Biomedical subjects

M Matsushima

Publications and source records attributed to M Matsushima.

At least 55 records · Page 3Linked to original sources

PubMed1999

Cell type-specific requirement of the MAPK pathway for the growth factor action of gastrin.

Gastrin (G17) has a CCKB receptor-mediated growth-promoting effect on the AR42J rat acinar cell line that is linked to induction of both mitogen-activated protein kinase (MAPK) and c-fos gene expression. We investigated the mechanisms that regulate the growth factor action of G17 on the rat pituitary adenoma cell line GH3. Both AR42J and GH3 cells displayed equal levels of CCKB receptor expression and similar binding kinetics of 125I-labeled G17. G17 stimulation of cell proliferation was identical in both cell lines. G17 stimulation of GH3 cell proliferation was completely blocked by the CCKB receptor antagonist D2 but not by the MEK inhibitor PD-98059 or the protein kinase C inhibitor GF-109203X, which completely inhibited G17 induction of AR42J cell proliferation. G17 induced a c-fos SRE-luciferase reporter gene plasmid more than fourfold in the AR42J cells, whereas it had no effect in the GH3 cells. In contrast to what we observed in the AR42J cells, G17 failed to stimulate MAPK activation and Shc tyrosyl phosphorylation and association with the adapter protein Grb2. Epidermal growth factor induced the MAPK pathway in the GH3 cells, demonstrating the integrity of this signaling system. G17 induced Ca2+ mobilization in both the GH3 and AR42J cells. The calmodulin inhibitor N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide inhibited AR42J cell proliferation by 20%, whereas it completely blocked G17 induction of GH3 cell growth. The Ca2+ ionophore ionomycin stimulated GH3 cell proliferation to a level similar to that observed in response to G17, but it had no effect on AR42J cell proliferation. Thus there are cell type specific differences in the requirement of the MAPK pathway for the growth factor action of G17. Whereas in the AR42J cells G17 stimulates cell growth through activation of MAPK and c-fos gene expression, in the GH3 cells, G17 fails to activate MAPK, and it induces cell proliferation through Ca2+-dependent signaling pathways. Furthermore, induction of Ca2+ mobilization in the AR42J cells appears not to be sufficient to sustain cell proliferation.

Adaptor Proteins, Signal Transducing↗

PubMed1999

Glycine-extended gastrin regulates HEK cell growth.

Posttranslational processing of progastrin to a carboxy terminally amidated form (G-NH(2)) is essential for its effect on gastric acid secretion and other biological effects mediated by gastrin/CCK-B receptors. The immediate biosynthetic precursor of G-NH(2), glycine-extended gastrin (G-Gly), does not stimulate gastric acid secretion at physiological concentrations but is found in high concentrations during development. G-NH(2) and G-Gly have potent growth stimulatory effects on gastrointestinal tissues, and G-NH(2) can stimulate proliferation of human kidney cells. Thus we sought to explore the actions of G-NH(2) and G-Gly on the human embryonic kidney cell line HEK 293. HEK 293 cells showed specific binding sites for (125)I-labeled Leu(15)-G17-NH(2) and (125)I-Leu(15)-G(2-17)-Gly. Both G-NH(2) and G-Gly induced a dose-dependent increase in [(3)H]thymidine incorporation, and both peptides together significantly increased [(3)H]thymidine incorporation above the level of either peptide alone. G-NH(2) and G-Gly were detected by radioimmunoassay in serum-free conditioned media. Antibodies directed against G-NH(2) and G-Gly lead to a significant reduction in [(3)H]thymidine incorporation. G-NH(2) but not G-Gly increased intracellular Ca(2+) concentration. We conclude that G-NH(2) and G-Gly act cooperatively via distinct receptors to stimulate the growth of a nongastrointestinal cell line (HEK 293) in an autocrine fashion.

