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Biomedical subjects

S Kubota

Publications and source records attributed to S Kubota.

At least 145 records · Page 8Linked to original sources
PubMed1996

Nucleo-cytoplasmic redistribution of the HTLV-I Rex protein: alterations by coexpression of the HTLV-I p21x protein.

The function of the Rex protein of human T-cell leukemia virus type I (HTLV-I) has been demonstrated to be very similar to the Rev protein of human immunodeficiency virus type 1 (HIV-1). Both of these retroviral regulatory proteins rescue unspliced viral RNAs from the nuclei of infected cells. The Rev protein of HIV-1 has been reported to shuttle between the nucleus/nucleolus and the cytoplasm. Here, we have found that Rex also relocated out of the nucleus in the presence of actinomycin D. This effect was demonstrated in dose- and time-course-dependent manners. In comparison with previous reports on HIV-1 Rev, these effects with Rex seemed to be similar, but less distinct, which may reflect precise differences in the subcellular localization and/or shuttling pathways of Rev and Rex. Interestingly, the endogenous truncated form of the Rex protein, p21x, significantly interfered with the intracellular translocation of Rex, when coexpressed in trans. As expression of p21x occurs in various HTLV-I-infected cells, p21x may play a role in the life-cycle of HTLV-I, through regulating the dynamic subcellular distribution of the viral trans-activator, Rex.

Animals↗
PubMed1996

Cis/trans-activation of the interleukin-9 receptor gene in an HTLV-I-transformed human lymphocytic cell.

The MT-2 cell-line, which had been established through in vitro cell to cell transmission of human T-cell leukemia virus type I (HTLV-I) among human primary lymphocytes, was shown to possess multiple copies of integrated proviruses, including defective proviral genomes. By analysing a genomic clone, we identified the integration site of a single HTLV-I long terminal repeat (LTR) in the interleukin-9 (IL-9) receptor (IL-9R) gene. The integrated HTLV-I-LTR was shown to be functional as a promoter and the integration site was located in an intron upstream of the first coding exon of the IL-9R gene. Upon analysis of total cellular RNA, specific expression of HTLV-I-LTR Il-9R chimeric mRNAs in MT-2 cells was demonstrated. Cloning and characterization of these cDNAs have identified HTLV-I-IL-9R chimeric splicing, using either intact or alternative splice sites within the IL-9R gene. The potential roles of multiple interactions between IL-9, IL-9R and HTLV-I in the monoclonal expansion and transformation of MT-2 cells are explored.

Base Sequence↗
PubMed1996

Tumor necrosis factor alpha and lymphotoxin stimulate human myeloblastic leukemia cell (ML-1) invasion through a reconstituted basement membrane (Matrigel) with concomitant induction of 92 kDa gelatinase secretion.

We have examined the effects of tumor necrosis factor alpha (TNF) and lymphotoxin (LT) on gelatinase (72 kDa and 92 kDa) and tissue inhibitor of metalloprotease 1 (TIMP1) secretion by human myeloblastic leukemia cells (ML-1) in vitro. TNF (0.1-30 ng/ml) significantly stimulated 92 kDa gelatinase secretion in a dose-dependent manner, but did not significantly stimulate 72 kDa gelatinase secretion. LT also significantly stimulated 92 kDa gelatinase secretion, but the stimulation was less effective compared to TNF. TNF, but not LT, concentrations at 30 ng/ml slightly stimulated TIMP1 secretion. Because 92 kDa gelatinase is thought to play a pivotal role in tumor invasion, we examined the effect of TNF or LT on ML-1 cell invasion through a reconstituted basement membrane (Matrigel). Exposure of ML-1 cells to TNF (3, 10, and 30 ng/ml) or LT (3, 10, and 30 ng/ml) stimulated ML-1 cell invasion through Matrigel in a dose-dependent manner in vitro. The data suggest that TNF- and LT-stimulated 92 kDa gelatinase secretion could play an important role in TNF- or LT-stimulated ML-1 cell invasion.

