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K Obata

Publications and source records attributed to K Obata.

At least 127 records · Page 7Linked to original sources
PubMed1995

Secretion of Alzheimer beta/A4 protein (1-40) and intracellular retention of beta/A4 protein (1-42) in transfected COS cells.

The amyloid beta/A4 protein (beta/A4) of Alzheimer disease is heterogeneous. A beta 1-40 (Asp1 to Val40) is rather soluble, whereas A beta 1-42 (Asp1 to Ala42) is more insoluble and can be the seed of deposition of A beta 1-40. Although beta/A4 itself could not be expressed transiently in COS cells by cDNA transfection, it was expressed after directly adding a signal sequence to its N-terminal end. The expressed A beta 1-40 was secreted, whereas A beta 1-42 was hardly secreted. A beta 1-40 secretion was not inhibited by metabolic inhibitors such as brefeldin A. The normally produced A beta 1-40 could not be retained intracellularly, whereas the abnormally generated A beta 1-42 might be accumulated intracellularly, resulting in cellular toxicity.

Amino Acid Sequence↗
PubMed1995

Increased levels of stromelysin-1 and tissue inhibitor of metalloproteinases-1 in sera from patients with rheumatoid arthritis.

OBJECTIVE: To evaluate the efficacy of stromelysin-1 (matrix metalloproteinase-3 [MMP-3]) and tissue inhibitor of metalloproteinases-1 (TIMP-1) in serum as markers for joint inflammation in rheumatoid arthritis (RA). METHODS: Levels of both macromolecules in sera from 97 healthy controls, 109 patients with RA, and 47 patients with osteoarthritis (OA) were measured by respective 1-step sandwich enzyme immunoassays. In the patients with RA, serum levels of MMP-3 and TIMP-1 were investigated in relation to laboratory and clinical measures of disease activity. In addition, the relationships between serum and synovial fluid (SF) levels in paired samples from individual patients were examined. RESULTS: Serum levels of both MMP-3 and TIMP-1 in RA patients were significantly higher than those in OA patients and in healthy controls (P < 0.001), and were shown to correlate with traditional systemic markers of inflammation including the erythrocyte sedimentation rate and C-reactive protein level, and with the Lansbury articular index. In addition, it was noted that serum levels of MMP-3 correlated with the corresponding values in paired SF samples obtained concurrently from patients with RA (rs = 0.588, P < 0.001), while such correlations were not found for TIMP-1 levels. CONCLUSION: Our results support the notion that levels of both MMP-3 and TIMP-1 in RA patient sera are increased in association with inflammation. Furthermore, the level of MMP-3 in serum provides a particularly useful marker of inflammatory activity in the joints of patients with RA.

Adult↗
PubMed1995

Levels of circulating collagenase, stromelysin-1, and tissue inhibitor of matrix metalloproteinases 1 in patients with rheumatoid arthritis. Relationship to serum levels of antigenic keratan sulfate and systemic parameters of inflammation.

OBJECTIVE: To measure serum levels of matrix metalloproteinase-1 (MMP-1), matrix metalloproteinase-3 (MMP-3), and tissue inhibitor of MMP-1 (TIMP-1) in patients with rheumatoid arthritis (RA) and in age-matched control subjects, and to determine how these correlate with serum levels of antigenic keratan sulfate (KS) and other biochemical and clinical indicators of disease activity. METHODS: Immunoassays were used to measure levels of MMP-1, MMP-3, TIMP-1, and antigenic KS. Radiologic and functional joint scores were based upon Steinbrocker's criteria. Erythrocyte sedimentation rates (ESR) and levels of C-reactive proteins (CRP) were measured. RESULTS: In RA patients, levels of MMP-3 and TIMP-1 were significantly increased, and strongly correlated with the ESR and CRP levels but not with radiologic or functional joint scores. Levels of antigenic KS were significantly lower in RA patients and correlated negatively with systemic parameters of inflammation and serum levels of TIMP-1. CONCLUSIONS: The increase in serum levels of MMP-3 and TIMP-1 appears to reflect systemic inflammation in RA. The inverse correlation between serum levels of TIMP-1 and antigenic KS suggests that an upregulation of TIMP-1 synthesis might be responsible for the apparent suppression of cartilage aggrecan catabolism in patients with severe inflammatory changes.

