Biomedical subjects

M Kasai

Publications and source records attributed to M Kasai.

At least 145 records · Page 8Linked to original sources

PubMed1998

Formation of a major DNA adduct of the mitomycin metabolite 2,7-diaminomitosene in EMT6 mouse mammary tumor cells treated with mitomycin C.

Treatment of EMT6 mouse mammary tumor cells with [3H]mitomycin C (MC) results in the formation of six major DNA adducts, as described earlier using an HPLC assay of 3H-labeled products of enzymatic hydrolysis of DNA isolated from MC-treated cells. Four of these adducts were identified as monofunctional and bifunctional guanine-N2 adducts in the minor groove of DNA. In order to establish relationships between individual types of MC-DNA adducts and biological responses it is necessary to identify all of the adducts formed in cells. To this end we have now identified a predominant, previously unknown adduct formed in MC-treated EMT6 cells as a derivative not of MC, but of 2,7-diaminomitosene (2,7-DAM), the major bioreductive metabolite of MC. Rigorous proof demonstrates that it is a DNA major groove, guanine-N7 adduct of 2,7-DAM, linked at C-10 to DNA. The adduct is relatively stable at ambient temperature, but is readily depurinated upon heating. Its isolation from MC-treated cells indicates that MC is reductively metabolized to 2,7-DAM, which then undergoes further reductive activation to alkylate DNA, along with the parent MC. Low MC:DNA ratios were identified as a critical factor promoting 2,7-DAM adduct formation in an in vitro model calf thymus DNA/ MC/reductase model system, as well as in MC-treated EMT6 cells. The 2,7-DAM-guanine-N7 DNA adduct appears to be relatively noncytotoxic, as indicated by the dramatically lower cytotoxicity of 2,7-DAM in comparison with MC in EMT6 cells. Like MC, 2,7-DAM exhibited slightly greater cytotoxicity to cells treated under hypoxic as compared to aerobic conditions. However, 2,7-DAM was markedly less cytotoxic than MC under both aerobic and hypoxic conditions. Thus, metabolic reduction of MC to 2,7-DAM represents a detoxification process. The differential effects of MC-DNA and 2,7-DAM-DNA adducts support the concept that specific structural features of the DNA damage may play a critical role in the cytotoxic response to a DNA-targeted chemotherapeutic agent.

Animals↗

PubMed1998

Psychological and physiological changes during isolation and confinement: I. Group dynamics and member interaction.

The isolated and confined environment of a laboratory setting was used as one of the analogous environments of long-term manned spaceflight. This setting has more advantages than the natural one in that the behavioral variables can be easily controlled. Thus, a hyperbaric chamber was used in this experiment. The subjects were five men from 22- to 26-years old who had never met before the experiment. Over a total 5-day isolation and confinement period as well as 2-day pre- and post-isolation periods, the subjects were given different kinds of physiological and psychological tasks. During these periods their behavior and communications were recorded in order to observe when the formation of a group begins and how it forms and functions. The formation of a sub-group of three men was observed on the third day of isolation, showing strong reciprocal connections among these subjects who developed bonds that were quite stable. It is, however, not known how this sub-group functioned within the large group, since the experimental period was possibly too short.

Adaptation, Physiological↗

PubMed1998

Allogeneic peripheral blood stem cell transplantation for standard-risk leukemia. A multicenter pilot study: Japanese experience. Japan Blood Cell Transplantation Study Group.

A multicenter phase II study of allogeneic peripheral blood stem cell transplantation (allo-PBSCT) was conducted. Twenty-two patients (median age, 36 years) with standard-risk leukemia were transplanted with G-CSF-mobilized PBSC from an HLA-identical sibling donor, and received cyclosporine and methotrexate for GVHD prevention. Median days to ANC >500/microl and platelets >50,000/microl were 12 (9-20) and 16 (11-32), respectively. Grade II-IV acute GVHD developed in 6/21 (29%) and extensive chronic GVHD in 12/20 (60%). These observations indicate that allo-PBSCT is characterized by rapid hematologic engraftment, no increase of acute GVHD and an increased risk of chronic GVHD, and can be used as an alternative to allogeneic bone marrow transplantation.

