Biomedical subjects
Publications and source records attributed to T Aoki.
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Long-Evans Cinnamon rats develop a necrotizing hepatitis characterized by excessive hepatic copper accumulation, defective holoceruloplasmin biosynthesis and impaired biliary copper excretion. To elucidate the molecular basis of this defect, a cDNA clone encoding the rat Wilson disease gene was isolated and used to examine gene expression in selected tissues from normal and Long-Evans Cinnamon rats. Although this cDNA readily detects Wilson transcripts in liver and other tissues from normal rats, such transcripts are entirely absent from tissues derived from the Long-Evans Cinnamon rat strain. These data therefore identify the Long-Evans Cinnamon rat as the first bona fide animal model of Wilson disease and suggest that this rat strain may be a valuable resource in the study of this genetic disorder.
We previously reported frequent loss of heterozygosity on chromosome 9p in esophageal carcinomas and suggested that a tumor suppressor gene located on this chromosomal arm might be involved in development of these cancers. Since recently published studies have shown that a gene mapped on chromosome 9p21, MTS1/CDK4I (multiple tumor suppressor 1/cyclin-dependent kinase 4 inhibitor), is frequently mutated in various types of tumors, we chose to examine esophageal squamous cell carcinomas for mutations in this candidate gene. DNA sequence analyses revealed somatic mutations of MTS1/CDK4I in 14 of 27 tumors examined; 8 were frame-shift mutations and 6 were missense mutations. These results suggested that the MTS1/CDK4I gene is a tumor suppressor the inactivation of which plays an important role during carcinogenesis of the squamous cell type of esophageal carcinoma.
Several episodes of "smelter disease", previously assumed to be caused by sulphur dioxide (SO2) poisoning, have been reported in workers replacing pipes in sulphuric acid manufacturing plants. One such incident, affecting 20 men, was recorded in Akita, Japan, in July, 1993, but the protection these workers used suggested that some cause other than SO2 needed to be looked for. 10 workers were affected despite wearing respirators with SO2 cartridges, the symptoms including dyspnoea, diarrhoea, colicky pain, muscle pain and eczema with erythema. Subsequently 10 other workers using face masks with supplied air were affected, though without respiratory symptoms. Sludge in the piping contained mercuric sulphate, and mercury fumes resulted when pipes were cut with gas burners. Blood and urine measurements confirmed heavy exposure to the metal, and simulation experiments in rats showed that skin absorption was likely too. The masks with supplied air ought to have excluded both SO2 and mercury fumes. The only way to avoid smelter disease reliably is to wear an encapsulated suit that prevents inhalation and skin absorption of industrial toxins.
BACKGROUND: Pulmonary fiber content of both asbestos and nonasbestos types were evaluated in Japanese patients with malignant pleural mesotheliomas. METHODS: Pulmonary fiber content was analyzed in 16 patients and 16 case-matched control subjects by transmission electron microscopy with energy-dispersive X-ray analysis using a low-temperature ashing procedure. RESULTS: The geometric mean content of total asbestos was significantly higher in the patients (22.0 x 10(6) fibers/g dry lung) than in the control subjects (2.24 x 10(6) fibers/g dry lung) (P < 0.01). When the asbestos content was analyzed by fiber type, the geometric means were also consistently and significantly higher among the patients compared with the control subjects (P < 0.01). Results were as follows: (1) amosite: patients 3.94 times 10(6) versus control subjects 0.23 x 10(6); (2) crocidolite: patients 3.56 times 10(6) versus control subjects 0.35 times 10(6); (3) total amphiboles: patients 16.0 times 10(6) versus control subjects 0.77 times 10(6); and (4) chrysotile: patients 3.76 times 10(6) versus control subjects 1.01 times 10(6). However, when individual total asbestos content was considered, 7 of the 16 patients (44%) had levels lower than the highest value noted among the control subjects. Pulmonary fiber content of patients and control subjects also revealed the presence of nonasbestos fibers. The geometric mean of nonasbestos fibers was significantly higher in the patients (87.3 x 10(6)) than in control subjects (33.8 x 10(6)) (P < 0.01). The major type of nonasbestos fibers in both groups was aluminum silicates. The mean of ratios of nonasbestos fiber contents to total asbestos contents in the patients and control subjects was 7.0 and 17.3, respectively. CONCLUSIONS: The results were mainly in agreement with the findings of earlier investigations, but fiber content of both chrysotile and nonasbestos fiber as well as those of amphibole asbestos were significantly higher in the patients than in the control subjects.
