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K Mitamura

Publications and source records attributed to K Mitamura.

At least 37 records · Page 2Linked to original sources
PubMed2000

Studies on neurosteroids XII. Determination of enzymatically formed catechol estrogens and guaiacol estrogens by rat brains using liquid chromatography-mass spectrometry-mass spectrometry.

Enzymic formations of catechol- and guaiacol-estrogens by rat brains have been investigated using classical- and catechol-estrogens as substrates, respectively. The incubation mixtures were pretreated with liquid-liquid and/or solid-phase extraction, and the products were identified by comparison with authentic samples using liquid chromatography-mass spectrometry (-mass spectrometry) [LC-MS (-MS)]. The enzymic activities were also determined by measuring the formed products with LC-MS.

Animals↗
PubMed2000

Detection of mutation of the p53 gene with high sensitivity by fluorescence-based PCR-SSCP analysis using low-pH buffer and an automated DNA sequencer in a large number of DNA samples.

Detection of mutations in genes responsible for hereditary diseases or tumors is important clinically. It is necessary to establish a simple technique for screening mutations in large numbers of samples. The polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) method has proved to be a useful technique for analyzing mutations or DNA polymorphisms. Non-radioisotopic versions using fluorescent dye and an automated DNA sequencer have also been exploited to extend this technique into the clinical field. We have examined mutations of exons 5-9 of the p53 gene in 112 colorectal, 28 esophageal and 33 hepatocellular carcinomas by fluorescence-based PCR-SSCP (F-SSCP) under various conditions. We found 64 types of mutations in 63, 17 and 12 cases of colon, esophageal and hepatocellular carcinomas by F-SSCP. We determined the sequence of all samples, and confirmed that all mutations were successfully detected by F-SSCP. With the low-pH buffer system, 61 types of mutants were detected, while 51 types were detected by TBE and 57 types were detected by TBE with glycerol gel. The polyacrylamide gel in TME or TBE without glycerol was tough and could be used repeatedly, but the glycerol containing gel was fragile and could not stand repeated use. Thus, use of a low-pH buffer in the electrophoresis of F-SSCP is simpler and better at detecting mutations than the conventional TBE buffer system. We believe that low-pH F-SSCP analysis is an efficient and powerful technique for examination of a large number of samples, in particular clinical specimens obtained by biopsy or surgery.

DNA Mutational Analysis↗
PubMed2000

A possible role for Ca(2+)/calmodulin-dependent protein kinase IV during pancreatic acinar stimulus-secretion coupling.

Ca(2+)/calmodulin-dependent protein kinases (CaMKs) are important intracellular mediators in the mediation of stimulus-secretion coupling and excitation-contraction coupling in a wide variety of cell types. We attempted to identify and characterize the functional roles of CaMK in mediating pancreatic enzyme secretion. Immunoprecipitation and immunoblotting studies using a CaMKII or CaMKIV antibody showed that rat pancreatic acini expressed both CaMKII and CaMKIV. Phosphotransferase activities of CaMKs were measured by a radioenzyme assay (REA) using autocamtide II, peptide gamma and myosin P-light chain as substrates. Although CaMKII and CaMKIV use autocamtide II as a substrate, peptide gamma is more efficiently phosphorylated by CaMKIV than by CaMKII. Intact acini were stimulated with cholecystokinin (CCK)-8, carbachol (CCh) and the high-affinity CCK-A receptor agonist, CCK-OPE, and the cell lysates were used for REA. CCK-8, CCh and CCK-OPE caused a concentration-dependent increase in CaMKs activities. When autocamtide II was used, maximal increases were 1.5-1.8-fold over basal (20.2+/-2.0 pmol/min/mg protein), with peaks occurring at 20 min after cell stimulation. In separate studies that used peptide gamma, CCK-8, CCh and CCK-OPE dose-dependently increased CaMKIV activities. Maximal increases were 1.5-2.4-fold over basal (30.7+/-3. 2 pmol/min/mg protein) with peaks occurring at 20 min after cell stimulation. Peak increases after cell stimulation induced by peptide gamma were 1.8-2.8-fold higher than those induced by autocamtide II. CCK-8, CCh and CCK-OPE also significantly increased phosphotransferase activities of myosin light chain kinase (MLCK) substrate (basal: 4.4+/-0.7 pmol/min/mg protein). However, maximal increases induced by MLCK substrate were less than 10% of those occurring in peptide gamma. Characteristics of the phosphotransferase activity were also different between autocamtide II and peptide gamma. When autocamtide II was used, elimination of medium Ca(2+) in either cell lysates or intact cells resulted in a significant decrease in the activity, whereas it had no or little effect when peptide gamma was used. This suggests that Ca(2+) influx from the extracellular space is not fully required for CaMKIV activity and Ca(2+) is not a prerequisite for phosphotransferase activity once CaMKIV is activated by either intracellular Ca(2+) release or intracellular Ca(2+) oscillations. The specific CaMKII inhibitor KN-62 (50 microM) had no effect on the CaMKIV activity and pancreatic enzyme secretion elicited by CCK-8, CCh and CCK-OPE. The specific MLCK inhibitor, ML-9 (10 microM), also did not inhibit CCK-8-stimulated pancreatic amylase secretion. In contrast, wide spectrum CaMK inhibitors, K-252a (1 microM) and KT5926 (3 microM), significantly inhibited CaMKIV activities and enzyme secretion evoked by secretagogues. Thus, CaMKIV appears to be an important intracellular mediator during stimulus-secretion coupling of rat pancreatic acinar cells.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed2000

