Biomedical subjects
N Yasui
Publications and source records attributed to N Yasui.
Immunohistochemical expression of Bcl-2, Bax and cytochrome c following focal cerebral ischemia and effect of hypothermia in rat.
Recent studies suggest that mild hypothermia significantly alleviate damage following cerebral ischemia though the precise mechanism is poorly defined. In the present study, middle cerebral artery occlusion (MCAo) was induced in Sprague-Dawley (SD) rats for 1 h followed by varying periods of reperfusion. Cerebral infarcts identified by hematoxylin & eosin (H&E) staining revealed extensive lesion in normothermic (NT) 37 degrees C and small lesion in hypothermic (HT) 33 degrees C group of rats. Immunohistochemical analysis revealed Bcl-2 was induced in many neurons of HT group, while Bax and cytochrome c was induced in few neurons. In situ detection of DNA fragmentation using 3'-OH end labeling method (terminal dUTP nick-end labelling (TUNEL)) indicated, higher number of TUNEL-positive cells in NT group, but significantly decreased in HT group. The expression pattern revealed many neurons at the penumbra region could survive in HT group whereas, many neurons are committed to die in NT group. Our results suggest that hypothermia is selectively interfering at more than one place and providing protection.
A RUNX2/PEBP2alpha A/CBFA1 mutation displaying impaired transactivation and Smad interaction in cleidocranial dysplasia.
Cleidocranial dysplasia (CCD), an autosomal-dominant human bone disease, is thought to be caused by heterozygous mutations in runt-related gene 2 (RUNX2)/polyomavirus enhancer binding protein 2alphaA (PEBP2alphaA)/core-binding factor A1 (CBFA1). To understand the mechanism underlying the pathogenesis of CCD, we studied a novel mutant of RUNX2, CCDalphaA376, originally identified in a CCD patient. The nonsense mutation, which resulted in a truncated RUNX2 protein, severely impaired RUNX2 transactivation activity. We show that signal transducers of transforming growth factor beta superfamily receptors, Smads, interact with RUNX2 in vivo and in vitro and enhance the transactivation ability of this factor. The truncated RUNX2 protein failed to interact with and respond to Smads and was unable to induce the osteoblast-like phenotype in C2C12 myoblasts on stimulation by bone morphogenetic protein. Therefore, the pathogenesis of CCD may be related to the impaired Smad signaling of transforming growth factor beta/bone morphogenetic protein pathways that target the activity of RUNX2 during bone formation.
PEBP2alphaA/CBFA1 mutations in Japanese cleidocranial dysplasia patients.
Cleidocranial dysplasia (CCD) is an autosomal dominant human bone disease whose genetic locus has been located on chromosome 6p21, where the PEBP2alphaA/CBFA1 gene essential for osteogenesis also maps. Previously, several heterozygous mutations in PEBP2alphaA/CBFA1 were found in CCD patients. In this study, we identified six different types of mutations in PEBP2alphaA/CBFA1 in Japanese CCD patients. Four cases were similar to those reported previously: two were nonsense mutations in the Runt domain, one was a hemizygous deletion, and the other was a missense mutation in the Runt domain which abolished the DNA-binding activity of Runx2/PEBP2alphaA/CBFA1. The remaining two mutations were novel: one had a heterozygous gt-to-tt mutation at the splice donor site (gt) between the exon3-intron junction, which resulted in abnormal exon3 skipping, and the other had a mutation in exon7, which led to the introduction of a translational stop codon in the middle of the transactivation domain. Thus, defects in either the DNA-binding domain or transactivation domain of Runx2/PEBP2alphaA/CBFA1 can cause CCD. The results not only provide a strong genetic evidence that mutations involving in PEBP2alphaA/CBFA1 contribute to CCD, but also provide a useful tool to study how Runx2/PEBP2alphaA/CBFA1 plays its pivotal role during osteoblastic differentiation.
[Humeral lengthening and deformity correction].
