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Biomedical subjects

A Watanabe

Publications and source records attributed to A Watanabe.

At least 199 records · Page 11Linked to original sources
PubMed2000

Subcutaneously inoculated cells and implanted pancreatic cancer tissue show different patterns of metastases in Syrian golden hamsters.

CONTEXT: We studied behavior of the subcutaneously implanted pancreatic tumors and the process of metastasis using syngeneic Syrian golden hamsters. DESIGN: HaP-T1, a cell line derived from nitrosamine-induced pancreatic cancer in Syrian golden hamsters was used for this experiment. Thirty-five animals were divided into two groups: subcutaneous cell inoculation and subcutaneous tissue implantation. The tumor tissue was obtained from subcutaneously implanted cancer cells. One month after implantation, the tumors were resected and studied histopathologically. The animals were followed-up weekly by palpation of the peripheral lymph nodes in order to identify local recurrence. After death, necropsy was performed. Liver, lungs and pancreas specimens were taken for histopathogical study and detection of K-ras point mutation using the PCR/RFLP method. RESULTS: The mean survival time in the subcutaneous cell inoculation group was 151+/-17.5 days, and in the subcutaneous tissue implantation group was 137 +/-12.9 days. During the follow-up, 13 subcutaneously cell inoculated hamsters (86.7%) had right axillary lymph node metastasis while subcutaneously tissue implanted hamsters did not show any palpable lymph nodes. After necropsy, 10 of the 20 subcutaneously tissue implanted animals (50%) showed metastases in the lungs at the histopathological level. However, 16 of the 20 subcutaneously tissue implanted animals (80%) showed K-ras point mutation in the lung specimens. The lungs of the animals of the subcutaneous cell inoculation group did not show any metastases. No metastases were found in the liver or the pancreas in either group. CONCLUSION: This study suggests that homologous subcutaneous cell inoculation and subcutaneous tissue implantation models showed completely different patterns of metastasis. These models may aid further research to clarify the mechanisms of metastasis in pancreatic cancer.

Animals↗
PubMed1999

Electrophysiological evidence for sequential activation of multiple brain regions during the auditory selective attention process in humans.

In an attempt to examine dynamic involvement of multiple brain regions in the auditory selective attention process, negative difference wave (Nd) generators were assessed using a high-resolution EEG system (128ch) and scalp current density (SCD) analysis. Ten normal volunteers participated in the study. Event-related potentials were recorded during a selective attention task. Sequential SCD mappings revealed that current sinks were located in the bilateral temporal regions at 160 ms subsequent to the onset of stimuli, shifting the dipole orientation more tangentially to the scalp at around 220 ms. Moreover, a current sink was demonstrated in the midfrontal region at around 320 ms. These findings confirm that different cortical regions are sequentially involved in the auditory selective attention process.

Adult↗
PubMed1999

Molecular cloning of the gene for p85 that regulates the initiation of cytokinesis in Tetrahymena.

Tetrahymena p85 is localized to the presumptive division plane before division furrow formation; its molecular weight in SDS-polyacrylamide gel electrophoresis differs in wild-type and temperature-sensitive cell-division-arrest mutant cdaA1 cells. At the restrictive temperature, p85 localization and division furrow formation are not observed in cdaA1 cells. In this study, we purified p85 and cloned a wild-type p85 cDNA. The deduced amino acid sequence of p85 was composed mainly of two kinds of repeat sequences. One of these contained regions homologous to a calmodulin-binding site and a part of actin, and the other contained a region homologous to a part of a cdc2 kinase homologue. Moreover, we cloned a cDNA encoding the cdaA1 p85. There was no difference in the predicted amino acid sequences of wild-type and cdaA1 p85, suggesting that the difference in molecular weight between p85 in wild-type and mutant cells is caused by a disorder of posttranslational-modification mechanisms of p85 in the cdaA1 cell.

Actins↗
PubMed1999

Involvement of glutamate release in substance P-induced phase delays of suprachiasmatic neuron activity rhythm in vitro.

