Biomedical subjects
Publications and source records attributed to S Kojima.
We report the case of a 33-year-old woman with eosinophilic cystitis, which masqueraded as invasive bladder cancer. The patient remained well with no recurrence or progression after vigorous transurethral resection of the lesion.
The neurofibromatosis type 1 (NF1) gene has been considered to be a tumor suppressor gene, since the NF1 gene product downregulates the ras oncogene product (p21(ras)). In addition, children with NF1 show an increased incidence of myelogenous leukemia, including juvenile chronic myelogenous leukemia (JCML). We studied 8 Japanese JCML patients without NF1 for mutations in exons 21-36 of the NF1 gene by using polymerase chain reaction/single-strand conformation polymorphism analysis. This region was chosen because it includes two main hotspots in the NF1 gene as well as the functionally important domain, GTPase-activating protein-related domain, which mediates the downregulation of ras activity. One of the 8 JCML patients exhibited a G to T transversion at the third nucleotide of the codon GAG for Glu at amino acid residue 1699, which results in the heterozygous conversion to Asp (E1699D). This variation was detected neither in 65 healthy volunteers nor 50 NF1 patients. No other variations were detected in our JCML patients. We suggest that NF1 gene mutation does not occur frequently in JCML without NF1.
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Drug absorption studies using dogs have been difficult because of different gastrointestinal(GI) conditions between dogs and humans, including dogs' shorter intestinal transit time and strong agitation force in the GI tract. We attempted to modify the agitation force and GI transit time in dogs using codeine. The agitation force was examined based on the in vitro/in vivo correlation for a CR tablet of acetaminophen showing agitation-dependent release. Codeine improved the GI condition better than atropine or loperamide, employed previously.
Crosslinked dextran hydrogels were prepared from glycidyl methacrylate (GMA)-derivatized dextran by gamma-irradiation initiation in the presence of insulin and lysozyme as model protein and peptide drugs. The diffusion coefficient of insulin and lysozyme in the hydrogels was determined successfully by a pulsed-field-gradient stimulated-echo method. A conventional NMR probe which can impose a relatively small magnetic field gradient (30 Gauss/cm) was used for the measurements. This method seems to be applicable to proteins and peptides with a molecular weight of about 10,000 or less. The diffusion coefficient determined by the NMR method almost agreed with that calculated from the drug release profiles, indicating the usefulness of the diffusion coefficient determined by the NMR method as a measure of the release rate from hydrogels.
Streptomyces griseus metalloendopeptidase II (SGMPII) is a unique protease, since it shows anomalous susceptibility to the proteinaceous "serine protease inhibitors" produced by Streptomyces, such as Streptomyces subtilisin inhibitor (SSI) and its homologous proteins. In this study, we analyzed the amino acid sequence of SGMPII by analyzing various peptide fragments produced enzymatically. The sequence of SGMPII, which is composed of 334 amino acids, showed no extensive similarity to SSI-insensitive metalloproteases produced by other species of Streptomyces, except for the amino acid residues essential for catalysis and zinc binding. However, SGMPII is 35-41% similar to thermolysin and its related metalloproteases, which are not inhibited by SSI, and the residues presumed to be critical for catalysis and zinc-binding are well conserved in SGMPII. Glu137 in a "His-Glu-Xaa-His" motif of SGMPII was identified as the residue modified by CICH2 CO-DL-(N-OH)Leu-Ala-Gly-NH2, an active-site-directed irreversible inhibitor of thermolysin-like metalloproteases. Based on the sequence comparison of SGMPII and other bacterial metalloproteases, we discuss the structural basis for the differences in substrate specificity and stability between SGMPII and other thermolysin-like proteases. A possible SSI-binding locus of SGMPII is also proposed.
Aqualysin I is an alkaline serine protease isolated from Thermus aquaticus YT-1, an extreme thermophile. We have measured the P1-specificity of aqualysin I, using wild-type and five P1-substituted derivatives of Streptomyces subtilisin inhibitor (SSI). SSIs efficiently inhibited the activity of aqualysin I, with low substrate specificity. Charge and hydrophobicity of side chain of the P1 amino acid residue showed no significant effect to the P1-specificity of this enzyme.
To study the neurophysiological functions of metallothioneins (MTs), localization of MT-I and -II was examined immunohistochemically in a variety of brain lesions in dogs, including infarct, laminar cortical necrosis, hemorrhage, invasive growth of tumour, inflammatory lesions in granulomatous meningoencephalitis and distemper encephalitis. MT-I and -II were demonstrated in both nucleus and cytoplasm of hypertrophic astrocytes in most brain lesions examined regardless of the type, size, localization and duration of the lesions. In addition, MT expression was stronger in a population of hypertrophic astrocytes localizing inside of the surviving brain tissue rather than those localizing at the boundary between the surviving brain tissue and necrotic area, where severe inflammatory changes were developing. These results suggest that MT-I and -II may play roles not only in protection of neurons from metals and free radicals ubiquitous in the inflammatory lesions but also in repair of injured neural tissues.
