Hes1 suppresses acute myeloid leukemia development through FLT3 repression.

In leukemogenesis, Notch signaling can be up and downregulated in a context-dependent manner. The transcription factor hairy and enhancer of split-1 (Hes1) is well-characterized as a downstream target of Notch signaling. Hes1 encodes a basic helix-loop-helix-type protein, and represses target gene expression. Here, we report that deletion of the Hes1 gene in mice promotes acute myeloid leukemia (AML) development induced by the MLL-AF9 fusion protein. We then found that Hes1 directly bound to the promoter region of the FMS-like tyrosine kinase 3 (FLT3) gene and downregulated the promoter activity. FLT3 was consequently upregulated in MLL-AF9-expressing immortalized and leukemia cells with a Hes1- or RBPJ-null background. MLL-AF9-expressing Hes1-null AML cells showed enhanced proliferation and ERK phosphorylation following FLT3 ligand stimulation. FLT3 inhibition efficiently abrogated proliferation of MLL-AF9-induced Hes1-null AML cells. Furthermore, an agonistic anti-Notch2 antibody induced apoptosis of MLL-AF9-induced AML cells in a Hes1-wild type but not a Hes1-null background. We also accessed two independent databases containing messenger RNA (mRNA) expression profiles and found that the expression level of FLT3 mRNA was negatively correlated with those of HES1 in patient AML samples. These observations demonstrate that Hes1 mediates tumor suppressive roles of Notch signaling in AML development, probably by downregulating FLT3 expression.

Treatment of hypertension in hemodialysis patients with nifedipine.

The hemodynamic effects of nifedipine were studied in nine patients with acute or chronic renal failure undergoing hemodialysis. Arterial blood pressure was lowered within 15 minutes of oral administration of nifedipine 10 mg. Mean arterial pressure and total systemic peripheral resistance were significantly decreased, whereas cardiac index and heart rate revealed slight but statistically insignificant changes. Changes in pulmonary artery diastolic pressure and mean right atrial pressure which were closely related to cardiac preload were not significant. However, the antihypertensive effect was enhanced in patients with lower pretreatment preload. Results show that oral nifedipine is effective in rapidly reducing blood pressure during or following hemodialysis.

Direct detection of hepatitis B virus gene integrated in the Alexander cell using fluorescence in situ polymerase chain reaction.

Prolonged infection of hepatitis B virus (HBV) is reported to cause hepatocellular carcinoma (HCC) via liver cirrhosis. However, it is still unknown whether the HCC is induced by the HBV DNA integration or by inflammatory stimulation during the phase of liver cirrhosis. The aim of this study is to determine the intracellular or intranuclear distribution of HBV DNA with a highly sensitive assay. Here we directly detected the integration of HBV DNA by fluorescence in situ polymerase chain reaction (FISPCR). Since FISPCR products directly incorporate rhodamine-4-dUTP, the nucleus of Alexander cells integrated with HBV gene reacted with the HBV primers emits obvious fluorescence. The fluorescence values which were measured with an imaging analyzer show a significant difference between Alexander cells as compared to the controls. In conclusion, the target sequences of HBV were specifically amplified as fluorescent DNA after the present FISPCR procedure. This method could provide a novel and simple strategy for determining the quantitative role of viral DNA integration in oncogenesis.

Hydrogen peroxide-induced cellular injury is associated with increase in endogenous fluorescence from rat gastric mucosal epithelial cell culture: A new method for detecting oxidative cellular injury by fluorescence measurement.

To develop a new method of detecting cellular injury caused by oxygen radicals, we studied endogenous fluorescence from the cultured cells of a rat gastric mucosal epithelial cell line. Measurement with an ultra-high sensitivity camera-image processor system under an inverted epifluorescence microscope showed that the fluorescence intensity of the cells increased time- and dose-dependently after the addition of hydrogen peroxide (H2O2), an oxygen radical precursor, to the medium. This increase was inhibited by the presence of catalase. Phase-contrast and fluorescence microscopy revealed that the fluorescence was emitted from granular substances in the cytoplasm of the injured cells. The spectral pattern of excitation and emission indicated that the fluorescent substances were flavins. In cell-free experiments, glutathione reductase which has flavin adenine dinucleotide (FAD) at the active site, increased in fluorescence after incubation with H2O2 in the presence of reduced glutathione and glutathione peroxidase. These findings indicate that FAD in the cytoplasm of cells injured by H2O2 increased in endogenous fluorescence according to the extent of injury, and suggest that fluorescence measurement may be a simple method in cellular toxicology to detect oxygen radical-induced injuries.

