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Biomedical subjects

T Ohshima

Publications and source records attributed to T Ohshima.

At least 163 records · Page 9Linked to original sources
PubMed1997

Species identification based on the point mutations of histo-blood group ABO genes by PCR-RFLP and direct sequencing.

Using the PCR-restriction fragment length polymorphism (RFLP) and direct sequencing methods, 181 bp DNA fragments at the ABO blood group locus of human and eight different primates were examined. Through PCR amplification and digestion of the product with Hha-1 restriction enzyme, each of the amplified fragments of human, chimpanzee and green monkey was cut into two fragments of 147 and 34 bp, and the corresponding fragment of the other primates was digested into three fragment of 115, 34 and 32 bp. The 181 bp fragments of chimpanzee and green monkey were digested with Mva-1 into three fragments of 82, 58 and 41 bp, and 69, 58 and 54 bp, while the fragments of human and the other primates were separated into two fragments of 123 and 58 bp. Thus, using Hha-1 and Mva-1, human PCR-amplified product of a part of the ABO gene could be discriminated from those of the other primates examined. In addition, by direct sequencing of the 181 bp DNA fragment, two Mva-1 recognition sites and one Hha-1 recognition site were found in the fragments of chimpanzee and green monkey, one Mva-1 site and one Hha-1 site were detected in humans, and one Mva-1 site and two Hha-1 sites in the other primates. These results corresponded well with the data of PCR-RFLP. This method has a good potential for species identification at the DNA level.

ABO Blood-Group System↗
PubMed1997

Eight cases of suicide by self-cutting or -stabbing: consideration from medico-legal viewpoints of differentiation between suicide and homicide.

The present article describes eight cases of suicide by self-cutting or -stabbing. Seven of the deceased had a history of psychiatric problems. In every case, sharp weapon injuries were observed at typical sites of self-inflicted injury. The self-inflicted injuries varied in severity from a single stab wound to multiple cut wounds. In four cases, hesitation marks were found. Among the remaining four cases without hesitation marks, only one had clothing injuries. Based on the scene investigation by police, it was deduced that each body had been found at or very near the place where the fatal injuries had been sustained. In two cases, toxicological analysis played an important role in discriminating suicide from homicide. In one case, a 52-year-old male stabbed himself fatally after potassium hydroxide ingestion, and in another, a 71-year-old male committed suicide by self-cutting and -stabbing after drinking an alkaline detergent solution. In five of the eight cases, unusual findings such as a single stab wound, multiple cut injuries, no hesitation mark or presence of clothing injuries were found. In these five cases, the psychiatric history of the deceased contributed to furthering our comprehension of these unusual findings. We believe that thorough toxicological analysis and scene investigation as well as autopsy are essential in discriminating between homicide and suicide. Furthermore, forensic pathologists and forensic physicians at the scene should obtain as much information as possible on the victim's previous character including previous psychiatric history in order to elucidate his or her manner of death.

Journal Article↗
PubMed1997

Construction of a new leucine dehydrogenase with preferred specificity for NADP+ by site-directed mutagenesis of the strictly NAD+-specific enzyme.

On the basis of sequence comparison between NAD+-dependent leucine dehydrogenase (LeuDH) from Thermoactinomyces intermedius and NADP+-dependent dehydrogenases, a set of amino acid residues that are supposed to determine the coenzyme specificity of LeuDH were assigned. Systematic replacement of these amino acids by others was done with the aim to switch its natural coenzyme specificity to a new one preferring NADP+. Single D203A, double D203A-I204R and triple D203A-I204R-D210R mutation enzymes were constructed. The wild-type LeuDH is inactive with NADP+. However, D203A single mutant exhibited dual specificity for NAD+ and NADP+ with essentially identical k(cat)/Km values for both coenzymes, but the values were three orders of magnitude lower than that of the wild-type enzyme. Introduction of positive charge at 204 together with the removal of the negative charge at 203 in the double mutant D203A-I204R provided the enzyme with significantly high affinity for NADP+. The best k(cat)/Km value for NADP+ was shown for the triple mutant D203A-I204R-D210R: more than 2% of the k(cat)/Km value of the wild-type enzyme. Thus, we succeeded in constructing a mutant LeuDH with a new coenzyme specificity preferring NADP+ which is highly active (specific activity, 19 micromol/mg/min).

