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Biomedical subjects

Masahide Takahashi

Publications and source records attributed to Masahide Takahashi.

At least 19 recordsLinked to original sources

[New porous gelatin particles for hepatic arterial embolization--investigation of passage through current microcatheters].

Hepatic arterial embolization (HAE) has been widely performed for unresectable hepatocellular carcinoma (HCC) in a selective manner using a microcatheter. Gelatin sponge is exclusively used as an embolic agent, though it has not yet been approved for an arterial blockade. Recently, new porous gelatin particles (GP) have been approved and are clinically available. In this study, passage of GP through microcatheters was investigated, as it is an important requirement for a new agent. Both 1-mm and 2-mm GP were injected through each of six current 2.0 Fr-2.7 Fr microcatheters by five board-certified interventional radiologists. In general, use of a 2.5 mL lock syringe was associated with higher subjective resistance score (5-10/10), longer injection time (20-30 sec), and lower injection pressure (1.0-2.5 MPa) with difficulty. When it was impossible to inject GP with a 2.5-mL lock syringe,injection was retested with a 1 mL lock syringe resulting in lower resistance score (1-6), shorter injection time (6-10 sec), and higher injection pressure (1.5-2.5 MPa) regardless of particle size. Thus,it is recommended to use a 1 mL lock syringe for injection of GP through a microcatheter. After passing through a 2.0 Fr microcatheter, 2 mm particles showed no significant deformation or fragmentation under observation by stereomicroscopy.

Catheterization↗

Enhancer of polycomb1, a novel homeodomain only protein-binding partner, induces skeletal muscle differentiation.

Homeodomain only protein, Hop, is an unusual small protein that modulates target gene transcription without direct binding to DNA. Here we show that Hop interacts with Enhancer of Polycomb1 (Epc1), a homolog of a Drosophila polycomb group gene product that regulates transcription, to induce the skeletal muscle differentiation. Yeast two-hybrid assay with the human adult heart cDNA library revealed that Hop can associate with Epc1. The amino-terminal domain of Epc1 as well as full Epc1 physically interacted with Hop in mammalian cells and in yeast. Epc1 is highly expressed in the embryonic heart and adult skeletal muscles. Serum deprivation induced differentiation of H9c2, a myoblast cell line, into skeletal myocytes, and Epc1 was up-regulated. Differentiation of H9c2 was induced by Epc1 overexpression, although it was severely impaired in Epc1-knockdown cells. Co-transfection of Hop potentiated Epc1-induced transactivation of myogenin and myotube formation. Hop knock-out mice elicited a decrease in myosin heavy chain and myogenin expressions in skeletal muscle and showed delay in hamstring muscle healing after injury. Differentiation was impaired in skeletal myoblasts from Hop knock-out mice. These results suggest that Epc1 plays a role in the initiation of skeletal muscle differentiation, and its interaction with Hop is required for the full activity.

Animals↗

Protective effect of hyperpigmented skin on UV-mediated cutaneous cancer development.

Recently, we crossed an original haired RET-transgenic mouse of line 242 with a hairless mouse and established a hairless RET-(HL/RET)-transgenic mouse line (242-hr/hr) with hyperpigmented skin but no tumors. In this study, we examined the effect of hyperpigmented skin in HL/RET-transgenic mice on UV irradiation-mediated cutaneous cancer development. UV irradiation to this mouse line never induced melanoma despite the presence of melanoma-inducible transgenic RET oncogenes. On the contrary, the hyperpigmented skin efficiently protected UV-mediated squamous carcinoma development in the skin. Probably underlying this result, hyperpigmentation protected the skin from damage and blocked the accompanying signal transduction for tyrosine phosphorylation of multiple cellular proteins and activation/phosphorylation of extracellular signal-regulated, c-Jun N-terminal, and p38 kinases. Thus, we demonstrated hyperpigmentation-mediated in vivo protection against UV irradiation-induced skin cancer.

Animals↗

Targeted mutation of serine 697 in the Ret tyrosine kinase causes migration defect of enteric neural crest cells.

The RET receptor tyrosine kinase plays a critical role in the development of the enteric nervous system (ENS) and the kidney. Upon glial-cell-line-derived neurotrophic factor (GDNF) stimulation, RET can activate a variety of intracellular signals, including the Ras/mitogen-activated protein kinase, phosphatidylinositol 3-kinase (PI3K)/AKT, and RAC1/JUN NH(2)-terminal kinase (JNK) pathways. We recently demonstrated that the RAC1/JNK pathway is regulated by serine phosphorylation at the juxtamembrane region of RET in a cAMP-dependent manner. To determine the importance of cAMP-dependent modification of the RET signal in vivo, we generated mutant mice in which serine residue 697, a putative protein kinase A (PKA) phosphorylation site, was replaced with alanine (designated S697A mice). Homozygous S697A mutant mice lacked the ENS in the distal colon, resulting from a migration defect of enteric neural crest cells (ENCCs). In vitro organ culture showed an impaired chemoattractant response of the mutant ENCCs to GDNF. JNK activation by GDNF but not ERK, AKT and SRC activation was markedly reduced in neurons derived from the mutant mice. The JNK inhibitor SP600125 and the PKA inhibitor KT5720 suppressed migration of the ENCCs in cultured guts from wild-type mice to comparable degrees. Thus, these findings indicated that cAMP-dependent modification of RET function regulates the JNK signaling responsible for proper migration of the ENCCs in the developing gut.

