[Submucosal tumor of the intestine].
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Humans↗
Biomedical subjects
K Yoshimoto
Publications and source records attributed to K Yoshimoto.
[Submucosal tumor of the intestine].
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Humans↗
[The molecular genetics of multiple endocrine neoplasia type 2A and 2B].
Multiple endocrine neoplasia (MEN) type 2A (MEN 2A) and type 2B (MEN 2B) are dominantly inherited with a predisposition to endocrine tumors. The responsible genes for MEN 2A and 2B have recently been localized to chromosome 10q 11.2 by genetic and physical mapping. The DNA segment encompasses the RET proto-oncogene. This is a receptor tyrosine kinase gene, which is expressed in medullary thyroid carcinoma and pheochromocytoma. Point mutations in the cysteine-rich domain of the RET were demonstrated in patients with MEN 2A. The cosegregation of these mutations and disease in MEN 2A families indicates that they possess a predisposition endocrine organs to develop into tumors. Biological assessment of the mutant forms in cell culture and transgenic mouse lines should provide further insight as to the role of the RET in the tumor development.
Animals↗
An end-trimming method to amplify adjacent cDNA fragments by PCR.
We describe a method for cDNA cloning by PCR, which we named "an end-trimming method." This method can be used for PCR amplification and cloning of unknown cDNA fragments adjacent to a short stretch of a known sequence by using a combination of a sequence-matched primer with an ATCG sequence added to the 5' end (5'-ATCG-primer) and an adaptor-(dT)17-primer (dT-primer). The fragments amplified by PCR using a 5'-ATCG-primer, which were modified to have a 5'-ATC overhang by blocking the G site, were exclusively cloned into pUC19 with the vector having a 3'-TAG-5' complementary overhang with a confined direction of inserted fragments. Practical application of this method for the determination of rat amidophosphoribosyltransferase cDNA resulted in successful cloning of adjacent cDNA fragments.
Animals↗
[Urinary bladder carcinoma was induced in a patient treated with cyclophosphamide for malignant lymphoma].
A case of urinary bladder carcinoma in a patient with malignant lymphoma (ML) involving the thyroid gland, gastrointestinal tract and the gallbladder is reported. The histological type of ML from the biopsy specimens of the thyroid gland and the stomach was non-Hodgkin's, diffuse large cell lymphoma of B cell origin. The patient had been treated for ML with cyclophosphamide (CPM), doxorubicin, vincristine and prednisolone effectively then maintained with low dose CPM. Two and a half years after the initial therapy, urinary bladder tumor diagnosed transitional cell carcinoma histologically was found. The patient had received 55 gr. of CPM totally. The dose of CPM was relatively smaller than the one in the other second cancer cases reported induced by CPM previously. It may be suggested that the urinary bladder carcinoma induced by CPM is related not only total dose of CPM but also the other factors. Thus usage of CPM must be cautious because of the secondary disease.
Carcinoma, Transitional Cell↗
Multiple fluorescence-based PCR-SSCP analysis.
Multiple fluorescence-based polymerase chain reaction single-strand conformation polymorphism (MF-PCR-SSCP) was developed. The target sequence was amplified by PCR using forward and reverse primers labeled with two different fluorescent dyes at their 5' ends. The amplified products were then heat-denatured, mixed with internal standard DNA markers labeled with a third fluorescent dye and applied to a temperature-controlled gel in an automated DNA sequencer, with a gel-temperature-controlling system. Mutations were detected as positional shifts of two-colored peaks in the electrophoretogram. The image data were analyzed by the computer program GENESCAN 672. The peak positions were standardized to internal DNA size markers. MF-PCR-SSCP analysis of 7 human tumor cell lines with 7 different single base mutations of the human K-ras oncogene detected all mutations even under the same electrophoresis conditions. Complete loss of heterozygosity was detected in two cell lines simultaneously. A gel temperature at 20 degrees C and polyacrylamide concentration of 10% gave the best separation. MF-PCR-SSCP is superior to the current PCR-SSCP in several ways: it does not involve radioactivity, migration patterns are standardized to internal standard DNA markers, there is a strict temperature-controlling system and the higher percentage of the gel enables better separation with resultant 100% detection of mutations most likely under one set of electrophoresis conditions.
Base Sequence↗
Neuron-specific glucose transporter (NSGT): CNS distribution of GLUT3 rat glucose transporter (RGT3) in rat central neurons.