Amino Acid Oxidoreductases↗
PubMed1997

Traumatic tear of the basilar artery associated with vertebral column injuries.

An unusual case of traumatic basal subarachnoid hemorrhage (SAH) due to a mechanical tear of the basilar artery is reported. A 70-year-old man who had been suffering from cerebrovascular dementia was found dead in a ditch. Externally, subcutaneous hemorrhage with abrasions was observed on the left side of the forehead, face, and lower jaw, together with small contusions in the left superciliary arch. Internally, a 3-mm-long transversal tear of the basilar artery was observed, and dislocations of both C6-C7 and T1-T2 as well as a small fracture of the processus spinosus of C5 were found. No pathological vascular lesions such as aneurysms and vasculitis, other than arteriosclerosis, were observed in the vertebral-basilar system. Ethanol was not detected in the intracardiac blood or in the urine. These findings indicate that when the man fell into the ditch, severe hyperextension occurred as a result of minor blunt forces to the face, and that the traumatic tear of the basilar artery was mechanically caused by overstretching due to hyperextension of the head. It is also suggested that due to his advanced age the muscle tone of the neck might have declined, impairing its defense action, and that head hyperextension might, therefore, occur rather more readily under such conditions.

Aged↗
PubMed1997

Effect of dibutyryl cyclic adenosine monophosphate on skeletal muscle reperfusion injury in the rat.

To clarify the action of dibutyryl cyclic adenosine monophosphate (DBcAMP) on reperfused ischemic muscle, experiments were conducted using the hindlimbs of 18 male Lewis rats. At the midportion of the thigh all tissues except for the femoral artery and vein were transected, and a route for continuous intravenous infusion was secured in a contralateral limb vein. After inducing total ischemia by clamping the femoral artery and vein with a vascular clamp for 4 h, the limb was reperfused for 1 h. Blood flow was then compared using the hydrogen gas clearance method in a group in which DBcAMP 10 mg was continuously infused from a vein in the contralateral hindlimb from 1 h prior to the induction of ischemia to 1 h after the completion of ischemia (DBcAMP group), a group in which saline was infused in the same manner (control group), and a group which was subjected to biopsy alone (biopsy group). The percent change in blood flow was significantly higher in the DBcAMP group than in the control group at 15 and 30 min after the release of the clamp. Adenosine triphosphate (ATP), phosphocreatine (PCr), and lipid peroxide (LPO) were measured in tissue samples obtained 1 h after reperfusion. Serum LPO was measured in blood samples collected at the same time. ATP values were higher in the DBcAMP group than in the control group. PCr was significantly higher in the DBcAMP group than in the control group. LPO levels in skeletal muscle tissue did not differ significantly between the DBcAMP and control groups. In contrast, serum LPO levels were significantly lower in the DBcAMP group than the control group. On morphologic analysis the control and DBcAMP groups showed normal vascular endothelial cells and absence of the 'no-reflow' phenomenon. These data confirm that in this reperfusion model the administration of DBcAMP enhances the viability of skeletal muscle cells. Moreover, mediated by an effect on vascular endothelial cells this agent is thought to be of help in mitigating the vascular endothelial cell injury occurring in acute ischemic injury. DBcAMP may be a useful agent in mitigating skeletal muscle ischemia-reperfusion injury.

Adenine Nucleotides↗
PubMed1997

Gene expression patterns of murine dentin matrix protein 1 (Dmp1) and dentin sialophosphoprotein (DSPP) suggest distinct developmental functions in vivo.