Nine patients with achondroplasia and one patient with Apert syndrome underwent the surgical lengthening of both humerus and simultaneous correction of both associated bone deformity. An unilateral external fixator was applied to the lateral aspect of the humerus with four half-pins and percutaneous predrilling osteotomy was performed at the apex of flexion deformity of the bone. During the waiting period before distraction, the flexion deformity of the distal humerus was corrected using an additional external fixator. Slow gradual distraction was subsequently carried out at a rate of 0. 25 mm every 6 hours. The average lengthening was 8 cm (range 7.5 to 9 cm), the overall treatment time 312 days (range 192 to 406 days), and the average healing index 39.0 days/cm. The average correction of the elbow flexion deformity was 20 degrees. We believe this treatment is useful to improve the function of the arms and the activity of daily living for the patients with bilateral short humeri.
Effects of repeated ingestion of grapefruit juice on the single and multiple oral-dose pharmacokinetics and pharmacodynamics of alprazolam.
The effects of repeated ingestion of grapefruit juice, an inhibitor of cytochrome P450 3A4 (CYP3A4), on the pharmacokinetics and pharmacodynamics of both single and multiple oral doses of alprazolam, a substrate of CYP3A4, were examined. In study 1, eight healthy volunteers ingesting 600 ml/day water or grapefruit juice for 10 days took a single oral 0.8-mg dose of alprazolam on the eighth day. Plasma drug concentrations were monitored up to 48 h after alprazolam dosing together with evaluation of psychomotor function. Grapefruit juice altered neither the plasma concentrations of alprazolam at any time points, any pharmacokinetic parameters, nor the majority of psychomotor function parameters in subjects. In study 2, 11 patients with anxiety disorders receiving alprazolam (0.8-2.4 mg/day) ingested grapefruit juice (600 ml/day) for 7 days. Blood samples were collected before and during grapefruit juice ingestion and 1 week after its discontinuation together with an assessment of clinical status. Grapefruit juice altered neither the steady-state plasma concentration of alprazolam nor the clinical status in patients. The present study shows that grapefruit juice is unlikely to affect pharmacokinetics or pharmacodynamics of alprazolam due to its high bioavailability.
Prolactin response to nemonapride, a selective antagonist for D2 like dopamine receptors, in schizophrenic patients in relation to Taq1A polymorphism of DRD2 gene.
The dopamine D2 receptor (DRD2) gene has a Taq1A restriction fragment length polymorphism yielding two alleles, A1 and A2. It has been shown that the subjects with less frequent allele, the A1 allele, have lower density and diminished function of DRD2 in the striatum, compared to those with no A1 allele. In the present study, the relationship between this polymorphism and prolactin response to nemonapride, an antipsychotic drug with selective and potent DRD2 antagonistic property, was investigated in 25 Japanese schizophrenic inpatients (13 males, 12 females). The daily dose of nemonapride was fixed at 18 mg, and the duration of treatment was 3 weeks. Taq1A genotypes were determined by the polymerase chain reaction method. Plasma prolactin concentrations were measured by enzyme immunoassay. The subjects were divided into four subgroups by gender and Taq1A genotypes, i.e., six males and eight females with the A1 allele, seven males and four females with no A1 allele. The (delta)prolactin (change from the pretreatment concentration) at 1 week was significantly (P<0.05) higher in females with the A1 allele (78.0 +/- 47.1 ng/ml) than in males with the A1 allele (33.4 +/- 14.0 ng/ml) or with no A1 allele (29.5+/-24.8 ng/ml). In addition, (delta)prolactin at 3 weeks was significantly (P<0.05) higher in females with the A1 allele (98.1+/-67.9 ng/ml) than in females with no A1 allele (33.4+/-24.6 ng/ml), males with the A1 allele (29.1+/-17.3 ng/ml) or males with no A1 allele (28.6+/-22.0 ng/ml). The present study thus suggests that female patients with the A1 allele show a greater prolactin response to nemonapride, who may have a high risk for adverse effects associated with neuroleptic-induced hyperprolactinemia.
Fluvoxamine but not citalopram increases serum melatonin in healthy subjects-- an indication that cytochrome P450 CYP1A2 and CYP2C19 hydroxylate melatonin.