Ninety-four esophageal squamous cell carcinomas were examined for loss of heterozygosity at several loci on the long arm of chromosome 17 (17q), using restriction fragment length polymorphism markers. Loss of heterozygosity was observed in 56 (62%) of 91 tumors that were informative with at least one marker. Comparison of these results with clinicopathological data indicated that the losses on chromosome 17q had occurred at an early stage of carcinogenesis. Detailed deletion mapping in these tumors revealed that the region commonly deleted was within the segment between loci defined by two markers at chromosomal band 17q21.3.
The Turcot syndrome (TS) is a rare, probably autosomal recessive, disorder characterized by development of primary neuroepithelial tumors of the central nervous system (CNS) and numerous adenomatous colorectal polyps. To examine the possible involvement of mutations of the APC gene, which is responsible for familial adenomatous polyposis (FAP), in Turcot syndrome, we examined DNAs from TS patients for alterations in this gene by means of ribonuclease protection analysis. Germ-line APC mutations were detected in each of three unrelated cases of TS, and additional (somatic) mutations were observed in colonic adenomas that had developed in one of these patients. However, no somatic mutations in APC were found among 91 neuroepithelial tumors (medulloblastoma, glioblastoma, astrocytoma, and oligodendroglioma), whether sporadic or associated with TS. These results suggest that the APC gene is associated with pathogenesis of one feature of TS, but that at least one other gene is responsible for the genesis of neuroepithelial tumors in the CNS.
To investigate genetic features of esophageal cancer, we have examined 93 squamous cell carcinomas of the esophagus for loss of heterozygosity (LOH), using 41 restriction fragment length polymorphism (RFLP) markers representing all autosomal chromosomes. Allelic losses at frequencies of at least 30% were observed at loci on chromosomal arms 3p (35%), 3q (30%), 5q (36%), 9p (57%), 9q (60%), 10p (33%), 13q (43%), 17p (62%), 17q (46%), 18q (38%), 19q (32%), and 21q (37%). These results suggest that several putative tumor suppressor genes, in addition to the cyclin D and TP53 genes that are sometimes mutated in esophageal carcinomas, may be associated with development and/or progression of esophageal cancer. By a comparison of LOH on each chromosomal arm with clinicopathological parameters, we have found a significant correlation between LOH on 19q and regional lymph node metastases. Interestingly, the frequency of LOH on 17q was significantly higher in tumors in female patients (12 of 14 cases) than in those in male patients (20 of 56 cases) (P = 0.0009 by Fisher's exact test). Furthermore, we examined for mutations of the APC gene on chromosome arm 5q. Screening of nearly one third of the APC coding region, including the MCR (mutation cluster region), revealed no alterations. Therefore, although allelic loss at the APC locus is frequent in squamous cell carcinomas of the esophagus, it is likely that a gene on 5q other than APC is involved in esophageal tumorigenesis.
To investigate genetic features in small and flat colorectal carcinomas that arise de novo, we searched for genetic alterations in six sporadic tumors by examining their APC, K-ras, and p53 genes. Two of the six tumors carried detectable mutations within the mutation cluster region (MCR) of the APC gene; both mutations were predicted to cause truncation of the gene product. Four tumors carried mutations of the p53 gene; three were missense mutations in exon 5, and the other was a 3-bp deletion in exon 6. However, neither codon 12 nor codon 13 of K-ras contained detectable mutation in any tumors. Hence, as "adenoma-carcinoma sequence" model of development of colorectal carcinoma, inactivation of the APC and p53 genes appear to be involved in development of the de novo type of colorectal carcinoma even though the adenoma stage is not observed.
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The respiratory chain of adult Paragonimus westermani, a lung fluke, was characterized in isolated mitochondria. The fluke mitochondria exhibited cyanide- and antimycin A-sensitive succinate oxidase activity at a rate of 16.8 nmol O2 min-1 mg-1 protein. The succinate oxidation was shown to be stimulated by ADP and linked to the formation of membrane potential. The specific activities of oxidoreductases composing the succinate oxidase system, i.e., succinate-ubiquinone and succinate--cytochrome c oxidoreductase (complex II and complex II-III, respectively) and cytochrome c oxidase (complex IV), were compared in mitochondria from adult Paragonimus, bovine heart (an aerobic tissue), and muscle of adult Ascaris suum which possesses an anaerobic respiratory chain. The activity values of complex II-III and complex IV were high, middle, and low for bovine heart, Paragonimus, and A. suum, respectively, whereas the activity of complex II was comparable among the three sources. The cytochrome contents of Paragonimus mitochondria as determined by difference absorption spectrophotometry ranged between those in Ascaris and bovine mitochondria for types c and aa3 cytochromes. Paragonimus mitochondria exhibited a high activity of NADH-fumarate reductase; the specific activity was about 18-fold higher in fluke submitochondria than in bovine heart submitochondria. Quinone analysis by HPLC and mass spectrometry showed that the fluke mitochondria contain both rhodoquinone-10 and ubiquinone-10 at concentrations of 0.572 and 0.321 nmol mg-1 mitochondrial protein, respectively. These data clearly show that mitochondria from adult P. westermani, unlike adult Ascaris mitochondria, possess both cyanide-sensitive succinate oxidase and NADH-fumarate reductase systems, indicating that the fluke mitochondria are facultatively anaerobic.
