Biomedical subjects

T Iwama

Publications and source records attributed to T Iwama.

At least 37 records · Page 2Linked to original sources

PubMed2001

Occurrence of transsulfuration in synthesis of L-homocysteine in an extremely thermophilic bacterium, Thermus thermophilus HB8.

A cell extract of an extremely thermophilic bacterium, Thermus thermophilus HB8, cultured in a synthetic medium catalyzed cystathionine gamma-synthesis with O-acetyl-L-homoserine and L-cysteine as substrates but not beta-synthesis with DL-homocysteine and L-serine (or O-acetyl-L-serine). The amounts of synthesized enzymes metabolizing sulfur-containing amino acids were estimated by determining their catalytic activities in cell extracts. The syntheses of cystathionine beta-lyase (EC 4.4.1.8) and O-acetyl-L-serine sulfhydrylase (EC 4.2.99.8) were markedly repressed by L-methionine supplemented to the medium. L-Cysteine and glutathione, both at 0.5 mM, added to the medium as the sole sulfur source repressed the synthesis of O-acetylserine sulfhydrylase by 55 and 73%, respectively, confirming that this enzyme functions as a cysteine synthase. Methionine employed at 1 to 5 mM in the same way derepressed the synthesis of O-acetylserine sulfhydrylase 2.1- to 2.5-fold. A method for assaying a low concentration of sulfide (0.01 to 0.05 mM) liberated from homocysteine by determining cysteine synthesized with it in the presence of excess amounts of O-acetylserine and a purified preparation of the sulfhydrylase was established. The extract of cells catalyzed the homocysteine gamma-lyase reaction, with a specific activity of 5 to 7 nmol/min/mg of protein, but not the methionine gamma-lyase reaction. These results suggested that cysteine was also synthesized under the conditions employed by the catalysis of O-acetylserine sulfhydrylase using sulfur of homocysteine derived from methionine. Methionine inhibited O-acetylserine sulfhydrylase markedly. The effects of sulfur sources added to the medium on the synthesis of O-acetylhomoserine sulfhydrylase and the inhibition of the enzyme activity by methionine were mostly understood by assuming that the organism has two proteins having O-acetylhomoserine sulfhydrylase activity, one of which is cystathionine gamma-synthase. Although it has been reported that homocysteine is directly synthesized in T. thermophilus HB27 by the catalysis of O-acetylhomoserine sulfhydrylase on the basis of genetic studies (T. Kosuge, D. Gao, and T. Hoshino, J. Biosci. Bioeng. 90:271-279, 2000), the results obtained in this study for the behaviors of related enzymes indicate that sulfur is first incorporated into cysteine and then transferred to homocysteine via cystathionine in T. thermophilus HB8.

Culture Media↗

PubMed2001

The clinical utility of contrast-enhanced 3D MR angiography for cerebrovascular disease.

The purpose of this study was to evaluate the usefulness of contrast-enhanced 3D MR angiography (CE-MRA) with an automated bolus-detection algorithm (SmartPrep technique) and the specialized phased-array coils for the patients suspected cerebrovascular disease. Forty-three patients with brain attack were examined with CE-MRA. A tracker volume of SmartPrep technique was placed along the ascending aorta in the coronal image. After the bolus injection of gadolinium, an increase in signal that corresponded to the arrival of gadolinium was used to trigger centric re-ordered spoiled gradient echo arterial selective MRA with imaging time of 20-40 sec. We were able to achieve a 100% successful triggering rate of SmartPrep technique and selectively arterial-phase carotid and vertebral arteries with almost no venous contamination could be delineated. These techniques enabled high resolution imaging of entire craniocervical arteries from aortic arch to the circle of Willis. This CE-MRA was useful to evaluate both occlusion of arteries and the collateral pathways and to measure stenosis and residual flow of dissection accurately. CE-MRA was a reliable less-invasive alternative to investigate the patients of cerebrovascular disease.

