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Y Yoshimura

Publications and source records attributed to Y Yoshimura.

At least 127 records · Page 7Linked to original sources
PubMed1999

Genomic analysis of male germ cell-specific actin capping protein alpha.

The Gsg3 gene which expresses specifically in haploid germ cells is a mouse testicular homolog of somatic cell type actin capping protein alpha (ACP alpha). We have obtained a mouse Gsg3 genomic clone using cDNA as a probe. Sequencing data showed that the Gsg3 gene was not interrupted by introns. The transcription initiation site of the gene was preceded not by a TATA box or GC rich promoter motifs, but by two consensus cAMP-response element (CRE) motifs at the putative position. Southern blotting analysis showed that Gsg3 is a single copy gene in the mouse, and conserved in mammals. Phylogenetic analysis showed that Gsg3 is a novel ACP alpha specific for haploid germ cells.

Amino Acid Sequence↗
PubMed1999

Wavelength-modulated diffraction: a new method for phase determination.

A new diffraction method has been developed in which the intensity of Bragg reflections is measured while changing continually the wavelength of the radiation over a range in the vicinity of the absorption edge of an atom contained in the crystal. It is shown that the intensity gradient with respect to the wavelength of the hkl reflection is in a simple relation to the real and imaginary parts of the structure factor of that reflection and, if the positions of the anomalously scattering atoms are known or properly assumed, the phase of F(hkl) can be derived by solving simultaneous linear equations. The procedure is particularly simple when the crystal is centrosymmetric. The method, called the wavelength-modulated diffraction (WMD) method, is free from the problem of intensity scaling encountered in other methods of phase determination. Synchrotron radiation is most suited to WMD measurements. Suggestions on how to measure the intensity gradient are given and the possible errors involved are discussed.

Journal Article↗
PubMed1999

Identification and characterization of a haploid germ cell-specific nuclear protein kinase (Haspin) in spermatid nuclei and its effects on somatic cells.

We have cloned the entire coding region of a mouse germ cell-specific cDNA encoding a unique protein kinase whose catalytic domain contains only three consensus subdomains (I-III) instead of the normal 12. The protein possesses intrinsic Ser/Thr kinase activity and is exclusively expressed in haploid germ cells, localizing only in their nuclei, and was thus named Haspin (for haploid germ cell-specific nuclear protein kinase). Western blot analysis showed that specific antibodies recognized a protein of Mr 83,000 in the testis. Ectopically expressed Haspin was detected exclusively in the nuclei of cultured somatic cells. Even in the absence of kinase activity, however, Haspin caused cell cycle arrest at G1, resulting in growth arrest of the transfected somatic cells. In a DNA binding experiment, approximately one-half of wild-type Haspin was able to bind to a DNA-cellulose column, whereas the other half was not. In contrast, all of the deletion mutant Haspin that lacked autophosphorylation bound to the DNA column. Thus, the DNA-binding activity of Haspin may, in some way, be associated with its kinase activity. These observations suggest that Haspin has some critical roles in cell cycle cessation and differentiation of haploid germ cells.

Amino Acid Sequence↗
PubMed1999

Molecular cloning and characterization of a surface antigen preferentially overexpressed on multiple myeloma cells.

HM1.24 antigen has been identified as a surface molecule preferentially expressed on terminally differentiated B cells, and its overexpression is observed in multiple myeloma cells. The HM1.24 antigen is, therefore, expected as a most potent target molecule for antibody-based immunotherapy for multiple myeloma. Here, we have identified the cDNA for human HM1.24 antigen and also analyzed its gene structure including the promoter region. The HM1.24 antigen is a type II membrane glycoprotein, which has been reported as a bone marrow stromal cell surface antigen BST2, and may exist as a homodimer on myeloma cell surface. Although a reason for the overexpression in myeloma cells is not understood, very interestingly, the promoter region of the HM1.24 gene has a tandem repeat of three cis elements for a transcription factor, STAT3, which mediates interleukin-6 (IL-6) response gene expression. Since IL-6 is a differentiation factor for B cells, and known as a paracrine/autocrine growth factor for multiple myeloma cells, the expression of HM1.24 antigen may be regulated by the activation of STAT3. Importantly, a humanized anti-HM1.24 antibody effectively lysed the CHO transformants which expressed HM1.24 antigen as high as human multiple myeloma cells, but not the cells with lower antigen expression. This evaluation shows that ADCC heavily depends on the expression level of target antigens and, therefore, the immunotherapy targeting the HM1.24 antigen should have a promising potential in clinical use.

