Search PubMed⌕ Search

Biomedical subjects

Y Yazaki

Publications and source records attributed to Y Yazaki.

At least 109 records · Page 6Linked to original sources
PubMed1999

Mouse jagged1 physically interacts with notch2 and other notch receptors. Assessment by quantitative methods.

The Delta/Serrate/LAG-2 (DSL) domain containing proteins are considered to be ligands for Notch receptors. However, the physical interaction between DSL proteins and Notch receptors is poorly understood. In this study, we cloned a cDNA for mouse Jagged1 (mJagged1). To identify the receptor interacting with mJagged1 and to gain insight into its binding characteristics, we established two experimental systems using fusion proteins comprising various extracellular parts of mJagged1, a "cell" binding assay and a "solid-phase" binding assay. mJagged1 physically bound to mouse Notch2 (mNotch2) on the cell surface and to a purified extracellular portion of mNotch2, respectively, in a Ca(2+)-dependent manner. Scatchard analysis of mJagged1 binding to BaF3 cells and to the soluble Notch2 protein demonstrated dissociation constants of 0.4 and 0.7 nM, respectively, and that the number of mJagged1-binding sites on BaF3 is 5,548 per cell. Furthermore, deletion mutant analyses showed that the DSL domain of mJagged1 is a minimal binding unit and is indispensable for binding to mNotch2. The epidermal growth factor-like repeats of mJagged1 modulate the affinity of the interaction, with the first and second repeats playing a major role. Finally, solid-phase binding assay showed that Jagged1 binds to Notch1 and Notch3 in addition to Notch2, suggesting that mJagged1 is a ligand for multiple Notch receptors.

Amino Acid Sequence↗
PubMed1999

Embryonic lethality and defective neural tube closure in mice lacking squalene synthase.

Squalene synthase (SS) catalyzes the reductive head-to-head condensation of two molecules of farnesyl diphosphate to form squalene, the first specific intermediate in the cholesterol biosynthetic pathway. We used gene targeting to knock out the mouse SS gene. The mice heterozygous for the mutation (SS+/-) were apparently normal. SS+/- mice showed 60% reduction in the hepatic mRNA levels of SS compared with SS+/+ mice. Consistently, the SS enzymatic activities were reduced by 50% in the liver and testis. Nevertheless, the hepatic cholesterol synthesis was not different between SS+/- and SS+/+ mice, and plasma lipoprotein profiles were not different irrespective of the presence of the low density lipoprotein receptor, indicating that SS is not a rate-limiting enzyme in the cholesterol biosynthetic pathway. The mice homozygous for the disrupted SS gene (SS-/-) were embryonic lethal around midgestation. E9.5-10.5 SS-/- embryos exhibited severe growth retardation and defective neural tube closure. The lethal phenotype was not rescued by supplementing the dams either with dietary squalene or cholesterol. We speculate that cholesterol is required for the development, particularly of the nervous system, and that the chorioallantoic circulatory system is not mature enough to supply the rapidly growing embryos with maternal cholesterol at this developmental stage.

Animals↗
PubMed1999

First systematic chiral syntheses of two pairs of enantiomers with 3,5-dihydroxyheptenoic acid chain, associated with a potent synthetic statin NK-104.

First systematic chiral syntheses of two pairs of enantiomers with 3,5-dihydroxyheptenoic acid chain, associated with a potent synthetic statin NK-104 are reported. A pair of syn diol isomers (NK-104 and its enantiomer) was obtained efficiently by diastereomeric resolution. The synthesis of a pair of anti diol isomers (3-epimer and 5-epimer) was accomplished effectively by the asymmetric aldol reaction followed by anti stereoselective reduction as key steps. Their purity determinations were effected by chiral HPLC analysis.

Heptanoic Acids↗
PubMed1999

Aberrant expression of cAMP-response-element-binding protein ('CREB') induces apoptosis.