Animals↗

The efficacy of sodium valproate and a MRA finding in confusional migraine.

Confusional migraine (CM) is recognized as a kind of pediatric migraine equivalent. We report that a 10-year-old girl had repeated attacks of confusional state and of migraine with aura. She suffered from total of six CM attacks and migraine attacks with aura almost once a month. Therefore, sodium valproate (VPA) was initiated to prevent these attacks. They have been controlled after increasing her dosage to 600mg/day. The intracranial magnetic resonance angiography (MRA) during the last CM attack indicated narrowing of left middle and posterior cerebral arteries. However, MRA during the attack free period at 41-day follow-up showed normal vessels. Electroencephalograms showed severe slowing without seizure discharges during the CM attacks. This case indicates that VPA is an effective treatment for the prophylaxis of CM and migraine with aura. MRA is considered helpful for the diagnosis of this kind of migraine in children because of noninvasive neuroimaging.

Anticonvulsants↗

Dok-4 regulates GDNF-dependent neurite outgrowth through downstream activation of Rap1 and mitogen-activated protein kinase.

During development of the central and peripheral nervous systems, neurite extension mediated via glial-cell-line-derived neurotrophic factor (GDNF) and its receptor RET is critical for neuronal differentiation. In the present study, we investigated the role of the RET substrate Dok-4 in neurite outgrowth induced by the GDNF/RET signaling pathway. In TGW neuroblastoma cells, which endogenously express both RET and Dok-4, depletion of Dok-4 through treatment with small interfering RNA resulted in a marked decrease in GDNF-stimulated neurite outgrowth. By contrast, exogenous expression of wild-type Dok-4 induced sustained p44/42 mitogen-activated protein kinase (ERK1/2) activation and enhanced neurite outgrowth. Expression of Dok-4 mutants in which the tyrosine residues at codons 187, 220 and 270, conserved between Dok-4, -5, and -6, were each replaced with a phenylalanine inhibited sustained ERK1/2 activation and neurite outgrowth. We also found that Dok-4 induced a significant activation of the small G protein Rap1 and that expression of a dominant active Rap1 mutant restored neurite outgrowth in Dok-4-depleted cells. By contrast, expression of a dominant negative Rap1 mutant impaired GDNF-stimulated neurite outgrowth from TGW cells. Finally, we found that neurite formation in cultured rat hippocampal neurons was enhanced by the expression of Dok-4. Together, our results suggest that Dok-4, through activation of the Rap1-ERK1/2 pathway, regulates GDNF-mediated neurite outgrowth during neuronal development.

Adaptor Proteins, Signal Transducing↗

Template-free magnesium oxide hollow sphere inclusion in organic-inorganic hybrid films via sol-gel reaction.

Magnesium oxide hollow spheres without a template core were conveniently prepared by stabilized bubble formation in a hybrid solution containing a magnesium acetate precursor, thus avoiding the complicated preparation process using a template. The hollow sphere could be aligned along the radial striation by spin coating, and its diameter from a micrometer to submicrometer dimension could be easily modified by the solution composition. It was also possible to control the open or closed hollow sphere by changing the solvent. Thus, the produced magnesium oxide hollow sphere is envisioned to have applications in many areas such as medicine, analysis, optics, and so on.

Magnesium Oxide↗

Viscoelastic and structural properties of a phenyl-modified polysiloxane system with a three-dimensional structure.

The relationships between the viscoelastic and structural properties of glass-forming materials with polysiloxane bonds, which serve as network formers, and phenyl groups, which act as network terminators, are examined based on shear viscoelasticity, (29)Si MAS NMR, and GPC measurements during the early stages of the network-forming process. The viscosities of the present samples do not depend on the frequency at temperatures up to 200 degrees C, suggesting that the origin of the viscous flow does not include intermolecular entanglement. According to the results of the strain dependence of the elastic modulus, the bridging-oxygen number, and molecular weight, the present polysiloxane system has a complex structure, or distribution of various-sized molecules composed of a polysiloxane network with various dimensionalities, and furthermore an elementary process of the viscosity is simple flow of these molecules. The structural factors that determine the viscosity and its temperature dependence are categorized into the molecular size and the intramolecular structure by using a theory based on the free-volume model. The relationship between the viscosity and the structure around the glass transition temperature is quantitatively examined and it is concluded that introducing larger numbers of Ph groups makes the viscosity less sensitive to structural factors.

