Search PubMed⌕ Search

Biomedical subjects

M Miwa

Publications and source records attributed to M Miwa.

At least 91 records · Page 5Linked to original sources
PubMed1998

High susceptibility of human cancer xenografts with higher levels of cytidine deaminase to a 2'-deoxycytidine antimetabolite, 2'-deoxy-2'-methylidenecytidine.

2'-Deoxy-2'-methylidenecytidine (DMDC) is a new 2'-deoxycytidine (dCyd) antimetabolite. The present study compared its antitumor activities with those of 2',2'-difluorodeoxy-cytidine (gemcitabine) in 15 human cancer xenograft models. DMDC was highly resistant to cytidine (Cyd) deaminase, which deaminates the dCyd analogues to inactive molecules, whereas gemcitabine was susceptible to the enzyme. Given p.o., high antitumor activity with therapeutic index of more than 10 was found with DMDC in 7 of 15 xenograft lines. In contrast, gemcitabine given i.v. or p.o. was highly effective in 4 of 15 human cancer xenograft lines. The antitumor spectrum of these compounds was quite different, although their molecular targets are reported to be similar. DMDC was highly effective in tumors with higher levels of Cyd deaminase activity, whereas it showed only slight activity in those with lower levels of Cyd deaminase. In contrast, gemcitabine appeared to be less effective in tumors with high levels of Cyd deaminase. We also investigated the correlation with the susceptibility to the two dCyd antimetabolites and dCyd kinase activity in tumors, but none was observed. Cyd deaminase activity was found to be high in tumor tissues from various types of human cancers thus far tested, such as colorectal cancer and non-small cell lung cancer. Such cancer types or individual patients who have tumors with high activity of the enzyme may be targets for DMDC therapy.

Animals↗
PubMed1998

[Serum factors in cancer patients affecting the antitumor effect of 5-fluorouracil].

A human colon cancer cell line, HCT-15, was found to proliferate in human sera. Sera of cancer patients undergoing continuous intravenous administration of 5-fluorouracil (5-FU) were tested for the antitumor effect by counting HCT-15 cells stained with Giemsa solution. 5-FU concentrations in those sera were not correlated with the antitumor effects. After HCT-15 cells were incubated in sera of cancer patients or healthy volunteers for 24 hours followed by removal of those sera, those cells were incubated in a culture medium containing 500 or 1000 ng/ml of 5-FU for 48 hours. It was shown that preincubation of HCT-15 cells with sera of cancer patients but not sera of healthy volunteers decreased the sensitivity of HCT-15 cells to 5-FU. These results suggest that there may be some factors in the serum of a cancer patient, which affect the antitumor effect of 5-FU.

Antimetabolites, Antineoplastic↗
PubMed1997

Cell-specific regulation of expression of plasma-type platelet-activating factor acetylhydrolase in the liver.

Platelet-activating factor (PAF) is a potent proinflammatory phospholipid mediator that causes hypotension, increases vascular permeability, and has been implicated in anaphylaxis, septic shock and several other inflammatory responses. PAF is hydrolyzed and inactivated by the enzyme PAF-acetylhydrolase. In the intact rat, a mesenteric vein infusion of lipopolysaccharide (LPS) served as an acute, liver-focused model of endotoxemia. Plasma PAF-acetylhydrolase activity increased 2-fold by 24 h following LPS administration. Ribonuclease protection experiments demonstrated very low levels of plasma-type PAF-acetylhydrolase mRNA transcripts in the livers of saline-infused rats; however, 24 h following LPS exposure, a 20-fold induction of PAF-acetylhydrolase mRNA was detected. In cells isolated from endotoxin-exposed rat livers, Northern blot analyses demonstrated that Kupffer cells but not hepatocytes or endothelial cells were responsible for the increased PAF-acetylhydrolase mRNA levels. In Kupffer cells, plasma-type PAF-acetylhydrolase mRNA was induced by 12 h, peaked at 24 h, and remained substantially elevated at 48 h. Induction of neutropenia prior to LPS administration had no effect on the increase in PAF-acetylhydrolase mRNA seen at 24 h. Although freshly isolated Kupffer cells contain barely detectable levels of plasma-type PAF-acetylhydrolase mRNA, when Kupffer cells were established in culture, PAF-acetylhydrolase expression became constitutively activated concomitant with cell adherence to the culture plates. Alterations in plasma-type PAF-acetylhydrolase expression may constitute an important mechanism for elevating plasma PAF-acetylhydrolase levels and an important component in minimizing PAF-mediated pathophysiology in livers exposed to endotoxemia.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗
PubMed1997

Integration of HTLV-1 provirus into mouse transforming growth factor-alpha gene.