Autofluorescence in indomethacin-induced gastric mucosal lesions in rats.

Autofluorescence observations enable scientists to sensitively identify various lesions. Non-steroidal anti-inflammatory drugs such as aspirin and indomethacin are well known to induce gastric mucosal injuries. Our purpose was to clarify whether the observation of mucosal autofluorescence could help us to recognize indomethacin-induced gastric lesion formation. Gastric mucosal fluorescence intensity and gastric lesion scores were time-sequentially measured after indomethacin treatment in rats. To identify the localization of autofluorescent substances, stomach cryosections were observed with an epifluorescence microscope. Fluorescent substances from damaged tissue were also analyzed by high-performance liquid chromatography. In addition, to elucidate whether oxidative stress directly generates fluorescent substances from heme, we investigated the reaction between hydrogen peroxide and hemoglobin in a cell-free system. Treatment with indomethacin induced gastric lesions by tissue peroxidation, with mucosal fluorescence intensity increasing time-dependently. The fluorescence products were mesoporphyrin and protoporphyrin, and they were localized in disrupted mucosal tissue. In the cell-free system, porphyrins were directly generated by hydrogen peroxide from hemoglobin. These findings indicate that indomethacin treatment increased the intensity of porphyrin fluorescence. Gastric mucosal lesion formation can be sensitively detected with fluorescence observations.

Bosentan, a novel synthetic mixed-type endothelin receptor antagonist, attenuates acute gastric mucosal lesions induced by indomethacin and HCl in the rat: role of endogenous endothelin-1.

Endothelin-1 has been reported to be responsible for gastric mucosal damage in various experimental models. We evaluated the role of endogenous endothelin-1 in the pathogenesis of gastric mucosal damage induced by indomethacin and HCl in the rat. Rats were given indomethacin (25 mg/kg) subcutaneously, and 15 min later, 0.2N HCl intragastrically. Gastric mucosal damage, gastric endogenous endothelin-1, and gastric mucosal hemodynamics were measured. The effects of bosentan, a mixed endothelin receptor antagonist, on gastric mucosal integrity and hemodynamics were assessed. Gastric endogenous endothelin-1 was significantly elevated at 20 min, gastric mucosal blood flow began to decrease significantly at 25 min, and gastric damage occupied 52.2% of the total glandular mucosa at 135 min after injection of indomethacin. Intragastric pretreatment with bosentan (5, 10, 30, and 60 mg/kg) significantly attenuated gastric damage, to 26.1%, 7.7%, 3.6%, and 1.6%, respectively, of the total glandular mucosa. Bosentan (60 mg/kg) prevented the initial decrease of blood flow and, even at 135 min, improved blood flow and hemoglobin oxygen saturation significantly. We suggest that indomethacin-induced endogenous endothelin-1 diminishes gastric mucosal blood flow and tissue oxygenation and ultimately causes gastric damage. Endogenous endothelin-1 may play an important role in the pathogenesis of the acute gastric mucosal lesions induced by indomethacin and HCl.

Pancreaticoduodenal artery aneurysms associated with celiac axis stenosis due to compression by median arcuate ligament and celiac plexus.

Celiac axis stenosis is frequently associated with pancreaticoduodenal artery aneurysms. Although the cause of stenosis was not clear in most of the reported cases, compression of the median arcuate ligament of the diaphragm was found to be responsible for the stenosis in 7 of 42 reported cases of this type of aneurysm. We report a case of aneurysm caused by compression of the median arcuate ligament of the diaphragm and celiac plexus. An asymptomatic 43-year-old Japanese man was admitted with a low echoic lesion in the uncus of pancreas. Computed tomographic scan and angiogram revealed stenosis of the celiac axis and two aneurysms in the inferior posterior pancreaticoduodenal artery. The celiac plexus and median arcuate ligament were divided surgically and normal flow was reestablished in the celiac axis. One of the aneurysms was resected and the afferent artery of the other aneurysm was ligated. In the setting of pancreaticoduodenal artery aneurysm associated with celiac axis stenosis, management of stenosis should be considered in addition to local treatment of the aneurysm. In this context, division of median arcuate ligament and celiac plexus or aorto-celiac bypass may normalize the flows in the pancreaticoduodenal arcade and could be effective in preventing aneurysm reformation.

Steroid-refractory severe ulcerative colitis responding to cyclosporine and long-term follow-up.