Growth patterns of superficially elevated neoplasia in the large intestine.

BACKGROUND: The growth pattern and malignant potential of superficially elevated neoplastic lesions remain controversial. A flat adenoma is classified as a superficially elevated neoplasm that characteristically demonstrates high-grade dysplasia despite its small size. In contrast, a nodule-aggregating (NA) tumor, which consists of multiple small aggregated nodules, can also be classified as superficially elevated neoplasia. METHODS: In this prospective study, 2720 consecutive patients undergoing total colonoscopy were examined for superficially for elevated lesions. Clinicopathologic characteristics, Ki-ras mutational status, and overexpression of p53 protein were compared in 25 NA tumors and 55 flat adenomas without a central depression (flat tumor). RESULTS: All flat tumors had a tubular pattern, whereas 21 of 25 NA tumors showed a villotubular or tubulovillous pattern. Ki-ras mutation was observed in 44% of NA tumors but in none of the flat tumors. Overexpression of p53 protein was found in 12% of NA tumors and 7% of flat tumors. CONCLUSIONS: NA tumors and flat tumors have different clinicopathologic and genetic characteristics, although both types of tumor are classified as superficially elevated lesions.

Adenocarcinoma↗
PubMed2000

Evaluation of balloon-occluded hepatic venography with carbon dioxide for portography and correlation between retrograde portogram and liver function in patients with liver cirrhosis.

Background/Aim: Hepatic venography with a positive-contrast medium has been reported as a method for evaluating liver disease. However, the contrast medium used in this method provides insufficient portal vein observation and may cause severe liver injuries. Carbon dioxide (CO(2)), a negative-contrast medium, may be able to depict the portal vein system with minimal hepatic toxicity. The aim of this study was to evaluate the usefulness and side-effects of balloon-occluded hepatic venography with CO(2) (CO(2) venography) and to evaluate the correlation between retrograde portogram and liver function in patients with cirrhosis. Subjects and methods: The subjects consisted of 23 biopsy-proven cirrhotic patients (male:female, 16:7; age, 58+/-12 years, range 34-80). The causes of cirrhosis were alcohol intake in ten, HCV infection in ten, HBV infection in one, primary biliary cirrhosis in one and Budd-Chiari syndrome in one. Of these patients, six were complicated with hepatocellular carcinoma (HCC). CO(2) venography was performed with an occlusion balloon catheter, and 25 ml of CO(2) was infused. CO(2) portograms were scored as follows: 0, no visualization of portal veins; 1, visualization of peripheral portal branches; 2, unilateral first portal branch; 3, bilateral first portal branches; 4, main portal vein; 5, left gastric vein, superior mesenteric vein and splenic vein. Hepatic venous pressure gradient (HVPG), cardiac functions, biochemical analysis, blood gas analysis and oxygen (O(2)) saturation monitoring were measured simultaneously. Arterio-portography was also performed. To evaluate the usefulness of CO(2) venography in patients with HCC accompanied by portal vein tumor thrombus (PVTT), three patients were also examined. Results: No significant changes in ALT, AST, O(2) saturation or blood gas data were observed after CO(2) venography. A statistically significant positive correlation was observed between CO(2) portogram scores and Child-Pugh scores (r=0.68, P=0.003). The correlations between CO(2) portogram scores and HVPG, and the forms of gastroesophageal varices in patients without PVTT and major shunts were not significant. The CO(2) portogram score was significantly higher in patients with alcoholic liver cirrhosis than in those with HCV-positive cirrhosis (P=0.04). Cavernous transformation and peripheral portal branches were demonstrated in patients with HCC accompanied by PVTT. These findings could not be observed by arterio-portography. Conclusion: CO(2) venography to obtain retrograde portogram is a safe and useful method for evaluating the portal vein system and liver function in patients with liver cirrhosis.

