Biomedical subjects

T Morita

Publications and source records attributed to T Morita.

At least 19 recordsLinked to original sources

PubMed1995

Regulation of the mouse histone H2A.X gene promoter by the transcription factor E2F and CCAAT binding protein.

We have molecularly cloned the genomic gene encoding the mouse histone variant H2A.X and characterized the promoter. The promoter region of the H2A.X gene was characterized by chloramphenicol acetyltransferase analysis using Balb/c 3T3 cells. Maximal promoter activity was found in the construct containing up to -282 base pairs H2A.X upstream region. Within this region, we found two sequences regulating the promoter activation; one was an E2F site and another was a CCAAT box. These sequences were also required for the DNA/protein binding activities. Thus, these activities corresponded to the promoter activities, implying that the promoter activity H2A.X gene was controlled by both the transcription factor E2F and H1TF2 through the E2F and CCAAT element. The CCAAT box binding activity was constitutive when cell cycle was progressed by release from G1 arrest, but transiently transfected chloramphenicol acetyltransferase activity slightly increased when cells entered S phase. Similarly, the level of the smallest form of E2F (free E2F) became higher when cells reentered the cell cycle, indicating that the free E2F was one capable of inducing the promoter activation. Thus, the free E2F and CCAAT DNA binding activity correlated with regulation of the promoter activity.

3T3 Cells↗

PubMed1995

Role of calcium(II) ions in the recognition of coagulation factors IX and X by IX/X-bp, an anticoagulant from snake venom.

IX/X-bp, an anticoagulant protein isolated from the venom of the habu snake Trimeresurus flavoviridis, has a structure homologous to the carbohydrate-recognition domains of C-type (Ca(2+)-dependent) animal lectins, and it binds to the gamma-carboxyglutamic acid (Gla) domains of coagulation factors IX and X in a Ca(2+)-dependent fashion. In the present study, we elucidated the role of Ca2+ ions in this binding. The binding of 125I-labeled IX/X-bp to both coagulation factors required about 1 mM Ca2+ ions in this at pH 7.5. A decrease in the pH to 6.5 had a striking negative effect on the binding, and the Ca(2+)-requirement curve was shifted rightward. We investigated the binding of Ca2+ ions to IX/X-bp directly by equilibrium dialysis and identified two independent binding sites with different affinities. At pH 7.5, the apparent Kd values for these sites were 25 and 200 microM, respectively. When the pH was decreased to 6.5, the affinity of the high-affinity binding site was reduced only slightly but that of the low-affinity site was reduced considerably. Moreover, it was evident from observations of Ca(2+)-induced changes in the intrinsic fluorescence that IX/X-bp underwent a conformational change upon binding of Ca2+ ions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Characterization of the testis-specific gene 'calmegin' promoter sequence and its activity defined by transgenic mouse experiments.

We have cloned the genomic DNA of calmegin [(1992) J. Biol. Chem. 269, 7744-7749] and analyzed its promoter region. It contained GC-rich sequences and potential binding sites for AP 2 and Sp 1, but lacked the TATA sequence. The 330 bp 5' flanking sequence of calmegin genomic DNA fused with the CAT gene was used for the study of promoter activity in transgenic mice. The CAT gene activity was detected exclusively in testes, indicating that the 330 bp calmegin 5' sequence was sufficient for the testis-specific expression. The existence of testicular nuclear factors specifically bound to the putative promoter sequence was also demonstrated.

Animals↗

PubMed1995

Identification of four different forms of syntaxin 3.

cDNAs for four different forms of syntaxin 3 were cloned from a mouse brain cDNA library, and the proteins encoded by these clones were named syntaxin 3A (previous syntaxin 3), 3B, 3C and 3D. Syntaxin 3B contained a different sequence in the carboxyl terminal region from that of syntaxin 3A. The amino terminal region of syntaxin 3C contained an 18 amino acid sequence instead of a 34 amino acid sequence present in syntaxins 3A and 3B. Syntaxin 3D consisted of only 86 amino acids and lacked any putative transmembrane segments. These forms of syntaxin 3 are probably generated by alternative splicing of the primary transcript of syntaxin 3 gene. Cytoplasmic portions of syntaxins 3A and 3B but not of syntaxin 3C or 3D bound to Munc-18/n-sec1.

Alternative Splicing↗

PubMed1995

Regulation of the tertiary structure and function of coagulation factor IX by magnesium (II) ions.

