Biomedical subjects

Keisuke Oboki

Publications and source records attributed to Keisuke Oboki.

14 recordsLinked to original sources

PubMed2006

Molecular mechanisms of mast cell development.

Mast cells are progeny of multipotential hematopoietic stem cells (MHSCs). MHSCs commit to the mast cell lineage in the bone marrow, and the mast cell-committed progenitors leave the bone marrow, migrate in blood, invade connective or mucosal tissue, and then proliferate and differentiate to connective tissue-type or mucosal mast cell. GATA-1, GATA-2, and PU.1 transcription factors seem to be involved i the commitment to mast cells, and MITF, a basic helix-loop-helix leucine zipper-type transcription factor, seems to be involved in the migration, phenotypic expression, and survival of mast cells. KIT ligand (KITL) is the most important cytoline for development of mast cells, and KIT is the receptor of KITL. Tissues of loss-of-function mutants of KIT, KITL, or MITF are deficient in mast cells.

Animals↗

PubMed2006

Gene expression profiling of human mast cell subtypes: an in silico study.

BACKGROUND: Human mast cells (MCs) were classified into at least two subtypes, i.e., tryptase- and chymase-positive MCs (MC(TC)) and tryptase-only-positive MCs (MC(T)). However, differences in global molecular expression between these subtypes are unknown. METHODS: We analyzed public microarray data of MC subtypes derived from various tissues and those of peripheral blood granulocytes by using hierarchical clustering methods to understand the global gene expression profiles. RESULTS: All the transcripts subjected to this clustering analysis were classified into two large clusters, i.e., MC-preferential or granulocyte-preferential. In the original works, MCs from tonsil, lung and skin had been cultured for more than several weeks to obtain highly viable and pure cell populations, and these MCs retained their typical profiles such as intensities of chymase protein expression. Most of the transcripts were commonly expressed by these MC subtypes. However, tonsil-derived MCs and skin-derived MCs but not lung-derived MCs expressed high levels of chymase (CMA1) as expected for the properties of MC(TC) and MC(T). These CMA1-high MCs and CMA1-low MCs respectively expressed distinct sets of transcripts as small gene clusters as well as CMA-1 even after being cultured in the absence of a tissue environment. CONCLUSIONS: The MC lineage seems to be far from the granulocyte lineages including basophils. CMA1-high MCs (MC(TC)) and CMA1-low MCs (MC(T)) can be regarded as differentiated MC subtypes. As such, importance of data analysis studies will be increasing along with the accumulation of global molecular data in the public database.

Cells, Cultured↗

PubMed2005

Micronuclei in EM9 cells expressing polymorphic forms of human XRCC1.

X-ray repair cross-complementing gene 1 (XRCC1) is involved in base excision repair (BER) through interaction with other BER enzymes, and polymorphisms in XRCC1 appear to increase the risk of various cancers. We evaluated how three XRCC1 polymorphisms, Arg194Trp, Arg280His and Arg399Gln, affect the extent of DNA damage and repair using the micronucleus assay. XRCC1 cDNAs containing the wild-type sequence and the three polymorphisms were overexpressed in EM9 cells, which lack the full sequence needed to perform XRCC1 functions. Normal human XRCC1 cDNA corrected the defect in EM9 cells. Only XRCC1 cDNA containing the Arg399Gln polymorphism did not fully correct the DNA repair defect in EM9 cells. These results indicate that the Arg399Gln polymorphism, but not the Arg194Trp or Arg280His polymorphism, influences the ability of XRCC1 to repair DNA. This study may provide a model that can be used to evaluate the functional significance of polymorphisms in DNA repair genes.

Animals↗

PubMed2005

Reduced expression of IL-12 receptor beta2 and IL-18 receptor alpha genes in natural killer cells and macrophages derived from B6-mi/mi mice.

The mi transcriptional factor (MITF) is a basic helix-loop-helix leucine zipper-type transcriptional factor. The mi mutant allele encodes an abnormal MITF, in which one out of four consecutive arginines is deleted in the basic domain. The VGA-9-tg (tg) allele is another mutant allele and considered to be a null mutant allele. C57BL/6 (B6)-mi/mi mice showed abnormal phenotypes of natural killer (NK) cells and macrophages, whereas B6-tg/tg mice did not. The expression levels of the genes for the interleukin-12 receptor (IL-12R) beta2 and IL-18Ralpha were reduced in both the NK cells and macrophages of B6-mi/mi mice, while the expression levels of the corresponding genes in B6-tg/tg mice were unaffected. The B6-mi/mi NK cells and B6-mi/mi macrophages showed impaired responses to stimulation with IL-12, IL-18, and IL-12 plus IL-18 stimulation. The abnormal NK cell and macrophage of B6-mi/mi mice appear to be due to decreased expression of the IL-12Rbeta2 and IL-18Ralpha genes.