Antibodies, Monoclonal↗
PubMed1996

Analysis of immunodominant piroplasm surface antigen alleles in mixed populations of Theileria sergenti and T. buffeli.

Previous studies revealed that isolates of Theileria sergenti consisted of at least 2 parasite populations bearing 2 different allelic forms of p33/32, an immunodominant piroplasm surface protein, and that the majority of T. sergenti-infected calves harbored mixed parasite populations of Ikeda (I)- and Chitose (C)-types. In this study we designed an additional oligonucleotide primer, which specifically amplified the p34 gene of T. buffeli (BI-type). DNAs from 3 field isolates in Japan were positive with this primer as well as those prepared from cattle imported from Australia and the T. buffeli Warwick stock, but restriction enzyme patterns of the PCR products from the former 3 were different from those of the latter. The nucleotide sequence of the gene amplified from Japanese isolates showed 89% homology at the nucleotide level and 88% homology at the amino-acid level with T. buffeli p34 genes. The results indicate that T. sergentilbuffeli distributed in Japan and Australia is a mixture of parasites with various combinations of 4 different allelic types.

Alleles↗
PubMed1996

PET analysis of a case of cerebrotendinous xanthomatosis presenting hemiparkinsonism.

We describe a 34-year-old Japanese woman presenting gait difficulty and Achilles tendon swelling. The patient was diagnosed as having cerebrotendinous xanthomatosis (CTX) based on the high serum cholestanol level and diminished enzymatic activity of 27-hydroxylase of fibroblasts from her skin. Her clinical presentation was atypical regarding the presence of hemiparkinsonism and absence of apparent cataract, dementia, and cerebellar ataxia. Although MRI studies could not detect any abnormality in the basal ganglia or midbrain, PET analysis using [18F]-6-fluoro-L-dopa revealed reduced uptake of dopamine into the putamen, suggesting the impairment of presynaptic dopaminergic neurons.

Adult↗
PubMed1996

(R-Z)-7,15-hexadecadien-4-olide, sex pheromone of the yellowish elongate chafer, Heptophylla picea.

An active component of the sex pheromone system of the yellowish elongate chafer, Heptophylla picea was identified by GC-EAD. Mass spectral data and hydrogenation revealed that the active compound was a hexadecadien-4-olide. It was not possible to determine the double bond positions by direct DMDS derivatization of the pheromone, but partial hydrogenation (diimide) followed by DMDS derivatization showed that the double bonds were located in positions 7 and 15. FTIR (tracer) of the pheromone corroborated the lactone structure (1772 cm-1) and showed a band characteristic of a terminal double bond at 3073 cm-1, and one of a double bond in the cis-configuration at 3002 cm-1. Chiral resolution of the pheromone, after hydrogenation, demonstrated that the natural lactone had the (R)-stereochemistry. Synthetic (R,Z)-7,15-hexadecadien-4-olide, prepared from L-malic acid in 14 steps, was identical to the natural product in MS, IR, retention times and biological activity. This is the first fatty acid derivative compound found as a sex pheromone of a Melolonthinae species and as far as biosynthesis is concerned this is the most complex pheromone constituent of a scarab species.

4-Butyrolactone↗
PubMed1996

Population dynamics of Theileria sergenti in persistently infected cattle and vector ticks analysed by a polymerase chain reaction.

Theileria sergenti Shintoku stock consists of 2 parasite populations bearing 2 allelic forms of p33/32, an immunodominant piroplasm surface protein. Parasite population changes during parasite passages among cattle and tick vectors, and during persistent infection in individual calves were analysed by using allele-specific polymerase chain reaction (PCR). The parasite DNAs were prepared from piroplasms from calves which had been infected with Shintoku stock by inoculation of sporozoite stabilates or parasitized erythrocytes, and from sporozoite stabilates which had been prepared from Shintoku stock-infected ticks. Changes in a dominant parasite population were demonstrated during transmission from calves to vector ticks and from infected ticks to calves. Parasite population changes were also apparent during persistent infection in cattle over several months, and this change is thought to occur under host immune pressure. The results of this study indicate that expression of diverse forms of p33/32 may play a role in parasite persistence within mammalian hosts and its transmission from tick vector.