Journal Article↗
PubMed2000

Studies on neurosteroids. Part XIII. Characterization of catechol estrogens in rat brains using liquid chromatography-mass spectrometry-mass spectrometry.

The existence of catechol estrogens in rat brains was clarified using liquid chromatography-atmospheric pressure chemical ionization-ion trap-mass spectrometry-mass spectrometry (LC-APCI-MS2). The catechol estrogens were extracted in the presence of ascorbic acid and then derivatized to acetates with acetic anhydride and pyridine. After a successive purification with silica gel mini-column chromatography, reversed-phase solid-phase extraction and preparative HPLC, the obtained fractions containing the catechol estrogen acetates were subjected to LC-APCI-MS2. 2-Hydroxyestrone, 2-hydroxyestradiol and their 4-hydroxy isomers were identified as acetates by comparison with authentic samples based on their chromatographic behavior and mass spectral data. The derivatization to acetate was useful for the treatment of labile catechol estrogens.

Acetates↗
PubMed2000

Involvement of calmodulin and protein kinase C in cholecystokinin release by bombesin from STC-1 cells.

The mouse intestinal neuroendocrine tumor cell line STC-1 secretes cholecystokinin (CCK) and other hormones. We investigated the role of Ca2+, calmodulin (CaM), and protein kinase C (PKC) in the regulation of CCK release from STC-1 cells. Phorbol 12-myristate 13-acetate (TPA) significantly stimulated CCK release. Staurosporine significantly inhibited CCK release from STC-1 cells stimulated by TPA in a dose-dependent manner. The absence of extracellular calcium completely inhibited CCK release from TPA-stimulated STC-1 cells. Neurotensin did not stimulate CCK release from these cells. W-7, a CaM antagonist, reduced CCK release from STC-1 cells stimulated by bombesin in a dose-dependent manner. These findings suggest that CaM and PKC play an important role in the regulation of CCK release from STC-1 cells stimulated by bombesin.

Animals↗
PubMed2000

TT virus infection in patients with fulminant hepatic failure.

OBJECTIVE: A new DNA virus, which has been designated the TT virus, was discovered in 1997. It is not clear whether TT virus is a cause of any of the types of hepatitis. We conducted a case-control study to test the hypothesis that the presence of TT virus is a necessary condition for the development of fulminant hepatic failure in people who have non-A, -B, or -C hepatitis. METHODS: We studied 55 patients with fulminant hepatic failure [28 men, 27 women, mean (+/- SD) age, 47 +/- 15 yr], 32 patients with acute hepatitis (18 men, 14 women, mean age, 38 +/- 15 yr), and 200 healthy subjects (106 men, 94 women, mean age, 42 +/- 14 yr). TT virus DNA was detected in sera by a nested polymerase chain reaction using a primer set for genotype 1. RESULTS: TT virus was more frequently detected in patients with fulminant hepatic failure [in 33 of 55 (60%); 95% confidence interval (CI), 47-73%] than in those with acute hepatitis [in 8 of 32 (25%); 95% CI, 10-40%; p = 0.0016] or in healthy subjects [in 50 of 200 (25%); 95% CI, 19-31%; p < 0.0001]. TT virus was detected at a significantly higher rate in non-A, -B, or -C fulminant hepatic failure [in 18 of 22 (82%); 95% CI, 66-98%] than in fulminant hepatic failure of A, B, or C type [45%, 28-62%, 15/33; p = 0.007] or in non-A, -B, or -C acute hepatitis [24%, 3-44%, 4/17; p = 0.0003]. The logistic regression analysis selected TT virus (p = 0.0009), age (p = 0.0116), and etiology (p = 0.0309) as independent variables associated with fulminant hepatic failure (coefficient of determination, 0.2335). CONCLUSIONS: TT virus comparatively plays a role in the pathogenesis of non-A, -B, or -C fulminant hepatic failure.