Amino Acid Sequence↗
PubMed1999

Protein phosphatase 1 is involved in the dissociation of Ca2+/calmodulin-dependent protein kinase II from postsynaptic densities.

Autophosphorylation-dependent translocation of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) to postsynaptic densities (PSDs) from cytosol may be a physiologically important process during synaptic activation. We investigated a protein phosphatase responsible for dephosphorylation of the kinase. CaM kinase II was shown to be targeted to two sites using the gel overlay method in two-dimensional gel electrophoresis. Protein phosphatase 1 (PP1) was identified to dephosphorylate CaM kinase II from its complex with PSDs using phosphatase inhibitors and activators, and purified phosphatases. The kinase was released from PSDs after its dephosphorylation by PP1.

Animals↗
PubMed1999

Possible factor for nonlinear pharmacokinetics of TAK-603, a new antirheumatic agent, in rats.

A new antirheumatic, TAK-603, shows nonlinear pharmacokinetics in both animals and humans. To elucidate the mechanism of these nonlinear pharmacokinetics, in vivo and in vitro metabolism of 14C-labeled TAK-603 ([14C]TAK-603) was studied using rats as these resemble humans in their metabolic profiles. After intravenous injection of [14C]TAK-603 to rats at doses of 1, 5, and 15 mg kg(-1), the total body clearance of unchanged drug decreased significantly with increasing dose, whereas the apparent distribution volume did not alter remarkably. Thus, saturation in the elimination processes was considered to be a factor responsible for the nonlinear pharmacokinetics. The disappearance of unchanged drug from the circulation, however, followed a dose-dependent first-order process, indicating that the nonlinearity observed was not merely due to saturation of the elimination capacity. In vitro studies using rat liver microsomes showed that TAK-603 competitively inhibited CYP-catalysed nifedipine oxidation and also that the demethylated metabolite M-I, the major metabolite in rats and humans, competitively inhibited the oxidation of nifedipine. These results suggested that inhibition by M-I of the metabolism of the parent drug (i.e. product-inhibition) may be the most likely factor responsible for the nonlinear pharmacokinetics of TAK-603.

Administration, Oral↗
PubMed1999

Effects of aging and sex steroids on the localization of T cell subsets in the ovary of chicken, Gallus domesticus.

The goal of this study was to determine the effects of aging and sex steroids on the frequency of T cells in hen ovary. Cryostat sections of ovarian tissues of immature and laying hens and those of immature hens treated with or without diethylstilbestrol (DES) or progesterone were immunostained for T cells using mouse anti-chicken CD3 (antigen of mature T cells), CD4 (antigen of helper T cells), and CD8 (antigen of cytotoxic T cells) monoclonal antibodies. Positive cells were observed under a light microscope and counted using a computer-assisted image analyzer. The frequency of CD3(+), CD4(+), and CD8(+) cells in the ovarian stroma and theca of primary follicles was significantly greater in young laying hens than in immature and old laying hens (P < 0.01). The CD4:CD8 ratio was significantly higher in the ovarian stroma of old laying hens than that of immature hens (P < 0.01), which was due to a greater decrease of CD8(+) cells than of CD4(+) cells. The frequency of CD3(+), CD4(+), and CD8(+) cells was significantly greater in the stroma and theca of primary follicles of DES-treated birds than in those of progesterone-treated and control birds (P < 0.01). Progesterone had no significant effect on the population of each subset of T cells. These results suggest that T cell frequency increases in association with sexual maturation and decreases thereafter during aging with an increase of CD4:CD8 ratio. Also, it is likely that estrogen is one of the factors which stimulates the influx of T cells in the hen ovary.

Aging↗
PubMed1999

Hysteroscopic selective salpingography.