Although the precise mechanisms of the conversion of predentin to dentin are not well understood, several lines of evidence implicate the noncollagenous proteins (NCPs) as important regulators of dentin biomineralization. Here we compared the in vivo temporospatial expression patterns of two dentin NCP genes, dentin matrix protein 1 (Dmp1), and dentin sialophosphoprotein (DSPP) in developing molars. Reverse transcription-polymerase chain reaction was performed on embryonic day 13 to 1-day-old first molars using Dmp1- and DSPP-specific primer sets. Dmp1 transcripts appeared at the late bud stage, while DSPP mRNA was seen at the cap stage. Expression of both genes was sustained throughout odontogenesis. In situ hybridization analysis revealed interesting differences in the expression patterns of these genes. While Dmp1 and DSPP showed coexpression in young odontoblasts before the start of mineralization, the expression of these genes was notably distinct at later stages. Dmp1 expression decreased in secretory odontoblasts after the appearance of mineral, while high levels of DSPP were sustained in odontoblasts. In early secretory ameloblasts, DSPP expression was transient and down-regulated with the appearance ofdentin matrix. Interestingly, Dmp1 expression became evident in ameloblasts during the maturative phase of amelogenesis. In contrast to Dspp expression that was tooth-specific, Dmp1 was expressed by osteoblasts throughout ossification in the skeleton. Probes directed to the "DSP" and "DPP" regions of the DSPP gene showed identical patterns of mRNA expression. These data show that the developmental expression patterns of Dmp1 and DSPP are distinct, implying that these molecules serve different biological functions in vivo.

Animals↗
PubMed1997

[A case of chronic subdural hematoma associated with idiopathic thrombocytopenic purpura (ITP)].

We reported a case of ITP associated with chronic subdural hematoma. A 51-year-old female was admitted to our hospital with left frontal pain on November 18, 1995. CT and MRI examinations revealed a left chronic subdural hematoma. Peripheral platelet count showed thrombocytopenia, 5,000/mm3. She was treated preoperatively by administration of steroids, a large dose of immunoglobulin and transfusion of platelet. Five days after admission, platelet count had elevated to 199,000/mm3, and evacuation of the hematoma through a burr hole under local anesthesia was performed successfully. No troubles were observed during and after surgical procedure. Intracerebral hemorrhages associated with coagulopathy are rather common, but only 6 cases of ITP associated with chronic subdural hematoma have been reported. The characteristic clinical features and treatment of these 6 cases including our own case was noted. The cases of patients less than 40 years old were usually treated by emergency operation with transfusion of platelet because of severe neurological deficits. However, in the cases of patients over 40 years old treated by elected operation after administration of steroids, a large dose of immunoglobulin and transfusion of platelet did not give rise to severe neurological deficits. Therefore, we emphasize that the treatment against thrombocytopenia should be recommended taking the age of patients into consideration.

Chronic Disease↗
PubMed1997

[Perioperative management of a patient with severe anemia caused by rare Jr (a-) blood].

We experienced the anesthetic management of a patient with severe anemia as low as 1.8 g.dl-1 of hemoglobin and 6.5 per cent of hematocrit before surgery. His blood had been determined as type B, anti-Jra antibody positive and the proper blood was unavailable. We gave mainly colloid fluids and partially blood products of different blood groups to maintain the circulating volume with monitoring central venous pressure peri-operatively. The operation finished relatively uneventfully and the patient was transferred to the common ward from the intensive care unit on the fifth postoperative day. We conclude that maintaining the patient's circulating volume to avoid shock should be given priority in a case of severe anemia.

Aged↗
PubMed1997

[Monitoring of inhibitors of the polymerase chain reaction for the detection of hepatitis C virus using the positive internal control].