Acute Disease↗
PubMed2000

[Clinical evaluation of rapid diagnostic kit detecting separately influenza A and B viruses].

The Directigen Flu A + B kit, a rapid diagnostic device for influenza virus A and B was evaluated. The nasopharyngeal aspirates were obtained from 239 patients who visited our hospital, between January and March, 2000, presenting flu-like symptoms. Influenza virus AH1: 77 and AH3: 51 were isolated from 128 specimens and none from 111 specimens. Directigen Flu A + B showed 115 specimens positive and 106 specimens negative. The sensitivity and specificity of this kit were 89.8% (115/128) and 95.5% (106/111) compared with viral isolation. Agreement on positive and negative interpretations between Direction Flu A and this kit was 97.9% (234/239). In the evaluation of this kit for influenza B virus, 60 frozen nasopharyngeal aspirates collected from February to April, 1999 were used. The sensitivity and specificity of this kit were 88.9% (16/18) and 88.1% (37/42) compared with viral isolation. Agreement on positive and negative interpretations between FLU OIA and this kit was 91.7% (55/60). The Directigen A + B demonstrated sensitivity and specificity equivalent to the conventional kits in nasopharingeal aspirates. This kit can also differentiate influenza A and B viruses, a feature which is useful for treatment using anti-viral agents such as amantadine and neuraminidase inhibitor. To date, the kit is the most effective tool for the rapid diagnosis of influenza.

Evaluation Studies as Topic↗
PubMed2000

[Sensitivity and specificity of rapid diagnosis kit detecting separately influenza A and B viruses].

Sensitivity and specificity of the Directigen Flu A + B kit, a rapid test for influenza virus A and B, were evaluated. This test detects influenza A and B viruses separately by EIA. Reactivity of the kit was tested using a total of 23 isolates: 13 isolates of human influenza virus A (H1N1, H3N2) and 10 isolates of human influenza virus B. All of the isolates were tested positive and no difference in reactivity was found in antigenic variables and subtypes. The kit was only reactive to influenza virus A and B, not reactive to other viruses. Typical influenza A and B strains were tested for detection limit. 7.8 x 10(3) pfu/ml was a detection limit for influenza virus A (H1N1: Beijing/262/95), 4.7 x 10(4) pfu/ml for influenza virus A (H3N2: Kitakyusyu/159/93), and 3.1 x 10(4) pfu/ml for influenza virus B (Guangdong/05/94). The Directigen Flu A + B kit was a easy-to-use, rapid detection device and the kit has sensitivity and specificity equivalent to other diagnostic devices, suggesting the kit are useful in medical institutions.

Evaluation Studies as Topic↗
PubMed2000

[Evaluation of the rapid detection test for influenza A and B viruses using neuraminidase activity].

The ZstatFlu test (ZymeTx, USA) is a rapid detection kit for influenza A and B viruses. This test is based upon the reaction between viral neuraminidase from influenza viruses and a chromogenic substrate. The clinical performance of the ZstatFlu test was determined by comparison with viral isolation in cell culture. A total of 176 respiratory specimens from 172 pediatric patients with influenza like illnesses during the 1998/99 season were tested. Influenza viruses were recovered from 97 specimens (type A: 6, type B: 91) in cell culture. ZstatFlu demonstrated 67.4% sensitivity (29/43) and 62.7% specificity (37/59) for throat swabs. Of the 22 ZstatFlu-positive, culture-negative throat swabs tested by RT-PCR, 18 were positive by RT-PCR. ZstatFlu showed 48.1% sensitivity (26/54) and 90.0% specificity (18/20) for nasopharyngeal aspirates. Of the two ZstatFlu-positive, culture-negative nasopharyngeal aspirates tested by HI titer of paired sera, one showed a 4-fold increase of HI titer. Nasopharyngeal aspirates therefore showed lower sensitivity than throat swabs at this test, different from EIA test kits such as Directigen FluA or FLU OIA. Overall, only 5 specimens were false positive by the ZstatFlu test. Therefore, this test demonstrated high specificity and positive predictive value. In conclusion, the ZstatFlu test is useful for the rapid detection of influenza A and B viruses to identify patients who need antiviral treatment.