The suprachiasmatic nucleus (SCN) has been identified as a mammalian circadian rhythm clock. Treatment with substance P (SP) at zeitgeber time 13-14 produced phase delays of circadian rhythm in spontaneous neural activity in SCN neurons in vitro. SP-induced phase delays are blocked by treatment with not only SP receptor antagonist, spantide, but N-methyl-D-aspartate receptor antagonist, MK-801. In the biochemical experiment, we demonstrated that SP-induced glutamate release from the SCN slices was observed by the high-performance liquid chromatography assay. The present results suggest that glutamate release may be involved in SP-induced phase delays.

Animals↗
PubMed1999

Efficacy of the mobilization of peripheral blood stem cells by granulocyte colony-stimulating factor in pediatric donors.

The advantages/disadvantages of the use of peripheral blood stem cells (PBSCs) for allogeneic transplantation still need to be clarified, particularly in children. We compared the kinetics, efficacy, and safety of PBSC mobilization by granulocyte colony-stimulating factor (G-CSF) and collection by apheresis between healthy pediatric and adult donors. A total of 19 pediatric (median age, 6 years) and 25 adult healthy donors (median age, 37 years) were given 10 micro/kg/day of G-CSF for 5 consecutive days for PBSC mobilization, which were harvested by apheresis on days 5 and/or 6. All of the donors tolerated the whole procedures. Serum trough levels of G-CSF determined by ELISA were significantly lower in the 16 pediatric donors evaluated than in adults (n = 16) on days 3 and 4 (P < 0.05). Although the WBC counts on days 4 and 5 were significantly higher in adults than in children (P = 0.006 and 0.004, respectively), the numbers of circulating CD34+ cells/unit of blood were identical. The number of blood CD34+ cells collected per unit of blood processed was identical in both donor populations. We propose that PBSCs could be effectively mobilized and collected in small children so that they could be donors for adult patients.

Adolescent↗
PubMed1999

Enhanced expression of a nuclease gene in leaves of barley plants under salt stress.

We isolated a cDNA clone, Bnuc1, encoding a nuclease I from leaves of salt-stressed barley (Hordeum vulgare L. cv. Haruna-nijyo) by the differential display method. Northern blot analysis revealed that the transcript of Bnuc1 gene was increased dramatically in barley leaves under salt stress. The expression of Bnuc1 gene was also increased by exogenously applied abscisic acid (ABA) in leaves, but not by gibberellic acid (GA) during seed germination. Furthermore, Bnuc1 gene was expressed more in old leaves than in young leaves during both salt stress and natural senescence. Salt-inducible nuclease activity possibly corresponding to the Bnuc1 gene was detected, and was much higher in old leaves than in young leaves under salt stress.

Amino Acid Sequence↗
PubMed1999

Identification of the protein components of mismatch binding complexes in human cells using a gel-shift assay.

In eukaryotes, mismatch recognition is thought to be mediated by two heterodimers, hMutSalpha (hMSH2+hMSH6), which preferentially binds to base-base mismatches and hMutSbeta (hMSH2+hMSH3), which binds to insertion/deletion loops. We studied these mismatch binding activities in several human cell lines with a gel-shift assay using various mismatch oligonucleotides as substrates. Both hMutSalpha and hMutSbeta activities could be detected in various human cell lines. In cells with amplified copies of the hMSH3 gene, a large increase in hMutSbeta and a reduction in hMutSalpha were observed. To identify the composition of each mismatch binding complex, the protein-DNA complexes were transferred from gel-shift polyacrylamide gel to a polyvinylidene difluoride membrane and were subjected to immunoblot analysis with an enhanced chemiluminescence protein detection system. The results clearly demonstrated that hMutSalpha detected by the gel-shift assay was composed of hMSH2 and hMSH6, while hMutSbeta was composed of hMSH2 and hMSH3. Our data, therefore, support a model whereby formation of hMutSalpha and hMutSbeta is mutually regulated. Combination of a gel-shift assay with immunoblotting (shift-Western assay) proved to be a highly sensitive technique and should be useful for studying the interactions between DNA and binding proteins, including DNA mismatch recognition.