Animals↗
PubMed1996

In vitro studies on a new method for islet microencapsulation using a thermoreversible gelation polymer, N-isopropylacrylamide-based copolymer.

Various materials for the semipermeable membrane for microencapsulation of islets, such as alginate complex and agarose, have been used. In this study, a thermoreversible gelation polymer, N-isopropylacrylamide based copolymer was used to make microencapsulated islets and was examined in vitro. The polymer has little or no cytotoxicity for human dermal fibroblasts. The characteristics of viscoelasticity below the soluble gel transition temperature (SGTT) and of thermoreversibility, the water soluble polymer below the SGTT (22 degrees C) becoming water insoluble upon heating, contributed to simplifying the encapsulation technique. We obtained viable islets at the center of the membrane with a thickness of approximately 20-50 microns, accounting for a 40% yield of encapsulated islets. Static glucose challenge test with microencapsulated islets revealed the insulin response to the concentration of glucose. The insulin concentrations of the culture medium in the microencapsulated islet group were the same as those in a similar free islet group up to 42 days. These results indicate that the morphological and functional stability of the new method for microencapsulation may be sufficient for it to be used for transplantation in diabetic animals.

Acrylamides↗
PubMed1996

Nuclear preservation and cytoplasmic degradation of human immunodeficiency virus type 1 Rev protein.

Rev, a major regulatory protein of human immunodeficiency virus type 1, has been demonstrated to shuttle between the nucleus and cytoplasm of infected cells. The fate of the Rev protein in living cells was evaluated by pulse-chase experiments using a transient Rev expression system. Sixteen hours after chasing with unlabelled amino acids, 45% of the labelled Rev was still present, which clearly indicates a long half-life of Rev in living cells. A Rev mutant which is deficient in the ability to migrate from the nucleus to the cytoplasm was degraded more slowly than the wild-type Rev protein. As well, another Rev mutant protein, which lacks a functional nucleolar targeting signal (NOS) and is unable to enter the cell nucleus, was rapidly degraded and undetectable 16 h after chasing. Nuclear-nucleolar targeting properties provided by a divergent NOS from a related retrovirus, which was used to substitute for the NOS of Rev, increased the intracellular half-life of this Rev mutant. Moreover, coexpression of an intracellular anti-Rev single-chain antibody (SFv), which has been shown to interfere with the nuclear translocation of Rev, accelerated the degradation of the wild-type Rev protein. Differential degradation of Rev in the nucleus and cytoplasm may play a critical role in determining and maintaining different stages of human immunodeficiency virus type 1 infection, in conjunction with the shuttling properties of the Rev protein.

Amino Acid Sequence↗
PubMed1996

Binding of intracellular anti-Rev single chain variable fragments to different epitopes of human immunodeficiency virus type 1 rev: variations in viral inhibition.