The identity of the glucose transporters (GLUT) expressed in neurons in situ has yet to be fully established. In the present study we have isolated the GLUT3 (RGT3) cDNA and produced anti RGT3 polyclonal antibody allowing us to investigate the cellular localization and tissue distributions of RGT3 mRNA and protein in the central nervous system of the rat by the methods of in situ hybridization and immunohistochemistry. Here we demonstrate the direct evidence that RGT3 is present in neurons in adult rat brain. In situ hybridization showed the expression of RGT3 mRNA mostly in the regions of hippocampus, cerebral cortex, striatum, and the granule cell layer of the cerebellum, indicating that RGT3 mRNA is predominantly expressed within neurons. Immunohistochemistry showed that RGT3 protein is widely distributed in the rat brain, and concentrated on the plasma membrane of neurons. Double labeling studies with anti-RGT3, glial fibrillary acidic protein (GFAP), and neuron specific enolase (NSE) antibodies revealed the specific expression of RGT3 protein in neurons. Thus, RGT3 is indicated to be a neuron specific glucose transporter isoform (NSGT), and suggested to play a functionally significant role in rat central neurons.
Amino Acid Sequence↗
Human heart-type cytoplasmic fatty acid-binding protein in serum and urine during hyperacute myocardial infarction.
We have previously reported that serum and/or urinary human heart-type cytoplasmic fatty acid-binding protein (HH-FABPc) can be used as an early indicator of myocardial injury (Clin Biochem 1991; 24: 195-201). To confirm the usefulness of HH-FABPc as an early diagnostic indicator of acute myocardial infarction (AMI), its serum and urinary levels were measured in samples obtained within 6 h after the onset of acute coronary syndrome related symptoms. Samples were collected from 97 patients, who were composed of 63 with AMI, 24 with unstable angina and 10 with chest pain syndrome. The positivity of serum and urinary HH-FABPc and cardiac creatine kinase isozyme MB (CK-MB) was analyzed in these samples. Serum HH-FABPc levels in AMI were above normal in 91.4% (64/70) of the samples tested within 3 h of the onset of symptoms and in 100% (111/111) of those tested at 3-6 h. Elevated urinary HH-FABPc levels in AMI were obtained in 88.9% (8/9) of samples at 0-3 h and in 75% (6/8) at 3-6 h. CK-MB activity in AMI was positive in 20% (8/40) and 66.3% (53/80) of serum samples at 0-3 h and 3-6 h, respectively. HH-FABPc was always positive when a serum sample was positive for CK-MB. Serum HH-FABPc at 0-6 h in chest pain syndrome and in unstable angina were positive in 17.8% (5/28) and 56.7% (34/60), respectively. The elevated HH-FABPc in serum and urine was noted much earlier than that of CK-MB during the hyperacute phase of AMI. HH-FABPc showed high positive value in unstable angina, but it was low in normal coronary patients having chest pain. However, HH-FABPc level in unstable angina and chest pain syndrome was lower than that of AMI. Thus, HH-FABPc may be a valuable indicator for the diagnosis of hyperacute myocardial infarction.
Adult↗
Rare mutations of the Gs alpha subunit gene in human endocrine tumors. Mutation detection by polymerase chain reaction-primer-introduced restriction analysis.
BACKGROUND: The Gs alpha (Gs alpha) gene can be activated to the putative oncogene gsp by specific point mutations at codons 201 or 227. Such mutations have been reported in growth hormone (GH)-secreting pituitary adenomas and thyroid tumors. To clarify the role of Gs alpha gene in human endocrine tumors, 197 tumors were screened for point mutations at codons 201 or 227 of the Gs alpha gene. METHODS: Mutations were detected by primer-introduced restriction analysis (PIRA) of the polymerase chain reaction (PCR) product of genomic DNA. RESULTS: These Gs alpha mutations were present in 4 of 53 pituitary adenomas (4 of 43 GH-secreting adenomas; 1 of these 4 was a GH- and prolactin-secreting adenoma from a patient with familial multiple endocrine neoplasia Type 1), 4 of 66 thyroid tumors (4 of 30 papillary carcinomas), and 1 of 19 adrenocortical adenomas (1 of 6 aldosterone-secreting adenomas). In contrast, none of these Gs alpha mutations were detected in parathyroid tumors, endocrine pancreatic tumors, or pheochromocytomas. CONCLUSIONS: Gs alpha mutations at these two loci may play a role in the pathogenesis of a small population of GH-secreting pituitary adenomas, papillary thyroid carcinomas, and adrenocortical adenomas, but that they are not involved in the pathogenesis of other types of endocrine tumors.
