Biomedical subjects

S Kubota

Publications and source records attributed to S Kubota.

At least 127 records · Page 7Linked to original sources

PubMed1998

Survey of Theileria parasite infection in cattle in Taiwan.

An survey of Theileria parasite infection in cattle in Taiwan was carried out by polymerase chain reaction (PCR). A total of 491 blood samples, 105 from southern area and 386 from northern area, were collected from bovine in 16 different farms. From northern area, Theileria piroplasms could be seen in only 4 of 105 blood samples microscopically. However, when p32/34 genes (encoding immunodominant piroplasm surface proteins) were amplified by PCR, 15 blood samples were detected positive. They were analyzed by using allele-specific primers of 3 allelic forms of p32/34 and all contained C type of T. sergenti. Four blood samples were found infected with both C and B (T. buffeli) type parasites. Examination of 386 blood samples from southern area of Taiwan did not reveal any Theileria parasite microscopically, as well as by PCR amplification.

Animals↗

PubMed1998

Alternative pre-mRNA splicing of the sterol 27-hydroxylase gene (CYP 27) caused by a G to A mutation at the last nucleotide of exon 6 in a patient with cerebrotendinous xanthomatosis (CTX).

A recently identified G to A mutation at the last nucleotide of exon 6 of the sterol 27-hydroxylase gene (CYP 27) in a patient with cerebrotendinous xanthomatosis (CTX) was shown here to cause alternative pre-mRNA splicing of the gene. Northern blot analysis of the patient's RNA revealed a broadened band in the human CYP 27 mRNA region compared to that of the normal sample, indicating that there may exist differently spliced mRNA species in the patient. RT-PCR produced three fragments in the patient, one was full-length size and the other two were of smaller sizes. Sequence analysis confirmed that the nucleotide of the full-length size was identical to that of the normal full-length cDNA, except for the G to A mutation at codon 362, which corresponds to the last nucleotide of exon 6. One of the smaller size species lacked exon 6 and the other was absent from the 3' terminal 88 bp of exon 6 due to the use of an activated cryptic 5' splice site in exon 6. The correctly spliced mRNA harbouring the G to A mutation was responsible for the deficiency of the sterol 27-hydroxylase activity, as confirmed by transfection experiment. Transfection of constructed minigenes, with or without the mutation, showed that correctly spliced mRNA was observed in the normal minigene while the mutant minigene was differently spliced. This is the first report of a G to A substitution at the last nucleotide of an exon resulting in both normal and abnormal pre-mRNA splicings, including exon skipping and activating of a coding region cryptic 5' splice site. The results reveal a new molecular basis for the CTX and provide information on aberrant splicing of pre-mRNA in multi-exon genes.

Adult↗

PubMed1997

Computer-assisted skull identification system using video superimposition.

This system consists of two main units, namely a video superimposition system and a computer-assisted skull identification system. The video superimposition system is comprised of the following five parts: a skull-positioning box having a monochrome CCD camera, a photo-stand having a color CCD camera, a video image mixing device, a TV monitor and a videotape recorder. The computer-assisted skull identification system is composed of a host computer including our original application software, a film recorder and a color printer. After the determination of the orientation and size of the skull to those of the facial photograph using the video superimposition system, the skull and facial photograph images are digitized and stored within the computer, and then both digitized images are superimposed on the monitor. For the assessment of anatomical consistency between the digitized skull and face, the distance between the landmarks and the thickness of soft tissue of the anthropometrical points are semi-automatically measured on the monitor. The wipe images facilitates the comparison of positional relationships between the digitized skull and face. The software includes the polynomial functions and Fourier harmonic analysis for evaluating the match of the outline such as the forehead and mandibular line in both the digitized images.

Adult↗

PubMed1997

Ornithine decarboxylase overexpression in mouse 10T1/2 fibroblasts: cellular transformation and invasion.