Binding Sites↗

PubMed1999

Allelic loss at 1p34, 13q12, 17p13.3, and 17q21.1 correlates with poor postoperative prognosis in breast cancer.

Allelic losses of tumor suppressor genes (TSGs), or the chromosomal regions harboring them, in tumor DNA may become useful postoperative prognostic indicators. To examine whether specific allelic losses might correlate with postoperative survival in a 5-year prospective follow-up, we tested tumors from a cohort of 264 breast cancer patients for allelic losses of 18 microsatellite markers representing either a known TSG or a region where genetic alterations are frequent in breast tumors. Patients whose tumors had lost an allele at 1p34, 13q12, 17p13.3, or 17q21.1 had significantly higher risks of postoperative mortality than those whose tumors retained both alleles at those loci (at 1p34, a 5-year mortality rate of 29% among patients with losses vs. 7% with retentions, P = 0. 0008; at 13q12, 31% vs. 10%, P = 0.0062; at 17p13.3, 24% vs. 13%, P = 0.026; and at 17q21.1, 31% vs. 13%, P = 0.0047). Furthermore, combined losses at 13q12 and 17p13.3 increased the predicted postoperative mortality risks by a factor of 9.6 (5-year mortality rate of 42% vs. 5% with retentions, P = 0.0001), and combined losses at 1p34 and 17p13.3 raised the predicted postoperative mortality risks by a factor of 8.6 (27% vs. 3%, P = 0.0064). We conclude that allelic losses at these loci can serve as negative prognostic indicators to guide postoperative management of patients. Genes Chromosomes Cancer 26:134-141, 1999.

Adult↗

PubMed1998

Evolution of strategies in the three-person iterated prisoner's dilemma game.

A three-person iterated prisoner's dilemma (3p-IPD) game is studied. The present simulation uses a lattice model of finite state automata. In particular, the similarity between a noisy 2p- and a noiseless 3p-IPD game is discussed. It is known that, due to the noise, Tit for Tat loses its robustness and is taken over by more complex strategies in a noiseless IPD game. But in the 3p-IPD game, even without noise, Tit for Tat loses its robustness and is also taken over by more complex strategies. It is found that similar strategies take over Tit for Tat in both situations. We thus remark that the role of noise in the two person game is replaced by the third player in the three-person game. As a result, the strategies diversify in both the noisy 2p- and the quiet 3p- IPD game. It is also found that game strategies in an automaton form can be understood as a combination of defensive and offensive substructures. A recognition of these substructures enables us to study the mechanism of robustness in the strategies of the 3p-IPD game.

Animals↗

PubMed1998

No evidence of correlation between polymorphism at codon 72 of p53 and risk of cervical cancer in Japanese patients with human papillomavirus 16/18 infection.

Human papillomavirus (HPV)-16 and -18 encode E6 oncoprotein, which binds to and induces degradation of the tumor suppressor protein p53. A common polymorphism of p53, encoding either proline or arginine at position 72, affects the susceptibility of p53 to E6-mediated degradation in vivo; Caucasian women homozygous for arginine 72 reportedly are about seven times more susceptible to HPV-associated carcinoma of the cervix than heterozygotes. To examine whether arginine 72 could be a risk factor for HPV-associated cervical carcinomas in the Japanese population, we used the same PCR-based assay to analyze p53 genotypes of HPV-positive invasive cervical carcinomas from 103 Japanese women versus 110 control samples. Inasmuch as we detected no significant difference in the frequencies of proline or arginine alleles between the two groups, p53 polymorphism at residue 72 does not seem to be involved in the development of HPV-associated cervical carcinomas in women of Japanese ethnicity.

Codon↗

PubMed1998

Structural and functional effect of Trp-62-->Gly and Asp-101-->Gly substitutions on substrate-binding modes of mutant hen egg-white lysozymes.