Human T cell leukemia virus type 1 (HTLV-1) provirus integration was investigated in mice inoculated with MT-2 cells, a HTLV-1 producing human T-cell line. From spleen DNA derived from a MT-2 cell-injected mouse, we cloned a HTLV-1 integration site by ligation-mediated PCR. The nucleotide sequence showed that HTLV-1 provirus was integrated into an intron of the mouse transforming growth factor-alpha gene in the reverse orientation. This result provides for the first time molecular evidence that mice can be infected with HTLV-1.

Amino Acid Sequence↗
PubMed1997

Position-independent and high-level expression of human alpha-lactalbumin in the milk of transgenic rats carrying a 210-kb YAC DNA.

The level of expression of transgenes in transgenic animals varies among lines, and is often much lower than that of endogenous genes (position effects). In order to surmount position effects and establish a more efficient production system of transgenic animals producing pharmaceutical proteins in their milk, transgenic rats carrying 210-kb YAC DNA containing the human alpha-lactalbumin gene were produced. Three transgenic lines transmitted the transgene to the next generation. They had one copy of the alpha-lactalbumin gene and secreted human alpha-lactalbumin in their milk at concentrations of 2.0-4.3 mg/ml. No position effect was seen. The transgene was expressed specifically in the mammary gland of the transgenic rats. The 210-kb region is thought to contain all the DNA elements required for proper expression of the human alpha-lactalbumin gene. The YAC carrying the human alpha-lactalbumin gene is a potential vector for the expression of foreign genes in the mammary gland.

Animals↗
PubMed1997

Contrasting effects of TGF-beta 1 and TNF-alpha on the development of dendritic cells from progenitors in mouse bone marrow.

Dendritic cells (DC) are a distinct population of leukocytes and specialized antigen-presenting cells for T cell responses. Prior work has shown that GM-CSF can induce the development of large numbers of DC from proliferating progenitors in mouse bone marrow. We have monitored the effects of potentially enhancing and suppressive cytokines in these cultures. In this system, many immature DC develop from proliferating precursors during the first six days of culture, and between days 6-8 maturation of typical nonadherent and nonreplicating DC takes place. The maturation is accompanied by a large increase in the expression of major histocompatibilities complex class II (MHC II) and B7-2/CD86, and in mixed leukocyte reaction stimulating activity. Tumor necrosis factor-alpha (TNF-alpha), previously shown to be required for development of human DC, was found to enhance the maturation of mouse DC in the last two days of culture. Transforming growth factor-beta 1 (TGF-beta 1), on the other hand, almost totally blocked DC maturation, but it had to be given in the first six days of culture when the DC were actively proliferating. TGF-beta 1 did not block the production of immature, MHC II-positive but B7-2/CD86-negative DC. Maturation would take place between days 6-8 as long as the cultures were depleted of Fc-receptor-bearing cells, or if TNF-alpha were added. In both instances, maturation was not blocked even when TGF-beta 1 remained in the culture. We conclude that the development of DC, in response to GM-CSF, can be modified by other cytokines. TGF-beta 1 is suppressive but only indirectly via Fc-receptor-bearing suppressive cells, presumably suppressive macrophages, while TNF-alpha enhances the final maturation of DC.

Animals↗
PubMed1997

Genomic organization and chromosomal localization of the human casein gene family.

Five yeast artificial chromosome (YAC) clones containing the human casein gene family were isolated and characterized to study the control mechanisms for the expression of these genes. Partial restriction analysis in conjunction with the chromosomal fragmentation method and fluorescence in situ hybridization (FISH) analysis were performed to construct a detailed physical map of the casein gene family and to determine the chromosomal localization of these genes. The isolated YAC clones 748F3, 750D11, 882G11, 886B3 and 960D2 were 1.2 Mb, 860 kb, 800 kb, 1.5 Mb and 1.5 Mb in size, respectively. The clones 748F3, 882G11, 886B3 and 960D2 contained the entire casein gene family, while the kappa-casein gene was absent in 750D11. The human alphaS1-, beta- and kappa-casein genes were found to be closely linked and arranged in the order alphaS1-beta-kappa. The distance between alphaS1 and beta, and between alphaS1 and kappa was approximately 10 and 300 kb, respectively. The beta-casein gene was oriented in the opposite direction to the alphaS1- and kappa-casein genes. The casein gene family was localized to chromosome 4q21.1 by FISH analysis.

Caseins↗
PubMed1997

lpr T cells have lower ability to maintain bcl-2 expression ex vivo.