Base Pair Mismatch↗
PubMed1999

The prognostic significance of amplification and overexpression of c-met and c-erb B-2 in human gastric carcinomas.

BACKGROUND: The c-met and the c-erb B-2 protooncogenes belong to a family of tyrosine kinase growth factor receptors. Abnormalities of these oncogenes and protein products have been reported in several cancers. The authors investigated the correlation between clinical factors and amplification or overexpression of the c-met and/or c-erb B-2 gene in Japanese patients with gastric carcinoma patients, with a focus on prognostic significance. METHODS: Amplification and overexpression of c-met and c-erb B-2 were investigated retrospectively in 128 gastric carcinoma patients by using immunohistochemistry and Southern blot hybridization. Survival analysis was performed with the Kaplan-Meier test, and the log rank test was used for statistical analysis. RESULTS: Overexpression of c-met and c-erb B-2 was observed in 46.1% and 16.4% of gastric carcinoma cases, respectively. Gene amplification of c-met and c-erb B-2 was detected in 10.2% and 11.7% of gastric carcinoma cases, respectively. Amplification and overexpression of c-met were correlated significantly with depth of tumor invasion and lymph node metastasis, whereas amplification and overexpression of c-erb B-2 were correlated significantly with histologic type. The survival rate of patients with amplification and/or overexpression of c-met or c-erb B-2 was significantly poorer than that of patients with no amplification or overexpression. Multivariate analysis revealed that c-met overexpression and lymph node metastasis were independent prognostic factors. CONCLUSIONS: These data suggest that overexpression and/or gene amplification of c-met and c-erb B-2 may be prognostic factors in gastric carcinoma.

Adult↗
PubMed1999

Deamidation and isoaspartate formation in smeared tau in paired helical filaments. Unusual properties of the microtubule-binding domain of tau.

An extensive loss of a selected population of neurons in Alzheimer's disease is closely related to the formation of paired helical filaments (PHFs). The most striking characteristic of PHFs upon Western blotting is their smearing. According to a previously described protocol (Morishima-Kawashima, M., Hasegawa, M., Takio, K., Suzuki, M., Titani, K., and Ihara, Y. (1993) Neuron 10, 1151-1160), smeared tau was purified, and its peptide map was compared with that of soluble (normal) tau. A CNBr fragment from soluble tau (CN5; residues 251-419 according to the 441-residue isoform) containing the microtubule-binding domain migrated at 15 and 18 kDa on SDS-polyacrylamide gel electrophoresis, whereas that from smeared tau exhibited two larger, unusually broad bands at approximately 30 and approximately 45 kDa, presumably representing dimers and trimers of CN5. In the peptide map of smeared tau-derived CN5, distinct peaks eluting at unusual locations were noted. Amino acid sequence and mass spectrometric analyses revealed that these distinct peptides bear isoaspartate at Asn-381 and Asp-387. Because no unusual peptides other than aspartyl or isoaspartyl peptide were found in the digests of smeared tau-derived CN5, it is likely that site-specific deamidation and isoaspartate formation are involved in its dimerization and trimerization and thus in PHF formation in vivo.

Alzheimer Disease↗
PubMed1999

Effect of Ca2+-alginate gel dissolution on release of dextran with different molecular weights.

Release of macromolecular drug from calcium-alginate gel beads was investigated. Fluorescein isothiocyanate-labeled dextrans with molecular weight ranging from 9400 to 145000 were used as a model macromolecular drug. Dextran release was observed to be molecular weight-dependent, with the release pattern changing from pseudo first-order for dextran with 9400 molecular weight to sigmoidal for dextran with molecular weight of 145000. Release of a lower molecular weight dextran is mainly governed by the drug diffusion through the calcium-alginate gel matrix. With increasing dextran molecular weights, dextran release was strongly influenced by the dissolution of alginate matrix through the exchange of Ca2+ ions which act as a cross-linker. Rapid and complete release was achieved for dextran with 145000 molecular weight in a "burst" fashion showing a initial lag time. Minimal dextran release was observed in pseudo-gastric fluid at pH 1.2, while rapid dextran release within a narrow time range was achieved in simulated intestinal fluid at pH 6. 8. These results strongly suggest that calcium-alginate gel is a useful vehicle for pulsatile release of macromolecular drugs in oral drug delivery.