In order to clarify the structural role of subsite B of hen egg-white lysozyme in hydrolytic activity towards a carbohydrate substrate, we analysed the structures of Trp-62-->Gly and Asp-101-->Gly mutant hen lysozymes, which have no side chain at positions 62 or 101, complexed with a substrate analogue, (N-acetyl-d-glucosamine)3 [(GlcNAc)3], using X-ray crystallography. The overall protein structures in the mutant lysozyme complexes were almost identical to those in the wild type. In the crystals of all the mutant complexes, the (GlcNAc)3 molecule, which is an inhibitor of wild-type lysozyme, had no inhibitory effect, but was hydrolysed as a substrate. One of the products, (GlcNAc)2, the reducing end of which is an alpha-anomer, was bound in an unproductive binding mode, protruding from the active-site cleft, and was able to act as an inhibitor. Hydrolysis of the synthetic substrate by the mutants occurred in a beta-anomer-retaining manner, and so the alpha-anomer product was converted from the beta-anomer product. Thus the interactions of Asp-101 and Trp-62 in subsite B are not essential for the catalytic mechanism, but co-operatively enhance the affinity of the substrate in the productive binding mode, other than the inhibitor in the unproductive mode.

Amino Acid Substitution↗

PubMed1998

Complete genomic structure DNA polymorphisms, and alternative splicing of the human AF-6 gene.

In our previous work, detailed deletion mapping of ovarian cancers indicated that a 300-kb region of chromosome 6q27 was likely to contain one or more putative tumor suppressor genes associated with development of this type of cancer. DNA sequencing in the region disclosed the presence of AF-6, a gene that had been identified as the ALL-1 fusion partner involved in acute myeloid leukemias with t(6;11)(q27;q23) translocations. In the work reported here, we determined the complete genomic sequence of the AF-6 gene, including exon-intron boundaries, and found six DNA polymorphisms. One of them, an insertion/deletion polymorphism, determined the presence or absence of seven amino acids in the AF-6 product. We also identified two alternatively spliced forms of the gene; the two novel transcripts would encode additional C-terminal peptides in comparison to the reported protein. Sequencing of seven cosmid clones that covered the entire gene revealed 32 exons (not including one exon involved in the insertion/deletion polymorphism), spanning approximately 140 kb of genomic DNA. These results may contribute to an understanding of the mechanism causing chromosomal translocations in leukemic cells.

Alternative Splicing↗

PubMed1998

Structural analysis of mutant hen egg-white lysozyme preferring a minor binding mode.

Trp62 in hen egg-white lysozyme has general features observed in protein-carbohydrate interactions, a stacking interaction toward nonpolar surface of the substrate sugar residue B and a hydrogen bonding network with the residue C. Our previous report (I. Kumagai, K. Maenaka, F. Sunada, S. Takeda, K. Miura, Eur. J. Biochem. 212 (1993) 151-156.) showed that the substitution of Trp62 changed the substrate binding modes; especially, the Trp62His mutant exhibited the drastic change of the binding mode and preferred to a minor binding mode of the wild-type enzyme. In order to clarify the relationship between functional and structural changes of the Trp62His mutant, we analyzed the structure of the Trp62His mutant hen lysozyme complexed with the substrate analogue, (GlcNAc)3, by X-ray crystallography. The overall protein structure in the mutant lysozyme complex was almost identical to that in the wild-type. His62 shared almost the same plane as the indole ring of Trp62 of the wild-type. Although the (GlcNAc)3 molecule which is an inhibitor against the wild-type lysozyme was cocrystallized, the Trp62His mutant did not put it in the sites A-B-C but hydrolyzed it as a substrate. One of the products, (GlcNAc)2, whose reducing end is alpha-anomer, was bound in another binding mode sticking out from the active-site cleft. The hydrolytic activity against the synthetic substrate showed that the mutant was a beta-anomer retaining enzyme, so the alpha-anomer product was converted from the beta-anomer product. Therefore, the Trp62His mutant showed the remarkable change of the substrate binding modes not by alteration of the catalytic system but possibly by subtle rearrangement of general features of protein-carbohydrate interactions between His62 and the sugar residues B and C.