Adolescent↗

PubMed1998

[Outcomes of 50 leukemia patients who received bone marrow transplants from unrelated donors].

Fifty leukemia patients were given bone marrow transplants (BMTs) from unrelated donors at Meitetsu Hospital. We studied the outcomes of their transplants from two perspectives: leukemia disease stage and acute graft versus host disease (GVHD). The probability of disease-free survival for standard-risk, high-risk, and super-high risk patients was 65%, 29%, and 8%, respectively. The main causes of death were septicemia, cardiac and renal failure, and relapse of leukemia in the high- and super-high risk patients, and grade III-IV acute GVHD in the standard-risk patients. The incidence of grade II-IV and grade III-IV acute GVHD was 32% and 17%, respectively. All 7 patients in whom grade III-IV severe acute GVHD developed died. We conclude that better control of acute GVHD and treatment of early stage complications are clearly important to improving the outcome of BMTs from unrelated donors, especially for high and super-high risk patients.

Acute Disease↗

PubMed1998

[Treatment of aplastic anemia with anti-thymocyte globulin and cyclosporine].

Antithymocyte globulin (ATG) is an effective immunosuppressive therapy for aplastic anemia (AA). We administered ATG combined with cyclosporine (CyA), to 9 patients (4 men and 5 women; median age, 55 years). AA was severe in 8 patients and moderate in 1. The ATG and CyA regimen was the initial treatment for 3 patients, but sequential treatment for the other 6, who were refractory to other agents. Peripheral T lymphocytes, including CD4-positive and CD8-positive cells, decreased rapidly after treatment. Although 1 patient died of pulmonary hemorrhage during the 6-month period following treatment with this combined regimen, 3 showed a favorable response, 2 moderate response, and 3 no response at all. Adverse drug reactions, including transient fever and rash, were not severe. These findings suggested that ATG and CyA in combination are a safe and effective immunosuppressive regimen for initial and refractory patients with AA.

Adult↗

PubMed1997

Potentiation of depolarization-induced calcium release from skeletal muscle triads by cyclic ADP-ribose and inositol 1,4,5-trisphosphate.

Two second messengers, cyclic ADP-ribose (cADPR) and inositol 1,4,5-trisphosphate (IP3), potentiated the Ca2+ release from sarcoplasmic reticulum induced by transverse tubular membrane depolarization monitored in a triadic vesicle prepared from skeletal muscle. However, without depolarization they could not trigger the Ca2+ release. On the contrary, only cADPR potentiated caffeine-induced Ca2+ release. Because Ca2+ releases potentiated by cADPR and IP3 were inhibited by 1 microM ruthenium red and 100 microM ryanodine, probably these second messengers potentiated the Ca2+ release through ryanodine receptor Ca2+ channels. These results suggest that in skeletal excitation-contraction coupling, cADPR and IP3 play a role as a potentiator or a modifier in vivo, but both modification pathways are different from each other.

Adenosine Diphosphate Ribose↗

PubMed1997

Selective antigen presenting activity of cortical thymic epithelial cells against CD4+ T cells associated with both lack of costimulatory molecules and inefficient presentation of MHC-peptide ligands.

Selective activation among several effector functions of a T cell clone, DB14, specific for pigeon cytochrome c 43-58 (p43-58) and restricted to I-Ab/d was induced by antigen (Ag) presentation with nonprofessional Ag-presenting cells (APC), cortical thymic epithelial cells (c-TEC) (B7-1- CD40-), whereas full activation of the DB14 was induced with another nonprofessional APC, I-Ab L cell (B7-1+ CD40+). In the present study, to elucidate the mechanism underlying the selective activation of DB14 cells by c-TEC, we established c-TEC transfected with human CD40 alone (huCD40-c-TEC) or both human CD40 and murine B7-1 (huCD40/mB7-1-c-TEC), and compared the APC functions with those of the original c-TEC and I-Ab L cell. IFN-gamma production but not the proliferative response of DB14 was elevated by Ag presentation with huCD40-c-TEC as compared with unmanipulated c-TEC. On the other hand, upon stimulation with Ag plus huCD40/mB7-1-c-TEC both a significant proliferative response and IFN-gamma production were induced in DB14. However, the level of these responses did not reach that induced in the presence of I-Ab L cell. A similar pattern of APC functions was demonstrated with the other B7-independent T cell clone, PAB3, or T cell hybridomas (DBhy22 and BD7-5) which are basically independent of costimulation for the activation. The present finding along with our previous report that several structural differences of I-Ab molecules are present between c-TEC and I-Ab L cell suggests that the distinct APC activity of c-TEC is attributable not only to a lack of B7-1 and CD40 but also to inefficient presentation of MHC-peptide complex on the c-TEC.

