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Biomedical subjects

T Sakaki

Publications and source records attributed to T Sakaki.

At least 163 records · Page 9Linked to original sources
PubMed1996

Intramedullary gadolinium-DTPA enhancement in a patient with cervical spondylotic myelopathy and an associated vascular lesion. Case report.

Intramedullary enhancement of the cervical spinal cord is rare in chronic compression disease. An accompanying vascular lesion should be considered in such a case. A 59-year-old man presented with severe cervical spondylotic myelopathy. Gadolinium-diethylenetriamine pentaacetic acid (Gd-DTPA) enhanced magnetic resonance imaging showed intramedullary enhancement at the C5-6 level, which was the most severely compressed level. A right ascending cervical arteriogram demonstrated a spinal arteriovenous fistula (AVF) fed mainly by the C-6 radicular artery and draining from the posterior medullary vein. Surgery was performed to decompress the myelopathy and to obliterate the AVF. Postoperative MR imaging with Gd-DTPA enhancement showed immediate and complete disappearance of the previous enhancement. The intramedullary enhancement presumably resulted from the intraparenchymal hemodynamics due to the AVF.

Journal Article↗
PubMed1996

Molecular engineering study on electron transfer from NADPH-P450 reductase to rat mitochondrial P450c27 in yeast microsomes.

We have reported the localization on yeast microsomes for a modified P450c27 (mic-P450c27) that contains the microsomal targeting signal of bovine P450c17 in front of the mature form of rat mitochondrial P450c27 (Sakaki, T., Akiyoshi-Shibata, M., Yabusaki, Y., and Ohkawa, H. (1992) J. Biol. Chem. 267, 16497-16502). In this study, we found that mic-P450c27 could be reduced by NADPH in the yeast microsomes without supplement of its physiological redox partners, adrenodoxin and NADPH-adrenodoxin reductase. In order to elucidate the direct electron transfer from NADPH-P450 reductase to mic-P450c27, we carried out simultaneous expression of mic-P450c27 and yeast P450 reductase. The reduction rate of mic-P450c27 was increased by overproduction of yeast P450 reductase, roughly in proportion to the reductase content in the microsomes. In addition, we constructed a fused enzyme between mic-P450c27 and yeast P450 reductase. The reduction rate of heme iron in the fused enzyme was too rapid to be measured. These recombinant yeast microsomes showed a notable 27-hydroxylation activity toward 5beta-cholestane-3alpha,7alpha, 12alpha-triol in the absence of adrenodoxin and adrenodoxin reductase. Finally, we purified mic-P450c27 from the recombinant yeast microsomes and reconstituted the hydroxylation system in liposomal membranes using the purified mic-P450c27 and yeast NADPH-P450 reductase. Mic-P450c27 was reduced by NADPH and showed its monooxygenase activity on the reconstituted system. Therefore, yeast NADPH-P450 reductase alone was found to transfer two electrons from NADPH to mic-P450c27. These results clearly show that mic-P450c27 not only localizes on the microsomes but also functions as a microsomal cytochrome P450 that accepts electrons from NADPH-P450 reductase.

Animals↗
PubMed1996

Cloning and expression in Escherichia coli and Saccharomyces cerevisiae of a novel tobacco cytochrome P-450-like cDNA.

A cDNA library constructed from poly(A)+ RNA of tobacco BY2 cells treated with 2,4-dichlorophenoxyacetic acid was screened by using a synthetic oligonucleotide corresponding to the heme binding region of avocado CYP71A1. A cloned 2-kb cDNA designated as cTBP contained an open reading frame of 1593 bp encoding a protein of molecular size of 58916. The deduced amino acid sequence included a cysteine residue corresponding to fifth ligand of heme-Fe at 497th. The coding sequence was expressed under the control of tac promoter and rrnB terminator in Escherichia coli to yield 7 to 10 nmol P450 equivalent per litre of the culture in the presence of delta-aminolevulinic acid. The modified coding sequences in which NH2-terminal residues 2-25 were replaced by the NH2-terminal 18 amino acid residues of microsomal bovine CYP17 were also expressed under the control of ADH promoter and terminator in Saccharomyces cerevisiae to yield 29 and 30 pmol of P450 equivalent/mg protein in the microsomal fraction, respectively. On co-expression of each of the modified coding sequences and yeast NADPH-cytochrome P-450 oxidoreductase gene, the yeast microsomes exhibited 7-ethoxycoumarin O-deethylase activity. Based on these results, tobacco cTBP was found to encode a novel P450-like species with a monooxygenese activity related to xenobiotic metabolism.