Adult↗
PubMed1995

Increase in synapsin I phosphorylation implicates a presynaptic component in septal kindling.

Synaptic plasticity in the CNS is thought to be an important component of learning and memory. Kindling is an animal model of synaptic plasticity in which repetitive local electrical stimulation eventually leads to a generalized motor seizure. Once established, the sensitivity of kindled animals to this epileptic condition is long-lasting. An increase in synaptic efficacy appears to underlie the plastic changes observed in kindling but the molecular mechanisms involved remain unknown. Here we demonstrate that the phosphorylation state of synapsin I, a synaptic vesicle-associated protein which has been implicated in the regulation of neurotransmitter release, is significantly increased in hippocampus and parietal cortex of rats two weeks after the establishment of septal kindling. Furthermore, K(+)-evoked release of L-glutamate is significantly increased in synaptosomes prepared from cerebral cortex of kindled animals. Thus, changes within the presynaptic nerve terminal may contribute, at least in part, to the long-lasting modification in neuronal function induced by kindling.

Animals↗
PubMed1995

A synaptic vesicle-associated protein (SVP-38) as an early indicator of delayed neuronal death.

Sixteen gerbils were subjected to 5 min of forebrain ischemia. Their brains were processed for immunohistochemical staining using monoclonal antibodies against a synaptic vesicle-associated protein 38 (SVP-38) and microtubule-associated protein 2 (MAP2) after recirculation times of 10 min, and 1, 4, and 7 days. After 10 min recirculation, SVP-38 immunoreactive dots were observed only in the CA1 region of the hippocampus. After 1 day recirculation, SVP-38 immunostaining was diffuse and weak throughout the hippocampus, despite preservation of MAP2 immunoreactivity. After 4 and 7 days recirculation, SVP-38 immunoreactivity had been restored in the whole hippocampus, despite the complete loss of MAP2 immunoreactivity due to delayed neuronal death. Our results demonstrate an immediate and significant change in the immunoreactivity of a synaptic vesicle-associated protein at the beginning of the process of delayed neuronal death. Thus, changes in the immunoreactivity of synaptic vesicle-associated proteins such as SVP-38 appear to be one of the earliest indicators of the onset of neuronal death.

Animals↗
PubMed1995

Cloning and sequence analysis of cDNA for a possible DNA-binding protein 5E5 in the nervous system.

Monoclonal antibody 5E5 recognized an intranuclear antigen of neurons in the rat. We isolated 5E5cDNA and determined its nucleotide and deduced amino acid sequences. The 5E5cDNA had an open reading frame of 825 amino acids and its amino acid sequence showed no significant homology to any protein or to any DNA binding motif so far known. 5E5 protein had an abundance of basic amino acids, especially arginine, and included a glycine-rich region and a proline cluster. Monoclonal antibody 12H raised against 5E5cDNA fusion protein recognized an intranuclear substance in rat brain sections and a single protein band of about 98 kDa in the brain nuclear extract fraction on immunoblotting. DNA-cellulose column chromatography indicated that 5E5 protein might have DNA-binding ability. Transfection studies indicated that 5E5 protein expressed in COS-1 is localized in cell nuclei. These results suggest that 5E5 protein is a possible DNA-binding protein which is expressed especially in neurons.

Amino Acid Sequence↗
PubMed1995

Isolation and characterization of the IS3-like element from Thermus aquaticus.

We have cloned and characterized a genetic element (1187 bp) that is responsible for the induction of thermotolerance as well as ompC expression in E. coli. This element (ISLtaq1) was isolated from Thermus aquaticus. DNA and protein data bases were searched with this element (ISLtaq1), which suggested it to be very similar to IS150 belonging to the IS3 family. ORF1, found on ISLtaq1, which encodes 100 amino acids (aa), had a DNA-binding motif: a helix-turn-helix and a leucine zipper. In fact, when the ORF1 protein was overproduced in E. coli, thermotolerance as well as ompC expression was induced.

Adaptation, Biological↗
PubMed1995

Urinary detection of type IV collagen and its increase in glomerulonephritis.