Autoimmune-prone lpr mice develop lymphoproliferative disorders, whereas their lymphocytes show accelerated apoptosis in culture. To elucidate whether the bcl-2 protein, a repressor of apoptosis, is critical to the discrepancy between in vivo and in vitro survival, we examined bcl-2 expression in T cells from +/+ and lpr mice during culture. The expression levels of bcl-2 in cultured T cells from lpr mice were significantly down-modulated compared to those from +/+ mice and freshly obtained T cells. Besides, the reduction of bcl-2 protein levels was inhibited in T cells cultured in the presence of T cell receptor (TCR) signalling. These results suggest that lpr T cells might be susceptible to apoptosis in vitro due to down-modulation of bcl-2 by withdrawal of TCR signalling.

Journal Article↗
PubMed1997

Japanese families with autosomal dominant pure cerebellar ataxia map to chromosome 19p13.1-p13.2 and are strongly associated with mild CAG expansions in the spinocerebellar ataxia type 6 gene in chromosome 19p13.1.

Autosomal dominant cerebellar ataxia is a group of clinically and genetically heterogeneous disorders. We carried out genomewide linkage analysis in 15 families with autosomal dominant pure cerebellar ataxia (ADPCA). Evidence for linkage to chromosome 19p markers was found in nine families, and combined multipoint analysis refined the candidate region to a 13.3-cM interval in 19p13.1-p13.2. The remaining six families were excluded for this region. Analysis of CAG-repeat expansion in the alpha1A-voltage-dependent calcium channel (CACNL1A4) gene lying in 19p13.1, recently identified among 8 small American kindreds with ADPCA (spinocerebellar ataxia type 6 [SCA6]), revealed that 8 of the 15 families studied had similar, very small expansion in this gene: all affected individuals had larger alleles (range of CAG repeats 21-25), compared with alleles observed in neurologically normal Japanese (range 5-20 repeats). Inverse correlation between the CAG-repeat number and the age at onset was found in affected individuals with expansion. The number of CAG repeats in expanded chromosomes was completely stable within each family, which was consistent with the fact that anticipation was not statistically proved in the SCA6 families that we studied. We conclude that more than half of Japanese cases of ADPCA map to 19p13.1-p13.2 and are strongly associated with the mild CAG expansion in the SCA6/CACNL1A4 gene.

Adult↗
PubMed1997

New method to calculate creatinine generation rate using pre- and postdialysis creatinine concentrations.

The creatinine (Cr) generation rate reflects the muscle mass, a possible indicator of protein nutritional status. Thus, in this study, we developed equations for calculating the Cr generation rate. Depner and Daugirdas recently developed a method for determining the protein catabolic rate (PCR) from the pre- and postdialysis blood urea nitrogen concentrations. We modified their method to develop equations for calculating the total Cr generation rate from the measured predialysis Cr concentration and estimated postrebound concentration. The total Cr generation rate is defined as the sum of the intrinsic Cr generation rate and the extrinsic Cr generation rate (i.e., the generation rate of Cr derived from food). In the present study, the postrebound Cr concentration was estimated on the basis of postdialysis Cr concentration and the K/V for Cr. The intrinsic Cr generation rate was obtained by subtracting the extrinsic Cr generation rate, which was estimated on the basis of the PCR, from the total Cr generation rate calculated. The intrinsic Cr generation rate determined with this method was virtually the same as that obtained using the postrebound Cr concentration, the concentration immediately before the next hemodialysis (HD) session, and the PCR. The intrinsic Cr generation rate determined with the present method did not vary with changes in the HD prescription (i.e., with an increase in blood flow rate, a prolongation of the HD duration time, or a change in dialyzer membrane area). The present study also indicated that the intrinsic Cr generation rate decreased with age in both males and females.

Adult↗
PubMed1997

Inhibition of syncytium formation by antisense oligonucleotide phosphorothioates complementary to tax mRNA of human T-cell leukemia virus type 1 (HTLV-1).

HTLV-1 infection is known as the factor to cause adult T-cell leukemia (ATL). Antisense oligonucleotide phosphorothioates against tax gene and control oligonucleotide phosphorothioates were synthesized. Antisense oligonucleotide was complementary to the region of initiation codon of tax gene. Two control oligonucleotides were tax sense and random. HTLV-1-positive human T-cell line, C91/PL and HTLV-1 non-infected human glioma cell line, U251-MG were co-cultured in the presence of antisense or control oligonucleotides for 24 hours. Oligonucleotides used in this study were not toxic at 10 microM concentration. Antisense oligonucleotide against tax gene inhibited 59% the syncytium formation assay at 10 microM concentration.