The indispensable role of Ca2+ ions in the maintenance of the functional tertiary structures of vitamin K-dependent coagulation factors has been definitively established but the participation of Mg2+ ions, another alkaline-earth metal that is present abundantly in blood plasma, in such a process is not yet understood. We show here that the Ca(2+)-stabilized conformation of coagulation factor IX undergoes a further conformational change upon binding of Mg2+ ions using three independent structural probes. The probes we used were (i) IX/X-bp, a snake venom anticoagulant that recognizes the Gla domains in coagulation factors IX and X, (ii) conformation-specific polyclonal antibodies against bovine factor IX, and (iii) monoclonal antibodies against the Gla domain of human factor IX. The binding of all these probes had an absolute requirement for Ca2+ ions, and Mg2+ ions alone were ineffective. However, when added together with Ca2+ ions, Mg2+ ions at physiological concentrations greatly augmented the binding of these probes to factor IX; the required concentration of Ca2+ ions was much reduced, and the affinity of each probe for factor IX was increased even in the presence of an excess of Ca2+ ions. These results suggest the presence of a Mg(2+)-specific binding site that does not interact with Ca2+ ions in factor IX. Furthermore, Mg2+ ions potentiated the susceptibility of factor IX to activation by factor XIa, concomitant with their effect on the conformation. Similarly, the required Ca2+ concentration was reduced by Mg2+ ions, and the rate of conversion to factor IXa was increased by Mg2+ ions in the presence of an excess of Ca2+ ions. At a saturating concentration of Ca2+ ions (5 mM), addition of 1 mM Mg2+ reduced the apparent Km value for factor IX from 0.31 to 0.18 microM, and in the presence of a physiological concentration of Ca2+ ions (1 mM), the reduction in Km by Mg2+ ions was far more striking (from 0.91 to 0.24 microM). The apparent Vmax values were hardly affected by Mg2+ ions. Our present data reveal a hitherto novel physiological role of the Mg2+ ions in plasma. Not only Ca2+ ions but also Mg2+ ions are important regulators of the stabilization of the native conformation of factor IX as well as of its efficient activation.

Animals↗

PubMed1995

Cloning and sequencing of the gene coding for dextranase from Streptococcus salivarius.

We cloned and sequenced the dextranase (Dex) (1,6-alpha-glucanhydrolase; EC 3.2.1.11)-encoding gene from Streptococcus salivarius (Ss) strain M-33. Recombinant clones from an Ss genomic library specifying Dex activity were identified as colonies surrounded by transparent halos on blue dextran plates. One of the clones had a 4.3-kb KpnI fragment containing the gene coding for an 826-amino-acid polypeptide with a molecular mass of 87.9 kDa, which corresponds well to that of native Dex from the Ss culture supernatant. There was no sequence homology between the gene encoding Ss Dex and the gene encoding dextran glucosidase of S. mutans, or between their protein products.

Amino Acid Sequence↗

PubMed1995

The cerebral hemodynamic response to electrically induced seizures in man.

The hemodynamic response to seizure has long been a topic for discussion in association with the neuronal damage resulting from convulsion. Electroconvulsive therapy (ECT) is an appropriate clinical model for the investigation of the cerebral physiology of seizure. In this study, we monitored the oxygenation state of brain tissue using near infrared (NIR) spectrophotometry, and flow velocity at the middle cerebral artery (MCA) using transcranial Doppler ultrasonography (tc-Doppler) in ninety cases where flow velocity at the middle cerebral artery (MCA) using transcranial Doppler ultrasonography (tc-Doppler) in ninety cases where ECT was prescribed to patients suffering from endogenous depression. Under general anesthesia with thiopental and succinyl choline, an electrical current was applied bilaterally at the minimal energy level. Throughout the therapy, end-tidal CO2 tension was maintained at 30-35 mmHg, and the SpO2 value was maintained above 98% by manual ventilation assistance. The total- and oxy-hemoglobin contents in the brain were reduced during the electrical shock, and then recovered to the pre-shock value (total-hemoglobin; 44.13 +/- 12.88 s after the shock, oxy-hemoglobin; 88.62 +/- 11.69 s after the shock). Subsequently, these values further increased beyond the preshock value. On the other hand, the deoxy-hemoglobin content increased for 90.73 +/- 15.88 s during and after the electrical shock, and decreased afterward. Reduction of cytochrome aa3 began 3.04 +/- 0.51 s after the electrical shock, and this was reoxygenated at 171.88 +/- 12.95 s after the shock.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗

PubMed1995

Chronic nerve root compression. Pathophysiologic mechanism of nerve root dysfunction.