OBJECTIVE: To evaluate the effectiveness of hysteroscopic selective salpingography (HSS) as a method for diagnosing the tubal proximal occlusion shown by hysterosalpingography (HSG). DESIGN: Prospective study. SETTING: Outpatient Department of Obstetrics and Gynecology, Social Insurance Saitama Chuo Hospital, Urawa, Japan. PATIENT(S): A total of 572 infertile women underwent HSG. Forty-seven of 50 women with unilateral or bilateral proximal tubal occlusion demonstrated by HSG underwent HSS. INTERVENTION(S): Hysteroscopic selective salpingography was performed for the diagnosis of tubal occlusion in cases in which the proximal tubal occlusion was shown by HSG. MAIN OUTCOME MEASURE(S): Number of patients who underwent HSS and pregnancy rate after HSS. RESULT(S): Twenty-seven (79.4%) of 34 patients with unilateral occlusion diagnosed by HSG were shown to have normal patency by HSS. Of 12 women with bilaterally normal patent tubes confirmed by HSS, 8 (66.7%) achieved normal pregnancies within 1 year. Seven (53.8%) of 13 patients with bilateral occlusion found by HSG were shown to have normally patent tubes by HSS. CONCLUSION: The simple method of HSS was clinically effective for evaluating the presence of proximal tubal occlusion.

Adult↗
PubMed1999

Are there inositol 1,4,5-triphosphate (IP3) receptors in human sperm?

Calcium signaling plays important roles in mammalian fertilization, such as the calcium wave in egg and the acrosome reaction (AR) in sperm. The calcium wave is accompanied by a transient increase of cytosolic calcium, which is mediated by inositol 1,4,5-triphosphate (IP3) induced calcium release (IICR). During AR, it is commonly recognized that influx of extracellular calcium causes a dramatic increase of cytosolic calcium. Participation of IICR in the phenomena, however, remains unclear. To investigate whether IICR also participates in calcium mobilization during AR, the present study examined the existence of the IP3 receptor family (IP3R type 1, 2 and 3) in human sperm and their changes during the reaction. Immunoblot analyses showed the existence of IP3R types 1 and 3 in human sperm, but IP3R type 2 was undetectable. The expression of IP3R type 1 was diminished after AR and was also detected in the vesiculated membrane fragment which was released by the fusion of the plasma membrane and the outer acrosomal membrane during AR. In contrast, the expression of IP3R type 3 on the blot was similar before and after the reaction. Immunohistochemical observations suggested that IP3R type 1 localized in the anterior portion of the sperm head and the expression was decreased after AR. IP3R type 3 was observed in the posterior portion of the sperm head, midpiece, and tail, and little change was found even after AR. The [3H]-IP3 binding assay suggested stoichiometric interactions between IP3 and IP3Rs of non-acrosome reacted and acrosome reacted human sperm.

Acrosome Reaction↗
PubMed1999

The humanized anti-HM1.24 antibody effectively kills multiple myeloma cells by human effector cell-mediated cytotoxicity.

A mouse monoclonal antibody, anti-HM1.24 (IgG2a/kappa), binds to a surface antigen preferentially overexpressed on multiple myeloma (MM) cells, and exhibits potent antitumor cell activity against MM cells by antibody-dependent cell-mediated cytotoxicity (ADCC). To develop an antibody-based immunotherapy against MM, a humanized anti-HM1.24 antibody, in which all FRs correspond to naturally processed human FRs, has been successfully constructed with the aid of both the hybrid variable region and two-step design methods. This humanized anti-HM1.24 antibody (IgG1/kappa) is able to effectively induce ADCC against human myeloma KPMM2 and ARH77 cells in the presence of human PBMCs as effectively as a chimeric anti-HM1.24 antibody. The humanized anti-HM1.24 antibody, therefore, could be expected as a potent immunotherapeutic agent for MM patients.

Amino Acid Sequence↗
PubMed1999

Ubiquitin-like polypeptide inhibits the proliferative response of T cells in vivo.