We have reported previously that cAMP-response-element-binding protein (CREB) was phosphorylated in a cell-cycle-dependent manner, showing that it was phosphorylated at early S-phase at casein kinase II target sites. To assess the possible involvement of CREB in cell cycle progression, CREB expression vector was transiently transfected into various cells. Unexpectedly we found that transfection with CREB expression vector resulted in an abundance of dead cells. Morphological examination revealed that these cells had undergone apoptosis. The coincidence of CREB overexpression and apoptosis induction at the individual cell level was confirmed by a immunohistochemical study. To confirm that overexpression of CREB was the cause of apoptosis, a dominant-negative mutant of CREB, KCREB, was co-expressed with the wild type. The co-existence of KCREB effectively rescued CREB-mediated apoptosis in a dose-dependent manner, verifying that apoptosis was truly a specific effect of overexpressed CREB and not an artifact of the transfection procedure. Deletion analysis indicates that neither the Q1 transactivation domain, which functions in transcription, nor the kinase-inducible domain, in which a cluster of various kinase targets exists, is necessary; however, the Q2 transactivation domain is required for the induction of apoptosis. A more precise study indicates that the four-residue stretch Glu-Glu-Ala-Ala at the most C-terminal region of the Q2 domain is especially important for the induction of apoptosis. Thus overexpressed CREB induces apoptosis by transmitting certain signals from the C-terminal portion of the Q2 domain. Possible roles of cell-cycle-regulated phosphorylation and also an elevation of the intracellular cAMP level in CREB-induced apoptosis are suggested.

Amino Acid Sequence↗
PubMed1999

Endothelin-1 induces expression of fetal genes through the interleukin-6 family of cytokines in cardiac myocytes.

We here examined the role of the interleukin-6 (IL-6) family of cytokines in endothelin-1 (ET-1)-induced hypertrophic responses using cultured cardiac myocytes of neonatal rats. ET-1 induced expression of IL-6 and leukemia inhibitory factor (LIF) genes. ET-1-induced LIF gene expression was abolished by inhibition of protein kinase C activity. ET-1 activated the promoter of atrial natriuretic peptide and beta-type myosin heavy chain genes through the tyrosine kinase pathway and IL-6 receptor gp130. These results suggest that the IL-6 family of cytokines mediates ET-1-induced expression of some fetal genes in cardiac myocytes.

Animals↗
PubMed1999

p85/p110-type phosphatidylinositol kinase phosphorylates not only the D-3, but also the D-4 position of the inositol ring.

Activation of p85/p110-type phosphatidylinositol (PI) kinase has been implicated in various cellular activities. This PI kinase phosphorylates the D-4 position with a similar or higher efficiency than the D-3 position when trichloroacetic acid-treated cell membrane is used as a substrate, although it phosphorylates almost exclusively the D-3 position of the inositol ring in phosphoinositides when purified PI is used as a substrate. Furthermore, the lipid kinase activities of p110 for both the D-3 and D-4 positions were completely abolished by introducing kinase-dead point mutations in their lipid kinase domains (DeltaKinalpha and DeltaKinbeta, respectively). In addition, both PI 3- and PI 4-kinase activities of p110alpha and p110beta immunoprecipitates were similarly inhibited by either wortmannin or LY294002, specific inhibitors of p110. Insulin induced phosphorylation of not only the D-3 position, but also the D-4 position. Indeed, overexpression of p110 in Sf9 or 3T3-L1 cells induced marked phosphorylation of the D-4 position to a level comparable to or much greater than that of D-3, whereas inhibition of endogenous p85/p110-type PI kinase via overexpression of dominant-negative p85alpha (Deltap85alpha) in 3T3-L1 adipocytes abolished insulin-induced synthesis of both. Thus, p85/p110-type PI kinase phosphorylates the D-4 position of phosphoinositides more efficiently than the D-3 position in vivo, and each of the D-3- or D-4-phosphorylated phosphoinositides may transmit signals downstream.

1-Phosphatidylinositol 4-Kinase↗
PubMed1999

BTEB2, a Krüppel-like transcription factor, regulates expression of the SMemb/Nonmuscle myosin heavy chain B (SMemb/NMHC-B) gene.