Macrophage scavenger receptor A (MSR-A) deficient mice MSR-A(-/-) were infected by the intraperitoneal injection of the Plasmodium berghei NK65 strain in the erythrocytic stage. The MSR-A(-/-) mice died significantly earlier than the control mice (P=0.060). In the surviving mice, two peaks of parasitemia were observed: the first 5-7 days and the second at 2-3 weeks after infection. Death of all MSR-A(-/-) mice occurred at either peak of parasitemia, suggesting that MSR-A protects mice from severe infection. This model may be useful for the study of molecular mechanisms of macrophage functions in malaria infection.
We report a case of small cell carcinoma of the kidney in a 61-year-old female. Chief complaints were left lumbago, gross hematuria and high fever. The computed tomography revealed a large invasive tumor in the left renal pelvis with renal pedicle lymph nodes swelling. Systemic chemotherapy with methotrexate, vinblastine, doxorubicin and cisplatin (M-VAC) showed a considerable effect and left nephrectomy was performed. But she died within 3 months postoperatively. Histologically, the tumor was composed of hyperchromatic small cells with increased N/C ratio. Immunohistochemical studies revealed positive staining for chromogranin A, synaptophysin, NSE, EMA, cytokeratin, and argyrophilic (Grimelius) silver impregnation stain. Electron microscopy revealed neurosecretory granules also. In conclusion, the present case was diagnosed as small cell carcinoma of the kidney with only 12 similar cases reported in the world literature.
The collaborative study participated by seven laboratories was carried out to develop a dissolution standard for evaluating vibration levels of dissolution apparatuses using enteric-coated granules of cefalexin (EG). Dissolution apparatuses could be divided into two groups according to their vibration levels and the dissolution test results of EG by the rotating basket method at 50 rpm. The critical value of acceleration was about 0.05 m/s2. The upper limit of normal dissolution rates of EG was calculated from the results of the rotating basket method at 50 rpm obtained from low vibration apparatuses. All high vibration apparatuses used in this study were distinguished by the limit from low vibration apparatuses, although most of them were not distinguished by current USP calibrators. These results suggest that EG would be useful as a calibrator for detection of apparatuses on high vibration levels.
Assessment of shelf-life equivalence among batches, packaging or formulations of pharmaceutical products, which was based on the range of shelf-life estimates, was proposed as an alternative method to an analysis of variance (ANOVA). The power of this analysis was not significantly affected by assay error, whereas that of ANOVA decreased markedly as assay error increased. Thus, ANOVA exhibites a tendency to overlook the stability variation from stability data of a larger assay error, but this is not the case for the proposed method.
We have isolated and determined the complete nucleotide sequence of canine metallothionein-III (MT-III) cDNA. The predicted amino acid sequence of the canine MT-III showed a high homology (93%, 87% identity) to that of human and mouse MT-III. The canine MT-III had 2 insertions relative to known mammalian MT-I and MT-II: a threonine after the 4th amino acid and a block of 6 amino acids near the carboxyl terminus. Expression of the canine MT-III mRNA was found exclusively in the central nervous system, where neurons in the olfactory bulb, hippocampus and cerebral cortex showed predominant signals.
BACKGROUND: The vasoreactivity after direct percutaneous transluminal coronary angioplasty (PTCA) in patients with previous myocardial infarction remains unknown. We examined the constrictor response to ergonovine of the infarct-related coronary artery in comparison with that of noninfarct-related coronary artery after angioplasty. METHODS: Ergonovine was administered intravenously to 17 patients with previous myocardial infarction (group I) and to 21 patients with stable angina (group II) 1 year after PTCA. The effects of ergonovine on lumen diameter were analysed quantitatively at the PTCA segment, nonPTCA segment (proximal to the PTCA segment), and nonPTCA artery. RESULTS: The ergonovine-induced decrease in minimal lumen diameter at the PTCA segment was significant in group I (decrease from 2.12 +/- 0.56 to 1.39 +/- 0.74 mm, P < 0.01), but not in group II (decrease from 1.60 +/- 0.35 to 1.43 +/- 0.33 mm, NS). Patients in group I showed a constrictor response at the nonPTCA artery (decrease in diameter from 2.54 +/- 0.90 to 1.94 +/- 0.77 mm, P < 0.01), and a tendency to constrict at the nonPTCA segment (2.56 +/- 0.67 to 2.11 +/- 0.66 mm, P = 0.06), whereas those in group II showed no significant constrictor response to ergonovine at any of the three segments examined. The changes in diameter at the three segments in patients in group I were significantly greater than those in group II (all P < 0.01). Subtotal coronary spasm at the PTCA segment was provoked only in three patients in group I (18%). CONCLUSIONS: The constrictor response to ergonovine of the infarct-related coronary artery was enhanced compared with that of the noninfarct-related coronary artery. This difference in coronary vasoreactivity at the angioplasty segment may be due to previous hypersensitivity of the smooth muscle.