BACKGROUND: Ornithine decarboxylase (ODC) plays a pivotal role in the synthesis of polyamines, a group of chemical compounds that are essential for cell growth. Recent reports have shown that ODC overexpression may be involved in malignant transformation of immortalized NIH 3T3 cells. We have demonstrated that ODC-overproducing mouse breast cancer cells are more invasive in vitro than control cells. However, little information is available concerning the relationship between ODC overexpression, tumor invasion, and metastasis and the signal transduction pathways involved in ODC-induced transformation and invasion. PURPOSE: Our purpose was twofold: 1) to determine whether ODC overexpression is directly involved in tumor cell invasion and 2) to determine whether ODC overexpression induces mitogen-activated protein (MAP) kinase activities that are associated with cell growth and transformation. METHODS: We transfected C3H clone 8 mouse 10T1/2 fibroblasts with an expression vector that carries a complementary DNA encoding rat ODC. Neomycin-resistant cells that overproduced ODC (4-6.5 times the control levels) were isolated. The transformed phenotype of these cells was determined by assessing colony formation and anchorage-independent growth in soft agar. The invasiveness of the cells was studied by means of an invasion assay that used Matrigel-coated filters in Boyden chambers. The MAP kinase activity of the cells was assayed by an in-gel kinase assay, using myelin basic protein as the substrate. RESULTS: Overexpression of ODC induced not only cell transformation and anchorage-independent growth in soft agar but also invasiveness through a Matrigel-coated filter. The ODC-overproducing transfectants showed enhanced MAP kinase activity that paralleled the magnitude of cell invasiveness. CONCLUSIONS: ODC plays a pivotal role not only in cell transformation but also in cancer cell invasion. ODC overexpression enhanced MAP kinase activity. IMPLICATIONS: Our results demonstrate a connection between the polyamine/ODC and the MAP kinase signal transduction pathways and suggest that MAP kinase may play a pivotal role in ODC-induced cell transformation and invasion.

Animals↗

PubMed1997

Anti-alpha3 integrin antibody induces the activated form of matrix metalloprotease-2 (MMP-2) with concomitant stimulation of invasion through matrigel by human rhabdomyosarcoma cells.

Proteolytic enzymes, such as matrix metalloproteases (MMPs), play a pivotal role in cancer invasion and metastasis. Invasive human rhabdomyosarcoma cells (RD) secreted proMMP-2 (72-kDa progelatinase). We found that anti-alpha3 and -alpha2 integrin antibodies induced the activated form of MMP-2 and enhanced proMMP-2 secretion by RD cells. The effect of anti-alpha2 integrin antibody was less prominent than that seen with anti-alpha3 integrin antibody. Moreover, we have found that anti-alpha3 and -alpha2 integrin antibodies enhanced RD-cell invasion through matrigel (reconstituted basement membrane) by 2.6- and 2.0-fold respectively this process was abrogated by neutralizing antibody to MMP-2. These data suggest that signaling events induced by anti-alpha3 integrin antibody may be involved in RD-cell invasion as a result of modulation of matrix-metalloprotease expression.

Antibodies, Anti-Idiotypic↗

PubMed1997

Control of Theileria sergenti infection by vaccination.

Bovine piroplasmosis caused by Theileria sergenti is a major cause of economic loss in grazing cattle in Japan. Infected calves show chronic anaemia with intraerythrocytic piroplasms and occasionally die in severe cases. We found that parasite stocks and isolates consist of genetically and antigenically mixed populations. To differentiate parasite populations bearing 3 allelic forms of p32/34, an immunodominant piroplasm surface protein, 3 sets of oligonucleotide primers were designed to amplify either of 3 alleles by polymerase chain reaction (PCR). By using this allele-specific PCR, we found that the majority of T. sergenti-infected calves in Japan harbored mixed parasite populations bearing C and I type parasites. To control Theileria infection, we produced 2 vaccine candidates: recombinant baculovirus p32 and synthetic peptide containing Lys-Glu-Lys (KEK) motif. Immunization with either recombinant p32 or synthetic peptide containing KEK sequences with Freund's complete adjuvant resulted in low parasitemia and reduced the clinical symptoms compared to control calves. Interestingly, the parasite with the p32 allelic form corresponding to the one used as the immunogen was suppressed.

