Biomedical subjects
Publications and source records attributed to J Yamamoto.
Structures of 17 minor forms of aureobasidins (Abs), Abs B-R, were elucidated by mass fragmentation and amino acid analysis. The fragmentation patterns by FAB-MS spectroscopy of Abs A-E seemed to follow predictable rules, so we used the rules to elucidate the 13 other Abs. All Abs consisted of eight amino acids and one hydroxy acid.
Aureobasidin A, a new antifungal antibiotic, was isolated from the culture medium of Aureobasidium pullulans R106. Aureobasidin A was a cyclic depsipeptide consisting of eight alpha-amino acid units and one hydroxy acid unit. The structures of the units were found by acid hydrolysis of the antibiotic to be 2(R)-hydroxy-3(R)-methylpentanoic acid, beta-hydroxy-N-methyl-L-valine, N-methyl-L-valine, L-proline, allo-L-isoleucine, N-methyl-L-phenylalanine, L-leucine, and L-phenyl-alanine. The sequence of the units was identified by NMR and FAB-MS of the products from the alkaline hydrolysis of aureobasidin A.
A 3.8-kb EcoRI-fragment containing the lysA gene [diaminopimelate (DAP) decarboxylase] of Bacillus subtilis has been cloned into B. subtilis phage phi 105 and its nucleotides sequenced. The nucleotide sequence of a 3,762 bp stretch contained three open reading frames (ORF1, ORF2, and ORF3) in one orientation and another open reading frame (ORF4) in the opposite orientation. ORF2 coded for the lysA gene based on the complementation of a B. subtilis lys auxotroph and on the fact that the predicted amino acid sequence (440 amino acids with a molecular weight of 48,876) of ORF2 shared a 29.7%, 38.3%, and 32.9% identity with the sequences of Escherichia coli, Corynebacterium glutamicum and Pseudomonas aeruginosa lysA genes, respectively. ORF1, ORF3, and ORF4 did not correspond to E. coli lysR. Based on the comparison of the B. subtilis lysA sequence with a sequence of the DAP-decarboxylase gene cloned into pUB110 (Yamamoto et al., Nucleic Acids Res., 17, 10105 (1989], it was found that the lys gene in the plasmid was fused with the dnaN gene in its COOH-terminal region.
Extrahepatic division of the right hepatic vein, after division of the inferior vena cava ligament and, if necessary, of the adrenal gland, was performed during 87 liver resections, and the results compared with those of intraparenchymal division of the right hepatic vein in 32 similar hepatectomies. The selection of the method reflected the surgeon's preference, and was unrelated to the weight of the resected specimen. The incidences of injury to the adrenal gland, inferior vena cava and short hepatic veins were comparable with the two methods, 28 versus 26%, but massive bleeding occurred in 13 vs. 6%, and the mean operative bleeding was 3.8 vs. 1.8 liters (p less than 0.01) during intrahepatic and extrahepatic dissection, respectively. Tumors close to the junction of the right hepatic vein with the inferior vena cava cannot be removed radically with intraparenchymal division of the right hepatic vein. For these reasons, we prefer extrahepatic right hepatic vein division, which was technically possible in 75 of the 84 cases in whom it was attempted.
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For quantitative evaluation of acute myocardial infarction, In-111 antimyosin Fab myocardial imaging (InAM) was performed in 17 patients with myocardial infarction who underwent Tl-201 (TL) and Tc-99m pyrophosphate (PYP) myocardial imaging in acute phase. For calculating the infarct size, voxel counter method was used for analysis in PYP and InAM, and extent and severity score were used on bull's-eye polar map in TL. The most appropriate cut-off level ranged from 65 to 80% by the fundamental experiment using cardiac phantom. The cut-off level of 0.70 (InAM) and 0.65 (PYP) were used for clinical application of voxel counter analysis. The infarct size calculated by InAM and PYP was compared with wall motion abnormality index by echocardiography (WMAI), TL extent score, TL severity score, peak CK and sigma CK. Infarct size by InAM showed the following correlations with other indices. PYP: r = 0.26 (ns), TL extent score: r = 0.72 (p less than 0.01), TL severity score: r = 0.65 (p less than 0.05), WMAI: r = 0.69 (p less than 0.05). The infarct size by PYP did not show any correlations with these indices. Therefore, the infarct size by InAM showed better correlations with TL and WMAI than that of PYP. So InAM was considered superior to PYP for quantitative evaluation of acute myocardial infarction.