Journal Article↗

A PKC-mediated backup mechanism of the MXXCW motif-linked switch for initiating tyrosine kinase activities.

The cysteine in the M/IXXCW motif is conserved in all but one (threonine in place of cysteine) of the human protein tyrosine kinases (PTKs). We showed that all RET-PTC-1 mutants in which the C in this motif (C376) was replaced with glycine, lysine, threonine or serine lost their activity in vitro. However, the C376T/S mutants showed normal tyrosine phosphorylation in vivo (in cells). Further analyses reveled that protein kinase C (PKC) initiated the activities of the C376T/S mutants in cells. We conclude that the M/IXXCW motif-mediated mechanisms which initiate PTK activities are partially replaced by a PKC-mediated mechanism.

Amino Acid Motifs↗

RET and neuroendocrine tumors.

The RET proto-oncogene encodes a receptor tyrosine kinase that is a main component of the signaling pathway activated by the glial cell line-derived neurotrophic factor family ligands. Gene targeting studies revealed that signaling through RET plays a crucial role in neuronal and renal organogenesis. It is well-known that germline mutations in RET lead to the human inherited diseases, multiple endocrine neoplasia type 2 (MEN 2) and Hirschsprung's disease, and that somatic rearrangements of RET cause papillary thyroid carcinoma. Due to marked advances in understanding of the molecular mechanisms of the development of MEN 2, a consensus on MEN 2 management associated with RET status is being reached and currently put into general use as a guideline. In this review, we summarize progress in the study of RET from bench to bedside, focusing on pathophysiology of neuroendocrine tumors.

Adrenal Gland Neoplasms↗

Analyses of ultraviolet-induced focus formation of hREV1 protein.

Translesional DNA synthesis (TLS) is one of the DNA damage tolerance mechanisms that allow cells with DNA damage to continue DNA replication. Each of the mammalian Y-family DNA polymerases (Pol eta, Pol iota, Pol kappa, and REV1) has been shown to carry out TLS by itself or in combination with another enzyme in vitro. Recently, the C-terminal region of mammalian REV1 (the total 1251 residues in human) was found to interact with Pol eta, Pol iota, and Pol kappa, as well as with the REV7 subunit of another TLS enzyme, Pol zeta. Thus, it is proposed that REV1 plays a pivotal role in TLS in vivo. We here describe our study on the localization of human REV1 protein (hREV1) in nondamaged and ultraviolet (UV)-irradiated cells. Ectopically expressed hREV1 in mammalian cells was localized to the nucleus and exhibited dozens of tiny foci in approximately 3% of nondamaged cells. The percentage of focus-forming cells markedly increased after UV irradiation in a time- and dose-dependent manner. The focus formation was associated with UV-induced DNA damage. Interestingly, although the hREV1 foci in S-phase cells colocalized with PCNA foci, suggesting the association of hREV1 with the replication machinery, hREV1 focus formation was observed not only in the S phase but also outside S phase. Furthermore, it was found that the hREV1 focus formation after UV irradiation required a region near the C-terminal (826-1178).

Animals↗

RET receptor signaling: dysfunction in thyroid cancer and Hirschsprung's disease.

Gain-of-function mutations within the receptor tyrosine kinase gene RET cause inherited and non-inherited thyroid cancer. Somatic gene rearrangements of RET have been found in papillary thyroid carcinoma and germline point mutations in multiple endocrine neoplasia (MEN) types 2A and 2B and familial medullary thyroid carcinoma (FMTC). Conversely, loss-of-function mutations are responsible for the development of Hirschsprung's disease, a congenital malformation of the enteric nervous system. Comparison between normal RET signaling activated by the RET ligand glial cell line-derived neurotrophic factor (GDNF) and abnormal RET signaling caused by various mutations has led to a deeper understanding of disease mechanisms. The focus of the present review is on recent progress in the study of RET signaling dysfunction in human diseases.

Animals↗

Girdin, a novel actin-binding protein, and its family of proteins possess versatile functions in the Akt and Wnt signaling pathways.