Animals↗

PubMed1998

Expression of basic fibroblast growth factor and its receptor mRNA in retinal tissue following ischemic injury in the rat.

BACKGROUND: Our purpose was to determine the time-dependent changes of expression of basic fibroblast growth factor (bFGF) and its receptor in pressure-induced retinal ischemia in rats. METHODS: Retinal ischemia was induced in Wistar rats by increasing the intraocular pressure to 110 mmHg for 45 min by cannulation into the eyes. At the end of the ischemic period, reperfusion of the retinal vasculature was confirmed. Localization of bFGF and FGF receptor-1 (FGF-R) mRNAs were evaluated by in situ hybridization at various times after reperfusion. The reverse-transcription polymerase chain reaction (RT-PCR) method was used to detect the expression of bFGF and FGF-R mRNA in the sensory retina. RESULTS: In normal sensory retina, bFGF and FGF-R mRNAs were observed in the ganglion cell layer and inner nuclear layer. bFGF gene expression in the sensory retina increased within 24 h, particularly at 6-12 h. FGF-R gene expression increased earlier than that of bFGF. By RT-PCR, expression of bFGF gene reached a peak at 6-24 h, and FGF-R reached a peak at 3-12 h. These RT-PCR results are comparable to those of in situ hybridization. CONCLUSIONS: These results demonstrate that transient retinal ischemia leads to the induction of bFGF mRNA synthesis, and suggest that bFGF has a protective role, e.g., a defense mechanism for the sensory retina.

Animals↗

PubMed1998

VNTR (variable number of tandem repeat) sequences as transcriptional, translational, or functional regulators.

VNTR (variable number of tandem repeat) markers, also called single-copy minisatellites, were originally isolated from human DNA as highly informative restriction fragment length polymorphisms for mapping purposes. Evidence has lately emerged that some VNTR sequences play significant roles in the regulation of transcription, and that some may also influence the translational efficiency or stability of mRNA, or modify the activity of proteins by altering their structure. Some apparent associations of VNTR sequences with personality traits or with susceptibility to diseases have strengthened the likelihood that these tandemly-repeated genomic elements are of physiological and biological importance. In this review, we summarize recent progress in efforts to clarify mechanisms involving VNTR sequences.

Gene Expression Regulation↗

PubMed1998

Transient neurologic syndrome in one thousand forty-five patients after 3% lidocaine spinal anesthesia.

UNLABELLED: Recent reports have discussed the potential risk of transient radicular irritation (TRI) after spinal anesthesia with lidocaine. Because we have not encountered such neurologic sequelae with the high incidence reported, we prospectively examined the incidence of TRI after spinal anesthesia with lidocaine. One thousand forty-five adult patients (aged 47 +/- 15 yr) receiving spinal anesthesia with 3% hyperbaric lidocaine (1.0-1.5 mL) for anorectal surgery were consecutively studied. After the induction of spinal anesthesia, all patients were placed in the prone position for surgery. Patients were evaluated for neurologic symptoms in the buttocks, thighs, or lower extremities using a checklist to standardize data collection. Although there was no complaint of neurologic symptoms on Postoperative Day (POD) 1, four patients (0.4%) reported aching, hypesthesia, numbness, or dull pain of both lower extremities and buttocks by the morning of POD 3. In three patients, the symptoms resolved without any treatment by POD 5, whereas in one patient, numbness of the lower extremities lasted until POD 7. We conclude that a combination of lidocaine with surgical position or leg manipulation during surgery might be a major contributing factor in the development of transient neurologic syndrome. IMPLICATIONS: Recent studies have reported the potential risk of transient neurologic syndrome after lidocaine spinal anesthesia. The current study reports a low incidence of such sequelae in 1045 patients undergoing anorectal surgery during the prone position. Other factors, such as surgical position, may be important in the development of this syndrome.