A 72-year-old man was found to have a 12 mm solid lesion in the pancreatic tail and an 8 mm cystic lesion in the body of the pancreas by computed tomography carried out during a routine follow-up study of his adult-onset diabetes mellitus. A distal pancreatectomy was performed revealing the pancreas to have an adenocarcinoma in the tail and a conglomeration of intraductal papillary adenoma in the body. A review of the literature disclosed similar coexistences of cystadenoma and carcinoma in four patients, none of which was documented with preoperative imaging features such as we had.
To elucidate changes with time in T1-201 scintigraphy after coronary revascularization, T1-201 stress myocardial scintigraphy was performed at least twice during the follow-up period (from one to 12 months) in 58 patients with ischemic heart disease (12 with angina, and 46 with myocardial infarction) who had undergone PTCA or A-C bypass surgery. The perfusion defects were classified in 4 grades, and scintigraphic changes over grade 1 were judged significant. We evaluated; 1) time of scintigraphic improvement after revascularization, 2) presence of reverse redistribution, and 3) assessment of coronary restenosis. Scintigraphic improvement was observed in 21 of 58 patients during a 3- to 12- month follow-up period, 7 of whom improved within one month. Reverse redistribution after coronary revascularization was observed in 8 of the 58 patients (14%), including 6 who showed scintigraphic improvement in 3 to 12 months (2 were not examined). Among 29 patients whose coronary angiogram and Tl-201 scintigram were compared, 11 had angiographic evidence of restenosis and 4 of them showed deterioration of scintigraphic findings (sensitivity 57%, specificity 68%, and accuracy 66%). In conclusion, scintigraphic improvement was observed over various periods (immediately after and up to 12 months) after coronary revascularization. Reverse redistribution appears to be a predictor of good prognosis. Coronary restenosis cannot always be reliably assessed by Tl-201 scintigraphy.
To assess the diagnostic accuracy, extent, and characteristics of 111In-antimyosin Fab scintigraphy (In-AM) in acute myocardial infarction (AMI), we studied In-AM in 17 patients with AMI and compared with In-AM, 99mTc-PYP and 201Tl scintigraphy. Intensity of In-AM uptake was classified into 3 grades. Fourteen of 17 patients (82%) showed positive uptake of In-AM. The locations of infarct area diagnosed by In-AM were in accordance with those by electrocardiography. There was a good correlation between the extent score of In-AM planar and that of SPECT (r = 0.72), In-AM SPECT and Tl SPECT (r = 0.79), In-AM planar and PYP planar (r = 0.92), In-AM SPECT and PYP SPECT (r = 0.76), respectively (p less than 0.01). Thus, In-AM is a useful method for diagnosis of AMI.
Ultrasonography plays an important role in the early detection of hepatocellular carcinoma. Ultrasonography detected 53% of 287 patients with small (less than or equal to 5 cm) hepatocellular carcinomas. Among 486 patients with hepatocellular carcinomas, sensitivities of intra-operative ultrasonography in detecting 451 small primary hepatocellular carcinoma nodules, intrahepatic metastasis in 330 patients with small hepatocellular carcinoma and 63 tumor thrombi were 98, 48 and 67%, respectively. The sensitivity of intra-operative ultrasonography in detecting small primary tumors was 10% better than ultrasonography, computed tomography, and angiography. The sensitivity of intra-operative ultrasonography in intrahepatic metastasis and tumor thrombus was two to three times better than pre-operative examination. Intra-operative ultrasonography was useful in detecting nonpalpable tumors and in guiding the transection of the liver, biopsy, and cryosurgery. Moreover, intra-operative ultrasonography made possible new hepatectomy procedures: systematic subsegmentectomy and hepatectomies which preserve the inferior right hepatic vein. Systematic subsegmentectomy guided by intra-operative ultrasonography resulted in better survival rates than the limited resection in patients with small hepatocellular carcinoma two years after hepatectomy; by the sixth year, this difference was significant (p less than 0.05). Ultrasonography and intra-operative ultrasonography are indispensable in the early detection, accurate diagnosis, operative guidance and postoperative care of hepatocellular carcinoma.