Brain↗

PubMed2001

Cerebral ischemic complications following intracranial bleeding in patients with moyamoya disease--three case reports.

Three patients with bleeding-type moyamoya disease suffered ischemic complications following their first episode of intracranial bleeding. The patients presented with intraventricular hemorrhage and suffered cerebral infarction on the 14th, 16th, and 11th days after the ictus, respectively. The clinical records revealed no obvious hypotension before progression to infarction and suggested that dehydration accelerated by the administration of hyperosmotic drugs was a contributing factor to the infarction. Two patients underwent superficial temporal artery-middle cerebral artery anastomoses in the chronic stages and have never experienced further ischemic complications. The other patient died despite receiving intensive care. Hemodynamic reserve may be severely impaired in patients with bleeding-type moyamoya disease. Adequate control of hydration is important to prevent cerebral infarction after intracranial bleeding in patients with moyamoya disease.

Adolescent↗

PubMed2000

A novel stereoselective preparation of various vinyl sulfide derivatives using beta-alkylthioalkenylselenonium salts

The treatment of alkynylselenonium salt and various thiophenol derivatives with a catalytic amount of triethylamine gave beta-arylthioalkenylselenonium salts in good yields. The alkenylselenonium salts thus prepared reacted with nucleophiles such as acetylides, thiolates, and alkoxides to produce (Z)-beta-arylthio-alpha-functionalized ethenes in high yields. The vinylselenonium salts bearing a hydroxy group on a beta-side chain caused intramolecular cyclization upon treatment with sodium hydride to produce medium-membered heterocyclic compounds containing sulfur and oxygen atoms. The reactions giving (Z)-beta-arylthio-alpha-functionalized ethenes would proceed via the formation of selenurane intermediates followed by the ligand coupling reaction.

Journal Article↗

PubMed2000

Racemic and asymmetric Diels-Alder reactions of 1-(2-oxazolidinon-3-yl)-3-siloxy-1,3-butadienes.

Achiral and chiral 1-(2-oxazolidinon-3-yl)-3-siloxy-1,3-butadienes were prepared from readily available starting materials. Although more stable than the parent 1-amino-3-siloxy dienes, the 1-(2-oxazolidinon-3-yl)-3-siloxy-1,3-butadienes are still very reactive in Diels-Alder reactions, somewhat more than 1,3-dialkoxy-1, 3-butadienes (e.g., Danishefsky's diene). The cycloadditions of the achiral and chiral dienes with several different dienophiles were examined. The reactions proceeded in good yield, with modest to high endo selectivity. The chiral dienes exhibited excellent facial selectivity in cycloadditions with alpha-substituted acroleins, maleic anhydride and N-phenylmaleimide. Upon reduction and hydrolysis of the cycloadducts, substituted cyclohexenones were obtained with ee's ranging from 22% to >98%.

Acrolein↗

PubMed2000

A mutant Escherichia coli tyrosyl-tRNA synthetase utilizes the unnatural amino acid azatyrosine more efficiently than tyrosine.

Alloproteins, proteins that contain unnatural amino acids, have immense potential in biotechnology and medicine. Although various approaches for alloprotein production exist, there is no satisfactory method to produce large quantities of alloproteins containing unnatural amino acids in specific positions. The tyrosine analogue azatyrosine, l-beta-(5-hydroxy-2-pyridyl)-alanine, can convert the ras-transformed phenotype to normal phenotype, presumably by its incorporation into cellular proteins. This provided the stimulus for isolation of a mutant tyrosyl-tRNA synthetase (TyrRS) capable of charging azatyrosine to tRNA. A plasmid library of randomly mutated Escherichia coli tyrS (encoding TyrRS) was made by polymerase chain reaction techniques. The desired TyrRS mutants were selected by screening for in vivo azatyrosine incorporation of E. coli cells transformed with the mutant tyrS plasmids. One of the clones thus isolated, R-6-A-7, showed a 17-fold higher in vivo activity for azatyrosine incorporation than wild-type TyrRS. The mutant tyrS gene contained a single point mutation resulting in replacement of phenylalanine by serine at position 130 in the protein. Structural modeling revealed that position 130 is located close to Asp(182), which directly interacts with tyrosyladenylate. Kinetic analysis of aminoacyl-tRNA formation by the wild-type and mutated F130S TyrRS enzymes showed that the specificity for azatyrosine, measured by the ratios of k(cat)/K(m) for tyrosine and the analogue, increased from 17 to 36 as a result of the F130S mutation. Thus, the high discrimination against azatyrosine is significantly reduced in the mutant enzyme. These results suggest that utilization of F130S TyrRS for in vivo protein biosynthesis may lead to efficient production of azatyrosine-containing alloproteins.