The monoclonal nonspecific suppressor factor (MNSF), a lymphokine produced by murine T cell hybridoma, possesses pleiotrophic Ag-nonspecific suppressive functions. Recently, we demonstrated that the recombinant form of the ubiquitin-like segment (rUbi-L) of MNSFbeta, a 15.6 kDa-protein consisting of a polypeptide with 36% homology with ubiquitin fused to the ribosomal protein S30, presented an antigen-nonspecific immunoregulatory action in a manner similar to native MNSF. Although this cytokine has been characterized in vitro, little is known about its effects in vivo. Thus, we investigated whether rUbi-L shows a suppressor activity in vivo. The proliferative response of Con A (5 microg/ml)-stimulated splenocytes of mice treated with rUbi-L (500 ng/body) was notably decreased in a dose-dependent manner (max. 57+/-20%). In contrast, administration of high dose ubiquitin (50 microg/body) showed a little, but significant, effect (30+/-7%). Interestingly, concomitant addition of ubiquitin inhibited Ubi-L-induced suppression. Mice injected with rUbi-L without gelatin did not show any suppressive effect. NA4 (1microg/body), a neutralizing monoclonal antibody against rUbi-L, abolished the Ubi-L-mediated suppression. Therefore, ubiquitin-like polypeptide may be implicated in the immune responses in vivo.

Animals↗
PubMed1999

Estimation of single channel conductance underlying synaptic transmission between pyramidal cells in the visual cortex.

Axon collaterals originating from pyramidal cells are one of the most abundant presynaptic elements in the neocortical circuits. To understand a quantitative aspect of synaptic transmission between pyramidal cells, we attempted to estimate single channel conductance by applying non-stationary noise analysis to unitary excitatory postsynaptic currents. Simultaneous recordings were carried out in two pyramidal cells of superficial layers in visual cortical slices. Unitary postsynaptic currents, which were evoked by action potentials of presynaptic cells impaled with conventional sharp electrodes, were recorded from postsynaptic cells with whole-cell patch clamp techniques. Estimated single channel conductance was 12.8 3.8(S.D.) pS for kittens and 10.4 +/- 1.5 pS for rats. Dividing these values by the conductance for unitary postsynaptic currents, we calculated the number of non-N-methyl-D-aspartate receptor channels activated during the postsynaptic currents. The obtained estimates were 52 (kittens) and 41 (rats). To further estimate the number of channels involved in each quantal event, we analysed amplitude histograms of miniature and spike-evoked excitatory postsynaptic currents. The derived number of estimates from these two kinds of histograms agreed quite well; about 20 channels were required for individual quantal events. Assuming open probability of non-N-methyl-D-aspartate receptor channels to be 0.7, our results suggest that the number of channels available for synaptic transmission between individual pyramidal cells would be 74 (kittens) and 59 (rats). We propose that at pyramidal-pyramidal synapses, the number of open channels is several times smaller than that previously reported for the synapses between geniculo-cortical afferent and layer IV spiny stellate cells.

Action Potentials↗
PubMed1999

Free radical scavenging activity of grape seed extract and antioxidants by electron spin resonance spectrometry in an H(2)O(2)/NaOH/DMSO system.

The scavenging effects of grape seed extract (GSE) on free radicals formed in an H(2)O(2)/NaOH/DMSO system were examined using a spin-trapping electron spin resonance (ESR) method and compared with other natural antioxidants, ascorbic acid, dl-alpha-tocopherol, and beta-carotene. GSE reduced greatly the ESR signal intensity of superoxide radical-5,5-dimethyl-1-pyrroline-N-oxide (DMPO) adducts. GSE also exhibited weak scavenging activity on hydroxyl radical and a little scavenging activity on methyl radical. Ascorbic acid exhibited strong superoxide and hydroxyl radical scavenging activities, but it increased the amount of methyl radical at high concentration. dl-alpha-Tocopherol reduced the amount of superoxide anion, especially the amount of methyl radical. However, it slightly reduced the amount of hydroxyl radical. beta-Carotene reduced the amount of hydroxyl radical and methyl radical, but it also slightly reduced superoxide anion. In the case of combination use of beta-carotene and dl-alpha-tocopherol, all radical species were suppressed. Combination of GSE and dl-alpha-tocopherol also could reduce all radical species. beta-Carotene and dl-alpha-tocopherol could reduce the methyl radical formation induced by ascorbic acid.

