Biomedical subjects

Hisakazu Takatsuka

Publications and source records attributed to Hisakazu Takatsuka.

9 recordsLinked to original sources

PubMed2026

Post-mortem diagnosis of CACT deficiency with a novel SLC25A20 variant.

Carnitine-acylcarnitine translocase deficiency is a severe neonatal metabolic disorder caused by SLC25A20 variants. We report a case of sudden neonatal death in which post-mortem CT revealed diffuse fatty liver, and subsequent genetic analysis identified compound heterozygous variants in SLC25A20, including a known variant, c.824G>A p.(Arg275Gln), and a novel nonsense variant, c.334C>T p.(Gln112Ter). These findings demonstrate the utility of post-mortem genetic analysis in determining the cause of sudden neonatal death.

Journal Article↗

PubMed2006

Collaborative action of NF-kappaB and p38 MAPK is involved in CpG DNA-induced IFN-alpha and chemokine production in human plasmacytoid dendritic cells.

CpG DNA induces plasmacytoid dendritic cells (pDC) to produce type I IFN and chemokines. However, it has not been fully elucidated how the TLR9 signaling pathway is linked to these gene expressions. We examined the mechanisms involving the TLR9 and type I IFN signaling pathways, in relation to CpG DNA-induced IFN-alpha, IFN regulatory factor (IRF)-7, and chemokines CXCL10 and CCL3 in human pDC. In pDC, NF-kappaB subunits p65 and p50 were constitutively activated. pDC also constitutively expressed IRF-7 and CCL3, and the gene expressions seemed to be regulated by NF-kappaB. CpG DNA enhanced the NF-kappaB p65/p50 activity, which collaborated with p38 MAPK to up-regulate the expressions of IRF-7, CXCL10, and CCL3 in a manner independent of type I IFN signaling. We then examined the pathway through which IFN-alpha is expressed. Type I IFN induced the expression of IRF-7, but not of IFN-alpha, in a NF-kappaB-independent way. CpG DNA enabled the type I IFN-treated pDC to express IFN-alpha in the presence of NF-kappaB/p38 MAPK inhibitor, and chloroquine abrogated this effect. With CpG DNA, IRF-7, both constitutively and newly expressed, moved to the nuclei independently of NF-kappaB/p38 MAPK. These findings suggest that, in CpG DNA-stimulated human pDC, the induction of IRF-7, CXCL10, and CCL3 is mediated by the NF-kappaB/p38 MAPK pathway, and that IRF-7 is activated upstream of the activation of NF-kappaB/p38 MAPK in chloroquine-sensitive regulatory machinery, thereby leading to the expression of IFN-alpha.

Blotting, Western↗

PubMed2006

Human Mpv17-like protein is localized in peroxisomes and regulates expression of antioxidant enzymes.

M-LP (Mpv17-like protein) is a protein that was initially identified in mouse tissues and shows high sequence homology with Mpv17 protein, a peroxisomal membrane protein involved in the development of early-onset glomerulosclerosis [R. Iida, T. Yasuda, E. Tsubota, H. Takatsuka, M. Masuyama, T. Matsuki, K. Kishi, M-LP, Mpv17-like protein, has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up-regulates expression of the manganese superoxide dismutase gene, J. Biol. Chem. 278 (2003) 6301-6306]. Here we report the identification and characterization of a human homolog of the M-LP (M-LPH) gene. The M-LPH gene is composed of four exons, extends over 14kb on chromosome 16p13.1, and is expressed as two alternatively spliced variants comprising four and three exons, respectively, which include open-reading frames encoding two distinct isoforms composed of 196 (M-LPH1) and 147 (M-LPH2) amino acids, respectively. These two variants were expressed ubiquitously in human tissues, however only M-LPH1 was detected at the protein level. Dual-color confocal analysis of COS-7 cells transfected with a green fluorescent protein-tagged M-LPH1 demonstrated that M-LPH1 is localized in peroxisomes. In order to elucidate the function of M-LPH1, we examined the mRNA levels of several enzymes involved in the metabolism of reactive oxygen species in COS-7 cells and found that transfection with M-LPH1 down-regulates expression of the plasma glutathione peroxidase and catalase genes. These results show the existence of the human homolog of M-LP and its participation in reactive oxygen species metabolism.