OBJECTIVE: The nocturnal serum melatonin (MT) level increases after ingestion of fluvoxamine (FLU)-- a selective serotonin re-uptake inhibitor (SSRI) with antidepressive properties. The mechanism behind the MT increase is unknown. Citalopram (CIT) is another SSRI. It is not known whether CIT affects the serum MT level. It may well be that these two compounds affect serum MT levels differently, inasmuch as the ways they inhibit cytochrome P450 (CYP) enzymes in the liver differ markedly. FLU inhibits CYP1A2 potently, and to some extent also CYP2C19, whereas CIT is without such an effect. CYP enzymes are probably involved in the hepatic metabolism of MT. If FLU, but not CIT, inhibits liver enzymes involved in the metabolism of MT, different serum MT concentrations should probably ensue. The objective of this investigation was to test this hypothesis. METHODS: Seven healthy subjects participated in three different experiments, which were performed in random order 6-8 days apart. In experiment A, placebo was given, in experiment B 40 mg CIT and in experiment C 50 mg FLU. All doses were given orally at 1600 hours. Serum MT concentrations were determined at regular intervals between 1600 hours and noon next day (20 h). Plasma concentrations of CIT were measured repeatedly in experiment B, and plasma FLU concentrations in experiment C. MT areas under the curve representing the 20-h period (MT-AUC(0-20)) were compared in the three experiments, and differences were statistically evaluated. RESULTS: FLU augmented the MT-AUC(0-20) by a factor of 2.8 compared with the effect of placebo (P < 0.01), whereas CIT was without significant effect. More MT was excreted in the urine after ingestion of FLU than after placebo. In contrast, CIT did not influence the MT excretion. A clear relationship was found between serum levels of MT and plasma concentrations of FLU. CONCLUSION: The serum MT level increased markedly after ingestion of FLU but not after CIT. The exact mechanism behind this finding is unknown, but decreased hepatic metabolism of MT by either CYP1A2 or CYP2C19, or both, is probable. Although exogenous MT, causing high MT concentration in plasma, has sleep-promoting properties in man, it is at this stage unknown whether serum MT concentrations in the range found in this study have similar effects. This has to be given further attention in additional studies.
Congenital transverse deficiency of the tibia and fibula: a report of two cases.
We report two similar, but unrelated, patients with congenital bilateral partial deficiencies of the tibia and fibula associated with intact feet. In both patients, the tibia and fibula were absent on initial radiographs, while the femur and the tarsal bones were well developed and there was bilateral teratologic dislocation of the hips. Ultrasound and magnetic resonance imaging (MRI) studies suggested the presence of cartilaginous remnants of the tibia and fibula. There were multidirectional instabilities in the knees and ankles. The clinical and radiological features of these cases are distinct from those of congenital longitudinal deficiency of the tibia, in which the fibula is always preserved, and from longitudinal deficiency of the fibula, in which the tibia is present and the foot is usually involved. We suggest that the bilateral partial deficiencies of the tibia and fibula associated with the intact foot and teratologic dislocation of the hips is a single-entity disorder, possibly categorized as an intercalary transverse deficiency of the lower limb.
Cerebral blood flow and oxygen metabolism during mild hypothermia in patients with subarachnoid haemorrhage.
Cerebral blood flow and O2 metabolism during hypothermia (33-34 degrees C) was evaluated in 5 patients with aneurysmal subarachnoid haemorrhage by positron emission tomography (PET). Their preoperative clinical condition was WFNS scale IV or V. The patients received surface cooling postoperatively, and were maintained in a hypothermic state during transfer for radiological examination. Positron emission tomography revealed a decrease in cerebral blood flow and O2 metabolic rate. Cerebral blood flow was 34.8+/-15.1 ml/100 ml/min and the O2 metabolic rate was 1.85+/-0.61 ml/100 ml/min in areas of the middle cerebral artery ipsilateral to the ruptured aneurysms, whereas these values were 30.8+/-7.1 and 2.21+/-0.45 ml/100 ml/min, respectively, on the contralateral side. This represents a decrease of 37+/-27% compared to normal cerebral blood flow and 52+/-16% compared to normal O2 metabolic rate (p < 0.02) in the ipsilateral areas, and decreases of 44+/-13% and 43+/-12%, respectively, on the contralateral side. The present results reflected the luxury perfusion state in almost all cases and provide the first PET evidence of decreased cerebral blood flow and metabolic rate of O2 during hypothermia in humans.
A technique of percutaneous multidrilling osteotomy for limb lengthening and deformity correction.