We report a case of steroid-refractory ulcerative colitis, treated with cyclosporine, in a 38-year-old woman with a 13-year history of ulcerative colitis. No remission was achieved with treatments that included intravenous hyperalimentation, sulfasalazine, and intensive parenteral prednisolone therapy for 4 weeks. Intravenous infusion of cyclosporine was performed because the patient refused to undergo surgery. Her condition improved dramatically and colectomy was avoided. She has been maintained on oral cyclosporine and azathioprine since steroids were discontinued, and she has remained in clinical and endoscopic remission for 2 years. The side effects were not significant, but mild paresthesia in both hands and mild hypertension, which was controlled by anti-hypertensives. Cyclosporine seems to be an effective treatment for patients with steroid-refractory severe active ulcerative colitis in whom colectomy seems inevitable. We believe further clinical trials of the treatment are warranted.

The tissue distribution of 2,3,7,8-chlorine substituted dibenzo-p-dioxins in humans who died of cancer.

The tissue distribution of 2,3,7,8-chlorine substituted dibenzo-p-dioxins was conducted in 11 patients who died of cancer. The concentration of octachlorodibenzo-p-dioxin (octa-CDD) was the highest in each organ and tissue and hepta-CDD was also found at relatively high levels, second only to OCDD. The levels of 1,2,3, 7,8-penta-CDD and 1,2,3,6,7,8-hexa-CDD in the spleen were the highest, respectively. 2,3,7,8-Tetra-CDD was also detected and its concentration was the highest in the gonad (0.8-3.2 pg/g-range). From the 2,3,7,8-TCDD toxic equivalent calculations, the highest equivalent value was obtained from a 54-year-old female who died of cancerous goiter. This individual had the highest concentrations of 2,3,7,8-substituted penta- and hexa-CDDs among the 11 patients.

Phosphoramidon, an endothelin converting enzyme inhibitor attenuates local gastric ischemia-reperfusion injury in rats.

Recently increased production of endothelin-1 has been implicated in the pathogenesis of gastric ischemia-reperfusion injury. We have investigated the effects of endothelin converting enzyme inhibition on local gastric ischemia-reperfusion injury in rats by using two metalloprotease inhibitors, phosphoramidon and thiorphan. In presence of exogenous 0.15M HCI intragastrically, local ischemia was induced by the clamping of left gastric artery for 15 min and reperfusion was done for 30 min. In separate group of rats, phosphoramidon (10-60 mg/kg) or thiorphan (60 mg/kg) were given as i.v. bolus injection immediately before the induction of ischemia. Phosphoramidon dose dependently attenuated the macroscopic and microscopic mucosal injuries while thiorphan did not. These results indicate that phosphoramidon-sensitive endothelin converting enzyme activity is highly present in stomach and phosphoramidon, by inhibiting the conversion of big endothelin-1 to endothelin-1 attenuated the gastric mucosal damage in this model.

Phosphoramidon, an inhibitor of endothelin-converting enzyme, prevents indomethacin-induced gastric mucosal damage in rats.

Endothelin-1 (ET-1) is produced from inactive precursor big ET-1 by endothelin-converting enzyme-1 (ECE-1), a membrane-bound metalloprotease, structurally similar to another metalloprotease, neutral endopeptidase 24.11 (NEP). Although both phosphoramidon and thiorphan are metalloprotease inhibitors, the ECE activity is inhibited by phosphoramidon but not by thiorphan, a specific inhibitor of NEP. Therefore, to investigate whether an ECE inhibitor can prevent indomethacin-induced gastric mucosal damage in rats, we compared the effects between the two metalloprotease inhibitors on both gastric mucosal integrity and the levels of ET-1 and big ET-1 in gastric tissue. Phosphoramidon significantly decreased ET-1 levels, causing a concomitant big ET-1 increase and dose-dependently attenuated indomethacin-induced gastric mucosal damage. By contrast, thiorphan neither changed the ratio of ET-1/big ET-1 nor attenuated the damage. In conclusion, the prevention of gastric mucosal damage by an ECE inhibitor indicates that endogenous ET-1 may play an important role in the pathogenesis of indomethacin-induced gastric mucosal damage.

Quantitative scintigraphic analysis of 123I-MIBG by polar map in patients with dilated cardiomyopathy.