Girdin (GIRDers of actIN filament, also reported as APE, GIV, or HkRP1) is a novel protein expressed ubiquitously in mammals and was recently identified as a binding partner of the serine/threonine kinase Akt. We found that Girdin is an actin-binding protein involved in both the remodeling of the actin cytoskeleton and in cell motility. Recent studies have uncovered new and varied functions of Girdin. For example, it prolongs the activation of Akt and regulates DNA replication in response to insulin signaling. Girdin also associates with heterotrimeric G proteins and dynamin (a large GTPase), which are involved in membrane transport. We found that Akt phosphorylates Girdin in response to growth factors such as epidermal growth factor (EGF) in fibroblasts. Furthermore, phosphorylated Girdin accumulates at the leading edge of migrating cells, suggesting its role in Akt-dependent cell migration or tumor invasion. To date, two paralogues of Girdin have been identified in mammals. One of them, Daple (also named HkRP2), is a binding partner and regulator of Dishevelled, an important cytoplasmic component of the Wnt signaling pathway. Another is a protein encoded by FLJ00354 (also named HkRP3), which has not been functionally characterized. These members are assumed to be dimers with large polypeptide chains (220-250 kDa) that associate at their central long coiled-coil domains. The amino acid sequences of the members show about 32-46% overall identity, with the greatest variation toward the carboxyl terminal domains that specify the binding partners. Here we propose roles for this family of proteins in diverse cellular processes.

Animals↗

Microspherule protein 1, Mi-2beta, and RET finger protein associate in the nucleolus and up-regulate ribosomal gene transcription.

The nucleolus is the site of ribosomal DNA (rDNA) transcription and ribosome production. In exploring the role of nucleolar protein MCRS1 (microspherule protein1)/MSP58 (58-kDa microspherule protein), we found that Mi-2beta, a component of a nucleosome remodeling and deacetylase (NuRD) complex, RET finger protein (RFP), and upstream binding factor (UBF) were associated with MCRS1. Yeast two-hybrid assays revealed that MCRS1 bound to the ATPase/helicase region of Mi-2beta and the coiled-coil region of RFP. Interestingly, confocal microscopic analyses revealed the co-localization of MCRS1, Mi-2beta, RFP, and the rRNA transcription factor UBF in the nucleoli. We also found that MCRS1, Mi-2beta, and RFP were associated with rDNA using a chromatin immunoprecipitation assay. Finally, we showed that MCRS1, Mi-2beta, and RFP up-regulated transcriptional activity of the rDNA promoter and that ribosomal RNA transcription was repressed when MCRS1, Mi-2beta, and RFP expression was reduced using siRNA. These results indicated that Mi-2beta and RFP, known to be involved in transcriptional repression in the nucleus, co-localize with MCRS1 in the nucleolus and appear to activate the rRNA transcription.

Adenosine Triphosphatases↗

PIAS proteins are involved in the SUMO-1 modification, intracellular translocation and transcriptional repressive activity of RET finger protein.

Ret finger protein (RFP) is a nuclear protein that is highly expressed in testis and in various tumor cell lines. RFP functions as a transcriptional repressor and associates with Enhancer of Polycomb 1 (EPC1), a member of the Polycomb group proteins, and Mi-2beta, a main component of the nucleosome remodeling and deacetylase (NuRD) complex. We show that RFP binds with PIAS (protein inhibitor of activated STAT) proteins, PIAS1, PIAS3, PIASxalpha and PIASy at their carboxyl-terminal region and is covalently modified by SUMO-1 (sumoylation). PIAS proteins enhance the sumoylation of RFP in a dose-dependent manner and induce the translocation of RFP into nuclear bodies reminiscent of the PML bodies. In addition, co-expression of PIAS proteins or SUMO-1 strengthened the transcriptional repressive activity of RFP. Finally, our immunohistochemical results show that RFP, SUMO-1 and PIASy localize in a characteristic nuclear structure juxtaposed with the inner nuclear membrane (XY body) of primary spermatocytes in mouse testis. These results demonstrate that the intracellular location and the transcriptional activity of RFP are modified by PIAS proteins which possess SUMO E3 ligase activities and suggest that they may play a co-operative role in spermatogenesis.

Animals↗

GDNF-inducible zinc finger protein 1 is a sequence-specific transcriptional repressor that binds to the HOXA10 gene regulatory region.

The RET tyrosine kinase receptor and its ligand, glial cell line-derived neurotrophic factor (GDNF) are critical regulators of renal and neural development. It has been demonstrated that RET activates a variety of downstream signaling cascades, including the RAS/mitogen-activated protein kinase and phosphatidylinositol-3-kinase(PI3-K)/AKT pathways. However, nuclear targets specific to RET-triggered signaling still remain elusive. We have previously identified a novel zinc finger protein, GZF1, whose expression is induced during GDNF/RET signaling and may play a role in renal branching morphogenesis. Here, we report the DNA binding property of GZF1 and its potential target gene. Using the cyclic amplification and selection of targets technique, the consensus DNA sequence to which GZF1 binds was determined. This sequence was found in the 5' regulatory region of the HOXA10 gene. Electrophoretic mobility shift assay revealed that GZF1 specifically binds to the determined consensus sequence and suppresses transcription of the luciferase gene from the HOXA10 gene regulatory element. These findings thus suggest that GZF1 may regulate the spatial and temporal expression of the HOXA10 gene which plays a role in morphogenesis.

Animals↗