STUDY DESIGN: The pathophysiologic changes of the spinal nerve root caused by chronic compression were assessed in the lumbar spine of the adult dog. METHODS: The seventh lumbar nerve root was banded with a Silastic tube 5 mm long and a 3 mm internal diameter, which is slightly larger than the diameter of the nerve root. Histologic and electrophysiologic studies as well as assessment of the blood-nerve barrier function in the nerve root were conducted at 24 hours through 12 months. RESULTS: The earliest findings were thickening of the dura mater and arachnoid membrane around the affected nerve root corresponding to the alteration of the blood-nerve barrier in the nerve root after 1 month. After 3 months, large myelinated fibers decreased in number and small newly formed fibers increased in the periphery of the fascicle. At 6 months, endoneurial fibrosis and Wallerian degeneration of nerve fibers became obvious. Compound action potentials and sensory nerve conduction velocity decreased by 3 and 12 months, respectively. Decrease in amplitude of the compressed action potential was identical to the period of decrease in large myelinated fibers, and nerve conduction velocity did not decrease as long as some large myelinated fiber remained until 12 months after tubing. CONCLUSION: Intraradicular edema caused by alteration of the blood-nerve barrier is the most important factor in the nerve root dysfunction due to chronic compression.

Animals↗

PubMed1995

Stress-response (heat-shock) protein 90 expression in tumors of the central nervous system: an immunohistochemical study.

This retrospective study deals with the expression of stress-response (heat-shock) protein 90 (srp 90) in a series of 148 human brain tumors. Immunohistochemical procedures were employed; cells of the human breast cancer line MCF7 exposed to hyperosmolar stress served as positive controls. Deposits of reaction products were found in the cytoplasm and they displayed a granular pattern. srp 90 was detected in 14/31 meningiomas and 5/10 breast cancer metastases to the brain. The protein was also present in 6/13 glioblastomas and 7/18 astrocytomas. In addition, a positive reaction was found in 2/10 medulloblastomas, 2/14 primitive neuroectodermal tumors, 1/11 pituitary tumor, 2/21 schwannomas and 2/11 lung tumor metastases; however, oligodendrogliomas and primary malignant lymphomas were not stained. The srp 90 was detected in Western blots of meningioma tissue homogenates. No significant immunohistochemical reaction was seen with sections of normal human cerebra, brain stem, cerebella, pituitary glands and spinal cords. These results document the expression of srp 90 by a variety of primary and metastatic intracranial tumors.

Autopsy↗

PubMed1995

The effects of intraportal prostaglandin E1 administration on hepatic warm ischemia and reperfusion injury in dogs.

To determine the route of prostaglandin E1 (PGE1) administration which would have the greatest protective effect against hepatic warm ischemia, two experiments were performed using dogs. The pharmacokinetics of PGE1 were investigated in a preliminary study, after which, the effects of PGE1 in a 90-min warm ischemic liver model were examined. The dogs were divided into three groups of ten, according to the treatment given: group A was an untreated control group, group B received PGE1 intravenously, and group C received PGE1 intraportally. The PGE1 was infused continuously at a rate of 0.02 microliters/kg/min before and after ischemia. All the dogs in groups A and B died within 24 h of induced ischemia. Whereas, six of the ten dogs in group C survived for over 3 days. The arterial ketone body ratio was not maintained in groups A and B, but it was in group C. Furthermore, in group C the serum lipid peroxide level, which reflects hepatocellular membrane damage, was maintained at a lower level than that in the other groups after ischemia. Electron microscopy revealed sinusoid destruction and changes in both the plasma membrane and parenchymal cell mitochondria in groups A and B, while in group C these structures were well preserved. These findings confirmed that intraportally administered PGE1 improved the hepatic microcirculation and stabilized the hepatocellular membranes. Our results indicate that intraportal administration of PGE1 has a greater protective effect than intravenous administration against warm ischemic liver injury.

Alprostadil↗

PubMed1995

Establishment and characterization of a new cell line from human bladder cancer (JMSU1).