Alanine↗

PubMed2000

Somatic mutations of LKB1 and beta-catenin genes in gastrointestinal polyps from patients with Peutz-Jeghers syndrome.

Peutz-Jeghers syndrome (PJS) is characterized by multiple gastrointestinal hamartomatous polyps, mucocutaneous melanin deposition, and increased risk of cancer, mainly in the gastrointestinal tract. We examined mutations of the LKB1, beta-catenin, APC, K-ras, and p53 genes in 27 gastrointestinal hamartomatous polyps from 10 patients in nine PJS families. Of these hamartomatous polyps, one intestinal polyp had an adenomatous lesion, and one gastric polyp contained adenomatous and carcinomatous lesions. Germ-line mutations of the LKB1 gene were detected in six PJS families. Somatic mutations of the LKB1 gene were found in 5 polyps, whereas loss of heterozygosity (LOH) at the LKB1 locus at 19p was seen in 14 other polyps. In adenomatous lesions microdissected from hamartomatous polyps, both beta-catenin mutation and 19p LOH were detected. Furthermore, a carcinomatous lesion in a gastric hamartomatous polyp was found to contain a mutation of the p53 gene and LOH at the p53 locus in addition to LOH at the LKB1 locus and a beta-catenin mutation. K-ras mutations were detected in a few polyps, whereas no APC mutation or 5q LOH was detected in hamartomatous polyps. These results suggest that gastrointestinal hamartomatous polyps in PJS patients develop through inactivation of the LKB1 gene by germ-line mutation plus somatic mutation or LOH of the unaffected LKB1 allele, and that additional mutations of the beta-catenin gene and p53 gene convert hamartomatous polyps into adenomatous and carcinomatous lesions.

AMP-Activated Protein Kinase Kinases↗

PubMed2000

Allelic loss at SMAD4 in polyps from juvenile polyposis patients and use of fluorescence in situ hybridization to demonstrate clonal origin of the epithelium.

Juvenile polyposis syndrome (JPS; Online Mendelian Inheritance in Man2 174900) is a rare Mendelian disorder in which individuals have typical hamartomatous polyps within the gastrointestinal tract. The stromal element of the polyps has classically been thought to be the proliferative component, although epithelial malignancies (largely gastrointestinal cancers) occur more frequently than expected in JPS patients. Germ-line mutations in SMAD4 (DPC4) account for about a third of JPS cases. It has been postulated that the apparent paradox of a stromal lesion predisposing to epithelial malignancy can be resolved by the "landscaper" effect: an abnormal stromal environment affects the development of adjacent epithelial cells, and the resulting regeneration of damaged epithelium leads to an increased risk of cancer. We have found allele loss at the SMAD4 locus on 18q in polyps from JPS individuals with a germ-line SMAD4 mutation, showing that SMAD4 is acting as a tumor suppressor gene in JPS polyps, as it does in sporadic cancers of the gastrointestinal tract. Interphase fluorescence in situ hybridization showed deletion of one copy of SMAD4 in the epithelial component of JPS polyps, but not in the inflammatory infiltrate. Fluorescence in situ hybridization also suggested that a single copy of SMAD4 was present in stromal fibroblasts of JPS polyps. Thus, biallelic inactivation of SMAD4 occurs in both the epithelium and some of the stromal cells in these lesions, suggesting a common clonal origin. Epithelial malignancies almost certainly develop in juvenile polyposis through direct malignant progression of the epithelial component of the hamartomas. SMAD4/DPC4 probably acts as a "gatekeeper" tumor suppressor in juvenile polyps, and there is no need to invoke a "landscaper hypothesis."