2,4-Dichlorophenoxyacetic Acid↗
PubMed1996

Alterations of retinoblastoma, p53, p16(CDKN2), and p15 genes in human astrocytomas.

BACKGROUND: Alterations of the suppressor genes, such as the retinoblastoma (RB), p53, p16(CDKN2), and p15 genes, have been reported in human gliomas. These genes have been suggested as the cell cycle regulatory genes at the G1-S checkpoint. METHODS: Alterations of the RB, p53, p16(CDKN2), and p15 genes in human astrocytomas were screened by single strand conformation polymorphism analysis of polymerase chain reaction products (PCR-SSCP analysis) and then confirmed by dideoxy sequencing. In addition, the expression of RB and p16 protein was examined by Western blot analysis. RESULTS: Aberrations of the RB gene were found in 3 of 23 surgical astrocytoma specimens (13%). Mutations were found at codon 754 in exon 22 (Val-->Gly), codon 519 in exon 17 (Thr-->Pro), and one base deletion at codon 903 resulting in stop codon at codon 905 in exon 26. These mutational locations were all near the regions associated with the functional domains of the RB gene. Aberrations of the p53 gene were found in 4 cases (17.4%). These mutations were found at codons 146 (Trp-->Gly) and 165 (Gln-->His) in exon 5, codon 73 (Val-->Glu) in exon 4, and codon 313 (Ser-->Asn) in exon 9. In addition, alterations of the p16(CDKN2) gene were found, with 5 cases (21.7%) having homozygous deletions, and 2 cases (8.7%) harboring point mutations. No p15 gene alteration was detected. The expression of p16 protein was undetectable in 10 cases (43.5%) by Western blot analysis, demonstrating an inverse correlation with the expression of RB protein. CONCLUSIONS: A few cases had overlapping alterations, and the incidence of one or more RB, p53, or p16(CDKN2) changes appeared to be relatively high in human astrocytomas. These results suggest that cell cycle regulatory gene alterations may play an important role in the development of gliomas.

Astrocytoma↗
PubMed1996

Amelioration of ischemic brain damage by the preischemic administration of propentofylline (HWA285) in a rat focal ischemia.

The protective effect of the propentofylline (HWA285) in a rat focal ischemia was investigated. The preischemic administration of HWA285 (10 mg/kg i.p.) significantly reduced the volume of ischemic damage both in the neocortex and the striatum. The postischemic administration of HWA285 failed to ameliorate ischemic damage. These findings suggest the therapeutic potential for ischemic core of the striatum. The neuroprotective mechanism may be related to its inhibition of both glutamate and dopamine release in the early period of focal ischemia.

Animals↗
PubMed1996

Multiple forms of human P450 expressed in Saccharomyces cerevisiae. Systematic characterization and comparison with those of the rat.

We systematically characterized the levels and substrate specificity of P450s from humans and rats to extrapolate drug metabolism data from experimental animals to humans. Human P450s (CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C18, 2D6, 2E1, and 3A4) were expressed in Saccharomyces cerevisiae and purified. Rat P450s were purified from hepatic microsomes of rats. We investigated the catalytic activities of purified P450s in a reconstituted system. Human CYP2B6 and rat CYP2B1 had high lidocaine N-deethylation activity. Human and rat CYP2D forms had high debrisoquine 4-hydroxylation activity. Human CYP3A4 and rat CYP3A2 had high testosterone 2 beta- and 6 beta-hydroxylation activities in a modified reconstituted system with a lipid mixture. The hydroxylation site of testosterone by CYP2B6 (16 alpha- and 16 beta-positions) agreed with that by rat CYP2B1. Human CYP2E1 had the highest lauric acid (omega-1)-hydroxylation activity and also had catalytic properties similar to those of rat CYP2E1. Human CYP2A and 2C forms had catalytic properties in testosterone metabolism different from those of rats. Antibodies raised against purified P450s were used to measure the levels of hepatic P450s. The level of CYP3A4 was the highest in human hepatic microsomes, comprising 30-40% of the total P450. CYP2C9 comprised 10-20% of the total. The levels of CYP1A2, 2A6, 2C8, 2D6, and 2E1 were moderate (5-15% of total P450). CYP2B6 content was very low. The information of this study is useful for drug metabolism and toxicological studies.