Type IV collagen is the major component of the glomerular extracellular matrix. The nature of type IV collagen antigens in the urine from various forms of glomerulonephritis was studied using enzyme-linked immunosorbent assay for monoclonal antibodies to the 7S domain and some of the non-7S and non-NC1 domains. The size distribution of antigenic material in urinary samples from a patient with IgA nephropathy was analyzed by gel filtration chromatography. The results demonstrated a single peak at 340 kDa, suggesting the measurement of almost intact type IV collagen molecules. The urinary concentration of type IV collagen in patients with membranous nephropathy and chronic renal failure was significantly increased compared with that of normal controls. Measurement of the urinary concentrations of type IV collagen is useful in the diagnosis of glomerular disease, particularly in cases where invasive methods are contraindicated and clinically membranous nephropathy is suspected.

Adolescent↗
PubMed1995

[Stromelysin-1 (MMP-3) level in the sera from patients with rheumatoid arthritis and other connective tissue diseases--clinical significances in early onset rheumatoid arthritis].

Stromelysin-1 (MMP-3) is a metalloproteinase that degrades articular cartilage matrix in patients with rheumatoid arthritis (RA). We measured MMP-3 in the sera from patients with RA and other connective tissue diseases using specific sandwich EIA and studied its clinical significance in early onset RA. MMP-3 level in healthy control (n = 170) was significantly higher in male than in female. The level of MMP-3 in RA was significantly and dramatically higher than in healthy control, osteoarthritis, systemic lupus erythematosus, progressive systemic sclerosis, primary sjogren's syndrome, mixed connective tissue disease, gouty arthritis and traumatic arthritis. Serum MMP-3 significantly correlated with serum BUN or serum creatinine levels in SLE patients but not in RA patients. In early onset RA, serum MMP-3 level was significantly elevated. Furthermore, when the relationship between the serum MMP-3 level and X-ray findings of the joints in RA was studied, it was found that MMP-3 level was elevated even in stage I or II and that there was no statistical differences between stage I or II and stage III or IV, suggesting that serum MMP-3 level is elevated in the early stage of initial inflammatory process when only mild cartilage degradation is seen. These results suggest that measurements of serum MMP-3 is an important tool for establishing diagnosis of early onset RA, and that serum MMP-3 level may be a marker of cartilage destruction and of estimating therapeutic efficacy in early onset RA.

Aged↗
PubMed1995

Coordinate regulation of matrix metalloproteases and tissue inhibitor of metalloproteinase expression in human synovial fibroblasts.

We examined the common signal transduction mechanisms governing collagenase (MMP-1), stromelysin-1 (MMP-3), and tissue inhibitor of metalloproteases (TIMP-1) gene expression in human synovial fibroblasts for insight into the pathophysiology of arthritis. MMP-1, MMP-3, and TIMP-1 expression and synthesis were induced in cultured human synoviocytes with recombinant human interleukin 1 beta in the absence or presence of either chemical inhibitors of protein kinase A and C (PKA, PKC), or prostaglandin E2, or cyclic AMP (cAMP) mimetics. We used enzyme immunoassays (EIA) to determine MMP-1, MMP-3, and TIMP-1 antigen levels in spent culture medium and Northern hybridization to measure steady state mRNA expression levels. Extracellular signals (e.g., IL-1, phorbol myristic acetate) that result in the activation of cytoplasmic PKC augment in tandem the expression and synthesis of MMP-1, MMP-3, and TIMP-1 in human synovial fibroblasts. In addition, such signals induce nuclear transcription factors (e.g., activator protein 1) that bind to common gene regulatory elements and augment promoter activity of MMP-1, MMP-3, and TIMP-1 gene promoter constructs. In contrast, signals that activate PKA oppose PKC mediated signals, in that the expression of MMP-1, MMP-3, and TIMP-1 are suppressed. Experimental data suggest that the expression of MMP-1, MMP-3, and TIMP-1 are coordinated through a series of common cytoplasmic signal transducing pathways, cis regulatory elements, and nuclear trans acting factors.

Cells, Cultured↗
PubMed1994

Partially oxygenated sickled cells: sickle-shaped red cells found in circulating blood of patients with sickle cell disease.