Alginates↗
PubMed1999

Role of lysine 39 of alanine racemase from Bacillus stearothermophilus that binds pyridoxal 5'-phosphate. Chemical rescue studies of Lys39 --> Ala mutant.

The lysine residue binding with the cofactor pyridoxal 5'-phosphate (PLP) plays an important role in catalysis, such as in the transaldimination and abstraction of alpha-hydrogen from a substrate amino acid in PLP-dependent enzymes. We studied the role of Lys39 of alanine racemase (EC 5.1.1.1) from Bacillus stearothermophilus, the PLP-binding residue of the enzyme, by replacing it site-specifically with alanine and characterizing the resultant K39A mutant enzyme. The mutant enzyme turned out to be inherently inactive, but gained an activity as high as about 0.1% of that of the wild-type enzyme upon addition of 0.2 M methylamine. The amine-assisted activity of the mutant enzyme depended on the pKa values and molecular volumes of the alkylamines used. A strong kinetic isotope effect was observed when alpha-deuterated D-alanine was used as a substrate in the methylamine-assisted reaction, but little effect was observed using its antipode. In marked contrast, only L-enantiomer of alanine showed a solvent isotope effect in deuterium oxide in the methylamine-assisted reaction. These results suggest that methylamine serves as a base not only to abstract the alpha-hydrogen from D-alanine but also to transfer a proton from water to the alpha-position of the deprotonated (achiral) intermediate to form D-alanine. Therefore, the exogenous amine can be regarded as a functional group fully representing Lys39 of the wild-type enzyme. Lys39 of the wild-type enzyme probably acts as the base catalyst specific to the D-enantiomer of alanine. Another residue specific to the L-enantiomer in the wild-type enzyme is kept intact in the K39A mutant.

Alanine↗
PubMed1999

Growth inhibition of human pancreatic cancer cell lines by anti-sense oligonucleotides specific to mutated K-ras genes.

About 90% of human pancreatic cancers carry K-ras point mutation, which may play an important role in tumorigenesis. We investigated the inhibitory effects of anti-sense oligonucleotides targeting K-ras point mutation on the growth of cultured human pancreatic cancer cells. Eight human pancreatic cancer cell lines were screened for K-ras codon 12 point mutations by PCR-RFLP analysis and direct sequencing. Then, 3 cell lines with the major types of K-ras point mutation, i.e.,HuP-T1, HuP-T3 and PANC-1, and 1 without mutation, BxPC-3, were used for the experiments. Seventeen mer anti-sense oligonucleotides were designed, targeting the point mutation of K-ras codon 12, and transfected into the cells by the liposome-mediated method. Cell-growth activities were estimated by MTT assay. Levels of K-ras mRNA expression were determined using quantitative RT-PCR, and K-ras p21 protein synthesis was evaluated with Western blotting. Mutation-matched anti-sense oligonucleotides effectively inhibited the growth of these pancreatic cancer cell lines, except for BxPC-3, by suppressing K-ras mRNA expression and K-ras p21 protein synthesis. Moreover, mutation-matched anti-sense oligonucleotides showed stronger anti-proliferative effects than did mutation-mismatched ones. Our results suggest that anti-sense therapy specific to point mutations of K-ras mRNA is a practical approach to selective suppression of tumor growth, with little effect on normal cells.

Cell Division↗
PubMed1999

Clinical implications of serum anti-p53 antibodies for patients with gastric carcinoma.