Intracellular immunization to target the human immunodeficiency virus type 1 (HIV-1) regulatory protein Rev has been explored as a genetic therapy for AIDS. Efficient intracellular expression of rearranged immunoglobulin heavy and light chain variable regions of anti-Rev monoclonal antibodies, with various vectors, and subsequent inhibition of HIV-1 replication have been previously reported by our laboratories. To further understand the molecular mechanisms and effects that intracellular anti-Rev single chain variable fragments (SFvs) have against HIV-1, via blocking of Rev function, two anti-Rev SFvs which specifically bind to differing epitopes of the Rev protein have been cloned. One SFv binds to the Rev activation domain, and the second SFv binds to the distal C terminus of Rev in the nonactivation region. Further studies now demonstrate that both anti-Rev SFvs lead to variable resistance to HIV-1 infection. Although binding affinity assays demonstrated that the SFv which specifically recognizes the Rev activation domain (D8) had an extracellular binding affinity significantly lower than that of the SFv specific to the nonactivation region (D1O), the SFv D8 demonstrated more potent activity in inhibiting virus production in human T-cell lines and peripheral blood mononuclear cells than did SFv D10. Thus, extracellular binding affinities of an SFv for a target viral protein cannot be used to directly predict its activity as an intracellular immunization moiety. These data demonstrate potential approaches for intracellular immunization against HIV-1 infection, by efficiently blocking specific motifs of Rev to after the function of this retroviral regulatory protein. These studies extend the understanding of the effects, on a molecular level, of SFvs binding to critical epitopes of Rev and further suggest that rational design of SFvs, with interactions involving specific viral moieties which mediate HIV-1 expression, may hold promise for the clinical application of genetic therapies to combat AIDS.

Amino Acid Sequence↗
PubMed1996

Relation between depressed cardiac response to exercise and autonomic nervous activity in mildly symptomatic patients with idiopathic dilated cardiomyopathy.

We investigated whether the depressed cardiac response to adrenergic stimulation is accompanied with impaired autonomic function in mildly symptomatic patients with idiopathic dilated cardiomyopathy (DCM). Twenty-seven patients with DCM (New York Heart Association class I or II) and 7 normal control subjects underwent exercise radionuclide ventriculography and 24-h ambulatory ECG. The following frequency components of heart rate variability were calculated: the areas under the low (low frequency component [LF], 0.04 to 0.15 Hz), high (high frequency component [HF], 0.15 to 0.40 Hz), and total frequency portions of the spectrum. HF and HF% (the ratio of HF to total power) were calculated as indexes of specific vagal influences, and LF% (the ratio of LF to total power) and the ratio of LF to HF were of sympathetic tone. The left ventricular ejection fraction (LVEF) increased by more than 5% in all normal control subjects during exercise, whereas 17 (63%) of patients failed to show more than a 5% increase in LVEF. The profile of the mean hourly HF% and LF/HF showed circadian variations in normal control subjects but not in patients. The HF and HF% during sleep were significantly lower and the LF/HF during sleep was higher in patients than in normal control subjects. In patients, the LVEF during exercise minus LVEF at rest was significantly correlated with HF, LF%, and LF/HF during sleep, and with the ratios of the mean values during early morning to the mean daytime values for those spectral indexes. Our results demonstrated that mildly symptomatic patients with DCM showed an attenuated cardiac response to exercise and altered autonomic function, and their close relationship, suggesting that autonomic nervous activity contributes to cardiac desensitization in DCM.

Adrenergic Fibers↗
PubMed1996

[Method of calculating TDF biological equivalent for optimal treatment dose in fractionated intracavitary irradiation of carcinoma of the uterine cervix].

Intracavitary irradiation therapy for carcinoma of the uterine cervix used with high or low dose rate irradiation is fractionated in Japan. The optimal treatment dose is determined according to the biological effect on both diseased and healthy tissues. The equations of modified NSD and TDF biological equivalents were recalculated from Arai's clinical data, which were used to examine the optimal time-dose-fractionation relationship for high and low dose rate intracavitary irradiation on squamous cell carcinoma of the cervix uteri. The optimal time-dose-fractionation relationship at point A is expressed as follows: D = NSD N0.26 T0.06 where NSD is 17.75 get for high dose rate and 31.78 get for low dose rate. TDF = K n d1.47 x-0.09 where K is 1.46 for high dose rate and 0.62 for low dose rate. The range of the optimal total dose to point A given by one fraction per week was 30.7 Gy for 4 fractions and 38.3 Gy for 8 fractions in high dose rate irradiation. In the case of low dose rate irradiation, the optimal total dose given by one fraction per week and the dose rate of 75.0 cGy/h was 55.0 Gy for 4 fractions. The maximum dose difference between our result and Arai's was about +/- 10%. The dose modification ratio for high dose rate and low dose rate is 1.79.