Animals↗

PubMed1997

Genomic structure and chromosomal localization of the gene encoding translin, a recombination hotspot binding protein.

Human Translin is a novel DNA end binding protein that specifically recognizes consensus sequences at the breakpoint junctions in chromosomal translocations, mostly involving immunoglobulin (Ig)/T-cell receptor gene segments, in human lymphoid neoplasms. We have recently cloned a full-length human Translin cDNA using peptide sequence information obtained from the purified protein. From comparisons of the amino acid sequences in humans and other vertebrates, we conclude that Translin has been highly conserved during evolution, especially at the leucine zipper motif and the basic region, which is thought to be the DNA binding domain. Analysis of the human and mouse Translin genes revealed that they have identical genomic structures consisting of six exons, five introns, and a GC-rich upstream region. In situ hybridization and physical mapping of somatic cell hybrids allowed localization of the gene to human chromosome 2q21.1.

Amino Acid Sequence↗

PubMed1997

Design and synthesis of potent antitumor 5,4'-diaminoflavone derivatives based on metabolic considerations.

Recently, we reported that 5,4'-diaminoflavone (1) exhibits potent and specific growth-inhibitory activity against the estrogen receptor (ER)-positive human breast cancer cell line MCF-7. However, when compound 1 was incubated with S-9 mix, its metabolites were observed. Moreover, addition of S-9 mix to the medium caused the drastic decrease in activity of compound 1. Since the 6-, 8-, and 3'-positions were considered to be metabolized oxidatively in vivo from MO calculations, a series of 5,4'-diaminoflavone derivatives substituted at such putative metabolic positions with various functional groups were synthesized aiming at the metabolically stable derivatives. Among them, 5,4'-diamino-6,8,3'-trifluoroflavone (14d) exhibited strong growth-inhibitory activity against MCF-7 cells even in the presence of S-9 mix. Moreover, orally administered compound 14d completely suppressed the growth of MCF-7 inoculated into nude mice, and the effect was more potent than that of compound 1. In addition to ER-positive breast cancer cells, compound 14d exhibited growth-inhibitory activity against a panel of human cancer cell lines including a part of ER-negative breast, endometrial, ovarian, and liver cancers. From these results, fluorine introduction to the putative metabolic positions of compound 1 was elucidated to be effective in the enhancement of the in vivo antitumor activity, probably due to the block of the metabolic deactivation.

Animals↗

PubMed1997

A voltage- and K+-dependent K+ channel from a membrane fraction enriched in contractile vacuole of Dictyostelium discoideum.

We obtained a membrane fraction enriched in the contractile vacuole by aqueous-polymer two-phase partitioning and its channel activities were analysed by incorporating it into artificial planar lipid bilayers. In asymmetrical KCl solutions (cis, 300 mM/100 mM, trans), we observed single-channel currents of a highly K(+)-selective channel with slope conductance of 102 pS and reversal potential of -20.4 mV, which corresponded to PK+/PCl- = 7. They showed bursts separated by infrequent quiescent periods. At 0 mV the mean open time was 2.0 ms. Among monovalent cations, Na+ and Li+ were impermeable, whereas Rb+ showed permeability equivalent to that of K+, although the unitary conductance was apparently reduced when the current flowed from the Rb+ containing side, suggesting that Rb+ is a permeant blocking ion. The open probability within bursts remained constant at approx.0.6 as long as the holding potential was positive on the cis side with respect to the trans side, but it decreased to 0 at negative potential. This channel was blocked by submillimolar concentrations of quinine and 30 mM TEA+. The open probability-voltage relationship showed a striking dependency on the KCl concentration on either side. This channel may play a role in water transport in this organelle.

