ile and Ile/Val (40.4%) or CYP1A1; A and B (40.5%). We conclude that the incidence of the p53 mutations does not seem to be significantly affected by only CYP1A1 or GSTM1 polymorphisms in lung cancer patients.

Metastasis to the inguinal lymph node from early gastric cancer is rare. We report a case of 63-year-old Japanese male with gastric mucosal cancer which metastasized to the left inguinal lymph nodes 4 years after a curative gastrectomy. The importance of routine examination of inguinal lymph nodes for cases with gastric cancer is suggested.

The human gene MAGE encodes tumor specific peptide antigens and consists of at least 12 families. Some antigens coded by the MAGE genes may be potentially useful for cancer specific immunotherapy. There is, however, so far little information on the expression of these gene families in human esophageal carcinomas. We investigated the expression of MAGE-1, -2, -3, -4, -6, -8, -9, -10, -11, and -12 genes in 24 human esophageal carcinoma cell lines, and in 50 pairs of tumor and corresponding normal tissue specimens from the human esophagus by means of reverse transcription polymerase chain reaction. The expression rate varied from 13% of MAGE-6 and 8 to 79% of MAGE-4 in the esophageal carcinoma cell lines, and from 6% of MAGE-6 to 62% of MAGE-4 in clinical tumor samples. The most frequently and the least expressed gene were the MAGE-4 and MAGE-6 genes, respectively, in both the cell lines and the clinical samples. Forty-seven of the 50 clinical tumors expressed at least one MAGE gene. No significant clinicopathologic difference between the tumor cases was observed, regardless of the presence or absence of MAGE gene expression. The findings of this study thus demonstrated that the MAGE gene family is frequently expressed in clinical samples as well as in the cell lines of human esophageal carcinomas. Therefore, to identify the MAGE gene family may be useful, not only for esophageal tumor specific immunotherapy but for molecular diagnostic usage as well.

The expression of pyrimidine nucleoside phosphorylase (PyNPase) mRNA in tumor (T) and normal (N) biopsy specimens obtained from 41 cases of colorectal carcinoma was examined by the reverse transcriptase/polymerase chain reaction (RT-PCR). The higher T/N ratio of the expression of PyNPase mRNA correlated significantly with the presence of lymph vessel invasion (p=0.039), the positive lymph node metastasis (p=0.014) and the advanced stage of the disease (p=0.014). There was a significant con-elation between the results determined by enzyme activity and those determined by RT-PCR (p=0.005). The findings suggested that the determination of PyNPase mRNA by RT-PCR may give useful information on tumor aggressiveness of colorectal carcinoma and this method can be used instead of enzyme activity.

We determined if any extracellular matrix degradative proteases were possibly associated with metastatic potentials of human pancreatic cancer cells. Liver metastatic abilities of six human pancreatic cancer cell lines were examined in nude mice, and divided into two high (2 lines) and low (4 lines) metastatic cell lines. Of six cell lines, matrix metalloproteinases-1 (MMP-1) was overexpressed in two high metastatic cell lines: and MMP-2 was overexpressed in one high metastatic cell line. Of the four low metastatic lines, two cell lines had relatively higher mRNA levels of tissue inhibitors of metalloproteinases-3 (TIMP-3). MMP activities due to MMP-1 and MMP-2 might be positively associated with liver metastasis of pancreatic cancer cells. Moreover, expression of TIMP-3 might be partly involved in the low metastatic potentials of pancreatic cancer.

Recent studies have shown that microsatellite instability (MSI) play an important role in the development of various types of cancer. To clarify the clinicopathologic significance of MSI in colorectal carcinoma (CRC), the presence of MSI was examined in 54 Japanese cases of CRC using the polymerase chain reaction-based method. The incidence of MSI in CRC cases was 13 out of 54 cases (24%). CRC with MSI also showed a significant tendency not to have lymph node metastasis (P<0.05), although neither the survival nor the prognosis of the cases examined in this study were available due to the short period of follow-up. The present study showed that the incidence of MSI in Japanese CRC was 24% and suggests that CRC with MSI may behave in a less malignant manner.

We examined the optimal duration of whole body hyperthermia (WBH, 41.5 degrees C) when administered simultaneously with cis-diamminedichloroplatinum(II) (CDDP) using F344 rat fibrosarcoma model. The antitumor efficacy was measured by tumor growth delay (TGD) and nephrotoxicity was evaluated by the day 5 blood urea nitrogen (BUN) associated with different duration of WBH (0.5, 1, 1.5, 2 or 3 h) with a constant dose of CDDP (2 mg/kg i.v. bolus). There was significant increase in TGD from 2.8 days to 5.4 days when the WBH duration was increased from 0.5 to 1 h (p<0.001). CDDP-induced nephrotoxicity was also enhanced by WBH. We estimated specific therapeutic efficacy (STE) for each treatment using the ratio of antitumor effect measured by TGD to nephrotoxicity calculated from the degree of increment of BUN. CDDP combined with 1 h WBH produced the best STE of 2.2. We conclude, that to maximize the therapeutic gain, a 1 h duration is optimal in the present setting of thermochemotherapy.

Vesnarinone, (3,4-dihydro-6-[4-(3,4-dimethoxybenzoyl)-1- piperazinyl]-2(1H)-quinolinone), a quinolinone derivative, is a positive inotropic agent. We examined the cytotoxicity either by vesnarinone alone or in combination with doxorubicin (DXR), in vitro. The cytotoxic effect of vesnarinone against HL-60 cells did not increase, even at concentrations as high as (50 mu g/ml). The cytotoxicity of DXR, however, was enhanced after being combined with 30 mu g/ml of vesnarinone. The intracellular level of DXR increased when DXR was administered after incubation with vesnarinone and the efflux of DXR was delayed when the cells were incubated in the presence of vesnarinone after DXR exposure. Flow cytometry showed that the combination of DXR and vesnarinone increased the cell population below the G(0)/G(1) region. Vesnarinone induced DNA ladder formation, but only when these cells were incubated for 72 h, while in addition, when DXR was combined with vesnarinone, the DNA ladder formation was enhanced. Based on the above findings, we thus conclude that the cytotoxicity of DXR was enhanced when combined with vesnarinone.

Telomeres of human chromosomes consist of a repeated TTAGGG sequence, and at the terminal of this repeat sequence, the 3' strand is longer than the 5' strand. In this study, we characterized single and double stranded telomere binding proteins (ssTBPs and dsTBPs) by gel mobility shift assay and South-Western blotting assay. At least two protein components with molecular weights of 29 and 33 kDa were bound to a single stranded telomeric sequence, and also two proteins with molecular weights of about 44 kDa and 70 kDa were bound to a double stranded telomeric sequence. A competition assay demonstrated that the binding properties of ssTBPs and dsTBPs were specific to the telomeric sequence. We further cloned a ssTBP cDNA (ssTBP-1) by screening a lambda-gt11 expression library and identified ssTBP-1 as a human heterogeneous nuclear ribonucleoprotein (hnRNP) Al on the basis of cDNA sequence. We also found that the expression of the hnRNP Al gene significantly decreased during in vitro passage of human microvascular endothelial cells.