We have recently developed a technique of per-cutaneous multidrilling osteotomy for limb lengthening and deformity correction. The bone is drilled percutaneously, using a special drill guide, and osteotomy is accomplished by connecting the multiple drill holes with a small chisel. The bone segments are subjected to slow progressive distraction with an external fixation device. We have lengthened 33 limbs in 22 patients with congenital or post-traumatic limb shortening and/or bone deformities. All the patients underwent the proposed lengthening and/or correction of the bone deformities through a single-treatment procedure. None of the lengthened segments resulted in nonunion. This technique can prevent undesirable bone cracks and preserve soft tissue around the osteotomy site, and is also applicable to other fields of orthopedic surgery.
Multicentric synchronous osteosarcoma: a case report with autopsy findings.
We report a typical case of multicentric synchronous osteosarcoma. An 8-year-old girl was referred to us. At diagnosis she had a dominant bone lesion in the left proximal tibia and other lesions in the pelvis, thoracic spine, right femoral shaft, and left lower limb, without any lung lesions. We administered high doses of cisplatin and doxorubicin hydrochloride, but they were not effective. A pulmonary metastatic lesion was first detected by chest X-ray 4 months after the diagnosis, and she died of pulmonary insufficiency within 1 year after the first presentation.
A novel type X collagen gene mutation (G595R) associated with Schmid-type metaphyseal chondrodysplasia.
Metaphyseal chondrodysplasia of the Schmid type (MCDS) is a skeletal dysplasia affecting the long bone metaphyses; it is characterized by short stature, bowlegs, and coxa vara. The spine is generally not involved. Here we report a novel missense mutation of the type X collagen gene in a sporadic case of MCDS. The mutation was a heterozygous single base-pair transition of G-to-A at nucleotide 1783, which predicted a substitution of glycine by arginine at codon 595 (G595R) in the carboxyl-terminal noncollagenous domain. Interestingly, another mutation of the same codon, in which glycine is substituted by glutamic acid (G595E), was previously reported to cause spondylometaphyseal dysplasia (SMD), another group of skeletal dysplasias with involvement of the spine in addition to the long tubular bones. The novel G595R mutation identified in the present study supports the concept of type X collagenopathy.
Mild hypothermia mitigates post-ischemic neuronal death following focal cerebral ischemia in rat brain: immunohistochemical study of Fas, caspase-3 and TUNEL.
Mild hypothermia is considered to have a protective effect during ischemic neuronal cell death. The present study provides experimental evidence for this beneficial role of mild hypothermia using reversible middle cerebral artery occlusion (MCAo) in a Sprague-Dawley (SD) rat model. MCAo was induced in rats for 1 h followed by reperfusion at different periods. Hematoxylin-eosin (HE) staining in normothermic (NT) 37 degrees C and hypothermic (HT) 33 degrees C groups of rats confirmed cerebral infarcts. The mean per cent infarct area was significantly reduced in the HT group of rats. Immunohistochemical analysis was done using anti-Fas and caspase-3 antibodies. The immunohistochemical expression of Fas and caspase-3 was demonstrable as early as 5 h after reperfusion, but the expression pattern maximized at 24 h after reperfusion. The expression of Fas and caspase-3 proteins showed a clear decrease in the HT group over the NT group. In situ detection of DNA fragmentation was done using the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling method (TUNEL). TUNEL-positive cells were first observed at 5h after reperfusion and progressively increased by 24h. A higher number of TUNEL-positive cells was found in the NT group, but they were significantly decreased in the HT group. Further, DNA fragmentation was confirmed by size fractionation in agarose gel. These findings demonstrate a positive relation between the expression of Fas, caspase-3 and TUNEL-positive cells.
Structure and photodimerizations of 1-alkylthymines in single crystals.
1-Alkylthymine gave four kinds of crystals depending on the solvents used. Photodimerizations of 1-alkylthymine in single crystals were found to depend on the crystal structure. In inactive crystals, the terminal methyl group of alkyl chains approached to the double bond of thymine, and inhibited rotation of thymine bases during photodimerization.
Therapeutic spectrum of nemonapride and its relationship with plasma concentrations of the drug and prolactin.