Adult↗
PubMed1997

Pathogenesis and prevention of HTLV-1-associated diseases.

HTLV-1 is an important factor involved in various diseases including adult T-cell leukemia/lymphoma and HTLV-1 associated myelopathy/tropical spastic paraparesis. Amount of HTLV-1 provirus integrated in human peripheral blood mononuclear cells might be a candidate for a risk factor in the manifestation of HTLV-1 associated diseases. Experimental animal models would be useful to dissect the pathogenesis of HTLV-1 associated diseases. We present rat and mouse models of HTLV-1 infection. Using these animal models, we could clarify the intrauterine transmission of HTLV-1, and have found that both genetic background and HTLV-1 infection are important in pathogenesis of HAM/TSP-like rats. We also discuss the preventive measures of HTLV-1 transmission by use of antisense oligonucleotides.

Adult↗
PubMed1997

Establishment of HTLV-1 carrier mice by injection with HTLV-1-producing T cells.

We injected with HTLV-1-producing T-cells, MT-2, into C3H/HeJ and Balb/c mice intraperitoneally within 24 hours after birth and at one week old. At 3 months old, HTLV-1 provirus was detected in peripheral blood mononuclear cells in both mice. It was also detected in lymph nodes, thymus, spleen and liver of those mice. The antibody response against HTLV-1 Gag antigen was detected in some of Balb/c mice. These findings suggest that the C3H/HeJ and Balb/c mice were infected with HTLV-1 persistently. The HTLV-1-infected mice should be helpful for studying the pathological state of HTLV-1 carriers and for an establishment of a small animal model of HTLV-1 related diseases including ATL.

Animals↗
PubMed1997

PARP cleavage in the apoptotic pathway in S2 cells from Drosophila melanogaster.

Caspase activities and two cDNA sequences have been identified in Drosophila melanogaster. To study the molecular events following the activation of the apoptotic pathway in D. melanogaster, S2 cells were treated with etoposide and the timing of the apoptotic events, such as caspase activation, mitochondrial pore opening, and loss of membrane asymmetry, was determined. Poly(ADP-ribose) polymerase (PARP) is known to be cleaved in the early phase of apoptosis in vertebrate systems. Little is known about the involvement of PARP cleavage in apoptosis in invertebrates. If PARP inactivation is a general event, this could mean that DNA repair enzymes need to be cleaved for the death pathway to be completed. We have found that in etoposide-treated cells, PARP protein is processed, but the nature of the cleavage is not known. Further experiments must be conducted and the peptide fragments must be sequenced to relate protease activities with PARP cleavage.

Animals↗
PubMed1996

Inactivation of p16/CDKN2 and p15/MTS2 genes in different histological types and clinical stages of primary ovarian tumors.

To define the involvement of p16/CDKN2 and p15/MTS2 inactivation in ovarian tumorigenesis and the association of these inactivation events with histological types and clinical stages of ovarian tumors, we analyzed homozygous deletion and somatic mutation of p16/CDKN2 and p15/MTS2 genes, as well as hypermethylation of the 5'-CpG island of the p16/CDKN2 gene, in 49 primary ovarian tumors and 6 ovarian carcinoma cell lines. We found homozygous deletions of p16/CDKN2 and p15/MTS2 in 6 (12%) and 5 (10%) primary tumors, respectively. Somatic mutation of p16/CDKN2 was found in only 1 primary tumor, but mutation of p15/MTS2 was not detected in any sample. None of the 28 primary tumors or 6 cell lines was hypermethylated at the 5'-CpG island of p16/CDKN2. The incidence of inactivation of p16/CDKN2 in primary tumors was significantly higher in the advanced stages (7 of 29) than in the early stages (0 of 14). Seven of 9 alterations in p16/CDKN2 and p15/MTS2 were observed in serous (3 of 12), endometrioid (3 of 9) and clear-cell (1 of 4) carcinomas. However, only normal sequences of these genes were detected in mucinous carcinomas. Loss of heterozygosity (LOH) at the IFNA locus was detected in 1 of 19 (5%) tumors, but no change at the D9S171 locus was observed in 17 tumors. These results suggest that: (i) homozygous deletion is the main mechanism of inactivation of p16/CDKN2 and p15/MTS2 in ovarian tumorigenesis; (ii) inactivation of p16/CDKN2 and p15/MTS2 may be the histological type-specific events involved in ovarian tumorigenesis; and (iii) inactivation of p16/CDKN2 is potentially involved in the progression of ovarian tumors in advanced stages.

Carcinoma↗