In the direct detection of pathogens by polymerase chain reaction (PCR) from clinical samples, false negative results due to the presence of inhibitor are problematic. In order to monitor such an inhibitor, we evaluated the detection of the positive internal control in PCR assay for HCV-RNA in serum samples positive for anti-HCV antibodies, of which 41 samples were positive for HCV-RNA by competitive RT-PCR assay. The positive internal control was coamplified with HCV-RNA and hybridized to the specific probe on magnetic beads and then hybrids were detected by colorimetric measurement using automatic PCR machine (COBAS AMPLICOR). Detection of the positive internal control was negatively correlated with viral copy number in sera assayed by competitive RT-PCR. Five of 52 samples (9.6%) with high HCV-RNA copy number (10(7) or 10(8) copies/ml) showed negative results for the internal control. The negative results for the internal control turned out to be positive when the sera were diluted and re-assayed, suggesting competitively inhibitory effects of high viral copy number on amplification of the internal control. Addition of heparin in the serum sample showed an inhibitory effect with a dose dependent manner on the detection of both HCV-RNA and the internal control, with a more effect on the lower copy number of HCV. On the other hand, addition of hemoglobin in the sample with concentration of up to 450mg/dl had no inhibitory effect on the detection of either HCV-RNA or the internal control. Coamplification and detection of the positive internal control was demonstrated to be useful to estimate effects of inhibitors, which may be present in clinical samples, in the detection of HCV-RNA by PCR.

False Negative Reactions↗
PubMed1997

An autopsy case of suicidal strangulation with four looped rubber bands.

A rare case of suicidal strangulation with four looped rubber bands is reported. A 57-year-old female was found dead in the vestibule of her house, with four looped rubber bands wrapped around her neck. Her face was markedly congestive, and lots of petechial hemorrhages were seen on the face and in the palpebral conjunctive. At the medico-legal autopsy, the ligature mark was 1.7 cm in maximum width at the anterior part of the neck and accompanied by such vital reactions as linear hemorrhage, but there were no other injuries to account for her death. Many petechial hemorrhages were also found in the visceral pleurae and epicardium. There was hemocoagulum in the cardiac blood, and the other organs were as well markedly congestive. Gas-chromatographic analysis revealed ethanol; 0.83 mg/ml in the heart blood and 1.37 mg/ml in the urine. No other drugs or poisons were found with Toxi-Lab and REMEDi-HS systems. In addition, police investigation denied the possibility of homicide and disclosed her living will stating that she had suffered from a chronic heart disease. This case was of suicidal strangulation with looped rubber bands.

Asphyxia↗
PubMed1997

False identification of urinary ephedrine as methamphetamine by gas chromatography/mass spectrometry with a DB-17 column.

A 17-year-old motorcyclist was killed in a traffic accident. The urine sample collected at autopsy was screened by the Toxi-Lab system; it showed a possible presence of either methamphetamine (MA) or ephedrine (Eph). Gas chromatography-mass spectrometry (GC-MS) was carried out for a trifluoroacetyl (TFA)-derivatized sample after extraction with an Extrelut column. A peak was detected at the same retention time as that of the authentic Eph by GC-MS with a DB-17 wide-bore column; the mass spectrum of the peak was, however, very similar to that of MA. The presence of Eph and absence of MA were proved by GC-MS using a DB-1 column. The Eph in the urine was further confirmed by high performance liquid chromatography (HPLC)-MS using an ODS column. By the REMEDi HS system, Eph and methylephedrine were determined in the urine, and their concentrations (hydrochloride salt) were 0.752 and 2.13 micrograms/ml, respectively. Ethanol was detected as well in the blood (1.55 mg/ml) and urine (2.01 mg/ml) by a pulse heating method. Nicotine, caffeine and dihydrocodeine were also qualitatively detected in the urine by Toxi-Lab, HPLC-MS and REMEDi HS systems. The present case warned that urinary Eph can be misidentified as MA by GC-MS with an intermediately polar column after TFA derivatization.

Accidents, Traffic↗
PubMed1996

In vivo cisplatin resistance of rat ascites hepatoma AH66.