Adenomatous Polyposis Coli↗

PubMed2000

Development of invasive adenocarcinoma in a long-standing diverted ileal J-pouch for ulcerative colitis: report of a case.

We report a case of a 50-year-old male with ulcerative colitis who developed well-differentiated adenocarcinoma in the ileal J-pouch, which had been defunctioning for 18 years. The extension of the carcinoma in the pouch suggested that it had recently appeared in the pouch. Monitoring by endoscopic examination and biopsy or pouch excision seems to be an appropriate action if a pouch is out of the fecal stream.

Adenocarcinoma↗

PubMed2000

Molecular evidence for multicentric development of thyroid carcinomas in patients with familial adenomatous polyposis.

Familial adenomatous polyposis is characterized by multiple colorectal adenomas and an increased incidence of colorectal carcinomas. Patients also develop various extracolonic tumors, of which, thyroid carcinoma is common in young females. The occurrence of multiple carcinomas in one thyroid is frequently observed, although some carcinomas are solitary. To clarify whether each carcinoma develops independently or metastatically spreads from the first one formed, we analyzed the adenomatous polyposis coli (APC) gene mutation in each carcinoma. We found that each carcinoma had a different somatic mutation of the APC gene. This is molecular confirmation for the multicentric development of thyroid carcinomas in familial adenomatous polyposis through biallelic inactivation of the APC gene.

Adenomatous Polyposis Coli↗

PubMed2000

The relationship between frequencies of extracolonic manifestations and the position of APC germline mutation in patients with familial adenomatous polyposis.

BACKGROUND: Familial adenomatous polyposis (FAP) patients develop various extracolonic lesions; however, the relationship between germline mutation of the APC gene and extracolonic manifestations is mostly unknown. To examine the genotype-phenotype relationship, we compared the APC mutation and clinical data. METHODS: Germline mutations from codon 157 to 1465 of the APC gene were identified in 39 families of FAP and clinical data were collected from 80 patients of these families. Germline mutations were classified into two groups: mutations from exon 4 to 9 (codon 157 to 416, Group 1) and those from exon 10 to 15H (codon 564 to 1465, Group 2). The complication rates of extracolonic manifestations were compared between these two groups. RESULTS: Frequencies of duodenal polyps and gastric adenomas in Group 2 were higher than those in Group 1 (p < 0.0001 and p < 0.0004, respectively) and development of osteoma was more frequent in Group 2 (p = 0.01). The number of colorectal polyps and retinal pigments also correlated with the germline mutation, which was consistent with previous reports. However, such correlations were less obvious with regard to gastric fundic polyps, desmoid tumors, soft tissue tumors and colorectal cancer. CONCLUSION: There are two types with regard to extracolonic manifestations of FAP: one is more severely affected according to the position of germline mutation of the APC gene and the other is not affected.

Adenoma↗

PubMed2000

Mutational analysis of ligand recognition by tcp, the citrate chemoreceptor of Salmonella enterica serovar typhimurium.

The chemoreceptor Tcp mediates taxis to citrate. To identify citrate-binding residues, we substituted cysteine for seven basic or polar residues that are chosen based on the comparison of Tcp with the well-characterized chemoreceptors. The results suggest that Arg-63, Arg-68, Arg-72, Lys-75, and Tyr-150 (and probably other unidentified residues) are involved in the recognition of citrate.

Amino Acid Sequence↗

PubMed2000

Analysis of genetic and phenotypic heterogeneity in juvenile polyposis.