Adult↗

PubMed1998

Multiplex mutation screening of the BRCA1 gene in 1000 Japanese breast cancers.

To detect BRCA1 mutations in Japanese breast cancer patients, we screened 1,000 unselected primary cancers for mutations in exon 11, which accounts for 61% of the entire BRCA1 coding sequence. Using a method based on multiplex single-strand conformational polymorphism (SSCP) analysis of multiple restriction fragments generated by restriction-enzyme digestion of amplified DNA, we identified eight mutations. All eight were germline mutations; four of them were non-sense mutations or small deletions resulting in premature stop codons, and the other four were missense mutations. The Japanese carriers of these mutant BRCA1 alleles had developed breast cancers at ages ranging from 45 to 62, five of them bilaterally.

Aged↗

PubMed1998

Identification, genomic organization, and alternative splicing of KNSL3, a novel human gene encoding a kinesin-like protein.

Proteins of the kinesin superfamily are microtubule-dependent molecular motors that play important roles in organelle transport and cell division. Through genomic sequencing and use of the RT-PCR technique, we have identified and characterized KNSL3 (kinesin-like 3), a novel member of the kinesin-like protein family in humans. We determined its genomic organization and detected four alternatively spliced transcripts. KNSL3 was expressed ubiquitously, but sizes and relative amounts of the major products were different in each of the tissues examined. Alternative splicing, along with the multiplicity of genes in the molecular family that includes KNSL3, produce diversity among the C-terminal ends of kinesins. These observations may contribute to an understanding of the specificity of different kinesins with respect to organelle binding.

Alternative Splicing↗

PubMed1998

Puberty, IDDM, and death in Japan. Diabetes Epidemiology Research International Study Group.

OBJECTIVE: To investigate the impact of age at onset on the prognosis of childhood IDDM in Japan. RESEARCH DESIGN AND METHODS: The study population consisted of 987 prepubertal-onset and 345 pubertal-onset IDDM cases who were registered by two nationwide surveys and who were diagnosed between 1965 and 1979. The living status was identified as of 1 January 1990 with the ascertainment rate of 96.5%. Prognosis was evaluated by cause- and age-specific mortality rates and life-table analyses; in addition, an attained-age-matched case-control study was analyzed by conditional logistic regression. RESULTS: The adjusted mortality rates per 100,000 person-years for the pubertal-onset cases were significantly higher than those of the prepubertal-onset cases (835 [95% CI 573-1,168] vs. 391 [283-526]). Life-table analyses revealed that the survival rate of the pubertal-onset cases was lower than that of the prepubertal-onset cases for each observation period. Life-table analyses after the age of 15 years indicated that the prognosis of pubertal cases was almost the same as that of the prepubertal cases despite having a shorter duration of diabetes. The attained-age-matched case-control study indicated that the pubertal onset was a potent determinant of death. CONCLUSIONS: The prognosis of pubertal-onset IDDM was considerably poorer than that of prepubertal-onset IDDM.

Accidents↗

PubMed1998

Isolation, mapping, and functional analysis of a novel human cDNA (BNIP3L) encoding a protein homologous to human NIP3.

We have isolated a novel cDNA that encodes a product showing significant sequence homology (56% identity) to human NIP3, a protein thought to interact with adenovirus E1B19kD and human BCL2 proteins. This cDNA contains an open reading frame of 657 nucleotides encoding a 219 amino acid polypeptide. The gene, designated BNIP3L, was expressed in all 16 normal human tissues examined; we mapped it to chromosome band 8p21 by fluorescence in situ hybridization. Introduction of the BNIP3L gene into six different cancer-cell lines caused significant growth suppression in each of them, while no such effect occurred when the antisense cDNA or the vector DNA was transfected, indicating that BNIP3L may function as a tumor suppressor.

Amino Acid Sequence↗

PubMed1998

[Short-term intravesical instillation of pirarubicin (THP) in prophylactic treatment after transurethral resection of superficial bladder tumor].