Animals↗

PubMed1997

The translin ring specifically recognizes DNA ends at recombination hot spots in the human genome.

We previously showed that consensus sequences exist at the chromosomal breakpoints in lymphoid malignancies and that these sequences are specifically recognized by a novel DNA binding protein, Translin. In the present study, the native form of Translin was established to be a ring-shaped structure by electron microscopy and crystallographic studies. It was also determined that this multimeric Translin formed by the subunits is responsible for its binding to target sequences situated only at single-stranded DNA ends. Furthermore, DNA-damaging reagents were found to initiate a signaling pathway for the active nuclear transport of Translin. The results support the hypothesis that staggered breaks occur at recombination hot spots and Translin has a pivotal function in recognition of the generated single-stranded DNA ends.

Binding Sites↗

PubMed1997

Selective formation of silent synapses on immature postsynaptic cells in cocultures of chick neurons of different ages.

Glutamatergic synapses usually contain two types of ionotropic glutamate receptor, N-methyl-D-aspartate receptors (NMDARs) and non-NMDA receptors (non-NMDARs), and the ratio of these receptors is thought to be critical for synaptic plasticity. To determine whether or not the ratio of these receptors at synaptic sites is controlled by the developmental stage of postsynaptic neurons, we applied a dual whole-cell recording technique to a culture of dissociated chick cerebral neurons of different ages. We found that formation of synapses that contained both types of receptor required maturation of postsynaptic neurons. Moreover, during the early development of postsynaptic neurons, NMDARs were selectively present at synaptic sites prior to the presence of non-NMDARs, even though both types of receptor were expressed in functional form in the neuronal membranes.

Age Factors↗

PubMed1997

Cloning of cDNA encoding the rice 22 kDa protein of Photosystem II (PSII-S) and analysis of light-induced expression of the gene.

Cloning of rice cDNA encoding the chlorophyll-binding 22 kDa protein of Photosystem II (PSII-S) and the light-induced expression of the gene are reported. One of the light-responsive cDNA clones, isolated by screening with a light-specific subtracted cDNA probe, was shown to encode PSII-S of rice. Genomic Southern analysis suggested that the PSII-S gene, psbS, is a single-copy gene in rice. A brief exposure to red light induced a severalfold increase in the steady state level of PSII-S transcripts in etiolated seedlings. The red light effect was reversed by far-red light, suggesting involvement of phytochrome in the PSII-S gene regulation. Prolonged exposure (3 h) to blue light, however, revealed a much stronger effect than red light on the accumulation of PSII-S transcripts in the etiolated seedlings. In dark-adapted green plants, prolonged exposure to blue light induced re-accumulation of transcripts encoding PSII-S, whereas red light had little effect.

Amino Acid Sequence↗

PubMed1997

Isolation and characterization of a cDNA encoding a Translin-like protein, TRAX.

Translin is a DNA binding protein which specifically binds to consensus sequences at breakpoint junctions of chromosomal translocations in many cases of lymphoid malignancies. To investigate its functional significance at such recombination hotspots, we examined whether Translin interacts with other proteins using a yeast two-hybrid system and identified an associated 33 kd protein partner, TRAX, with extensive amino acid homology. The TRAX protein was established to contain bipartite nuclear targeting sequences in its N-terminal region, suggesting a possible role in the selective nuclear transport of Translin protein lacking any nuclear targeting motifs.

Amino Acid Sequence↗

PubMed1997

Effects of hypotonic stress on the survival of mouse oocytes and embryos at various stages.