Animals↗
PubMed1996

Characteristics of intracranial aneurysms associated with moyamoya disease. A review of 111 cases.

A retrospective analysis of 111 patients with aneurysms associated with Moyamoya disease is presented. The subjects comprised of our 12 cases and 99 other well-documented cases. These 111 cases had 131 aneurysms. There were 48 males and 63 females. The average age was 40.3 years. The clinical manifestations were intracranial haemorrhage in 99 cases (89%), and ischaemic events in 9 cases (8%), but no mention was made of these in the last three cases (2%). The Hunt and Kosnik grades were grade 1 in 8%, grade 2 in 23%, grade 3 in 31%, grade 4 in 35%, and grade 5 in 3%. Of the 131 aneurysms, 73 (56%) were found distributed around the circle of Willis, 24 (18%) in the basal ganglia, 29 (22%) on collateral vessels, and 5 (4%) on other vessels. Forty-six percent of the cases were treated surgically, 51% conservatively, and 3% by endovascular procedures. The surgical procedures for the aneurysms were; neck clipping in 49%, aneurysmectomy in 18%, wrapping of the aneurysm in 11%, coating or cautery of the aneurysm in 7%, and revascularization only in 11%. The outcomes were Glasgow Outcome Scale 1 in 30%, 2 in 22%, 3 in 11%, 4 in 1%, and 5 in 25%. The main reasons for the unfavourable outcome were initial poor clinical grade and rebleeding. Follow-up angiography of 25 aneurysms demonstrated that all aneurysms in the basal ganglia or on the collateral vessels disappeared. We recommend surgical intervention for aneurysms associated with Moyamoya disease to prevent rupture or rebleeding, especially for aneurysms around the circle of Willis. However, direct surgery is not recommended for aneurysms found in the basal ganglia or on the collateral vessels.

Adolescent↗
PubMed1996

Rapid natriuresis and preventive hypervolaemia for symptomatic vasospasm after subarachnoid haemorrhage.

To prevent symptomatic cerebral vasospasm, we have used hypervolaemia (HV) or volume expansion in patients with aneurysmal subarachnoid haemorrhage (SAH) in recent years. In these patients we could not perform effective fluid and sodium (Na) replacement because of rapid and overwhelming water and Na loss. Although this phenomenon is characteristic under hypervolaemic states, we regard it important to elucidate the mechanism underlying initiation of vasospasm after aneurysmal SAH. Patients with aneurysmal SAH, operated on within 24 hours of onset, were analysed prospectively. We selected 17 patients in good pre-operative condition. Intravascular volume expansion was accomplished with plasma fractionate or albumin and crystalloid solutions in all patients. We divided the 17 patients into two groups; symptomatic spasm group (S-group) consisting of 4 cases developing transient ischaemic symptoms and non-symptomatic spasm group (NS-group) consisting of 13 cases. In S-group, rapid and marked natriuresis developed characteristically before the onset of ischaemic symptoms. The differences in daily Na balance between the two groups were significant on the 3rd and 5th days (p < 0.05). The mean cumulative Na balance in S-group during the 10 days of the study (-375 +/- 159 mEg) was higher than that of NS-group (-24.4 +/- 225 mEq) (p < 0.05). Rapid natriuresis preceded the development of ischaemic symptoms, and was important as a trigger for symptomatic vasospasm after SAH. We considered that hormonal disorders were implicated in this phenomenon, and atrial natriuretic peptide (ANP), antidiuretic hormone (ADH), renin, and aldosterone were each measured three times during the period, with no significant differences, found between the two groups. It was speculated that another potent natriuretic factor, similar to ANP, induced a rapid selective natriuresis resulting in symptomatic vasospasm.

Aged↗
PubMed1996

Expanding laminoplasty for cervical myelopathy-spinous process roofing technique.

We have carried out expanding laminoplasty using spinous process roofing technique for patients presenting with cervical myelopathy. The technique is a modified Kurokawa's method. The result shows that laminoplasty with this technique provides better postoperative neck movement with a simpler surgical procedure. The clinical material consists of 54 cases with more than 3 years follow-up. Long-term investigation was performed focusing on the following three points, (1) bony fusion rates, (2) radiographic neck movement, (3) neurological improvement. The results indicate good postoperative bony fusion in all cases. Postoperative neck movement was also well preserved in the majority of cases.