We conducted a prospective randomized controlled study on the prophylactic effects of short-term intravesical instillation of pirarubicin (THP) against recurrence to determine the effective administration schedule. All patients gave their informed consent. The subjects included bladder cancer patients who had pTa or pT1, and G1 or G2 cancer, and became tumor-free after transurethral resection of the bladder tumor (TUR-BT). After dissolving 30 mg of THP into 5 ml of distilled water, physiological saline was added to adjust the total volume to 50 ml, which was then instilled into the bladder, and was retained for 5 minutes. The schedule of instillation was for daily for 7 consecutive days from the day of TUR-BT and subsequently once a week for 10 weeks, 17 times in total for Group I, and once every two weeks for 6 months (12 times) starting 2 weeks after TUR and subsequently once a month until one year had passed after surgery (6 times), 18 times in total for Group II. The total number of cases was 69 (36 in Group I, 33 in Group II). The tumor-free ratios determined by the Kaplan-Meier analysis were 93.9% in Group I and 72.7% in Group II for one year, and 86.8% in Group I and 59.5% in Group II for two years. There was a statistically significant difference in the tumor-free ratios between the two groups by the generalized Wilcoxon test and the Log rank test (p = 0.0145 and 0.0107, respectively). Multivariated analysis using Cox's comparison hazard model produced p-values of 0.0002, 0.0007, 0.0009 and 0.0040 in the order of therapeutic mode, initial onset/recurrence, stage and number of tumor. Adverse events that forced discontinuation of the therapy for a while occurred in 4.3%. These results demonstrated that short-term intensive intravesical instillation of THP immediately after TUR-BT was a safe and effective therapy.

Administration, Intravesical↗

PubMed1997

Novel hepatitis C virus protease inhibitors: thiazolidine derivatives.

This study evaluated the inhibitory effects of thiazolidine derivatives on hepatitis C virus (HCV) protease and other human serine proteases. The inhibition efficacy was tested with a reversed-phase high-performance liquid chromatography (HPLC) assay system using a NS3-NS4A fusion protein as the HCV protease and a synthetic peptide substrate that mimics the NS5A-5B junction. Nine thiazolidine derivatives showed more than 50% inhibition at 50 microg/ml. The most potent derivative was RD4-6250, with 50% inhibition at a concentration of 2.3 microg/ml; this concentration was lower than those of other protease inhibitors reported previously. The most selective derivative was RD4-6205, with 50% inhibition at a concentration of 6.4 microg/ml, a lower concentration than those on other serine proteases (chymotrypsin, trypsin, plasmin, and elastase). These results suggest that the RD4-6205 skeleton is an important structure for inhibitory activity on the HCV protease NS3-NS4A.

Enzyme Inhibitors↗

PubMed1997

Surface of bacteriorhodopsin revealed by high-resolution electron crystallography.

Bacteriorhodopsin is a transmembrane protein that uses light energy, absorbed by its chromophore retinal, to pump protons from the cytoplasm of bacteria such as Halobacterium salinarium into the extracellular space. It is made up of seven alpha-helices, and in the bacterium forms natural, two-dimensional crystals called purple membranes. We have analysed these crystals by electron cryo-microscopy to obtain images of bacteriorhodopsin at 3.0 A resolution. The structure covers nearly all 248 amino acids, including loops outside the membrane, and reveals the distribution of charged residues on both sides of the membrane surface. In addition, analysis of the electron-potential map produced by this method allows the determination of the charge status of these residues. On the extracellular side, four glutamate residues surround the entrance to the proton channel, whereas on the cytoplasmic side, four aspartic acids occur in a plane at the boundary of the hydrophobic-hydrophilic interface. The negative charges produced by these aspartate residues is encircled by areas of positive charge that may facilitate accumulation and lateral movement of protons on this surface.

Bacteriorhodopsins↗