The therapeutic spectrum of nemonapride, a new substituted benzamide, and its relationship with plasma concentrations of the drug and prolactin were investigated by a fixed-dose study (18 mg/day for 3 weeks) in 31 patients with acutely exacerbated schizophrenia. Of 31 patients, 25 (80.6%) were responders who showed a reduction in symptoms (percentage of improvement) of 50% or more after 3 weeks. The mean values of percentage of improvement in scores on the total Brief Psychiatric Rating Scale (BPRS) and the five subscale symptoms were 71.5% for total, 73.2% for Positive, 86.0% for Excitement, 53.9% for Negative, 84.2% for Cognitive, and 67.5% for Anxiety-Depression. Responders had higher percentage of improvement in positive (84.6 +/- 17.0% vs. 25.9 +/- 15.7%; p < 0.001) and anxiety-depression (76.9 +/- 18.8% vs. 28.5 +/- 39.9%; p < 0.005) symptoms than did nonresponders after 3 weeks. The percentage of improvement in total BPRS after 2 weeks was well correlated with that after 3 weeks (Spearman rank correlation coefficient: r(s) = 0.711; p < 0.01). There was an inverted U-shaped relationship between plasma drug concentrations (nemonapride plus desmethylnemonapride) and percentage of improvement in total BPRS symptoms after a 3-week treatment (y = 46.9 + 73.9x - 44.2x2; p < 0.001). These findings suggest that nemonapride has a broad therapeutic spectrum in the treatment of acute schizophrenia. The improvements in scores for the Positive and Anxiety-Depression subscale symptoms are regarded as determinant factors for total response to nemonapride. An assessment of clinical status after 2 weeks and plasma drug monitoring may be useful for the prediction of the final outcome for patients.
Effect of a genetic polymorphism of CYP1A2 inducibility on the steady state plasma concentrations of haloperidol and reduced haloperidol in Japanese patients with schizophrenia.
The effect of a genetic polymorphism of inducibility of cytochrome P450 (CYP) 1A2 on the steady state plasma concentrations (Css) of haloperidol and reduced haloperidol was studied to clarify if these Css are dependent on the CYP1A2 activity. The subjects were 101 Japanese schizophrenic inpatients receiving oral haloperidol 12 mg/d. The Css of haloperidol and reduced haloperidol were measured in duplicate by high performance liquid chromatographic method, and were corrected to the mean body weight. A point mutation from guanine (wild-type) to adenine (mutated-type) at position -2964 in the 5'-flanking region of CYP1A2 gene was identified by polymerase chain reaction (PCR)-fragment length polymorphism method. Based on the present results, i.e., significant effects of CYP2D6 genotypes on the Css of haloperidol and reduced haloperidol, analyses were separately performed in two groups, i.e., patients with 0 mutated allele of the CYP2D6 (41 cases) and those with 1 or 2 mutated alleles (60 cases). Subjects in each CYP2D6 genotype group consisted of 4 subgroups according to smoking habit and the presence of the mutated allele of the CYP1A2. Neither the Css of haloperidol nor that of reduced haloperidol significantly differed among the 4 subgroups in either CYP2D6 genotype group. The present study thus suggests that the CYP1A2 activity does not play an important role in controlling the Css of haloperidol or reduced haloperidol.
Novel mutations of the cathepsin K gene in patients with pycnodysostosis and their characterization.
Pycnodysostosis is a rare autosomal recessive skeletal dysplasia characterized by short stature, osteosclerosis, acroosteolysis, bone fragility, and skull deformities. Recently, mutations in the gene encoding cathepsin K (CK), a lysosomal cysteine protease localized exclusively in osteoclasts, were found to be responsible for this disease. We analyzed genomic DNA from four unrelated Japanese patients with this disorder and identified three different mutations of their CK genes: a previously reported missense mutation (A277 V), a novel single base deletion mutation (531 del T) causing a frame shift from codon 142 that results in a premature termination codon, and a novel missense mutation (L9P) in the signal peptide region. To investigate whether the L9P mutation disrupts signal peptide function and decreases protein synthesis, mutant and wild-type CK complementary DNAs driven by the cytomegalovirus promoter were transfected into COS-7 cells, and their gene products were detected by immunohistochemistry and Western blotting. Expression of the mutant protein was markedly reduced, suggesting decreased mature CK production in this patient, which may have been due to dysfunction of the signal peptide. These results provide evidence that a structural change in the signal peptide of the CK protein was involved in the pathogenesis of pycnodysostosis.