Acromegaly↗
Memory effectors: a potent, IL-4-secreting helper T cell population that develops in vivo after restimulation with antigen.
Resting CD4+ memory T cells from the spleens of mice primed more than 6 wk earlier with KLH in adjuvant produce IL-2 and IL-3, but only low levels of IL-4 and IFN-gamma, and undetectable IL-5 after specific restimulation in vitro. In contrast, when previously primed mice are boosted by i.v. injection of soluble KLH, a CD4+ T cell population develops in the spleen that can produce greatly increased levels of IL-4, and variably increased IL-2, IFN-gamma, and/or IL-3. The ability of these memory effector Th cells to be restimulated to secrete cytokines is maximal by 3 days after in vivo boosting with Ag, and declines rapidly thereafter. Memory effector Th are induced in the spleens of KLH-primed mice that have been depleted of precursor T cells by thymectomy more than 25 wk previously, suggesting that they are derived from a memory T cell population. Cell separation on the basis of CD45RB or MEL-14, surface markers that are down-regulated on memory CD4+ cells, indicates that memory effector Th are in the CD45RBlo and MEL-14- subsets of CD4+ T cells. The frequency of KLH-specific splenic CD4+ T cells with the capacity to secrete cytokines in limiting dilution cultures is very low in unprimed mice (1/3 x 10(4) to 1/9 x 10(4)), but increases as much as 50-fold after priming. IL-2, but not IL-4, can be detected in the supernatants of stimulated unprimed CD4+ T cells. Both IL-2- and IL-4-producing cells are readily detected after priming, and frequencies of IL-2-secreting cells (1/250 to 1/4000) are up to ninefold greater than those for IL-4 secreting cells. Boosting elicits variable (two to eightfold) increases in the frequencies of both IL-2- and IL-4-producing CD4+ T cells. However, high IL-4 titers associated with memory effector Th function can be obtained when significant frequency changes are not detected after boosting. The data suggest that reactivation of resting memory Th in vivo induces their differentiation into effector Th with the capacity to secrete much higher levels of IL-4. Although the capacity to produce high levels of IL-4 is associated with both primary and memory CD4+ effector populations, memory effector Th are the higher producers.(ABSTRACT TRUNCATED AT 400 WORDS)
Animals↗
Molecular cloning of rat amidophosphoribosyltransferase.
The cDNA of rat amidophosphoribosyltransferase (EC 2.4.2.14, ATase), which is the supposed regulatory allosteric enzyme of de novo purine nucleotide biosynthesis, has been cloned by polymerase chain reaction. The predicted open reading frame encodes a protein of 517 amino acids with a deduced molecular weight of 57,436 including a supposed 11-amino acid propeptide. The 16 amino acid residues next to the propeptide were identical to the N-terminal amino acid microsequence of a purified rat liver ATase, which is consistent with the cleavage of the propeptide from the proenzyme in rat liver. The derived amino acid sequence is the first sequence reported for a mammalian ATase and it exhibits 81, 41, 36, and 31% identity with the sequences of chicken, Bacillus subtilis, Escherichia coli, and Saccharomyces cerevisiae ATases, respectively. The molecular weight (M(r)) of 57,436 suggests a tetrameric structure of native ATase with a M(r) of 240,000-248,000. Southern blot analysis suggested that the ATase gene exists as a single copy in the rat genome. Northern blot analysis revealed that ATase is expressed at a high level in brain, heart, liver, and stomach. The ATase mRNA in brain, heart, and stomach was 3.5 kilobases (kb) and in liver the 3.5-kb band was observed as well as an additional band of 4.2 kb. Reverse transcription-polymerase chain reaction analysis showed that ATase is ubiquitously expressed in all tissues examined. Comparison with chicken ATase showed that 2 cysteine residues for an iron-sulfur cluster were conserved. Three conserved and two non-conserved consensus phosphorylation sites for cAMP-dependent protein kinase were found.
Amidophosphoribosyltransferase↗
Molecular cloning of human amidophosphoribosyltransferase.