Animals↗

PubMed2004

Involvement of connective tissue-type mast cells in Th1 immune responses via Stat4 expression.

Mast cells are the sentinels of immune systems and, like other immuno-competent cells, they are produced by hematopoietic stem cells. We analyzed the expression of signal transducer and activator of transcription 4 (Stat4), and investigated its role in mast cells. Murine mast cells are usually divided into 2 distinct populations by their distribution and contents of their granules: mucosal mast cells (MMCs) and connective tissue-type mast cells (CTMCs). Stat4 protein was detected in CTMCs but not in MMCs. The absence of Stat4 expression in cultured mast cells was due to the presence of Stat6. In T-helper (Th) cells, Stat4 plays an important role in Th1 shift by inducing a set of genes, such as interferon gamma (IFN-gamma) and interleukin-18 receptor alpha subunit (IL-18Ralpha). As in Th1 shift, we found that Stat4 trans-activated these genes in the Stat4-expressing cultured mast cells, namely, microphthalmia transcription factor (MITF)-deficient cultured MMCs, Stat6-deficient cultured MMCs, and cultured CTMCs. Stat4 also enhanced expression of nitric oxide synthase 2 (NOS2) in CTMCs, which brought about increased levels of NO-dependent cytotoxic activity. These data indicate that expression of Stat4 in CTMCs plays an important role on Th1 immune responses.

Animals↗

PubMed2004

MITF is necessary for generation of prostaglandin D2 in mouse mast cells.

Mast cells generate eicosanoids that are linked to asthma and other inflammatory diseases. A basic-helix-loop-helix leucine zipper transcription factor termed MITF is essential for the development of mast cells. Although other substances also linked to inflammatory reactions (such as various proteases and serotonin) require MITF for their expression, the role of MITF in eicosanoid generation has not been studied. We examined eicosanoid generation in bone marrow-derived mast cells (BMMCs) of tg/tg mice that lack MITF. Most eicosanoids generated by BMMCs are either prostaglandin (PG) D2 or leukotriene C4. The former is synthesized via the cyclooxygenase pathway, whereas the latter is synthesized via the 5-lipoxygenase pathway. In response to stimulation with IgE and antigens, BMMCs of tg/tg mice synthesized leukotriene C4 normally. However, neither immediate nor delayed PGD2 production was detected in these BMMCs. This indicates that MITF is a transcription factor that specifically activates the cyclooxygenase pathway, but not the 5-lipoxygenase pathway. Significant decreases in expression of hematopoietic PGD2 synthase (hPGDS, a terminal synthase for PGD2) were observed at both mRNA and protein levels in tg/tg BMMCs. MITF transactivated the hPGDS gene via a CACCTG motif located in the promoter region. MITF appeared to be essential for generation of PGD2 by enhancing expression of the hPGDS gene in BMMCs.

Amino Acid Motifs↗

PubMed2004

Roles of MITF for development of mast cells in mice: effects on both precursors and tissue environments.

The mutant tg/tg mice, which do not express mi transcription factor (MITF), lack mast cells in most tissues. Since MITF is expressed in both mast cells and tissues where mast cells develop, there is a possibility that the tg/tg mice may show abnormalities in both mast cell precursors and tissue environments. We examined this possibility by bone marrow and skin transplantation. When bone marrow cells of tg/tg mice were transplanted to W/W(v) mice that possess normal tissue environment, mast cells did not develop in all tissues examined. The number of developing mast cells in the skin of W/W(v) mice was much lower when grafted to tg/tg recipients than when grafted to normal (+/+) recipients. These results indicated that mast cell precursors of tg/tg mice were defective. When bone marrow cells of +/+ mice were transplanted, the number of developing mast cells was significantly lower in examined tissues of tg/tg recipients than in those of W/W(v) recipients, suggesting that the tissue environment for mast cell development was defective in tg/tg mice. MITF appeared essential for the function of both mast cell precursors and tissue environments for their development.

Animals↗

PubMed2004

Number of mast cells in the peritoneal cavity of mice: influence of microphthalmia transcription factor through transcription of newly found mast cell adhesion molecule, spermatogenic immunoglobulin superfamily.