We have recently characterized the promoter region of the rabbit embryonic smooth muscle myosin heavy chain (SMemb/NMHC-B) gene and identified the 15-bp sequence, designated SE1, located at -105 from the transcriptional start site as an important regulatory element for its transcriptional activity in a smooth muscle cell (SMC) line. In this study, we attempted to isolate cDNA clones encoding for the transcription factors that control the expression of the SMemb gene through binding to this cis-regulatory element. We screened a lambdagt11 cDNA library prepared from C2/2 cells, a rabbit-derived SMC line, by using a radiolabeled concatenated oligonucleotide containing SE1 as a probe. Sequence analysis revealed that one of the cDNA clones corresponds to the rabbit homologue of basic transcriptional element binding protein-2 (BTEB2), which has previously been identified as one of the Krüppel-like transcription factor. Gel mobility shift assays and antibody supershift analyses with nuclear extracts from C2/2 cells indicate that BTEB2 is a major component of nuclear factor:SE1 complexes. Furthermore, a glutathione S-transferase-BTEB2 fusion protein binds to the SE1 in a sequence-specific manner. In support of the functionality of BTEB2 binding, basal promoter activity and BTEB2-induced transcriptional activation were markedly attenuated by the disruption of the SE1. In adult rabbit tissues, BTEB2 mRNA was most highly expressed in intestine, urinary bladder, and uterus. BTEB2 mRNA levels were downregulated in rabbit aorta during normal development. Moreover, immunohistochemical analysis indicated a marked induction of BTEB2 protein in the neointimal SMC after balloon injury in rat aorta. These results suggest that BTEB2 mediates the transcriptional regulation of the SMemb/NMHC-B gene and possibly plays a role in regulating gene expression during phenotypic modulation of vascular SMC.

Age Factors↗
PubMed1999

Frequent false-positive results of Aspergillus latex agglutination test: transient Aspergillus antigenemia during neutropenia.

BACKGROUND: Two serologic assays, Aspergillus latex agglutination testing (LA) and plasma (1-->3)-beta-D-glucan (BDG) measurement, are used when invasive pulmonary aspergillosis (IPA) is suspected. Despite the high specificity of these assays, false-positive results are frequent for neutropenic patients. This study was conducted to evaluate the efficacy of LA and BDG and to investigate the cause of the false-positive results. METHODS: Eighty-eight consecutive patients with hematologic malignancies who underwent intensive chemotherapy were tested weekly with LA and BDG. RESULTS: Sixteen of 88 patients were diagnosed as having IPA. The sensitivity, specificity, and positive predictive values were 23%, 98%, and 64% for LA and 27%, 88%, and 52% for BDG, respectively. Of 11 patients who became positive for LA only during neutropenic periods, 2 patients developed IPA. In contrast, six of eight patients who became positive for LA during nonneutropenic periods developed IPA. Transient Aspergillus antigenemia was more frequently encountered during neutropenia (2.9%) than during nonneutropenic periods (0.2%). The plasma BDG concentration increased at the nadir of neutropenia in 36 of 45 patients who had no signs of IPA, and it exceeded the level of 20 pg/mL in 2 patients. CONCLUSIONS: Both BDG and LA have a low sensitivity and a high specificity for IPA. However, the false-positive rate of LA increases during neutropenic periods. Caution should be exercised in interpreting the results of these blood tests, especially when patients are neutropenic.

Adolescent↗
PubMed1999

Improvement of impaired myocardial vasodilatation due to diffuse coronary atherosclerosis in hypercholesterolemics after lipid-lowering therapy.

BACKGROUND: Diminished myocardial vasodilatation (MVD) in hypercholesterolemics without overt coronary stenosis has been reported. However, whether the diminished MVD of angiographically normal coronary arteries in hypercholesterolemics can be reversed after lipid-lowering therapy is not known. METHODS AND RESULTS: A total of 27 hypercholesterolemics and 16 age-matched controls were studied. All patients had >1 normal coronary artery, and those segments that were perfused by anatomically normal coronary arteries were studied. Myocardial blood flow (MBF) was measured during dipyridamole loading and at baseline using positron emission tomography and 13N-ammonia, after which MVD was calculated before and after lipid-lowering therapy. Total cholesterol was significantly higher in hypercholesterolemics (263+/-33.8) than in controls (195+/-16.6), and it normalized after lipid-lowering therapy (197+/-19.9). Baseline MBF (ml. min-1. 100 g-1) was comparable among hypercholesterolemics (both before and after therapy) and controls. MBF during dipyridamole loading was significantly lower in hypercholesterolemics before therapy (189+/-75.4) than in controls (299+/-162, P<0.01). However, MBF during dipyridamole loading significantly increased after therapy (226+/-84.7; P<0.01). MVD significantly improved after therapy in hypercholesterolemics (2.77+/-1.35 after treatment [P<0.05] versus 2. 02+/-0.68 before treatment [P<0.01]), but it remained significantly higher in controls (3.69+/-1.13, P<0.01). There was a significant relationship between the percent change of total cholesterol and the percent change of MVD before and after lipid-lowering therapy (r=-0. 61, P<0.05). CONCLUSIONS: Diminished MVD of anatomically normal coronary arteries in hypercholesterolemics can be reversed after lipid-lowering therapy.