Dimethyl Sulfoxide↗
PubMed1999

Evaluation of free radical scavenging activities of antioxidants with an H(2)O(2)/NaOH/DMSO system by electron spin resonance.

An H(2)O(2)/NaOH/DMSO system has been developed for the formation of three free radicals, and the application of the system was examined with the antioxidants ascorbic acid and tocopherol. Superoxide anion, hydroxyl radical,and methyl radical are simultaneously generated in this system. The scavenging activity of ascorbic acid and tocopherol for these radicals was estimated by 5, 5'-dimethyl-1-pyrroline-N-oxide spin trapping electron spin resonance. Both water-soluble and oil-soluble antioxidants could be evaluated by using this system. Ascorbic acid specifically inhibited the superoxide anion and hydroxyl radical, whereas tocopherol suppressed the methyl radical.

Antioxidants↗
PubMed1999

An accurate and rapid gender determination assay in single cells by the capillary polymerase chain reaction method.

PURPOSE: In preimplantation genetic diagnosis (PGD), a rapid and accurate assay has been required. We have therefore developed a capillary polymerase chain reaction (PCR) method using rapid thermal cycling programs to determine the gender of single amniocytes. METHODS: Single amniocytes from each amniotic fluid sample were isolated by micromanipulation and their gender was determined by a multiplex PCR assay in a capillary tube, using primers that amplify a 308-bp DXZ1 and a 154-bp DYZ1 repeat sequence on the X and Y chromosomes, respectively. RESULTS: All four thermal cycling programs, which took 180, 150, 120, and 90 min, were 100% accurate in diagnosing the gender of single amniocytes. No DNA contamination was observed in any samples. CONCLUSIONS: The multiplex PCR assay was rapid and accurate in diagnosing gender in single cells and may be clinically applicable in PGD.

Amnion↗
PubMed1999

Assessment of the dominant abnormal form is useful for predicting the outcome of intracytoplasmic sperm injection in the case of severe teratozoospermia.

PURPOSE: Our purpose was to investigate the relation between the dominant sperm anomaly and sperm morphology and the outcome of intracytoplasmic sperm injection (ICSI). METHODS: Two hundred ninety-five patients who underwent a total of 181 cycles of in vitro fertilization (n = 168) and/or 217 cycles of ICSI (n = 177) between July 1995 and May 1997 at Keio University Hospital were investigated. RESULTS: The rates of fertilization and pregnancy were 63.3 and 27.8%, respectively, in ICSI cycles with < or = 4% normal forms. When the percentage of strictly normal morphology was < or = 4, the fertilization rate was lower in the case of severely tapered head (13.0%; n = 4) than in the cases of other deformities in ICSI. The acrosomal defect made no difference in the fertilization rate with ICSI. CONCLUSIONS: The predominant abnormal form affects the ICSI outcome in the case of < or = 4% normal forms.

Adult↗
PubMed1999

Localization of macrophages in the chicken oviduct: effects of age and gonadal steroids.

The aim of this study was to localize macrophages in the hen oviduct and determine the effects of age and gonadal steroids on their population. Cryostat sections of oviducal tissues from immature hens (60 d of age), virgin young (175 d of age), and old (620 d of age) laying hens, and immature hens (84 d of age) treated with gonadal steroids were immunostained for macrophages. The population of macrophages was analyzed by an image analysis system under a light microscope. Macrophages were observed in the stroma and mucosal epithelium of all oviducal segments of immature and laying hens. The population of macrophages in the oviducal stroma increased with age. Young laying hens had a significantly higher population of macrophages than immature hens in the vagina. Old laying hens had a significantly higher population than immature hens in the infundibulum, magnum, and vagina, and than young laying hens in the magnum. In the immature hens treated with gonadal steroids, the macrophage population increased only in the stroma of shell gland of the progesterone-treated birds. These results indicate that macrophage population in the oviducal stroma increases in association with sexual maturation and aging. Their population in the stroma may be partially affected by progesterone.

Aging↗