The mi (microphthalmia) locus of mice encodes a transcription factor, MITF. B6-tg/tg mice that do not express any MITF have white coats and small eyes. Moreover, the number of mast cells decreased to one-third that of normal control (+/+) mice in the skin of B6-tg/tg mice. No mast cells were detectable in the stomach, mesentery, and peritoneal cavity of B6-tg/tg mice. Cultured mast cells derived from B6-tg/tg mice do not express a mast cell adhesion molecule, spermatogenic immunoglobulin superfamily (SgIGSF). To obtain in vivo evidence for the correlation of nonexpression of SgIGSF with decrease in mast cell number, we used another MITF mutant, B6-mi(vit)/mi(vit) mice that have a mild phenotype, ie, black coat with white patches and eyes of normal size. B6-mi(vit)/mi(vit) mice had a normal number of mast cells in the skin, stomach, and mesentery, but the number of peritoneal mast cells decreased to one-sixth that of +/+ mice. Cultured mast cells and peritoneal mast cells of B6-mi(vit)/mi(vit) mice showed a reduced but apparently detectable level of SgIGSF expression, demonstrating the parallelism between mast cell number and expression level of SgIGSF. The number of peritoneal mast cells appeared to be influenced by MITF through transcription of SgIGSF.

Animals↗

PubMed2004

Deficient eosinophil chemotaxis-promoting activity of genetically normal mast cells transplanted into subcutaneous tissue of Mitfmi-vga9/Mitfmi-vga9 mice: comparison of the activity and mast cell distribution pattern with KitW/KitW-vMice.

Despite the practical lack of mast cells in the skin tissue of WBB6F(1)-Kit(W)/Kit(W-v), the skin tissue of WBB6F(1)-Mitf(mi-vga9)/Mitf(mi-vga9) mice contains one third of mast cells than that of WBB6F(1)-+/+ mice. We attempted to investigate the function of the decreased but appreciable number of mast cells in the skin of WBB6F(1)-Mitf(mi-vga9)/Mitf(mi-vga9) mice. The substance P (SP)-induced eosinophil infiltration was examined using air-bleb assay. The air-bleb membrane was composed of the subcutaneous connective tissue. Unexpectedly, we found that the air-bleb membranes formed in the back of WBB6F(1)-Mitf(mi-vga9)/Mitf(mi-vga9) mice contained no mast cells. The WBB6F(1)-Mitf(mi-vga9)/Mitf(mi-vga9) mice showed impaired SP-induced eosinophil infiltration as observed in WBB6F(1)-Kit(W)/Kit(W-v) mice, indicating that mast cells detected in the dermis of WBB6F(1)-Mitf(mi-vga9)/Mitf(mi-vga9) mice did not help SP-induced eosinophil infiltration. Subcutaneous transplantation of cultured mast cells from WBB6F(1)-+/+ mice normalized SP-induced eosinophil infiltration in WBB6F(1)-Kit(W)/Kit(W-v) mice but not in WBB6F(1)-Mitf(mi-vga9)/Mitf(mi-vga9) mice. The greater number and the more dispersed distribution pattern of mast cells that appeared in the subcutaneous connective tissue of WBB6F(1)-Kit(W)/Kit(W-v) mice after the transplantation appeared to explain the difference between WBB6F(1)-Kit(W)/Kit(W-v) and WBB6F(1)-Mitf(mi-vga9)/Mitf(mi-vga9) mice.

Animals↗

PubMed2004

Strain-dependent inhibitory effect of mutant mi-MITF on cytotoxic activities of cultured mast cells and natural killer cells of mice.

MITF is a transcription factor encoded by the mi locus. MITF encoded by mi and Mi(or) mutant alleles (mi-MITF and Mi(or)-MITF, respectively) possessed an inhibitory effect, whereas the tg, mi(ew) and mi(ce) were null mutants. We examined the cytotoxic activities of cultured mast cells (CMCs) and natural killer (NK) cells of various MITF mutants in C57BL/6 (B6) background. Cytotoxic activities of CMCs and NK cells of B6-mi/mi and B6-Mi(or)/Mi(or) mice were remarkably reduced. In B6-tg/tg, B6-mi(ew)/mi(ew) and B6-mi(ce)/mi(ce) mice, however, the cytotoxic activity of CMCs was reduced only slightly and the NK activity was normal. The cytotoxic activity of CMCs paralleled with the expression level of granzyme B (Gr B) mRNA, and the NK activity with that of perforin (Pfn) mRNA. In contrast to the case of B6-mi/mi mice, cytotoxic activities of CMCs and NK cells were not impaired in WB-mi/mi mice. The expression of Gr B mRNA was not reduced in CMCs of WB-mi/mi mice, and that of Pfn mRNA was not reduced in NK cells of WB-mi/mi mice. WB-mi/mi mice appeared to have factor(s) compensating for the inhibitory effect of mi-MITF on the expression of Gr B and Pfn genes.

Animals↗

PubMed2002

Additive effect of mouse genetic background and mutation of MITF gene on decrease of skin mast cells.