To evaluate sympathetic dysfunction of the heart, one of the factors responsible for heart failure in patients with dilated cardiomyopathy (DCM), we performed 123I-metaiodobenzylguanidine (123I-MIBG) myocardial single photon emission tomography (SPET) in 22 cases of DCM. There were significant relationships between severity scores and defect scores for the early and delayed images. When the data acquired by 123I-MIBG myocardial SPET were compared with left ventricular function, the extent score (r = -0.754, P < 0.01) and severity score (r = 0.693, P < 0.01) for the early images were both significantly correlated with left ventricular ejection fraction (LVEF). For the delayed images also, a good correlation was obtained between LVEF and extent score (r = -0.704, P < 0.01) and between LVEF and severity score (r = -0.801, P < 0.01). the washout rate for the entire left ventricle, calculated from the early and delayed images, also correlated with LVEF (r = -0.643, P < 0.01). Since the 123I-MIBG defect on the early images was shown to be significantly related to left ventricular function, a decrease in neuronal uptake at sympathetic nerve endings appears in the main to account for the 123I-MIBG defect on the early images. The regional washout rate was increased compared to that in the control group (P < 0.01). It was particularly high in the inferior wall, suggesting that acceleration of turnover and poor retention in the area of the 123I-MIBG defect on the early images and then the entire left ventricle was strongly associated with the 123I-MIBG defect on the delayed images. Our results suggest that 123I-MIBG myocardial SPET may be useful in determining the severity of DCM.

Simultaneous multielemental analysis of daily food samples by inductively coupled plasma mass spectrometry.

Several toxic elements in total composite food samples of hospital diets were determined simultaneously by inductively coupled plasma mass spectrometry (ICP-MS). Trace element concentrations of Be, Cr, As, Cd, Sn, Sb, Ce, Hg and Pb changed widely ranging from a few ng.g-1 to less than 3 micrograms.g-1 in all 24 samples collected through a year. Be, Sn, and Hg were not detected in rice sample alone, and other trace elements, with the exception of Cr and As, were less than approximately 50% levels, as compared with mixed diet samples. The daily intakes of these toxic elements were in the ranges of a few to several hundreds of micrograms.

Polychlorinated dibenzo-p-dioxins and dibenzofurans in plywood combustion gas.

Polychlorinated dibenzo-p-dioxin and dibenzofuran (PCDD/DF) levels in plywood combustion gas were investigated for the effects of ammonium chloride (NH4Cl) and paint. For combustion systems in which neither NH4Cl nor paint were present in the plywood samples, total amounts of PCDD/DFs in the combustion gas were in the order of 35-529 ng/Nm3, and were higher in the systems with softwood trees than broadleaf ones, depending on the Cl concentrations. For the systems with added NH4Cl and no paint, and those without NH4Cl but were painted, higher PCDD/ DF rates were observed at combustion temperatures of 270 degrees C and 500 degrees C, respectively. However, for the systems with both NH4Cl and paint, their amounts in the range of 0.6-13 ng/Nm3 were the lowest in all systems. The PCDD/DF abundance profiles were similar to their patterns in pentachlorophenol (PCP). Furthermore, it was found that the 2,3,7,8-chlorine substituted penta- and hexa-CDDs contributed more than other 2,3,7,8-chlorine substitutes to toxicity equivalency (TEQ).

PCDD/DF formations by the heterogeneous thermal reactions of phenols and their TiO2 photocatalytic degradation by batch-recycle system.

Polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/DFs) formation by the thermal reactions of phenols with CuCl2 under oxygen flux were carried out in relation to their formation mechanisms. To evaluate the effect of photocatalytic degradation of titanium dioxide (TiO2) thin film prepared by the sol-gel method, the photocatalysis of PCDD/DFs in acetonitrile/water solution by batch-recycle system was conducted. For the thermal reaction system of powder mixtures of 2,4,5-trichlorophenol (2,4,5-TCP) and CuCl2, the formation rates were 8.1 microg/g-2,4,5-TCP/min for total PCDD/DFs and 6.9 microg/g-2,4,5-TCP/min for PCDDs, and total PCDD/DF rate was higher by approximately 40 fold compared to phenol vapor/oxygen/CuCl2 powder system. For the system of 2,4,5-TCP, PCDDs were mainly formed via ortho-phenoxyphenols (POP) intermediate by the condensation of 2,4,5-trichlorophenate. For PCDD/DF photocatalytic degradations, most PCDD congeners photodecomposed rapidly and the rates presented more than 70% (as dechlorination rates of 76% for PCDDs) at 24 h after irradiation, using PCDD/DFs formed with 2,4,5-TCP. The rate constants were in the order of 4.8-6.1 x 10(-3) min(-1), assuming the pseudo-first-order reactions for their low levels.

Clinical application of three-dimensional myocardial imaging: evaluation of efficacy of medical treatment on myocardial perfusion.