Amino Acid Sequence↗

PubMed1997

A germline restricted, highly repetitive DNA sequence in Paramyxine atami: an interspecifically conserved, but somatically eliminated, element.

In some species of hagfish, the phenomenon of chromosome elimination occurs during embryogenesis. However, only two repetitive DNA families are known to be represented in chromosomes that are eliminated from somatic cells of the Japanese hagfish Eptatretus okinoseanus. Using molecular analyses, another germ line-restricted, highly repetitive DNA family has been detected in another Japanese hagfish, Paramyxine atami. The repeat unit of this family, which is 83 bp long, has been designated "EEPa1", for Eliminated Element of P. atami 1. DNA filter hybridization using EEPa1 as a probe revealed that this family is shared among several species and is conserved in the germline DNA. Although eliminated, repetitive DNA that is shared interspecifically has not been reported in hagfish species, cases of chromatin diminution and chromosome elimination processes have been described previously in other organisms. The patterns and intensities of hybridization signals suggest that members of the repetitive DNA family defined by EEPa1 have undergone concerted molecular evolution.

Animals↗

PubMed1997

Depressed contractile response to exercise in diabetic patients in the absence of cardiovascular disease: relationship to adrenergic cardiac dysinnervation.

We investigated the presence of subclinical left ventricular dysfunction by determining left ventricular diastolic filling at rest and the left ventricular ejection fraction (LVEF) response to exercise with radionuclide angiography. The subjects were 40 patients with non-insulin-dependent diabetes who showed no evidence of cardiovascular disease based on clinical findings, electrocardiography and 201T1 perfusion scintigraphy. We also used 123I-metaiodobenzylguanidine (123I-MIBG) scintigraphy to investigate whether subclinical left ventricular abnormalities were related to adrenergic cardiac dysinnervation. The change on LVEF in response to exercise (delta LVEF) was < 5% in 1 of 20 normal controls and in 22 of 40 diabetic patients (P < 0.01). The peak filling rate was within normal limits in all controls but was abnormal (< 2.3 EDV s-1) in 5 of 40 patients (P = N.S.). Of the clinical and scintigraphic variables that correlated significantly with delta LVEF, the heart-to-mediastinum 123I-MIBG uptake ratio on the late planar images was the most important and independent predictor. Nearly half of the patients with non-insulin-dependent diabetes without apparent cardiovascular disease exhibited a depressed LVEF response to exercise. This subnormal response was significantly associated with diminished myocardial 123I-MIBG uptake, suggesting a causal relationship.

3-Iodobenzylguanidine↗

PubMed1997

Comparison of free and reversed pedicled posterior interosseous cutaneous flaps.

The posterior interosseous cutaneous flap is characterized by its thinness and the adjustable length of its vascular pedicle. This flap can be used both as a free and reversed vascular pedicled flap. We applied the free flap mostly for reconstruction of the digits, and the reversed vascular pedicled flap for soft-tissue defects of the proximal hand. Of 17 free flaps, 16 survived completely, and 1 developed partial necrosis. Eleven flaps were elevated as reversed vascular pedicled flaps, three of which were combined with the lateral arm flap to cover large defects. In these three combined flaps, the posterior radial collateral artery was anastomosed to an artery in the recipient site to enhance circulation; all three flaps survived completely. Operative time was 30 to 40 minutes shorter for the reversed vascular pedicled flaps. Two of the remaining eight reversed vascular pedicled flaps developed partial necrosis. The flap success rate was thus higher in the free flaps than in the reversed vascular pedicled flaps.

Adult↗

PubMed1997

A novel method to evaluate thromboresistance of drip chambers with filter used in extracorporeal blood circuits for hemodialysis.