A new human bladder cancer cell line designated JMSU1 has been established from malignant ascitic fluid of a 75-year-old Japanese man with bladder cancer, and maintained in culture for more than 7 years and over 240 passages. Inverted phase-contrast microscopy revealed that JMSU1 was composed of morphologically distinct cells (polygonal to spindle-shaped cells), showing morphological heterogeneity in vitro. Histological examination of xenografts showed poorly differentiated transitional cell carcinoma, resembling the original tumor. Immunohistochemical staining for cytokeratin and electron microscopic examination suggested that JMSU1 was of epithelial origin. Chromosome analysis gave a modal number of 69 with no Y chromosome. Isozyme analysis (LDH, G6PD, and NP) showed the mobility pattern of human type B. DNA fingerprint analysis demonstrated that there was no cross-culture contamination of JMSU1 during the passages. In conclusion, a newly established and well-characterized cell line, JMSU1, offers promising material for the investigation of the biological properties of bladder cancer.

Aged↗

PubMed1995

Rheological significance of tandem lesions of the coronary artery.

It is not known whether the individual lesions that constitute tandem lesions of the coronary artery are developmentally or rheologically related. Luminal changes and their rheological significance were examined by percutaneous angioscopy in 44 tandem lesions of 21 patients with ischemic heart disease. Angioscopically, individual narrowing of angiographically documented tandem lesions appeared only as a tangentially expressed prominent portion of an atherosclerotic spiral fold. The directions of the fold were counterclockwise in the proximal to middle segments and clockwise in the distal segment of the right coronary artery, clockwise in the proximal to middle segments and counterclockwise in the distal segment of the left anterior descending artery, and counterclockwise in the proximal to middle segments of the left circumflex artery. The bloodstream always ran along the spiral folds in the tandem lesions. The results suggest that angiographically documented tandem coronary lesions are merely a tangential expression of atherosclerotic spiral folds and that they may act to prevent blood turbulence by generating a spiral laminal flow.

Adult↗

PubMed1995

Repeated ketamine administration produces up-regulation of muscarinic acetylcholine receptors in the forebrain, and reduces behavioral sensitivity to scopolamine in mice.

To study the effects of repeated ketamine administration on central muscarinic acetylcholine receptors (mAchRs), ddY male mice were administered subcutaneous doses of 25 mg/kg ketamine every 3 days for a total of five times. Receptor binding assays of mAchR were carried out in the forebrain (FB), cerebellum (CB) and brainstem (BS), using [3H]quinuclidinyl benzilate ([3H]QNB) as a ligand. In addition, we examined whether repeated ketamine (12.5, 25 and 50 mg/kg) or saline (five times) could modify the hyperlocomotion induced by scopolamine (0.5 mg/kg, SC) (a muscarinic antagonist), using a behavior-pharmacological technique. Repeating the ketamine administration resulted in a significant increase in the receptor density value (Bmax) for [3H]QNB only in FB, dependent on the numbers of administrations (1270 +/- 33 fmol/mg protein for a single dose, 1620 +/- 59 for four treatments, 1738 +/- 70 for five treatments without any change in apparent affinity (defined as the reciprocal of the dissociation constant) (Kd). A competitive inhibition study of repeated (5 times) administration of ketamine failed to detect any subtype-specific changes in mAchRs. Repeated ketamine administration reduced the scopolamine-induced hyperlocomotion in a dose-related way, and the changes were significant at 50 mg/kg. Our results suggest that repeated ketamine administration produces an up-regulation of mAchRs, and this change may be associated with altered Ach transmission in the central nervous system.

Animals↗

PubMed1995

Angioscopic detection of residual pulmonary thrombi in the differential diagnosis of pulmonary embolism.

Definite diagnosis of pulmonary embolism (PE) by conventional methods such as angiography is frequently difficult. If residual thromboemboli incorporated into the pulmonary arterial wall or in the distal small segments are visible, differential diagnosis of PE versus primary pulmonary hypertension (PPH) can be made without open-chest pulmonary biopsy. Six patients suspected of having acute PE, 6 suspected of having chronic PE, and 4 with PPH diagnosed by pulmonary biopsy underwent percutaneous pulmonary angioscopy. In patients suspected of having PE, globular and mural thromboemboli were detected by both angioscopy and angiography in 4 and 1 patients, respectively. By angioscopy, emboli incorporated into the arterial wall were detected in 7 and microemboli obstructing the distal small segments were detected in 6. However, these emboli were detected by angiography in none. In patients with PPH, no embolus was detected by angioscopy and angiography. Angioscopically, however, stenoses were observed in the distal small segments in all patients. The results indicate that residual pulmonary thromboemboli in PE and stenoses of distal pulmonary arteries in PPH are detectable by percutaneous angioscopy, and therefore this method is feasible for differential diagnosis of PE.

Acute Disease↗

PubMed1995

Angiogenic therapy of acute myocardial infarction by intrapericardial injection of basic fibroblast growth factor and heparin sulfate: an experimental study.