Aged↗
PubMed1999

Heart-specific activation of LTK results in cardiac hypertrophy, cardiomyocyte degeneration and gene reprogramming in transgenic mice.

Leukocyte tyrosine kinase (LTK) is a receptor-type protein tyrosine kinase belonging to the insulin receptor superfamily. To elucidate its biological role, we generated transgenic mice expressing LTK under the control of cytomegarovirus enhancer and beta-actin promoter. The transgenic mice exhibited growth retardation and most of the transgenic mice died within several months after birth. Interestingly, although LTK was expressed in several major organs, the activation (tyrosine-phosphorylation, kinase activity, and multimerization) of LTK was observed selectively in the heart, where LTK was localized on intracellular membrane, presumably on endoplasmic reticulum. Echocardiography showed that the transgenic heart underwent severe concentric hypertrophy, which resulted in reduced cardiac output, low blood pressure, and increased heart rate. Histological examination of the heart exhibited focal degeneration of cardiomyocytes. These histological changes were considered to be due to apoptosis, based on the finding that the sarcolemmas of the degenerative cardiomyocytes were well preserved. In addition, expression of fetal genes, such as atrial natriuretic peptide and skeletal alpha-actin, was markedly induced in the transgenic heart. These results indicate that a certain tissue-specific mechanism of activating LTK exists in the heart and that the activated LTK resulted in cardiac hypertrophy, cardiomyocyte degeneration and gene reprogramming. These findings will provide novel insights into the activating mechanism and biological role of LTK in vivo.

Animals↗
PubMed1999

The molecular mechanism of cardiac hypertrophy and failure.

Mechanical stretch induced by high blood pressure is an initial factor leading to cardiac hypertrophy. In an in vivo study, an angiotensin II (AngII) type 1 receptor antagonist TCV116 reduced left ventricular (LV) weight, LV wall thickness, transverse myocyte diameter, relative amount of V3 myosin heavy chain, and interstitial fibrosis, while treatment with hydralazine did not. In an in vitro study using cultured cardiomyocytes, mechanical stretch activated second messengers such as mitogen-activated protein (MAP) kinase, followed by increased protein synthesis. Additionally, in the stretch-conditioned medium AngII and endothelin-1 concentrations were increased. Furthermore, the Na+/H+ exchanger activated by mechanical stretch modulated the hypertrophic responses of cardiomyocytes. The pathways leading to MAP kinase activation differed between cell types. In cardiac fibroblasts AngII activated MAP kinase via G beta gamma subunit of Gi, Src, Shc, Grb2, and Ras, whereas Gq and protein kinase C were critical in cardiomyocytes.

Angiotensin II↗
PubMed1999

Role of the renin-angiotensin system in cardiac hypertrophy.

Hypertrophy is an adaptive response of the heart to hemodynamic overload such as hypertension. However, it is generally accepted that cardiac hypertrophy is one of the most critical risk factors of heart disease. Therefore, for the treatment of hypertension it is important to understand the mechanism of cardiac hypertrophy and to establish effective pharmaceutical interventions. Mechanical stretch induced by hypertension is an initial factor leading to cardiac hypertrophy. In an in vivo study using spontaneously hypertensive rats, an angiotensin II type 1 receptor antagonist, TCV116, decreased left ventricular weight, left ventricular wall thickness, transverse myocyte diameter, relative amount of V3 myosin heavy chain, and interstitial fibrosis, whereas treatment with hydrolazine did not. In an in vitro study using cultured cardiomyocytes of neonatal rats, mechanical stretch activated second messengers, such as extracellular signal-regulated protein kinase (ERK), followed by increased protein synthesis. Additionally, in the stretch-conditioned medium, the levels of angiotensin II and endothelin-1 concentrations were increased. Moreover, the Na+/H+ exchanger activated by mechanical stretch modulated the hypertrophic responses of cardiomyocytes. To further elucidate whether angiotensin II is indispensable for mechanical stress-induced cardiac hypertrophy, mechanical stretch-induced ERK activation was examined in angiotensin II type 1a receptor knock-out mice. Although the addition of angiotensin II had no effects on the ERK activity in cardiomyocytes of angiotensin II type 1a receptor knockout mice, mechanical stretch induced a larger increase in the ERK activity in cardiac myocytes from these mice compared with cardiac myocytes of wild-type mice. These results suggest that mechanical stretch could induce hypertrophic responses in cardiac myocytes even in the absence of angiotensin II. The pathways leading to ERK activation differed between cell types. In cardiac fibroblasts, angiotensin II activated ERK via the G(beta)gamma subunit of Gi, Src, Shc, Grb2, and Ras, whereas Gq and protein kinase C were critical in cardiomyocytes.