Animals↗

PubMed2005

Quantitative change in mitochondrial DNA content in various mouse tissues during aging.

In order to systematically characterize age-related changes in the mtDNA content of various tissues during aging, we analyzed the mtDNA content of eight tissues from mice at five different ages from young to senescent by quantitative real-time PCR analysis. Obvious variations of mtDNA content among the tissues were detected: There was a 20-fold range in 2-week-old mice and a 50-fold range in 15-month-old mice. The mtDNA contents of the heart, lung, kidney, spleen and skeletal muscle increased gradually with age, whereas those of bone marrow and brain showed no age-related pattern. The expression patterns of mitochondrial transcription factor A (mtTFA) and mitochondrial single-strand DNA binding protein (mtSSB), possible regulatory factors of the mtDNA copy number, were not necessarily linked with the age-related pattern of the mtDNA content, suggesting the existence of other factors that affect the mtDNA content. The Western blot analysis of mtDNA-encoded cytochrome c oxidase subunit III (MTCO3) demonstrated that the expression levels of this protein in the heart and skeletal muscle increase with age in parallel with the mtDNA content. These findings confirm that the mtDNA content of tissues changes during aging.

Aging↗

PubMed2005

A novel alternative spliced Mpv17-like protein isoform localizes in cytosol and is expressed in a kidney- and adult-specific manner.

Mpv17-like protein (M-LP) has been identified as a new protein that shows high sequence homology with Mpv17 protein, a peroxisomal membrane protein involved in the development of early onset glomerulosclerosis. We previously showed that the originally identified M-LP isoform, designated M-LPL, is, like Mpv17, localized in peroxisomes, and that transfection with M-LPL up-regulates expression of the manganese superoxide dismutase (SOD2) gene [R. Iida, T. Yasuda, E. Tsubota, H. Takatsuka, M. Masuyama, T. Matsuki, K. Kishi, M-LP, Mpv17-like protein, has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up-regulates expression of the manganese superoxide dismutase gene. J. Biol. Chem. 278 (2003) 6301-6306.]. We report here the identification of a novel alternative splicing product of the M-LP gene, designated M-LPS. A comparison of the genomic sequence with the cDNA sequences and an analysis of 5'-flanking regions revealed that the two isoforms are generated by alternative usage of two promoters. M-LPS consists of the C-terminal half of M-LPL (90 amino acids) and therefore lacks the peroxisome targeting signal of membrane protein that exists near the N-terminus of M-LPL. Expression of green fluorescent protein-tagged M-LPS in COS-7 cells demonstrated that M-LPS localizes in the cytosol. In mice, M-LPS is expressed exclusively in kidneys after the age of 6 weeks. Moreover, quantitative real-time PCR analysis revealed that transfection with M-LPS up-regulates expression of the SOD2 gene and down-regulates expression of the cellular glutathione peroxidase (Gpx1) and plasma glutathione peroxidase (Gpx3) genes. Taken together, these results suggest different functional attributes of the two M-LP isoforms during aging and development.

Age Factors↗

PubMed2004

Improvement of a multiplex PCR system for DYS441, DYS442, DYS443, DYS444 and DYS445, and a population study in 340 Japanese males.

A multiplex PCR system for five Y-STRs (DYS441, DYS442, DYS443, DYS444 and DYS445) has been improved to increase the probability of obtaining a DNA typing result from aged samples. Newly designed PCR primers for amplification of the DYS441 and DYS442 loci and optimization of PCR conditions enabled successful typing from blood and semen stains that had been stored for more than seven years at room temperature. Analysis of 340 Japanese males revealed 7, 5, 6, 5 and 4 alleles at the DYS441, DYS442, DYS443, DYS444 and DYS445 loci, respectively, yielding 122 haplotypes with a cumulative haplotype diversity of 0.97.