The cDNA of human amidophosphoribosyltransferase (EC 2.4.2.14, ATase), which is the supposed regulatory allosteric enzyme of de novo purine nucleotide biosynthesis, has been cloned from human hepatoma (HepG2) cDNA library. The predicted open reading frame encodes a protein of 517 amino acids with a deduced molecular weight (Mr) of 57,398, which is consistent with the molecular mass of 56 kDa on SDS-polyacrylamide gel electrophoresis (SDS-PAGE) of the ATase subunit purified from human placenta. The derived amino acid sequence exhibits 93, 82, 41, 37, and 33% identity with the sequences of rat, chicken, Bacillus subtilis, Escherichia coli, and Saccharomyces cerevisiae ATases, respectively. Southern blot analysis suggested that the ATase gene exists as multiple copies. ATase mRNA (3.5 kb) is ubiquitously expressed in various human tissues. Comparison with rat and chicken ATases showed that two cysteine residues for an iron-sulfur cluster were conserved. Four consensus phosphorylation sites for cAMP-dependent protein kinase were found.
Amidophosphoribosyltransferase↗
Hemofiltrability of histamine H2-receptor antagonist, nizatidine, and its metabolites in patients with renal failure.
To study if an H2-receptor antagonist, nizatidine, and its metabolites [N-2-monodesmethylnizatidine (N-2-MDMN) and nizatidine sulfoxide (nizatidine S-Ox)] would be removed by an arteriovenous hemofiltration, the authors measured their plasma concentrations and amounts recovered in ultrafiltrate during 11 sessions of an intermittent hemofiltration performed in seven patients with renal failure who were given an oral administration of nizatidine (150 mg). The concentrations of the parent drug and its metabolites in plasma and ultrafiltrate were determined with a high-performance liquid chromatography with ultraviolet absorbance detection. The mean (+/- standard deviation [SD]) amounts of nizatidine removed by the procedure performed at the mean ultrafiltration rate of 18 (range, 11-25) mL/min over the mean duration of 179 (60 to 300) minutes accounted for 1.9 +/- 1.4% of the dose administered. The corresponding values for N-2-MDMN and nizatidine S-Ox were 0.3 +/- 0.2% and 0.2 +/- 0.2% of the molar dose of nizatidine, respectively. There was a significant correlation between the filtration rate and the hemofiltration clearance of nizatidine (r = .94, P < .001) or its active metabolite, N-2-MDMN (r = 0.83, P < .01), indicating that the sieving coefficient (Sc), an index of filtration efficiency, for these compounds is largely constant (0.59 and 0.67 for nizatidine and N-2-MDMN, respectively) under the current hemofiltration conditions.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
Plasma and urinary heart-type cytoplasmic fatty acid-binding protein in coronary occlusion and reperfusion induced myocardial injury model.
The leakage of heart-type cytoplasmic fatty acid-binding protein (H-FABPc) from injured myocardial cells has been reported. We have previously proposed that its plasma and urinary levels could be used as an early indicator of myocardial injury and also reflect the severity of myocardial injury. To confirm this hypothesis, the time course of changes of the plasma and urinary H-FABPc was investigated during myocardial injury induced by coronary artery occlusion and reperfusion in dogs. The plasma elimination kinetics and urinary excretion kinetics of H-FABPc were also analysed in dogs which were given a bolus injection of exogenous H-FABPc. The distribution of circulating H-FABPc was determined in mice by whole-body autoradiography using 125I-labelled H-FABPc. In myocardial injury model, plasma and urinary H-FABPc level showed rapid increase after reperfusion. The elimination kinetic study revealed that H-FABPc was mono-exponentially cleared from the circulation. The elimination rate constant (Ke) was 0.0275 +/- 0.0094/min (mean +/- S.D., n = 7) and the disappearance half-time (t1/2) was 27.5 +/- 8.4 min (mean +/- S.D., n = 7). Exogenous H-FABPc appeared in urine soon after administration, with the peak level being at 6.9 +/- 2.0 min (mean +/- S.D., n = 7). Whole-body autoradiography also demonstrated that 125I-H-FABPc accumulated rapidly in the kidneys. This study demonstrated that H-FABPc leaked rapidly from injured myocardium and rapidly appeared in plasma and urine. Infarct size was closely correlated with the calculated H-FABPc release (r = 0.89, r2 = 0.80, P < 0.01, n = 7) and with the amount of the urinary H-FABPc (r = 0.94, r2 = 0.88, P < 0.01, n = 7). These data suggest that measurement of the H-FABPc levels in plasma and urine might be useful for the early detection of myocardial injury and also for the assessment of infarct size.