To investigate the clinical applicability of the three-dimensional (3D) myocardial imaging using a newly developed system (the Application Visualization System-Medical Viewer), thallium-201 myocardial single photon emission computed tomography was performed in 19 patients with previous myocardial infarction before and after treatment with nisoldipine. We have developed a new method for automatically reconstructing 3D imaging for the stereoscopic evaluation of myocardial perfusion. The left ventricular myocardial volume with a radioisotope count > or = 50% of maximum was calculated by using the conventional surface rendering method. With these images, the effect of nisoldipine on myocardial perfusion was assessed and the myocardial volume with a radioisotope count > or = 50% of maximum was compared. In fifteen (88%) of 19 patients, myocardial perfusion increased in the infarct areas after nisoldipine treatment. Nisoldipine significantly increased the myocardial volume with a radioisotope count > or = 50% of maximum from 141 +/- 17 to 153 +/- 18 ml on the stress 3D imagings. These findings indicate that nisoldipine improved myocardial perfusion during exercise. 3D imaging provided stereoscopic assessment of the changes in myocardial perfusion following treatment with nisoldipine and also detected transient enlargement of the left ventricular lumen induced by exercise.

Studies on single-dose toxicity of hydrophobically modified hydroxypropyl methylcellulose in rats.

Single-dose toxicological studies of hydrophobically modified hydroxypropyl methylcellulose (HM-HPMC, hydroxypropyl methylcellulose modified with stearylglycidylether) were conducted. A dispersion of HM-HPMC was administered to rats orally or by dermal application at doses up to 900 mg/kg. After the oral administration, the mean body weight of the 900 mg/kg group on the first day after administration was slightly but significantly lower (P less than 0.05) than that of the control group, and one rat had loose stools at 30 min. after the administration. No other abnormalities were noted. In the case of dermal application, no abnormalities were observed. No rats died, and no abnormalities in their organs were found by either route. In conclusion, there was no observed toxicity of HM-HMPC after oral or dermal administration at single dose up to 900 mg/kg under the conditions of these studies.

Primary dermal and eye irritability tests of hydrophobically modified hydroxypropyl methylcellulose in rabbits.

Primary dermal and eye irritation tests of hydrophobically modified hydroxypropyl methylcellulose (HM-HPMC, hydroxypropyl methylcellulose modified with stearylglycidylether), a new cellulose derivative used as a thickener for topical pharmaceuticals and cosmetics, were conducted in rabbits. A dispersion of HM-HPMC (3%) was applied to intact and abraded skins and reactions were observed. A very slight erythema was observed in both skins and this polymer was categorized as a "mild irritant". In the eye irritation test, with a dispersion of the same concentration, it was categorized as "marginal" in unrinsed eyes and "negative" in rinsed eyes.

A three-month repeated oral administration study of a low viscosity grade of hydroxypropyl methylcellulose in rats.

The toxicity of the lowest viscosity grade of hydroxypropyl methylcellulose (HPMC) that is currently commercially available was investigated by means of a three-month repeated oral administration study in male and female Crj:CD (SD) IGS rats at doses of 505, 1,020 and 2,100 mg/kg/day. Body weights of males and females in the 2,100 mg/kg group were lower than those of the control group on and after day 28 of administration, but the differences were not statistically significant. The degree of suppression of body weight gain in males was higher than that in females. This tendency was similar to the results in other toxicity studies of HPMC that have been reported. Males in the 2,100 mg/kg group showed a tendency (not significant) for decreased food consumption and urine volume. Examinations of general signs, hematology, blood chemistry, ophthalmology, absolute and relative organ weights, autopsy and histopathology revealed only a few, apparently coincidental, statistically significant differences from the control, and no evidence of any dose-dependent changes was found. It was concluded that the lowest viscosity grade of HPMC showed extremely low toxicity under the conditions of this study, as has been found for higher viscosity grades.

A repeated-dose dermal toxicity study of hydrophobically modified hydroxypropyl methylcellulose in rats.

A six-month repeated-dose dermal toxicity study followed by a 30-day recovery test of hydrophobically modified hydroxypropyl methylcellulose (HM-HPMC), a new cellulose derivative used as a thickener for topical pharmaceuticals, was conducted using rats. Aqueous paste of HM-HPMC was applied to the skin of rats once daily at dose levels up to 60 mg/kg/day, which was the highest dose that could be administered. Items checked included general signs, urinalysis, hematology, ophthalmology, and histopathology. One rat died during the administration period owing to a malignant tumor in the hemopoietic system, which was not attributed to the test substance. Statistically significant differences were found in some test results, but those were not dose-dependent and were considered to be incidental or spontaneous. It was concluded that the test substance was not toxic upon chronic dermal administration at dose levels up to 60 mg/kg/day.