BACKGROUND: Juvenile polyposis syndrome (JPS) is characterised by gastrointestinal (GI) hamartomatous polyposis and an increased risk of GI malignancy. Juvenile polyps also occur in the Cowden (CS), Bannayan-Ruvalcaba-Riley (BRRS) and Gorlin (GS) syndromes. Diagnosing JPS can be problematic because it relies on exclusion of CS, BRRS, and GS. Germline mutations in the PTCH, PTEN and DPC4 (SMAD4) genes can cause GS, CS/BRRS, and JPS, respectively. AIMS: To examine the contribution of mutations in PTCH, PTEN, and DPC4 (SMAD4) to JPS. METHODS: Forty seven individuals from 15 families and nine apparently sporadic cases with JPS were screened for germline mutations in DPC4, PTEN, and PTCH. RESULTS: No patient had a mutation in PTEN or PTCH. Five different germline mutations were detected in DPC4; three of these were deletions, one a single base substitution creating a stop codon, and one a missense change. None of these patients had distinguishing clinical features. CONCLUSIONS: Mutations in PTEN and PTCH are unlikely to cause juvenile polyposis in the absence of clinical features indicative of CS, BRRS, or GS. A proportion of JPS patients harbour DPC4 mutations (21% in this study) but there remains uncharacterized genetic heterogeneity in JPS.

Adenomatous Polyposis Coli↗

PubMed1999

Frequent mutation of beta-catenin and APC genes in primary colorectal tumors from patients with hereditary nonpolyposis colorectal cancer.

Hereditary nonpolyposis colorectal cancer (HNPCC) is characterized by defective DNA mismatch repair, which results in genetic instability of tumors; however, only a few target genes have been recognized. Our previous study detected a low frequency of APC gene mutation (21%) in colorectal tumors from HNPCC patients, in contrast to a high frequency of APC gene alteration (>70%) in non-HNPCC tumors. Because both beta-catenin and ACP gene mutations have recently been shown to activate the same signaling pathway, we analyzed beta-catenin mutation in HNPCC tumors. A notable frequency of beta-catenin gene mutation (43%, 12 of 28) was found to occur in HNPCC colorectal tumors. Beta-catenin mutations were not detected in tumors with APC mutations. All beta-catenin mutations detected in HNPCC tumors existed within the regulatory domain of beta-catenin. Immunohistochemical staining of tumors with this mutation showed accumulation of beta-catenin protein in nuclei. These and previous data from our laboratory suggest that activation of the beta-catenin-Tcf signaling pathway, through either beta-catenin or APC mutation, contributes to HNPCC colorectal carcinogenesis in approximately 65% of cases.

Base Pair Mismatch↗

PubMed1999

Regiocontrolled synthesis of carbocycle-fused indoles via arylation of silyl enol ethers with o-nitrophenylphenyliodonium fluoride.

[formula: see text] A new, regiocontrolled synthesis of carbocycle-fused indoles has been developed. The two-step procedure involves first the regiospecific arylation of silyl enol ethers with o-nitrophenylphenyliodonium fluoride (1). Reduction of the nitro group on the aromatic ring with TiCl3 followed by spontaneous condensation of the aniline with the ketone then affords the indole products.

Indoles↗

PubMed1999

Predominant germ-line mutation of the hMSH2 gene in Japanese hereditary non-polyposis colorectal cancer kindreds.

By means of PCR-SSCP and direct sequencing, we detected 12 germ-line mutations of hMSH2 or hMLH1 in 37 Japanese hereditary non-polyposis colorectal cancer (HNPCC) kindreds, of whom 15 satisfied the Amsterdam and 22 the Japanese criteria. The germ-line mutation detection rate of hMSH2 was much higher than that of hMLH1 (11/37 vs. 1/37). The total mutation detection rate of hMSH2 and hMLH1 in the Amsterdam criteria group was significantly higher than that in the Japanese criteria group (9/15 vs. 3/22). Furthermore, the mean age of the HNPCC patients in the mutation-positive group was lower than that in the mutation-negative one; the rates of both vertical transmission and multiplicity of tumors in the mutation-positive group were higher than those in the mutation-negative one. In addition, the number of patients with microsatellite instability-positive cancers in the mutation-positive group was higher than that in the mutation-negative one. Our results suggest firstly that the hMSH2 gene plays a much more important role than hMLH1 in the carcinogenesis of Japanese HNPCC patients, secondly that the rate of hMSH2 and hMLH1 mutations is high in the kindreds satisfying the Amsterdam criteria and thirdly that both the clinical phenotypes (early onset, vertical transmission and multiplicity of tumors) and the microsatellite instability status are important for the genetic screening of HNPCC.