We have developed a novel method to simultaneously evaluate thromboresistance of two different types of drip chambers with a filter used in extracorporeal blood circuits for hemodialysis using canine arteriovenous shunt models. Bilateral bypasses between the femoral artery and vein on both sides of the animal were formed with 2 test circuits consisting of polyvinyl chloride tubing and two types of drip chambers with filters. Blood flow was derived into the drip chambers with a filter and returned to the femoral vein without using peristaltic pumps and systemic anticoagulants. By monitoring the quantitative changes of the blood flow rate through each drip chamber with a filter and measuring the time elapsed before the blood flow through its was completely obstructed by clots, thromboresistance of the drip chambers was evaluated under the same blood conditions. The drip chamber with a filter that had a small effective filter area, a big pore size, and slitlike pores showed better thromboresistance. This novel method proved useful in evaluating the thromboresistance of differently designed drip chambers with a filter.

Animals↗

PubMed1997

Predictive value of left ventricular response to exercise in patients with dilated cardiomyopathy--assessment by radionuclide ventriculography.

The objective of this study was to determine noninvasively the likelihood of recovery of the left ventricular ejection fraction (LVEF) in patients with dilated cardiomyopathy (DCM) verified by radionuclide ventriculography. Twenty patients with DCM were classified into two groups according to the LVEF by M-mode echocardiographic findings two years after ventriculographic examination. The LVEF recovered to > or = 50% in 10 patients (Group A), while it was sustained at < 50% in 10 patients (Group B). The clinical features of each group were compared, based on results of physical examination, radionuclide ventriculography, and other diagnostic tests performed on their referral to hospital. Only the systolic blood pressure differed significantly between the two groups, being slightly, but significantly, increased in Group A over that in Group B (P < 0.05). LVEF at rest by radionuclide did not differ (Group A: 31.5 +/- 10.3% vs Group B: 26.5 +/- 9.4%). Peak exercise EF-EF at rest (peak delta EF) in Group A was apparently increased (3.4 +/- 4.0%), while that in Group B was decreased (-4.4 +/- 5.2%). The positive peak delta EF had a highly predictive value of 90% for patients with DCM whose LVEF will recover to more than 50%. The recovery EF-EF at rest did not differ significantly between groups (Group A: 8.4 +/- 4.7 vs Group B: 3.9 +/- 2.3, P < 0.05). Other clinical parameters such as functional class, cardiothoracic ratio, LVEF by echocardiography, cardiac index by Swan-Ganz catheter examination, and histologic examination of biopsied endocardium were indistinguishable in the two groups. The authors conclude that peak delta EF of radionuclide ventriculography on exercise indicates a preservation of LVEF and predicts a good clinical recovery in patients with DCM.

Blood Pressure↗

PubMed1997

Myocardial contractile reserve as prognostic determinant in patients with idiopathic dilated cardiomyopathy without overt heart failure.

To assess the prognostic significance of myocardial contractile reserve in patients with idiopathic dilated cardiomyopathy (DCM) without overt heart failure (New York Heart Association functional class I or II), seventy-one patients underwent exercise radionuclide angiography in addition to clinical, radiographic, hemodynamic, and echocardiographic evaluations. Myocardial contractile reserve was assessed as left ventricular ejection fraction (LVEF) during peak exercise minus LVEF at rest (delta LVEF). During an average of 49 months, 18 patients died of the disease. Cox's proportional-hazards regression analysis showed that the delta LVEF was the most powerful and independent discriminator for survival (p = 0.0002). Ejection time (p = 0.0029) and cardiothoracic ratio (p = 0.017) were the second and third most predictive variables, respectively. Evaluation of the delta LVEF, which reflects residual myocardial contractile reserve, can provide important information about the prognosis of patients with DCM and mild symptoms.

Adolescent↗

PubMed1997

Malignant cystosarcoma phyllodes tumor of the breast mainly composed of chondrosarcoma: a case report.

Sarcoma of the breast is a very rare occurrence. We experienced a patient with malignant phyllodes tumor, in which a chondrosarcomatous element constituted 90% of the tumor volume. That bone or cartilage may be present in mammary neoplasms has been known for a long time. Most mammary tumors with areas of chondroid metaplasia show a benign histologic appearance. Cystosarcoma phyllodes displaying a chondrosarcomatous element is very rare. The results obtained by echographic and pathologic examinations strongly suggest that the chondrosarcomatous element originates from the fibrous stroma.