Postoperative liver failure following major hepatectomy is a frightened complication. In order to increase safety of major hepatectomy, a study evaluating preoperative portal embolization (PE) was performed. Between 1983 and 1990, PE was performed in 71 patients (41 with hepatocellular carcinoma (HCC), 8 with other liver tumors, and 22 with biliary carcinoma), in 63 cases to the main branch, prior to hepatectomy. Out of these 63 patients 42 (extended) hemihepatectomies were performed. For comparison 77 patients with the same extent of hepatectomy, but without PE, were studied. Liver volume was evaluated by CT examination. Portal venous pressure was elevated by 73 mmH2O just after embolization of the main portal branch. However, no complications were associated to the PE procedure except for temporary elevation of transaminases. Volume of the unembolized lobe increased by 8.8% in average following PE. Prominent volume gain was observed in the patients with prior arterial embolization and long observation period. In patients with PE prior to major hepatectomy, postoperative bilirubin values were significantly lower (p less than 0.01). In the group with PE, no fulminant liver failure was seen and no operative mortality was encountered, as compared to 11.7% (9/77) in the control group. Preoperatively performed PE was a safe procedure decreasing postoperative liver failure and mortality.
The primary structure of L-histidine decarboxylase (HDC: L-histidine carboxy-lyase, EC 4.1.1.22) from mouse mastocytoma P-815 cells has been determined by parallel analysis of the amino acid sequence of the protein and the nucleotide sequence of the corresponding cDNA. HDC contains 662 amino acid residues with a molecular mass of 74017, which is larger by about 21,000 Da than that of the previously purified HDC subunit (53 kDa), suggesting that HDC might be posttranslationally processed. The HDC cDNA hybridized to a 2.7 kilobase mRNA of mastocytoma cells. Homology was found between the sequences of mouse mastocytoma HDC and fetal rat liver HDC.
A trihexacontapeptide corresponding to the sequence 8-70 of eglin c and its related peptides were synthesized by the conventional solution method and their inhibitory activity against human leukocyte elastase, cathepsin G and alpha-chymotrypsin was examined. Although synthetic eglin c (41-49) inhibited cathepsin G and alpha-chymotrypsin (Ki = 4.0 x 10(-5) M and 2.0 x 10(-5) M, respectively) but not leukocyte elastase, the synthetic trihexacontapeptide potently inhibited cathepsin G, alpha-chymotrypsin and leukocyte elastase (Ki = 1.8 x 10(-9) M, 1.4 x 10(-9) M and 2.2 x 10(-9) M, respectively). The relationship between the structure of eglin c and the inhibitory activity against the above enzymes is also described.
A novel regulatory protein for the rho proteins (rhoA p21 and rhoB p20), belonging to a ras p21/ras p21-like small molecular weight (Mr) GTP-binding protein (G protein) superfamily, was purified to near homogeneity from bovine brain cytosol and characterized. This regulatory protein, designated here as GDP dissociation inhibitor (GDI) for the rho proteins (rho GDI), inhibited the dissociation of GDP from rhoB p20 and the binding of guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) to the GDP-bound form of rhoB p20 but not of that to the guanine nucleotide-free form. The Mr value of rho GDI was estimated to be about 27,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and from the S value, indicating that rho GDI is composed of a single polypeptide without a subunit structure. The isoelectric point was about pH 5.7. rho GDI made a complex with the GDP-bound form of rhoB p20 with a molar ratio of 1:1 but not with the GTP gamma S-bound or guanine nucleotide-free form. rho GDI did not stimulate the GTPase activity of rhoB p20 and by itself showed neither GTP gamma S-binding nor GTPase activity. rho GDI was equally active for rhoA p21 and rhoB p20 but was inactive for other ras p21/ras p21-like G proteins including c-Ha-ras p21, smg p25A, and smg p21. rho GDI activity was detected in the cytosol fraction of various rat tissues. These results indicate that, in mammalian tissues, there is a novel type of regulatory protein specific for the rho proteins that interacts with the GDP-bound form of the rho proteins and thereby regulates the GDP/GTP exchange reaction of the rho proteins by inhibiting the dissociation of GDP from and the subsequent binding of GTP to them. Since there is a GTPase-activating protein for the rho proteins stimulating the GTPase activity of the rho proteins in mammalian tissues, the rho proteins appear to be regulated at least by GTPase-activating protein and GDI in a dual manner.