Adult↗
PubMed1996

Correlation between spinal cord blood flow and arterial diameter following acute spinal cord injury in rats.

Simultaneous measurements of spinal cord blood flow and arterial diameter at areas adjacent to a site of spinal cord injury were carried out to determine changes in CO(2) reactivity and autoregulation. The spinal cord injury was made at T10 level by the epidural clip compression method. A spinal window was drilled at an area either 7 mm caudal or 7 mm rostral to the injury site for the measurement of spinal cord blood flow and arterial diameter at the same time. Spinal cord blood flow was decreased at both spinal windows, especially at the rostral window. Arterial diameter was also decreased significantly at both sites. The ischaemic zone evaluated histologically tended to expand more diffusely in the rostral direction than in the caudal direction. In the pre-injury stage, both CO(2) reactivity and autoregulation were present in the spinal cord. Following the clip injury, CO(2) reactivity and autoregulation were both impaired in the areas 7 mm adjacent to the impact site. Correlation coefficients suggested that the rostral spinal cord tended to sustain more injury than the caudal spinal cord. The histologically proven spinal cord ischaemia following the injury may have resulted from the decreased arterial diameter and impaired CO(2) reactivity and dysautoregulation of the spinal cord.

Animals↗
PubMed1996

Clinical features of the haemorrhage type moyamoya disease based on 31 cases.

We evaluated and analysed our own 31 cases of the haemorrhagic type of moyamoya disease to clarify the clinical features of this disease. The cases were divided into three groups. Group A consisted of 12 cases with aneurysms. Aneurysms on the circle of Willis were treated as ordinary saccular aneurysms. Group B consisted of 14 cases with intracerebral haemorrhage (ICH) without aneurysms. These were managed almost as spontaneous ICH. Group C consisted of 5 cases with intraventricular haemorrhage (IVH) without aneurysms or ICH. Twenty-two surgical procedures for aneurysms, ICH and IVH were done in 19 cases (62%). Nineteen procedures for preventing future strokes were undertaken in 11 cases (35%). The overall initial outcome was excellent in 12 cases (39%), good in 7 cases (23%), poor in 7 cases (23%), and death in 5 cases (15%). During the follow-up period (mean: 6.5 years), rebleeding occurred in two cases (8%), and ischaemic attacks in two cases (8%). The rate of rebleeding or ischaemic attacks was 1.19% per patient-year during the follow-up period. There was no ischaemic or rebleeding episode in cases treated by STA-MCA bypass with encephalomyosynagiosis (EMS) during the follow-up period. Management of the primary haemorrhage should be according to the clinical condition, type of haemorrhage, and source of haemorrhage. When the patient needs to undergo revascularization surgery to prevent future strokes, we recommend STA-MCA bypass with EMS instead of encephaloduro-arteriosynangiosis (EDAS).

Adult↗
PubMed1996

Characteristics and localization of rat submandibular gland proteoglycans.

The submandibular gland proteoglycans were investigated biochemically and immunohistochemically in male Sprague-Dawley rats. Proteoglycans were extracted with 4 M guanidine-HCl, followed by ultracentrifugation in a CsCl density gradient, and fractionated by ion-exchange chromatography and gel filtration. The molecular weight of PGs was estimated by SDS-PAGE and immunoblot analysis with monoclonal antibodies (HepSS-1 or 6-B-6). The glycosaminoglycan side-chains in the proteoglycan fractions were identified by electrophoresis on cellulose acetate membrane. Three proteoglycan fractions were obtained. One was a heparan sulphate proteoglycan that migrated as a diffuse band of about 210 kDa. The other two fractions contained at least two dermatan sulphate proteoglycans of 70-85 kDa and 40-50 kDa. Digestion of these two proteoglycans with chondroitinase ABC, but not heparitinase, produced two bands of 50 and 21 kDa, which were core proteins. The smaller dermatan sulphate proteoglycan may be a portion of the other, as the core protein of both bound to 6-B-6 antibody, and sugar chains of both were the same (20-30 kDa). Heparan sulphates recognized by antibody HepSS-1 were observed widely in the basement membrane, fibrous connective tissue, and striated and excretory ductal cells, while dermatan sulphate proteoglycans recognized by antibody 6-B-6 were located in the connective tissue surrounding striated and excretory ducts.

Animals↗
PubMed1996

Induction of Na+/myo-inositol cotransporter mRNA after focal cerebral ischemia: evidence for extensive osmotic stress in remote areas.