Growth factors such as basic fibroblast growth factor (bFGF) have been found to promote the survival and proliferation of endothelial cells. However, the mechanism by which growth factors control the regeneration and degeneration of the endothelial cells remained poorly understood. In this study, we demonstrated that apoptosis of murine aortic endothelial (MAE) cells was induced by deprivation of bFGF but required new RNA and protein synthesis. Furthermore, enforced expression of bcl-2 gene in MAE cells using gene transfer techniques decreased apoptosis induced by deprivation of bFGF. These findings suggest that bcl-2 interferes with a pathway for endothelial cell death that is induced by deprivation of bFGF.
A superoxide dismutase (SOD) from adult worms of Dirofilaria immitis was purified using ethanol-chloroform and acetone treatment, DE 52 cellulose, and Sephadex G-75 gel chromatography to obtain a 677-fold purification and a specific activity of 5483 units/mg of protein. The purified SOD was essentially homogeneous as judged by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and composed of two identical 18.5 kDa subunits. The purified SOD was inactivated completely by 3 mM potassium cyanide, and reduced to about 18% of the initial activity by incubation at 100 degrees C for 10 min and to 28% by treatment with 2% SDS. This means that D. immitis SOD differs markedly from the mammalian Cu/Zn SOD, although it is indeed a Cu/Zn SOD. Atomic absorption spectrometry revealed the presence of 0.80 mole of Cu and 0.78 mole of Zn per mole of subunit. The enzyme consisted of 22.6% acidic and 11.9% basic amino acids. No free amino acid was detected in the SOD by N-terminal amino acid sequencing.
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The pathogenesis of delayed sequelae of carbon monoxide (CO) exposure is still unknown. We repeatedly examined a 55-year-old woman with the interval form of CO poisoning, using proton magnetic resonance spectroscopy (MRS). When the clinical picture was severe, MRS revealed markedly lowered N-acetyl-aspartate (NAA)/creatine and phosphocreatine (Cr) ratio and slightly increased choline containing compounds (Cho)/Cr ratio. Subsequently, NAA and Cho/Cr ratio tended to return to normal, reflecting clinical improvement. Proton MRS shows the previously unrecognised neuronal activity in CO poisoning and precisely reflects the severity of symptoms. We stress the superiority of proton MRS over the conventional radiological examinations in CO poisoning.
To evaluate the chronic effects of theophylline on cardiac function, M-mode and pulsed Doppler derived variables were measured at rest and the suprasternal continuous wave Doppler measurement of ascending aortic flow was used during treadmill exercise testing. Subjects consisted of 13 children with stable asthma (mean 11.7 +/- 2.2 years) who were treated with theophylline for at least one year and 16 age-matched, untreated normal volunteers. In the resting state, the chronic administration of theophylline seemed to produce a slight increase in percent fractional shortening, outflow peak velocity and atrial contribution to ventricular filling in the asthmatic children as compared to normals, but these changes were not statistically significant. The asthmatic children showed significantly lower values than the controls in exercise induced changes in the peak velocity, stroke index and cardiac index, but not in the heart rate. Therefore, chronic administration of theophylline appears to have a minimal effect on resting cardiac function, but a possibly deleterious effect on the cardiac response to exercise testing.
The purpose of this study was to evaluate the high resolution magnetic resonance angiography (MRA) as pre-operative angiography for the detection of ruptured cerebral aneurysms. MRA was performed on 1.5 tesla system using the 3-dimensional time of flight (3D-TOF) method. The field of view was 16 cm or 20 cm and matrix size was 192 x 256 or 256 x 512. Twenty patients with ruptured cerebral aneurysms and 35 cases of non-ruptured aneurysms (incidental aneurysm) examined by both conventional angiography and MRA were included in this study. All the ruptured aneurysms were operated on based on the information obtained from conventional angiography and MRA. Aneurysms smaller than 3 mm in size were difficult to visualize on MRA. However, most ruptured aneurysms were clearly visualized because they were generally larger than 3 mm. The image quality of MRA was satisfactory for planning surgery. Screening for non-ruptured cerebral aneurysms using MRA is controversial. However, in ruptured aneurysms larger than 3 mm in size, MRA clearly revealed the aneurysm, and MRA findings were informative enough to plan surgery. It is concluded that this noninvasive examination can be selected as a first-choice examination especially in cases of ruptured aneurysm.