Adolescent↗
PubMed2000

[Depression during interferon therapy in renal cell cancer patients--comparison with chronic hepatitis C patients].

BACKGROUND AND PURPOSE: Depression is well known as one of the psychiatric complications of interferon (IFN) therapy in patients with chronic active hepatitis C. We compared the psychiatric status during interferon therapy between patients with renal cell cancer (RCC) and those with chronic active hepatitis C. METHODS: The psychiatric status of 40 patients with renal cell cancer, 36 who were subjected to radical nephrectomy and 4 who were not because of the presence of distant metastasis and other complicated underlying diseases, was assessed by psychiatrists before and at 2, 4, 12 and 24 weeks after the start of IFN therapy according to the major depressive episode diagnostic criteria listed in the DSM-III-R and the Hamilton Depression Scale (HDS). The mental aspect was measured using the Eysenk Personality Questionnaire (EPQ) before IFN therapy, and the results were compared with those obtained in 85 hepatitis C patients evaluated according to the same protocol for psychiatric assessment. RESULTS: Eight of the 40 patients were diagnosed as having depression before IFN therapy and 5 of them became even more depressed 2 weeks after the start of IFN therapy. Nine patients were found to be depressed after the start of IFN therapy, 7 of them were diagnosed before 4 weeks of therapy. All the patients needed treatment for depression: discontinuation of IFN therapy in 4, decrement of the IFN dose in 8, and psychiatric drug medication in 13. The patients who suffered from depression had significantly higher scores in neuroticism (EPQ) before IFN therapy than those who did not become depressed. There was no difference in age, sex or cancer stage between the two groups. The occurrence of depression in RCC patients was the same as that observed in hepatitis C patients receiving IFN therapy. But in RCC patients the depressive state was diagnosed earlier and was more severe than that observed in hepatitis C patients. CONCLUSION: Urologists should be aware of the psychiatric symptoms of RCC patients under IFN therapy just as in hepatitis patients. The depression in RCC patients was considered to be related to the cancerous disease itself which may explain its higher degree of severity, as compared to hepatitis C patients.

Aged↗
PubMed2000

The impact of winter epidemics of influenza and respiratory syncytial virus on paediatric admissions to an urban general hospital.

To demonstrate the impact of influenza epidemics on pediatric hospital admissions, admissions that were attributable to influenza and respiratory syncytial virus (RSV) infection to the pediatric ward of an urban general hospital in Japan were followed-up during a 4-month period from December to March 1991 through 1998. During the 1997-1998 influenza type A (H3N2) epidemic, a diagnosis of influenza type A (H3N2) was made in 26.3% of all patients admitted aged 15 years or lower. During the peak of the epidemic, as many as 50-70% of the admissions were attributable to influenza type A (H3N2). In the seven winters from 1991 to 1988, 14.0% of all admissions were associated with infection with influenza virus (mean age 4.4 years), and 17.5% were due to RSV. More patients were admitted to hospital for influenza than RSV infection in three of the seven seasons. Among the patients with influenza, 74.5% of the cases were previously healthy children. Influenza and RSV infection are leading causes of pediatric hospital admissions during the winter. Effective methods of prophylaxis are needed not only for high-risk patients, but for healthy young children.

Adolescent↗
PubMed2000

[Rapid diagnosis of influenza by EIA].