Chromosomes, Human, Y↗

PubMed2003

A novel multiplex PCR system consisting of Y-STRs DYS441, DYS442, DYS443, DYS444, and DYS445.

We have developed a new sensitive multiplex PCR system consisting of five male-specific and polymorphic tetranucleotide STRs--DYS441 (GDB: 10013873), DYS442 (GDB: 10030304), DYS443 (GDB: 10807127), DYS444 (GDB: 10807128), and DYS445 (GDB: 10807129) on the Y chromosome. Fifty pg DNA per 10 microL reaction volume was required for the correct typing of five STRs. Using this system, the five Y-STRs were correctly typed from blood and semen stains that had been stored for several years at room temperature.

Blood Stains↗

PubMed2002

M-LP, Mpv17-like protein, has a peroxisomal membrane targeting signal comprising a transmembrane domain and a positively charged loop and up-regulates expression of the manganese superoxide dismutase gene.

M-LP (Mpv17-like protein) has been identified as a new protein that has high sequence homology with Mpv17 protein, a peroxisomal membrane protein involved in the development of early onset glomerulosclerosis. In this study, we verified the peroxisomal localization of M-LP by performing dual-color confocal analysis of COS-7 cells cotransfected with green fluorescent protein-tagged M-LP and DsRED2-PTS1, a red fluorescent peroxisomal marker. To characterize the peroxisomal membrane targeting signal, we examined the intracellular localizations of several green fluorescent protein-tagged deletion mutants and demonstrated that, of the three transmembrane segments predicted, the first near the NH(2) terminus and NH(2)-terminal half of the following loop region, which is abundant in positively charged amino acids, were necessary and sufficient for peroxisomal targeting. To elucidate the function of M-LP, we examined the activities of several enzymes involved in reactive oxygen species metabolism in COS-7 cells and found that transfection with M-LP increased the superoxide dismutase activity significantly. Quantitative real-time PCR analysis revealed that the manganese SOD (SOD2) mRNA level of COS-7 cells transfected with M-LP was elevated. These results indicate that M-LP participates in reactive oxygen species metabolism.

5' Untranslated Regions↗

PubMed2002

CpG-DNA-induced IFN-alpha production involves p38 MAPK-dependent STAT1 phosphorylation in human plasmacytoid dendritic cell precursors.

Human plasmacytoid or CD4(+)CD11c(-) type 2 dendritic cell precursors (PDC) were identified as natural type I interferon (IFN)-producing cells in response to viral and bacterial infection. They represent effector cells of innate immunity and link it to the distinct adaptive immunity by differentiating into mature DC. It has been reported that oligodeoxyribonucleotides containing unmethylated CpG motifs (CpG DNA) stimulate PDC to produce IFN-alpha, but the molecular mechanisms involved remain unknown. We found that CpG-DNA-induced IFN-alpha production in PDC was completely impaired by the inhibitor of the p38 mitogen-activated protein kinase (MAPK) pathway. Expression of IFN regulatory factor (IRF)-7 was enhanced by CpG-DNA treatment, which was preceded by the phosphorylation of signal transducer and activator of transcription (STAT)1 on Tyr-701, as well as its enhanced phosphorylation on Ser-727. All of these events were also suppressed by the p38 MAPK inhibitor. STAT1, STAT2, and IRF-9, components of IFN-stimulated gene factor 3 (ISGF3), were recognized in the nuclear fraction of CpG-DNA-treated cells. Neither anti-IFN-alpha/beta antibodies (Ab) nor anti-IFNAR Ab suppressed STAT1 phosphorylation, enhancement of IRF-7 expression, or IFN-alpha production in the early phase of the culture. These results suggest that CpG DNA induces p38 MAPK-dependent phosphorylation of STAT1 in a manner independent of IFN-alpha/beta, which may cause ISGF3 formation to increase the transcription of the IRF-7 gene, thereby leading to IFN-alpha production in human PDC.

Cell Nucleus↗