Brachytherapy↗
PubMed1996

[A case of congenital AVM in temporoparietal muscle].

A rare case with coexistent congenital scalp AVM in the temporoparietal muscle and a saccular aneurysm of the main feeder is presented. A 47-year-old female was admitted with the complaint of a right gradually expanding and painful pre-auricular mass. Physical examination showed a round, tender and compressible tumor measuring 3 x 3 cm in the right pre-auricular region, and there was bruit on the mass. Pre- and Post-contrast CT scans showed a slightly enhanced extracranial mass in the right temporal subcutaneous region. MR imaging showed a flow void mass both on T1 and T2 weighted images. Selective external carotid angiogram revealed multiple feeders deriving from the superficial temporal artery (STA) and the abnormal vascular networks. And also the saccular aneurysm was detected in the main feeder of the STA. Under a diagnosis of scalp AVM, it was totally removed with the temporoparietal muscle, and the aneurysmal neck was ligated. Histopathological examination revealed AVM. She was discharged on the 17th postoperative day with no deficits. Based on our experience and a review of the literature, the treatment of choice is embolization for scalp AVM localized in the skin alone and surgical total removal for AVM embedded in the muscle.

Arteriovenous Fistula↗
PubMed1996

[An autopsy case of renal failure as its cause of death in a patient with primary hyperparathyroidism associated with chronic pancreatitis].

This paper presents a 59-year-old man who was admitted to our hospital because of abdominal pains in 1973. He had pancreatic calcification and showed high levels of serum amylase, Ca, and PTH. He was diagnosed as primary hyperparathyroidism with chronic pancreatitis. After excision of an ectopic parathyroid adenoma, serum Ca levels were decreased and normalized by dihydrotachysterol p.o. At the same time his symptoms disappeared. The exocrine and endocrine pancreatic functions, however, decreased gradually. Diabetes mellitus appeared in 1975 and he required insulin injection since 1983. In spite of the treatment, his diabetic control was poor. Seventeen years later in 1992, he showed hypertension and edema (nephrotic syndrome). Because of renal failure, he underwent hemodialysis and passed away due to myocardial infarction in 1993. Autopsy findings showed existence of diabetic nephropathy as the cause of renal failure. Clinical course of this patient suggests that severe complications occur even in pancreatic diabetes and that we have to control diabetes strictly in pancreatic diabetes as well as in primary diabetes.

Chronic Disease↗
PubMed1995

Analysis of a novel defective HTLV-I provirus and detection of a new HTLV-I-induced cellular transcript.

HTLV-I generally integrates at least one full-length copy in adult T-cell leukemia (ATL) cells. A group of patients without full-length provirus have a unique conserved truncation of the provirus which retains env-pX-3'LTR. Tumor cells of a patient from this group were genetically analyzed. Analysis of the 5' and 3' cellular flanking region adjacent to the provirus suggest that the defective provirus was integrated immediately downstream of a promoter of an unknown cellular gene. The activity of the promoter was weak but was responsive to Tax-like HTLV-I LTR. The provirus may have utilized it as a substitute for the 5'LTR and thus 3'LTR may have become an alternative promoter for the cellular gene, which may give similar viral-cellular interactions to that of general cases with full-length proviruses. Surprisingly, the 3' cellular flanking region which is thought to be controlled originally by the promoter is constitutively expressed specifically in an HTLV-I producing ATL cell line HUT1O2G, in which the corresponding region is not modified by provirus. The detection of this HTLV-I-induced transcript provides a probe to find an HTLV-I inducible unknown cellular gene that may be related to the pathogenesis of ATL.

Adult↗