Breast Neoplasms↗

PubMed1997

Novel homozygous and compound heterozygous mutations of sterol 27-hydroxylase gene (CYP27) cause cerebrotendinous xanthomatosis in three Japanese patients from two unrelated families.

The autosomal recessively inherited cholesterol metabolic disease, cerebrotendinous xanthomatosis (CTX), is caused by mutations in the sterol 27-hydroxylase gene. Three Japanese CTX patients from two unrelated families were studied genetically. By DNA sequence analysis a novel mutation of A for G substitution at amino acid position 372 (CGG 372Arg to CAG 372Gln) was identified in one of the CTX families. The mutation was also found in two patients from the other family, with a compound heterozygous pattern of A for G substitution at amino acid position 441 (CGG 441Arg to CAG 441Gln). The latter mutation was the same as previously reported by our group (J. Lipid Res. 1994. 35: 1031-1039). As the two mutations changed the restriction enzyme sites, rapid screening methods were developed for the detection of the carriers. Transfection of the two mutant cDNAs into COS cells resulted in markedly reduced sterol 27-hydroxylase activity. These results indicate that the two mutations are responsible for the deficiency of the sterol 27-hydroxylase activity in these patients. The features of mutations identified till now in Japanese CTX patients are also reviewed.

Cholestanetriol 26-Monooxygenase↗

PubMed1997

[Recurrent hypertensive intracerebral hemorrhage: characteristics and risk factors].

From January 1978 to December 1995, a total of 269 patients with hypertensive intracerebral hemorrhage were treated in our institution. We reviewed 23 patients with recurrence to analyze their characteristics and risk factors. RECURRENCE RATE: The recurrence rate was 8.6%, showing it is not so rare. HEMORRHAGIC SITE: In the first hypertensive intracerebral hemorrhage, the hemorrhagic site was putaminal in 60.9%, subcortex in 13.0%, thalamic in 8.7% and cerebellar in 8.7%. In the second episode, the site was putaminal in 39.1%, thalamic in 34.8%, cerebellar in 13.0%, and subcortex in 8.7%. Therefore, during recurrence, hemorrhage occurred relatively more frequently in the thalamus than in the initial site. INTERVAL TO REBLEEDING: The Interval between the first and second hemorrhage ranged from 2 months to 15 years (average 4 years and 7 months). BLOOD PRESSURE AND RECURRENCE: In patients in whom hypertensive intracerebral hemorrhage recurred in less than 3 years from the initial hemorrhage, blood pressure was well controlled by antihypertensive drugs during the interval. This indicates that factors other than blood pressure control are concerned in the prevention of recurrence. In patients in whom hemorrhage recurred 3 years or more after the initial episode, blood pressure was poorly controlled, indicating that strict antihypertensive therapy is necessary to prevent recurrence after a long interval. RISK FACTORS FOR RECURRENCE: The following risk factors for recurrence of hypertensive intracerebral hemorrhage have been reported in the literature: diastolic blood pressure > or = 100 mmHg; unstable-hypertension (blood pressure abruptly rising from the normal range) : uncontrollable hypertension; and hypoproteinemia. Of these, we emphasize that unstable hypertension is the most important risk factor, as illustrated in our series of patients.

Adult↗

PubMed1997

Vaccine development against Theileria parasite.

Bovine piroplasmosis caused by Theileria sergenti is a major cause of economical loss in grazing cattle in Japan. We found that parasite stocks and isolates consist of genetically and antigenically mixed population. To differentiate parasite populations bearing 2 allelic forms of p32, an immunodominant piroplasm surface protein, 2 sets of oligonucleotide primers were designed to amplify either of the 2 alleles by polymerase chain reaction (PCR). By using this allele-specific PCR, we found that the majority of T. sergenti-infected calves in Japan harbored mixed parasite populations with C and I type parasites. Amino acid sequence of p32 contains Lys-Glu-Lys (KEK) motif which is one of tripeptide necessary for malaria parasite to invade erythrocytes. We produced 2 vaccine candidates, recombinant baculovirus p32 and synthetic peptide containing KEK motif. Immunization of either recombinant p32 or synthetic peptide containing a KEK sequence with adjuvant resulted in low parasitemia and reduced the clinical symptoms compared to control calves. Interestingly, parasites with a p32 allelic form corresponding to one used as the immunogen were suppressed. Therefore, a cocktail vaccine containing KEK peptides derived from C and I type parasites is desired for control Theileria parasite infection in Japan.