Liver endothelial cells are important components of the tissue along the hepatic sinusoid. They are responsible for microcirculation in the liver and scavenger functions. It would therefore be important to include these cells in any hybrid type of artificial liver in addition to hepatocytes. However, it is difficult to culture these cells in vitro. The development of a liver endothelial cell line, which maintains the characteristics of the primary culture, would thus be of great benefit in the development of an artificial liver. In the present study we established immortalized liver endothelial cells from the liver of an H-2Kb-tsA58 transgenic mouse, which harbors the SV40 TAg gene. Hepatic sinusoidal cells isolated from H-2Kb-tsA58 mouse proliferated in the presence of gamma-interferon at 33 degrees C. Four clones were established, out of which clone M1 had the highest amounts of PGI2 production, as well as plasminogen activator activity and internalized acetylated low density lipoprotein. On culture dishes the M1 cells grew individually and spread. Sieve plates on the cell surface were not readily visible, but small pores were detected under electron microscopic observation. These results suggest that M1 clone cells originated from liver endothelial cells. Moreover it was possible to culture the immortalized liver endothelial cells in a radial-flow bioreactor for 5 days, with a maximum 6-keto prostaglandin F1alpha production of 25 microg per day. This suggests that immortalized liver endothelial cells and a radial-flow bioreactor can prove useful tools in the development an artificial liver.
Blood activities of hepatocellular enzymes such as lactate dehydrogenase (LDH), glutamic pyruvic transaminase (GPT) and glutamic oxalacetic transaminase (GOT) peaked at 12 hours after a single intraabdominal injection of ferric nitrilotriacetate (Fe(3+)-NTA) in rats. Enzymes such as alkaline phosphatase (ALP) and leucin amino peptidase (LAP) originating in the capillary bile ducts or bile secretory liver cells were also released into the blood between 6-24 hours after intraabdominal injection of Fe(3+)-NTA in rats. Furthermore, hyperoxidation of lipids occurred in rat hepatic cell membranes, reaching a peak 6 hours after intraabdominal injection of Fe(3+)-NTA. It was found that a single prior 0.5 Gy whole body X-ray irradiation significantly increased superoxide dismutase (SOD) activities and suppressed above-mentioned symptoms of transient hepatopathy in rats.
From their crystallographic comparisons, Fisher et al. (Biochemistry 34, 8960-8972, 1995) have proposed that in an important transition of myosin heads (M), M.ATP-->M.ADP.Pi, an interdomain rotation occurs in Gly468 (of chicken smooth muscle myosin) and that the rotated state is stabilized by newly-formed interdomain contacts including the salt link between Glu470 and Arg247 (of chicken smooth muscle myosin). Here, we have studied the effects of Gly468, Glu470, and Arg247 mutations on the hydrolysis of ATP. The G468A HMM did not show a significant ATPase activity, a stoichiometric initial phosphate burst, and tryptophan fluorescence enhancement attributed to bound ADP.Pi. The E470A HMM also did not show a significant ATPase activity and the phosphate burst, but the mutant gave tryptophan response attributed to bound ATP. The E470R/R247E HMM exhibited an ATPase activity and the phosphate burst which were comparable to those of the wild-type HMM, whereas neither the E470R HMM nor the R247E HMM showed such a significant ATPase activity and burst. We thus propose that both an unhindered rotation and a salt link that stabilizes the rotated state are necessary for ATP hydrolysis.
For palliative therapy of Duchenne muscular dystrophy (DMD), corticosteroids have been tried since 1970. According to recent reports, corticosteroids maintained muscular strength to some extent and prolonged period of ambulation. However, their mechanism of action is mostly unclear. In the present study, mdx mice were injected with 0.6 mg prednisolone 3 times a week for 30 weeks. Serum creatine kinase (CK) values remained 23% of controls. In muscle pathology of the quadriceps muscle, fibers with peripheral nuclei were increased, suggesting reduction of muscle necrosis. In pathological examination of liver, pyknotic cytoplasmic masses and formation of vacuoles were observed. Present study showed that prednisolone might attenuate muscle fiber necrosis at least for 30 weeks. Prednisolone may reduce secondary tissue reactions, which develop more serious muscle damage.