Myo-inositol is one of the major organic osmolytes in the brain. It is accumulated into cells through an Na+/ myo-inositol cotransporter (SMIT) that is regulated by extracellular tonicity. To investigate the role of SMIT in the brain after cerebral ischemia, we examined expression of SMIT mRNA in the rat brain after middle cerebral artery occlusion, which would reflect alteration of extracellular tonicity. The expression of SMIT mRNA was markedly increased 12 h after surgery in the cortex of the affected side and lasted until the second day. Increased expression was also found in the contralateral cingulate cortex. Up-regulated expression was found predominantly in the neurons in remote areas, although nonneuronal cells adjacent to the ischemic core also expressed this mRNA. These results suggest that cerebral ischemia causes extensive osmotic stress in brain and that the neuronal cells respond to this stress by increasing SMIT expression.

Animals↗
PubMed1996

Age changes in the rat temporomandibular joint articular disc: a biochemical study on glycosaminoglycan content.

The temporomandibular joint (TMJ) articular discs were removed from female Sprague-Dawley rats 3, 5, 10, 32, 90 and 130 weeks of age. Glycosaminoglycans (GAGs) were extracted from the discs by heat treatment, alkali treatment and digestion with Pronase E, and purified by precipitation with cetylpyridinium chloride and ethanol. The concentration of total GAG was highest in the 3 week extracts and tended to decrease with age. Dermatan sulphate was the predominant GAG detected in all age groups along with chondroitin sulphate, hyaluronic acid and heparan sulphate. The disaccharides obtained from chondroitin sulphate were delta Di-4S, delta Di-6S and delta Di-0S, with delta Di-4S being the predominant isomer followed by delta Di-6S for all ages of all the GAG examined. The concentration of chondroitin sulphate showed a decrease with age. Quantitative changes of GAG with age may be related to functional changes in TMJ discs.

Aging↗
PubMed1996

Disaccharide analysis of chondroitin sulfate in peri-implant sulcus fluid from dental implants.

We collected peri-implant sulcus fluid by capillary tubes from sites around titanium osseointegrated implants and determined the chondroitin sulfate released into the peri-implant sulcus fluid by high-performance liquid chromatography. Chondroitin sulfate was found in all peri-implant sulcus fluid samples, and its content was similar to that in gingival crevicular fluid obtained around natural teeth. The predominant unsaturated disaccharide isomer was delta Di-0S, followed by delta Di-4S. Delta Di-6S was present in trace amounts. The amount of delta Di-0S was greater in peri-implant sulcus fluid than in gingival crevicular fluid. Assaying chondroitin sulfate disaccharides in peri-implant sulcus fluid may be an effective method of monitoring the peri-implant condition of dental implants.

Chondroitin Sulfates↗
PubMed1996

High-performance liquid chromatography analysis of chondroitin sulphate isomers in human whole saliva in a variety of clinical conditions.

OBJECTIVES: Tests have been carried out to assess the level of unsaturated disaccharide isomers obtained from chondroitin sulphate in whole saliva, which contains chondroitin sulphate derived from gingival crevicular fluid (GCF). MATERIALS AND METHODS: Whole saliva was collected from periodontally diseased subjects (PDS), clinically healthy subjects (CHS) and edentulous subjects (ES). Glycosaminoglycans (GAG) were liberated by digestion with Pronase E, and precipitated with cetylpyridinium chloride and ethanol. The unsaturated disaccharides obtained by chondroitinase ACII digestion of the liberated GAG were analysed by high-performance liquid chromatography. The unsaturated disaccharides included delta Di-0S, delta Di-6S and delta Di-4S. RESULTS AND CONCLUSIONS: Analysis of data indicated that delta Di-0S, delta Di-6S and delta Di-4S were found in all PDS samples. The amount (ng ml-1 collected whole saliva) of delta Di-0S, delta Di-6S and delta Di-4S (P < 0.01) indicated significant differences between CHS and PDS whole saliva samples. The quantities of delta Di-0S and delta Di-4S (P < 0.01) indicated significant differences between PDS and ES whole saliva. The amount of delta Di-0S (P < .05) and delta Di-6S (P < 0.01) also indicated significant differences between CHS and ES whole saliva. These results indicate that chondroitin sulphate in PDS and CHS whole saliva is representative of that previously reported in gingival crevicular fluid and so provides a useful and alternative means of assessing the role of GAG as indicators of periodontal disease.

Adult↗