The rat ascites hepatoma AH66 cell line had higher resistance to cisplatin than a variant AH66F cell line in in vivo experiments, though in vitro sensitivities to cisplatin of both cell lines were similar in a medium containing 5% fetal calf serum (FCS). When AH cells were cultured in the medium containing 5% ascites fluid (ASF), the sensitivity of AH66 cells to cisplatin was significantly lower than that in the medium containing 5% FCS, but the sensitivity of AH66F cells was almost same in the case of 5% FCS. The metallothionein (MT) contents in AH66 cells, but not in AH66F cells, increased with the incubation time in the medium containing ASF. MT in AH66 cells was also induced by treatment with zinc ion, but the induction in AH66F cells was very low. These results indicate that AH66 cells have a high ability to induce MT and thereby may acquire in vivo resistance to cisplatin.

Animals↗
PubMed1996

Targeted disruption of the cyclin-dependent kinase 5 gene results in abnormal corticogenesis, neuronal pathology and perinatal death.

Although cyclin-dependent kinase 5 (Cdk5) is closely related to other cyclin-dependent kinases, its kinase activity is detected only in the postmitotic neurons. Cdk5 expression and kinase activity are correlated with the extent of differentiation of neuronal cells in developing brain. Cdk5 purified from nervous tissue phosphorylates neuronal cytoskeletal proteins including neurofilament proteins and microtubule-associated protein tau in vitro. These findings indicate that Cdk5 may have unique functions in neuronal cells, especially in the regulation of phosphorylation of cytoskeletal molecules. We report here generation of Cdk5(-/-) mice through gene targeting and their phenotypic analysis. Cdk5(-/-) mice exhibit unique lesions in the central nervous system associated with perinatal mortality. The brains of Cdk5(-/-) mice lack cortical laminar structure and cerebellar foliation. In addition, the large neurons in the brain stem and in the spinal cord show chromatolytic changes with accumulation of neurofilament immunoreactivity. These findings indicate that Cdk5 is an important molecule for brain development and neuronal differentiation and also suggest that Cdk5 may play critical roles in neuronal cytoskeleton structure and organization.

Animals↗
PubMed1996

Unexpected sudden death of a 12-year-old male with congenital single coronary artery.

A rare autopsy case of a 12-year-old male with a single coronary artery is reported in which he suddenly and unexpectedly died after a brief period of physical and mental excitation. The single coronary artery originated from the left aortic sinus, then branched out the left circumflex artery and afterwards descended as the left anterior descending artery, while the right coronary artery originated almost at a right angle from the left anterior descending artery. The entire heart as well as the cardiac conduction system depended exclusively on the single coronary artery for oxygenated blood supply, and the unbalanced blood distribution on his exertion probably led to sudden cardiac death. Moreover, in this case, both the deceased's mother (at the age of 20 years) and grandmother on the mother's side died suddenly, thus suggesting a possibility of maternal transmission of this congenital anomaly in his family.

Autopsy↗
PubMed1996

Molecular cloning and chromosomal mapping of the mouse gene encoding cyclin-dependent kinase 5 regulatory subunit p35.

A neural-specific activating subunit, p35, of cyclin-dependent kinase 5 (Cdk5) was recently reported to differ from other mammalian cyclins, suggesting a new type of regulatory subunit for Cdk activity. The mouse gene encoding p35, Cdk5r, was isolated from a mouse 129/SvJ genomic library, and the genomic structure of Cdk5r was characterized. The most notable features of Cdk5r are the absence of introns in the amino acid coding region and the high homology of amino acid sequence among species. The 5'-flanking region of Cdk5r contained no canonical TATA or CAAT box but had several putative promoter elements, including Sp1, AP2, MRE, and NGFIA. The mouse Cdk5r transcript was detected only in the brain by Northern blot analysis. Mouse Cdk5r was mapped to a position on mouse chromosome 11.

Amino Acid Sequence↗