Amino Acid Sequence↗

PubMed1996

Molecular cloning and expression of rabbit sterol 12alpha-hydroxylase.

Sterol 12alpha-hydroxylase is an important enzyme in bile acid biosynthesis, responsible for the balance between formation of cholic acid and chenodeoxycholic acid. The enzyme has been purified to apparent homogeneity from rabbit liver (Ishida, H., Noshiro, M., Okuda, K., and Coon, M. J. (1992) J. Biol. Chem. 267, 21319-21323), and we here describe the cloning and sequencing of a cDNA coding for this enzyme. After tryptic digestion of purified protein in a polyacrylamide gel, eight different peptides were isolated and sequenced. Using oligonucleotides deduced from the amino acid sequences, clones were isolated from a rabbit liver cDNA library. In addition to several overlapping clones, one full-length clone was obtained that coded for a polypeptide of 500 amino acids, corresponding to a molecular mass of 57 kDa. All of the eight peptides and the reported NH2-terminal amino acid sequence were matched against the sequence. The peptide sequence showed a 39% similarity with human prostacyclin synthase (CYP8) and 31% similarity with the rate-limiting enzyme in over-all synthesis of bile acids, the cholesterol 7alpha-hydroxylase (CYP7) of the rabbit. The similarity with most other sterol cytochrome P-450 hydroxylases was less. Thus, this species of cytochrome P-450 should belong to a group of its own, here denoted CYP12. Transfection of COS cells with the coding part of the cDNA resulted in a significant expression of sterol 12alpha-hydroxylase activity toward 7alpha-hydroxy-4-cholesten-3-one. Northern blotting showed that the enzyme was exclusively expressed in the liver. The major mRNA fraction in rabbit liver had a size of approximately 2.9 kilobases, and those found in rat and human liver were about 2.5 and 4.5 kilobases, respectively. Fasting of rats and mice led to a severalfold increase in both enzyme activity and mRNA levels. In contrast, starvation of rabbits had little or no stimulatory effect on enzyme activity and mRNA levels.

Amino Acid Sequence↗

PubMed1996

Genetic analysis of a Japanese cerebrotendinous xanthomatosis family: identification of a novel mutation in the adrenodoxin binding region of the CYP 27 gene.

Cerebrotendinous xanthomatosis (CTX), an autosomal recessive lipid-storage hereditary disorder, is caused by mutations in the sterol 27-hydroxylase gene (CYP 27). A 24-year-old female Japanese CTX patient and her parents were studied for a CYP 27 mutation. Multiple xanthomas were the main complaint of the patient and plasma cholestanol level was markedly elevated. Sterol analysis of a xanthoma biopsy confirmed cholesterol and cholestanol deposition, and the cholestanol accounted for 8.1% of the total sterols. Sterol 27-hydroxylase activity in fibroblasts derived from the patient was undetectable, while the activities in fibroblasts from her mother and father were 54% and 41% of the normal level, respectively. Direct sequence analysis showed a missense mutation of A for G substitution in the CYP 27 gene at codon 362 (CGT 362Arg to CAT 362His) with a homozygous pattern in the patient, and a heterozygous pattern in the parents. The mutation, which eliminates a normal HgaI endonuclease site at position 1195 of the cDNA and is located at the adrenodoxin binding region of the gene, is most probably responsible for the decreased sterol 27-hydroxylase activity in this Japanese CTX family. The combined data strongly support that the primary enzymatic defect in CTX is the disruption of sterol 27-hydroxylase and that the disease is inherited in an autosomal recessive trait.

Adrenodoxin↗