Angiotensin II↗
PubMed1999

Vascular endothelial growth factor induces activation and subcellular translocation of focal adhesion kinase (p125FAK) in cultured rat cardiac myocytes.

Vascular endothelial growth factor (VEGF) has been proposed to be among the candidate factors with the most potential to play a role in ischemia-induced collateral vessel formation. Recently, we found that VEGF activated the mitogen-activated protein kinase cascade in cultured rat cardiac myocytes. To elucidate how VEGF affects adhesive interaction of cardiac myocytes with the extracellular matrix (ECM), one of the important cell functions, we investigated the molecular mechanism of activation of focal adhesion-related proteins, especially focal adhesion kinase (p125(FAK)), in cultured rat cardiac myocytes. We found that the 2 VEGF receptors, KDR/Flk-1 and Flt-1, were expressed in cardiac myocytes and that KDR/Flk-1 was significantly tyrosine phosphorylated on VEGF stimulation. VEGF induced tyrosine phosphorylation and activation of p125(FAK) as well as tyrosine phosphorylation of paxillin; this was accompanied by subcellular translocation of p125(FAK) from perinuclear sites to the focal adhesions. This VEGF-induced activation of p125(FAK) was inhibited partially by the tyrosine kinase inhibitors genistein and tyrphostin. Activation of p125(FAK) was accompanied by its increased association with adapter proteins GRB2, Shc, and nonreceptor type tyrosine kinase p60(c-src). Furthermore, we confirmed that VEGF induced a significant increase in adhesive interaction between cardiac myocytes and ECM using an electric cell-substrate impedance sensor. These results strongly suggest that p125(FAK) is one of the most important components in VEGF-induced signaling in cardiac myocytes, playing a critical role in adhesive interaction between cardiac myocytes and ECM.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Molecular and histological evaluation of pancreata from patients with a mitochondrial gene mutation associated with impaired insulin secretion.

A mutation in mitochondrial DNA, which was originally identified in patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS), can be associated with a subtype of diabetes mellitus. To determine the molecular and histological basis of impaired insulin secretion in the subjects with this mutation, we studied autopsy pancreata specimens from eight subjects diagnosed as having MELAS. The 3243 bp mutation was identified in seven out of eight pancreata examined. Immunohistochemical studies demonstrated a reduction in total islet mass, and in the numbers of both B and A cells. No evidence of insulitis or apoptosis was found. These data suggested that the 3243 bp mutation may cause the reduction of islet cells, mainly through mechanisms other than autoimmune destruction.

Adolescent↗
PubMed1999

Pulsatile stretch activates mitogen-activated protein kinase (MAPK) family members and focal adhesion kinase (p125(FAK)) in cultured rat cardiac myocytes.

Recently, we demonstrated that pulsatile mechanical stretch induced rapid secretion of vascular endothelial growth factor (VEGF) by cultured rat cardiac myocytes in vitro. To investigate whether pulsatile stretch activates intracellular signaling in cardiac myocytes, we examined the activation of mitogen-activated protein kinase (MAPK) family members and focal adhesion kinase (p125(FAK)) in cultured rat cardiac myocytes. We found that pulsatile stretch rapidly phosphorylated p44/p42 MAPKs (extracellular signal-regulated protein kinase [ERK] 1/2), stress-activated protein kinase (SAPK), p38MAPK, and p125(FAK). The stretch-induced activation of ERKs was at least partly mediated by VEGF, which was shown to be induced by transforming growth factor (TGF)-beta, and was also partly dependent on tyrosine kinases as well as protein kinase C (PKC). These data provide the direct evidence that pulsatile stretch can activate intracellular signaling in cardiac myocytes and that this was at least partly mediated by VEGF, which may play a role in cardiac adaptation to mechanical overload.

Angiotensin II↗