To examine the sensitivity of mammalian oocytes and embryos to osmotic swelling, which can occur during the removal of cryoprotectant from cryopreserved cells, the effect of hypotonic stress on the survival of fresh and vitrified mouse oocytes/embryos at various stages was examined. Oocytes and embryos were suspended in phosphate-buffered saline (PBS) media of various hypotonicities for 30 min at 25 degrees C. They were then returned to isotonic PBS medium, and the survival was assessed by the apparent integrity of the blastomeres and/or the developmental potential during culture. The survival of stressed embryos at one- to eight-cell stages assessed by the appearance was close to that assessed by the developmental ability, suggesting that hypotonic stress causes physical damage in the cell membrane. Fresh oocytes and embryos were almost totally unaffected by exposure to a 0.5x isotonic solution at all developmental stages examined. However, the extent of injury resulting from exposure to 0.3 to 0.2x isotonic solutions varied and depended on the developmental stage of the embryos. For example, zygotes were the least sensitive and morulae were the most sensitive to the hypotonic stresses. Except for morulae, vitrified cells were more sensitive to hypotonic stresses than were fresh ones. However, in many cases, the sensitivity was reduced or eliminated when the oocytes and embryos were cultured for a short period before exposure to the hypotonic stress. Furthermore, the survival rate of some stressed embryos which had been equilibrated in vitrification solution without cooling was higher than the survival of embryos stressed immediately following vitrification. These results show that sensitivity to osmotic swelling is variable among oocytes and embryos. The results also show that cryopreserved cells just after warming are more sensitive to osmotic swelling than are fresh ones, and even swelling corresponding to that in 0.5x solution may decrease survival in some stages.

Animals↗

PubMed1997

Cryopreservation of mammalian embryos.

As an innovative method for embryo cryopreservation, vitrification not only reduced the cooling stage duration to a minimum, but also eliminated any injuries cased by extracellular ice, which is a major cause of cell injury. Therefore, if embryos are treated adequately, high survival can be obtained. As a component of a vitrification solution, a permeating cryoprotective agent is essential, and additional inclusion of a macromolecule and a small saccharide makes the solution more effective. The author's group composed a solution, designated EFS40, with ethylene glycol, Ficoll, and sucrose. This solution proved effective for the cryopreservation of various stages of embryos in many species. In this article, the author describes the detailed procedure for the vitrification of mouse morulae; related information is also described.

Animals↗

PubMed1997

ATP-sensitive anion channel from rat brain synaptosomal membranes incorporated into planar lipid bilayers.

An anion channel was incorporated from rat brain synaptic plasma membrane fractions into planar lipid bilayers. The single-channel conductance was found to be 48.5 pS in choline-Cl solution (300 microM cis/100 microM trans). The anion selectivity of the channel was rather low (PCl/Pcholine = 1.7). The gating rate of the channel did not change with membrane potential over the range of -50 mV to 50 mV. Several drugs, which are known as inhibitors of anion channels, were found to be efficient inhibitors for the synaptosomal anion channel. 4-Acetoamino-4'-isothiocyanostilbene-2,2'-disulfonic acid, ethacrynic acid, indanyloxyacetic acid, and 5-nitro-2-(3-phenylpropylamino) benzoic acid inhibited the channel from the cis side of the membrane, corresponding to the cytoplasmic side of the plasma membrane. We found that the channel is regulated by intracellular ATP at millimolar concentrations. Other nucleotides, ADP and GTP, inhibited the channel as well. Glibenclamide, which is known as an inhibitor of an ATP-regulated potassium channel, inhibited the channel at micromolar concentrations from the trans side of the membrane. It is likely that the synaptosomal anion channel is a member of the ATP-binding cassette superfamily.

Adenosine Triphosphate↗

PubMed1997

Long-lasting enhancement of synaptic activity in dissociated cerebral neurons induced by brief exposure to Mg2+-free conditions.

The long-lasting enhancement of periodic clusters of spontaneous excitatory postsynaptic currents (SEPSCs) was examined in dissociated chick cerebral neurons that had been transiently exposed to Mg2+-free solution for 15 min. Since the enhancement was diminished by blockade of synaptic transmission, it clearly depended on synaptic activities. A specific antagonist of N-methyl-D-aspartate receptors (NMDARs) also inhibited the potentiations. Furthermore, the presence of inhibitors of protein and RNA synthesis in the Mg2+-free solution blocked the potentiation. In the potentiated neurons, the frequency of miniature excitatory postsynaptic currents (mEPSPs) increased. In addition, a diffusible molecule(s) that promoted the potentiation appeared to be involved in this phenomenon, since the conditioned medium of Mg2+-free treated neurons enhanced synaptic activity in other dish.

6-Cyano-7-nitroquinoxaline-2,3-dione↗