To examine whether angiogenesis and myocardial salvage occur, 30 micrograms basic fibroblast growth factor (bFGF) and 3 mg heparin sulfate (HS) were injected through the right atrium into the pericardial cavity by a thin needle-tipped catheter in a canine model of acute myocardial infarction. One month later infarcted weight/left ventricle weight was 24% +/- 5.2%, 25% +/- 4.0%, 18% +/- 2.4%, and 10% +/- 1.8% (mean + SE) in saline solution, HS, bFGF alone, and bFGF plus HS groups, respectively. Vascular number in the infarcted area of the outer layer was 13 +/- 3.3, 20 +/- 2.2, 47 +/- 8.3, and 136 +/- 26.3/200 x 200 microns2 in saline solution, HS, bFGF alone, and bFGF plus HS groups, respectively. Thus the vascular number was the largest in the bFGF plus HS group. The vascular number was larger in the subepicardial than in the subendocardial infarcted areas. Vessels directed from the epicardium toward the subepicardial infarcted area were also observed. The transcatheter intrapericardial injection of bFGF plus HS caused angiogenesis and myocardial salvage. This method might bring about a selective therapeutic and preventive modality of myocardial infarction irrespective of coronary anatomy and contraindications for coronary interventions and surgery.

Animals↗

PubMed1995

Observation of atherosclerotic lesions by an intravascular microscope in patients with arteriosclerosis obliterans.

The magnification power of conventional fiberscopes for intravascular use is up to x30, and therefore they are not feasible for observation of fine structures of vascular changes. We devised a microscope that enables percutaneous transluminal observation of the vascular luminal changes of cellular order. Thus we examined its feasibility in patients. The microscope was 8F in diameter with a rod lens inside, and its magnification power was from x150 to x350. The microscope was introduced into the iliac or femoral artery for observation of vascular lesions during angioplasty in seven patients with arteriosclerosis obliterans. With the aid of vital staining individually damaged endothelial cells, fibrin threads, foam cells, collagen, elastic fibers, and structure of microthrombi were discriminated clearly. The results indicate that structures of human vascular lesions of cellular order can be observed percutaneously by the intravascular microscope.

Aged↗

PubMed1995

Prediction of acute coronary syndromes by percutaneous coronary angioscopy in patients with stable angina.

To pinpoint the link between plaque characteristics and acute coronary syndromes, we performed a 12-month prospective follow-up study in 157 patients with stable angina pectoris in whom regular coronary plaques were observed by percutaneous coronary angioscopy. Acute coronary syndromes occurred more frequently in patients with yellow plaque than in those with white plaques (11 of 39 vs 4 of 118; p = 0.00021). Moreover, the syndromes occurred more frequently in patients with glistening yellow plaques than in those with nonglistening yellow plaques (9 of 13 vs 2 of 26; p = 0.00026). Thrombus arising from the ruptured identical plaques was confirmed by angioscopy as the culprit lesion of the syndromes. The results indicate that acute coronary syndromes occur frequently and in a short time in patients with glistening yellow plaques and that angioscopy but not angiography is feasible for prediction of the syndromes.

Acute Disease↗

PubMed1995

Low pH leads to sister-chromatid exchanges and chromosomal aberrations, and its clastogenicity is S-dependent.

The effect of low pH on sister-chromatid exchanges (SCE), chromosomal aberrations (CA), and the cell cycle were investigated in Chinese hamster cells. The cells were treated in media over the pH range 7.2-5.4 during 24-h continuous or 3-h pulse treatments. In Chinese hamster ovary K1 cells, slight increases in SCE frequency were induced by 3-h pulse treatment with a 28-h recovery time. In Chinese hamster V79 379A cells, similar slight increases in SCE frequency were observed with both treatments. A severe delay in the cell cycle was noted in both cell types. DNA analysis with flow cytometry indicated that the cell cycle delay occurred in S phase. CA were observed in the first metaphase. Multiple fixation times over a 27-h period were used to determine whether or not CA could be induced in cells exposed to low pH medium in more than one part of the cell cycle. Only a few chromatid gaps were induced when the cells were fixed at 0-9 h after the 3-h treatment, most probably representing cells that were treated in their G2 or late S phase. CA were induced in cells fixed between 12 and 27 h after the 3-h treatment. These cells were most probably treated in early S phase, in G1, or in the previous G2/M. These results suggest that low pH clastogenicity is S-dependent.

Animals↗