There are several commercially available rapid tests by enzyme immunoassay for the laboratory diagnosis of influenza. The tests detect viral nucleoproteins with monoclonal antibodies. They are less sensitive than viral culture or PCR, but rapid and easy to perform, so they can be used at the point-of-care. We evaluated these kits and compared them with viral culture, and we concluded that they are reliable. When compared with viral culture, the sensitivity of the EIA tests with nasal specimens showed greater than 80%. EIA was more sensitive in detecting influenza viruses in nasal aspirates and nasal swabs than in throat swabs. The rapid detection kits facilitate the timely initiation of antiviral therapy and control of influenza outbreaks in institutions. They are clinically useful and cost-effective for the diagnosis of influenza.

Humans↗
PubMed1999

Studies on neurosteroids. IX. Characterization of estrogens in rat brains using gas chromatography-tandem mass spectrometry.

The characterization of the classical estrogens (estrone, estradiol, estriol) and guaiacol estrogens (2-hydroxyestrone 3-methyl ether, 4-hydroxyestrone 3-methyl ether) in rat brains was performed using gas chromatography-tandem mass spectrometry (GC-MS-MS). Estrogens were purified from Wistar strain rat brains by some chromatographic pre-treatments, such as solid-phase extraction, preparative thin-layer chromatography or preparative high-performance liquid chromatography. After the derivatization with O-methylhydroxylamine and/or N,O-bis(trimethylsilyl)trifluoroacetamide, estrogens were identified by comparison of their chromatographic behavior during GC-MS-MS operating in the product ion scan mode and comparison with the product ion MS spectra of an authentic sample. These evidences suggested that estrogens exist in rat brains as neurosteroids or neuroactive steroids.

Animals↗
PubMed1999

Relationship between Helicobacter pylori infection and histologic features of gastritis in biopsy specimens in gastroduodenal diseases, including evaluation of diagnosis by polymerase chain reaction assay.

We investigated the relationship between Helicobacter pylori infection and the histologic features of gastritis in gastroduodenal disease, and evaluated the diagnostic usefulness of the polymerase chain reaction (PCR) assay for the detection of H. pylori before and after eradication therapy. Endoscopic biopsy specimens from 81 patients with gastroduodenal disease were examined for the presence of H. pylori by culture and histologic examination. Histologic features of gastritis were classified according to the updated Sydney System, and results of the PCR assay were compared with those of histologic examination, using histologic scores. The density of H. pylori was significantly correlated with polymorphonuclear neutrophil activity and chronic inflammation. These findings suggest that the grades of infiltration of polymorphonuclear neutrophil cells and chronic inflammatory cells correspond to the density of H. pylori infection assessed by the updated Sydney System. Patients with positive results on PCR assay and negative results on histologic examination may have a low density of H. pylori because of severe atrophy in the gastric mucosa. Differences in results for the PCR assay and histologic examination were found in 2 of 12 patients in the detection of H. pylori after eradication therapy. According to the results of the PCR assay and histologic features before and after eradication, the gastric tissue-based PCR assay for H. pylori after eradication may be too sensitive to judge successful eradication of H. pylori.

Adolescent↗
PubMed1999

Hepatocyte growth factor stimulates cell growth and enhances the expression of transforming growth factor alpha mRNA in AsPC-1 human pancreatic cancer cells.

We investigated the effects of hepatocyte growth factor (HGF) and transforming growth factor alpha (TGF alpha) on cell growth in four human pancreatic cancer cell lines. Changes in the expression of mRNAs of HGF, c-met, TGF alpha, and epidermal growth factor receptor (EGFR) by treatment with HGF and TGF alpha were observed. Cell growth with growth factors was assessed with the MTT assay and compared with basal growth without growth factors. Although HGF stimulated cell growth in AsPC-1, COLO-357, and T3M4 cells, Panc-1 cells showed no response to HGF. TGF alpha stimulated the growth of all the above cells. The expression of c-met mRNA under nonstimulated conditions was detected with Northern blotting in all cells. Treatment with HGF slightly enhanced the expression of c-met mRNA only in COLO-357 cells. The intensity of EGFR expression was consistent, and HGF mRNA was not detected during induction experiments in any cell type. Concomitant treatment with HGF and TGF alpha exerted an effect that was additive or less on the growth of all cells. Expression of TGF alpha was enhanced by HGF treatment only in AsPC-1 cells. These results suggested that HGF and TGF alpha stimulated cell growth through a final common pathway of signal transduction.

Blotting, Northern↗