BACKGROUND: Mutations of p53 can lead to the production of anti-p53 antibodies in sera of cancer patients. Before this study, the value of preoperative serum anti-p53 antibodies in determining the prognoses of patients with gastric carcinoma had yet to be determined. METHODS: The authors used a highly specific enzyme-linked immunosorbent assay (ELISA) kit (Pharma Cell, France) to determine the preoperative presence of serum anti-p53 antibodies in 120 patients with gastric carcinoma. The relation between the positivity of serum anti-p53 antibodies and p53 abnormal staining of gastric carcinoma tissues was examined. Clinicopathologic characteristics and prognoses of these patients were given attention. RESULTS: Anti-p53 antibodies were detected in 19.2%(23 of 120) of these patients with gastric carcinoma. Among those who were positive for anti-p53 antibodies, female patients were predominant, the depth of invasion was greater, and liver metastasis was present, as compared with those who were negative for anti-p53 antibodies. Regarding other factors, there were no differences between those who were positive or negative for anti-p53 antibodies. Gastric carcinoma tissues had a 60.9% (14 of 23) positivity rate of p53 staining with anti-p53 antibodies and a 33.0% (32 of 97) negativity rate, and this difference was statistically significant (P < 0.05). The survival time of patients with anti-p53 antibodies in their sera was shorter than that of subjects with sera negative for anti-p53 antibodies (P < 0.05). The presence of anti-p53 antibodies was not an independent prognostic factor in multivariate analysis. CONCLUSIONS: Serum assay of anti-p53 antibodies is a rapid and readily facilitated test for predicting tumor advancement, depth of invasion, and liver metastasis, and it will show a poorer prognosis for surgically treated patients with gastric carcinoma.

Antibodies, Monoclonal↗
PubMed1999

Improvement in CNS protective treatment in non-high-risk childhood acute lymphoblastic leukemia: report from the Japanese Children's Cancer and Leukemia Study Group.

BACKGROUND: Prevention of central nervous system (CNS) leukemia by early introduction of therapy to this sanctuary site is an essential component of modern treatment strategy for acute lymphoblastic leukemia (ALL). However, the optimal form of preventive CNS therapy remains debatable. PROCEDURE: To address this issue, we evaluated the efficacy of CNS preventive therapy for 572 children with ALL who achieved complete remission in the Children's Cancer and Leukemia Study Group (CCLSG) ALL874 (1987-1990) and ALL911 (1991-1993) studies. They received risk-directed therapy based on age and leukocyte count. In the ALL 874 study, the non-high-risk (low-risk [LR] + intermediate risk [IR]) patients were randomly assigned to the conventional cranial irradiation (CRT) regimen (L874A and I874A) and the high-dose methotrexate (HDMTX) regimen without CRT (L874B and I874B). The former patients received 18-Gy CRT plus 3 doses of intrathecal (i.t.) MTX and the latter patients received 3 courses of HDMTX at 2 g/m2 plus 13 doses of ITMTX (L874B) or 4 courses of HDMTX at 4.5 g/m2 plus 1 dose of ITMTX (I874B). RESULTS: The 7-year probabilities (+/- SE) of CNS relapse-free survival were 97.3% +/- 2.6% (L874A, n = 41) vs. 90.3% +/- 5.3% (L874B, n = 39) (P = 0.25) in the LR patients, and 100% (I874A, n = 55) vs. 78.5% +/- 6.5% (I874B, n = 54) (P = 0.002) in the IR patients. The corresponding disease-free survival (DFS) rates were 79.4% +/- 6.5% vs. 74.4% +/- 7.3% (P = 0.62) in the LR group and 63.3% +/- 6.8% vs. 58.3% +/- 7.2% (P = 0.66) in the IR group. Thus, the HDMTX regimen could not provide better protection of CNS relapse as compared with the CRT regimen, although their overall efficacy was not significantly different. In the ALL 911 study, intensive systemic chemotherapy with extended i,t, injections of MTX plus cytarabine achieved a high CNS relapse-free survival (98% +/- 1.9% at 7 years) and a favorable DFS (85.5% +/- 5% at 7 years) in the IR patients. The patients in the high-risk (HR) group in both ALL874 and ALL911 studies received the 18-Gy or 24-Gy CRT with intensive systemic chemotherapy. Their 7-year probabilities of CNS relapse-free survival ranged from 88% to 95%, among which the T-ALL patients had a risk of CNS leukemia, which was 3-4 times higher compared with B-precursor ALL patients. CONCLUSIONS: These results indicate that long-term intrathecal CNS prophylaxis as well as appropriate systemic therapy for the non-high-risk patients can provide protection against CNS relapse equivalent to that provided by cranial irradiation.