The mi transcription factor (MITF) is a basic-helix-loop-helix leucine zipper transcription factor and is encoded by mi locus. The mi/mi mutant mice showed a significant decrease of skin mast cells in C57BL/6 (B6) genetic background but not in WB genetic background. Kit ligand (KitL) is the most important growth factor for development of mast cells, and the decrease of skin mast cells in B6-mi/mi mice was attributable to the reduced expression of c-kit receptor tyrosine kinase (KIT) that is a receptor for KitL. However, the expression level of KIT in WB-mi/mi mast cells was comparable with that of B6-mi/mi mast cells, suggesting that a factor compensating the reduced expression of KIT was present in WB-mi/mi mice. By linkage analysis, such a factor was mapped on chromosome 10. The mapped position was closely located to the KitL locus. Two alternative spliced forms are known in KitL mRNA: KL-1 and KL-2. Soluble KitL, which is important for development of skin mast cells, is produced more efficiently from KL-1 mRNA than from KL-2 mRNA. The KL-1/KL-2 ratio was higher in WB-mi/mi than in B6-mi/mi mice, suggesting that the larger amount of soluble KitL may compensate for the reduced expression of KIT in WB-mi/mi mice.

Alternative Splicing↗

PubMed2002

Dual abnormal effects of mutant MITF encoded by Mi(wh) allele on mouse mast cells: decreased but recognizable transactivation and inhibition of transactivation.

MITF is a basic helix-loop-helix leucine zipper-type transcription factor and is important for development of mast cells. MITF encoded by Mi(wh) allele (Mi(wh)-MITF) was mutated at a single amino acid of basic domain, and possessed a deficient but apparent DNA-binding ability. Here, we characterized the unique effects of Mi(wh)-MITF on the expression of mast cell-related genes. The expression level of mouse mast cell protease (mMCP)-4, -5, and -6 genes in Mi(wh)/Mi(wh) cultured mast cells (CMCs) was intermediate between levels of normal (+/+) CMCs and tg/tg CMCs, which did not express any MITFs. Mi(wh)-MITF appeared to show the positive transactivation effect through the remaining DNA-binding ability. On the other hand, the expression level of tryptophan hydroxylase gene was lower in Mi(wh)/Mi(wh) CMCs than in tg/tg CMCs, suggesting the inhibitory effect of Mi(wh)-MITF on the transactivation. Mi(wh)-MITF possessed dual abnormal effects on transactivation of mast cell-related genes.

Alleles↗

PubMed2002

Isoforms of mi transcription factor preferentially expressed in cultured mast cells of mice.

MITF is a basic helix-loop-helix leucine zipper transcription factor, which is important for normal phenotypic expression of mast cells. Three isoforms of MITF have been known in mice, MITF-A, -H, and -M. Since cultured mast cells (CMCs) are useful for studying the function of MITF, we examined isoforms of MITF expressed in CMCs using 5'-RACE, and found a new isoform of MITF, MITF-E. We assessed the relative mRNA amount of various MITF isoforms with reverse transcription-PCR. When the mRNA amount of MITF-E was used as a standard, that of MITF-M was approximately 10%, that of MITF-H was approximately 1%, and that of MITF-A was approximately 0.1%. Although MITF-E was the preferential isoform in CMCs, peritoneal mast cells expressed only MITF-M. The expression profile of MITF isoforms appeared to be influenced by the developing process of mast cells.

3T3 Cells↗

PubMed2001

Interaction and cooperation of mi transcription factor (MITF) and myc-associated zinc-finger protein-related factor (MAZR) for transcription of mouse mast cell protease 6 gene.

The mi transcription factor (MITF) is a basic-helix-loop-helix leucine zipper (bHLH-Zip) transcription factor that is important for the normal phenotypic expression of mast cells. Most transcription factors function in cooperation with other factors by protein-protein interactions. To search proteins interacting with MITF, we carried out a yeast two-hybrid screen and isolated Myc-associated zinc-finger protein related factor (MAZR) as a partner of MITF. When expressed with MITF in NIH/3T3 cells, MAZR was colocalized with MITF. The association of MAZR with MITF was further confirmed by a co-immunoprecipitation study and in vitro binding assay. The zinc-finger domain of MAZR and the Zip domain of MITF were essential for the interaction. MAZR was expressed in cultured mast cells and MST mastocytoma cells containing mouse mast cell protease (mMCP)-6 transcript abundantly. The overexpression of dominant negative MAZR in MST mastocytoma cells reduced the amount of mMCP-6 mRNA. The simultaneous transfection of MAZR and MITF significantly increased the promoter activity of the mMCP-6 gene, indicating that the MAZR and MITF synergistically transactivated the mMCP-6 gene. MAZR appeared to play important roles in the normal phenotypic expression of mast cells in association with MITF.

3T3 Cells↗