Racemic 7-r,8-t-dihydroxy-9-t,10-t-epoxy-7,8,9,10-tetrahydrobenzo[a] pyrene was reacted with yeast RNA. Modified nucleosides were isolated and resolved by high-performance liquid chromatography; nine adduct peaks were collected for analysis. The bases in these adducts were identified by comparing their retention times with those of adducts from poly(G), poly(A), and poly(C). These samples gave two major and two minor Guo adducts, four major Ado adducts, and at least four Cyd adducts. The relative efficiencies of adduct formation with the polyribonucleotides were poly(G) greater than yeast RNA greater than poly(A) greater than poly(C). Fluorescence measurements show that emission from Guo adducts is strongly quenched relative to that from Ado adducts. Liquid secondary ion mass spectrometry (LSIMS) of underivatized samples and electron-impact mass spectrometry (EIMS) of permethyl derivatives were used to confirm the base identities and establish the alkylation sites of the RNA adducts. Unique nitrogen-containing hydrocarbon fragments that were observed with all samples by EIMS establish that in each adduct analyzed the C-10 position of the hydrocarbon is linked to the exocyclic amino group of the base. This suggested that the multiple adducts formed with each base are diastereomers derived from cis/trans epoxide ring opening of the (+) and (-) enantiomers of the carcinogen. Several adducts exhibited molecular ions by both LSIMS and EIMS. Large fragments observed by EIMS usually resulted from the loss of CH3OH, CH3O., CH2O, CH3., and H. from the molecular ion. Major fragmentation pathways also resulted in formation of nucleoside, base, ribose, hydrocarbon, and base-hydrocarbon ions. Each of these major ions in turn resulted in further characteristic fragmentation patterns.
A novel HLA-B5 CREG gene, HLA-B SNA was cloned and the primary structure was determined. The sequence data showed that HLA-B SNA was identical to HLA-B51 except the alpha 1 domain in which one amino acid substitution at residue 74 and 5 amino acid substitutions associated with the Bw4/Bw6 epitopes were observed between these Ag. The comparison with other HLA-B locus genes suggested that HLA-B SNA evolved from HLA-B51 by gene exchange or recombination at the exon 2 between HLA-B51 and B8. A total of 10 of 14 HLA-B51-specific CTL clones showed significantly weak or no recognition of HLA-B SNA Ag. They also gave the same degree of a lysis of Hmy2CIR cells expressing the HLA-B35/51 chimeric Ag composed of the alpha 1 domain of HLA-B35 and other domains of HLA-B51 as that of Hmy2CIR cells expressing the HLA-B SNA Ag. These results demonstrated that amino acid substitutions within positions 77-83 associated with the HLA-Bw4/Bw6 epitopes have an influence on recognition of the HLA-B SNA antigen by HLA-B51-specific CTL.
To observe the rat brain blood flow with microspheres, a left ventricular catheter for sphere introduction was threaded into position via the right brachial artery. Cross-circulation with a donor rat was employed to prevent any mismatch between the infused and sampled blood volumes. Multiple measurements yielded stable flow data, and the responsiveness to hypercapnia was well preserved, irrespective of the dose of injected spheres (48 x 10(3) and 130 x 10(3)).
A protein stimulating the GTPase activity of rhoB p20, a ras p21-like GTP-binding protein (G protein), was partially purified from the cytosol fraction of bovine brain. This protein, designated as rhoB p20 GTPase-activating protein (GAP), did not stimulate the GTPase activity of other ras p21/ras p21-like G proteins including c-Ha-ras p21, smg p21 and smg p25A. The activities of c-Ha-ras p21 GAP and smg p21 GAP were also detected in the cytosol fraction of bovine brain and rhoB p20 GAP was separated from these GAPs. The activity of rhoB p20 GAP was eliminated by tryptic digestion or boiling. The Mr value of rhoB p20 GAP was estimated to be 150-200 x 10(3) and 37 x 10(3) by gel filtration and sucrose density gradient ultracentrifugation, respectively. These results indicate that there is rhoB p20 GAP in addition to c-Ha-ras p21 GAP and smg p21 GAP in bovine brain. In rat brain, about 50% of rhoB p20 GAP was found with the highest specific activity in the P2 fraction containing myelin, synaptosomes and mitochondria. In the P2 fraction, about 30% of rhoB p20 GAP was found in the P2C fraction containing mainly synaptosomes. rhoB p20 GAP was detected in the cytosol and particulate fractions of not only rat brain but also other rat tissues.