A previously uncharacterized type of sickled cell was found in venous blood of patients with sickle cell disease when blood was collected without exposure to air and fixed immediately with 1% glutaraldehyde solution equilibrated with 5% oxygen. These cells were either elongated, resembling irreversibly sickled cells (ISCs), or nonelongated, with a raisin-like shape. Both types assumed a normal discoidal shape upon full oxygenation. Since these cells exist only under partially oxygenated conditions, they are described as partially oxygenated sickled cells (POSCs). POSCs are morphologically distinct from partially deoxygenated sickled cells formed during deoxygenation by having rounded edges, while the latter have sharp edges. Transmission electron microscopy of POSCs revealed various amounts of misaligned Hb S polymers. Investigations in vitro demonstrated the formation of POSC-like cells by partial oxygenation of deoxygenated cells. Since POSCs contain intracellular fibers and sickle readily upon deoxygenation, they may have clinical and pathological significance.

Anemia, Sickle Cell↗
PubMed1994

Cleavage at the N-terminal site of Alzheimer amyloid beta/A4 protein is essential for its secretion.

To characterize the secretory pathway of Alzheimer amyloid beta/A4 protein (beta/A4), mutated cDNAs of amyloid precursor protein (APP) were expressed transiently in COS cells. Although the expression of C100 (Met671-Asn770) resulted in the secretion of beta/A4-like peptide, the cells expressing the mutated APPs with the longer N-terminal domain beyond beta/A4 sequence secreted little beta/A4-like peptide. The C-terminal domain truncated APPs, with a stop codon at the end of beta/A4 sequence or the predicted membrane spanning domain produced little beta/A4-like peptide. When beta/A4 was expressed with direct addition of the signal sequence, beta/A4-like peptide was found in the cell lysate but little of it was secreted. Hence, the secretion of beta/A4 was started initially by cleavage at the N-terminal site of beta/A4, although cleavage at its C-terminal site was also necessary for its secretion.

Alzheimer Disease↗
PubMed1994

Haloperidol activates tyrosine hydroxylase gene-expression in the rat substantia nigra, pars reticulata.

The cellular distribution of tyrosine hydroxylase (TH) and TH mRNA in the rat substantia nigra (SN) was investigated using immunohistochemistry (IMHC) and non-radioactive in situ hybridization histochemistry (ISH), respectively. Number and density of both TH immunoreactive and TH cRNA labeled cells were increased in the pars reticulata of the substantia nigra (SNr) 8 h after single administration of a dopamine antagonist haloperidol. At the same time number and density of TH positive cells remained unchanged in a ventro-medial, dorso-medial or lateral part of the pars compacta (SNc) and in the pars lateralis (SNl) of the substantia nigra. A D2 receptor-specific agonist, quinpirole, was without effect on either ISH or IMHC in any of these areas, including the SNr. These results reveal the existence of a population of TH-negative neurons in the SNr, in which TH gene-expression can be activated through a dopamine receptor-mediated mechanism, leading to detectable levels of both TH and TH mRNA. Furthermore they suggest that TH gene-expression in these neurons normally is inhibited by dopamine released from somata and dendrites in the SNr.

Animals↗
PubMed1994

Serum levels of collagenase, stromelysin-1, and TIMP-1. Age- and sex-related differences in normal subjects and relationship to the extent of joint involvement and serum levels of antigenic keratan sulfate in patients with osteoarthritis.

OBJECTIVE: To measure serum levels of collagenase (MMP-1), stromelysin-1 (MMP-3), and tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) in normal subjects and in patients with osteoarthritis (OA), and to assess how these correlate with biochemical and clinical indicators of disease activity in OA. METHODS: Specific immunoassays were used to measure MMPs, TIMP-1, and antigenic keratan sulfate (KS). The total area of cartilage affected by the disease was measured (expressed as an articular index). RESULTS: In the normal population (n = 118), the serum concentration of MMP-3, but not of MMP-1 or TIMP-1, increased with age and was approximately 2 times higher in males than in females. In the OA patients (n = 33), the serum levels of MMP-3, but not of MMP-1 or TIMP-1, were significantly elevated and correlated strongly with the articular index but poorly with objective and subjective functional capacity scores as well as with serum levels of antigenic KS and systemic parameters of inflammation. CONCLUSION: These findings illustrate the importance of matching patients and normal controls for age and sex in further studies of MMP-3 and are consistent with the hypothesis that MMP-3 might play an important role in the degradation of joint cartilage in OA. Further, serum levels of MMP-3 may prove useful for monitoring therapy for OA.