Colorectal Neoplasms, Hereditary Nonpolyposis↗

PubMed1999

Higher frequency of Smad4 gene mutation in human colorectal cancer with distant metastasis.

We have previously detected an increased frequency of loss of heterozygosity (LOH) on chromosome 18q during progression of colorectal carcinomas. To clarify the target of 18qLOH, mutation of Smad4 and Smad2 genes was analysed in 176 colorectal tumors with different stages, including liver metastasis, from 111 sporadic, 52 familial adenomatous polyposis (FAP) and nine hereditary nonpolyposis colorectal cancer (HNPCC) patients. Mutation of other Smad gene families in the TGF-beta signaling pathway was also examined. Twenty-one Smad4 mutations and one Smad2 mutation were detected, whereas mutation of Smad3, 6 and 7 genes was not detected. Smad4 mutations included seven frameshift, one inframe deletion, four nonsense and nine missense mutations, 95% of which resulted in alteration of Smad4 protein regions included in homo-oligomer and hetero-oligomer formation. Frequencies of tumors with Smad4 mutation were 0/40 (0%) in adenoma, 4/39 (10%) in intramucosal carcinoma, 3/44 (7%) in primary invasive carcinoma without distant metastasis, 6/17 (35%) in primary invasive carcinoma with distant metastasis, and 11/36 (31%) in distant metastasis (metastatic/non-metastatic: P=0.006 approximately 0.01). Loss of the other allele was observed in 19 of 20 (95%) invasive and metastasized carcinomas with Smad4 mutations. In four cases both primary and metastasized carcinomas in the same patients showed the same mutations. The present results suggest that Smad4 gene is one of true targets of 18qLOH, and that its inactivation is involved in advanced stages, such as distant metastasis, in human colorectal carcinogenesis.

Colorectal Neoplasms↗

PubMed1999

Allelic imbalance at the LKB1 (STK11) locus in tumours from patients with Peutz-Jeghers' syndrome provides evidence for a hamartoma-(adenoma)-carcinoma sequence.

Patients with Peutz-Jeghers' syndrome (PJS) develop hamartomatous gastrointestinal polyps and characteristic pigmentation, as a result of germline mutations in the LKB1 gene. The hamartomas in PJS were long considered to be without malignant potential. There is, however, accumulating epidemiological evidence to suggest that PJS predisposes to cancers at several different sites (colon, pancreas, breast, ovary, testis, and cervix), although large enough patient samples are rarely available to prove this. Allelic imbalance [allele loss, loss of heterozygosity (LOH)] has previously been reported in a small number of PJS polyps, suggesting that LKB1 acts as a tumour suppressor in these tumours. This study confirms allelic loss at LKB1 in PJS polyps and shows that LOH also occurs in cancers of the colon, breast, and cervix in PJS patients. Allele loss was additionally found in a colonic adenoma from a PJS patient, strongly suggesting the existence of a hamartoma-(adenoma)-carcinoma sequence in tumourigenesis. These results provide molecular evidence that PJS patients are predisposed to cancers at several sites, as a direct result of selection for loss of the 'wild-type' LKB1 allele in tumours. Given the rare involvement of LKB1 in sporadic cancers, these data also suggest that the indirect effect on cancer risk (or 'bystander effect') proposed for hamartomas in juvenile polyposis does not apply to carcinomas in PJS.

AMP-Activated Protein Kinase Kinases↗