Animals↗
Deglycation of glycated proteins with hydrazine analogues.
Glycated human serum albumin (glc HSA) and glc proteins in back hairs from diabetic rats were deglycated by incubation with hydrazine or aminoguanidine (guanylhydrazine) at physiological temperature (37 degrees C). Glc HSA decreased by 35% with 0.2 M hydrazine at day 20 and by 56% with 0.1 M aminoguanidine at day 10. The glc proteins in hairs decreased by 40-50% and 70-77% with hydrazine and aminoguanidine at more than 0.1 M by day 20, respectively. The deglycation of glc protein with aminoguanidine was slower than that with hydrazine. The present result suggests that the inhibitory effects by aminoguanidine on the late Maillard reaction may include a decrease in the level of glc proteins (Amadori compounds) due to deglycation.
Animals↗
Rapid postmortem changes of rat striatum dopamine, serotonin, and their metabolites as monitored by brain microdialysis.
Brain microdialysis was used to monitor changes in extracellular dopamine (DA), serotonin (5-HT), and their metabolite levels in the rat striatum at death by cervical dislocation. Maximum respective 450-fold and 150-fold increases in the extracellular output of DA and 5-HT were observed within the first 30 min of death. DA and 5-HT outputs remained elevated over the following 2 h at levels about 100-fold and 50-fold above pre-death values, respectively. In contrast with monoamine outputs, the outputs of the DA metabolite, 3,4-dihydroxyphenylacetic acid (DOPAC), and the 5-HT metabolite, 5-hydroxyindoleacetic acid (5-HIAA), rapidly decreased by 10% and 20%, respectively 1 h after death. 5-Hydroxytryptophol (5-HTOL) gradually decreased after death. Before death both the extracellular DOPAC/DA and 5-HIAA/5-HT ratios were about 400; after death these ratios dropped to 0.56 and 4.0, respectively at 30 min. These observations suggested that regulation of neurotransmitter releases through the neuronal membrane and metabolisms in the rat striatum were seriously disrupted at death. This finding may be helpful in the determination of death in the field of forensic medicine.
3,4-Dihydroxyphenylacetic Acid↗
Candidate recombinant vaccine for human B19 parvovirus.
Recombinant baculoviruses were used to produce human B19 parvovirus empty capsids composed of only VP2 and VP2 capsids containing 4%, 25%, 35%, or 41% VP1 protein. Immunogenicity of the purified capsids, formulated with or without adjuvant, was evaluated in mice, guinea pigs, and rabbits. Sera were analyzed for total anti-B19 parvovirus antibodies, antibodies specific to the region unique to the VP1 capsid protein, and virus neutralizing antibodies. A relationship was observed between the development of antibodies specific to sequences unique to the VP1 protein and virus neutralization. The polypeptide composition of the empty capsid immunogens appeared to be important for elicitation of potent virus neutralizing activity. VP2 capsid immunogens devoid of VP1 protein, or consisting of only 4% VP1, the composition of naturally occurring virions, were generally poor at eliciting high levels of virus neutralizing activity. Capsids consisting of > or = 25% VP1 protein efficiently and consistently provoked vigorous B19 virus neutralizing responses. Recombinant empty capsids enriched for the VP1 protein should serve as the basis for a human B19 parvovirus vaccine.
Aluminum Hydroxide↗
A novel colorimetric method for determination of glycated protein based on 2-keto-glucose release with hydrazine.
We tried to measure glycated proteins by a novel method based on colorimetry of 2-keto-glucose which is released from the glycated protein (ketoamine) on heating with hydrazine. Reaction conditions were optimized with glycated human serum albumin (glc HSA) as a model compound. Ketoamine reacted quantitatively with hydrazine on heating at 100 degrees C for 0.5 h, followed by heating with phenylhydrazine at 60 degrees C for 1 h. Glucose interference with the assay was eliminated by preincubation of the sample with glucose oxidase at 37 degrees C for 0.5 h. Time courses for the coloration of glc HSA and human serum showed a profile similar to that of N-p-tolyl-D-isoglucosamine under optimized reaction conditions. The lower limit for the assay of glc HSA was 0.7 microM. The serum level of glycated proteins measured by the present method correlated well with that (fructoamine value, microM) measured by the conventional method (nitroblue tetrazolium-reducing method) (r = 0.92, n = 35). In conclusion, the present method is a novel, highly sensitive and reliable one for measuring glycated proteins in biological samples.
Colorimetry↗