Aged↗
PubMed1994

Expression of matrix metalloproteinase 9 (92-kDa gelatinase/type IV collagenase) induced by tumour necrosis factor alpha correlates with metastatic ability in a human osteosarcoma cell line.

We have examined the correlation between matrix metalloproteinase (MMP) expression and metastatic properties of a low metastatic osteosarcoma cell line, osteosarcoma takase (OST), under stimulation by tumour necrosis factor alpha (TNF alpha). In vivo, OST cells exhibited significantly increased colonization in the lungs of nude mice in a dose-dependent manner when they were treated by TNF alpha prior to injection. In vitro, TNF alpha enhanced tumour cell invasion through the reconstituted basement membrane in a transwell chamber up to 2.5-fold. Gelatin zymography and sandwich enzyme immunoassays demonstrated marked production of MMP-9 [92-kDa gelatinase/type IV collagenase (gelatinase B)] but not MMP-2 [72-kDa gelatinase/type IV collagenase (gelatinase A)], MMP-3 (stromelysin-1) or MMP-7 (matrilysin). Motility of the tumour cells and adhesion to cultured endothelial cells were slightly increased by the TNF alpha treatment up to 1.6-fold and 1.4-fold, respectively, while the growth rate was decreased. These results suggest that upregulation of MMP-9 together with enhanced motility and endothelial adhesion contribute to the increased metastatic ability of OST cells induced by TNF alpha treatment.

Collagenases↗
PubMed1994

Results of adjuvant chemotherapy for invasive urothelial cancer with lymph-node metastasis.

From 1980 to 1991, 59 patients with advanced urothelial cancer (pathological stage, > pT3) underwent radical operations. Of these 59 patients, 33 had nodal involvement. This study focused on those 33 patients with nodal involvement. The primary site was the urinary bladder in 20 patients and the upper urinary tract (renal pelvis and/or ureter) in 13. In all, 13 patients underwent adjuvant chemotherapy with an M-VAC or M-VEC [methotrexate (M), vinblastine, doxorubicin (ADM) or epirubicin, and cisplatin (CDDP)] regimen, and another 8 patients were treated with other insufficient chemotherapies [CDDP + ADM or CDDP + ADM + etoposide (VP-16)]. A group of 12 patients did not receive any additional treatment. Most of the patients in the M-VAC and M-VEC groups received more than 2 cycles of the regimen (median, 3.2 cycles; range, 1-9 cycles). The overall 5-year survival rate of the M-VAC and M-VEC group was 31%, whereas the rate was 0 for the other insufficient-chemotherapy groups and the no-chemotherapy group. Of the 13 patients in the M-VAC group, 4 (31%) patients were alive without disease progression and 9 (69%) were dead due to progressive disease. In the other groups, only 1 patient was alive without progression. Our results suggest that adjuvant M-VAC or M-VEC chemotherapy may extend the median survival of patients with advanced urothelial cancer, but it failed to reduce the rate of cancer death.

Aged↗
PubMed1994

A randomized trial of early intravesical instillation of epirubicin in superficial bladder cancer. The Nagoya University Urological Oncology Group.

A total of 135 patients with superficial bladder cancer diagnosed as totally resectable were entered into a randomized multicenter trial to investigate the efficacy of early intravesical epirubicin instillation after resection in comparison with transurethral resection (TUR) alone. Epirubicin (40 mg/40 ml saline) was given within 24 h of TUR and once during the 1st week, weekly for 4 weeks and then monthly for 11 months. In all, 122 patients (90.4%) were eligible and 119 (88.1%) were evaluable. The interval to initial recurrence was significantly longer (P = 0.02) in the epirubicin group (36 months; 95% confidence interval, 32-40 months) than in the group receiving TUR alone (28 months; 95% confidence interval, 24-32 months). The recurrence rate per year in the epirubicin group was less than that in the TUR-alone group (0.13 versus 0.29 annual recurrences). Disease progression was observed in only one patient in the epirubicin-instillation group. The main toxicity encountered was bladder irritation (13.8%). These results demonstrate that early intravesical epirubicin instillation is efficacious in preventing local recurrence.

Administration, Intravesical↗