Adolescent↗
PubMed1999

Rapid recovery from anorexia nervosa after a life-threatening episode with severe thrombocytopenia: report of three cases.

We report the cases of three patients with anorexia nervosa (AN) who each recovered rapidly after experiencing a life-threatening episode with severe thrombocytopenia. All three cases were the typical restricting-type of AN, occurring in adolescence. They refused to be admitted to a hospital until their general condition had been severely deteriorated. Their lowest platelet counts were 2.9, 4.6, and 2.3 x 10(4)/mm3, respectively. Apparent hemorrhagic tendencies, such as purpura, gingival and nasal bleeding, and gastrointestinal bleeding were observed. The bone marrow examination showed apparent hypoplasia in two patients. No evidence of disseminated intravascular coagulation or autoantibody to platelets was detected. The platelet counts recovered rapidly by water and nutritional supplementation. The recovery from the AN itself was excellent in all three patients without specific psychotherapy.

Adolescent↗
PubMed1999

Eating problems in female Japanese high school students: a prevalence study.

OBJECTIVES: This study was conducted to estimate the prevalence of eating problems and to identify factors associated with the eating problems among Japanese high school girls. METHODS: This study used a cross-sectional design. The study population was 3,032 female students attending three high schools in Fukushima, Japan. The 26-item version of the Eating Attitudes Test (EAT-26) was administered to assess eating problems. Inquiries were also made regarding possible risk factors for the eating problems. RESULTS: Of the 2,685 subjects, 5.4% had a total EAT-26 score above the cutoff point (20 < or =). Logistic regression analysis demonstrated that older age, higher body mass index, a distorted body image, obsessive-compulsive tendency, and some familial issues were independently related to the eating problems. DISCUSSION: The prevalence of eating problems in the Japanese female population was low compared with reports from Western countries. In addition, distorted body image was suggested to have the greatest influence on eating problems.

Adolescent↗
PubMed1999

Functional B-cell response in intrahepatic lymphoid follicles in chronic hepatitis C.

Intrahepatic lymphoid follicle (ILF) formation is one of the most characteristic and commonly observed histological features in patients with chronic hepatitis C. However, little is known regarding whether follicles in the liver belong to functional lymphoid tissues, where B cells are activated, differentiated, and proliferated, or if the lymphocytes are merely infiltrated after recruitment from the secondary lymphoid organs. To ascertain this possibility, we examined the expression of markers for B-cell activation, differentiation, and proliferation in ILFs in patients with chronic hepatitis C using surgically resected specimens, and compared them with specimens of perihepatic lymph nodes by an immunohistochemical technique. Germinal center (GC) formation in the ILFs was frequently found in HCV-positive cases. The distribution of immunoglobulin M (IgM)-, IgD-, and IgG-positive cells and the expression patterns of Ki-67, CD23, or bcl-2 and bcl-6 gene products in the follicles with GC formation in the liver of patients with chronic hepatitis C were similar to those of lymph nodes, indicating that B cells are activated, proliferated, and differentiated in the ILFs with GC formation in patients with chronic hepatitis C. Oligoclonal expansion of B cells in the livers with ILFs was confirmed by an analysis of immunoglobulin heavy chain (IgH) gene rearrangement determined by polymerase chain reaction (PCR). These data strongly suggest that ILFs with GC formation, which are frequently found in patients with chronic hepatitis C, may functionally be the same as those found in lymph nodes with respect to B-cell expansion and maturation.

Adult↗