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Hiroyuki Sano

Publications and source records attributed to Hiroyuki Sano.

At least 19 recordsLinked to original sources
PubMed2006

The 47kDa Akt substrate associates with phosphodiesterase 3B and regulates its level in adipocytes.

We have previously described a novel putative 47kDa substrate for the protein kinase Akt (designated AS47) in 3T3-L1 adipocytes. In the present study, we have found by co-immunoprecipitation that AS47 was associated with cyclic nucleotide phosphodiesterase 3B (PDE3B) in lysates of 3T3-L1 adipocytes. The patterns of expression of AS47 and PDE3B upon 3T3-L1 adipocyte differentiation, among mouse tissues, and in adipocytes with and without the transcription factor C/EBPalpha were virtually coincident. Partial knockdown of AS47 in 3T3-L1 adipocytes with shRNA resulted in a similar reduction in PDE3B protein. These results indicate that AS47 exists in a complex with PDE3B in adipocytes and that the amount of AS47 protein regulates the amount of PDE3B.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed2006

[A case of acute pulmonary thromboembolism accompanied by anomalous inferior vena cava with azygous continuation].

A 38-year-old man with atopic dermatitis presented with right chest pain and dyspnea. Previously, he had received 2mg of betamethasone daily, to prevent rejection of the right transplanted cornea, for 24 days. His body temperature was 37.4 degrees C, peripheral leucocyte count measured 12,000/mm3, and C-reactive protein was 6.3 mg/dl. A computed tomogram of the chest revealed infiltration in the right lower lung field, and he was then treated for pneumonia. The second day he fell down one flight of stairs due to a syncopal attack and received a head injury. At this point his vital blood pressure was 102/55 mmHg, heart rate was 130/min and SpO2 under breathing room air was 76%. These findings indicated possible acute pulmonary thromboembolism. Enhanced computed tomography revealed pulmonary arteries occluded by massive thrombosis and anomalous inferior vena cava with azygous continuation. To decrease the risk of further cerebral bleeding, anti-coagulation therapy was administered with only 24,000 IU/day of heparin. Following treatment, the patient completely recovered. We reported this rare case of acute pulmonary thromboembolism accompanied by anomalous inferior vena cava with azygous continuation.

Acute Disease↗
PubMed2005

Dexamethasone inhibits phosphorylation of histone H3 at serine 10.

Glucocorticoids are the most effective anti-inflammatory drugs used in the treatment of inflammatory diseases. While phosphorylation of histone H3 at serine 10 (p-Ser10) is one of the histone modifications related to transcription of some inflammation-related genes, the effect of glucocorticoids on p-Ser10 is not established. Here, we investigated the ability of dexamethasone (Dex) to inhibit p-Ser10 expression in response to tumor necrosis factor (TNF-alpha) in the human lung adenocarcinoma cell line A549 and the SV-40-transformed human airway epithelial cell line BEAS-2B. By Western blot analysis in BEAS-2B cells, the expression of p-Ser10 was repressed by pretreatment with Dex, an effect not seen in A549 cells. Flow cytometric analysis at a single-cell level in A549 cells indicated that TNF-alpha treatment caused early induction of p-Ser10 at 15 min, which was inhibited significantly by pretreatment with 10(-5) M Dex. By immunostaining, the p-Ser10 signal appeared as granules in TNF-alpha-treated cells at same sites of phosphorylated RNA polymerase II. In contrast, the signal was scattered in the nuclei of Dex-pretreated cells. These findings suggested that Dex limits airway inflammation by inhibiting p-Ser10 expression and redistributing p-Ser10 away from transcription sites.

Anti-Inflammatory Agents↗
PubMed2005

AS160, the Akt substrate regulating GLUT4 translocation, has a functional Rab GTPase-activating protein domain.

Recently, we described a 160 kDa protein (designated AS160, for Akt substrate of 160 kDa) with a predicted Rab GAP (GTPase-activating protein) domain that is phosphorylated on multiple sites by the protein kinase Akt. Phosphorylation of AS160 in adipocytes is required for insulin-stimulated translocation of the glucose transporter GLUT4 to the plasma membrane. The aim of the present study was to determine whether AS160 is in fact a GAP for Rabs, and, if so, what its specificity is. We first identified a group of 16 Rabs in a preparation of intracellular vesicles containing GLUT4 by MS. We then prepared the recombinant GAP domain of AS160 and examined its activity against many of these Rabs, as well as several others. The GAP domain was active against Rabs 2A, 8A, 10 and 14. There was no significant activity against 14 other Rabs. GAP activity was further validated by the finding that the recombinant GAP domain with the predicted catalytic arginine residue replaced by lysine was inactive. Finally, it was found by immunoblotting that Rabs 2A, 8A and 14 are present in GLUT4 vesicles. These results indicate that AS160 is a Rab GAP, and suggest novel Rabs that may participate in GLUT4 translocation.

Adipocytes↗
PubMed2005

Synip phosphorylation does not regulate insulin-stimulated GLUT4 translocation.

Insulin causes the rapid translocation of the glucose transporter GLUT4 from intracellular sites to the plasma membrane in fat and muscle cells. There is considerable evidence that the signaling to this trafficking process is downstream of the insulin-activated protein kinase Akt. One Akt substrate that connects signaling to trafficking is a 160 kDa GTPase activating protein for Rabs. Another potential connecting substrate is the protein Synip, which associates with the SNARE syntaxin4. A recent study presents evidence that Akt phosphorylates Synip on serine 99, at least in vitro, and proposes that this phosphorylation enables GLUT4 translocation by causing the dissociation of Synip from syntaxin4. In the present study we show that marked overexpression of Synip mutant S99A, which lacks this phosphorylation site, has no effect on insulin-stimulated GLUT4 translocation in 3T3-L1 adipocytes. This finding is strong evidence that phosphorylation of Synip on serine 99 is not required for GLUT4 translocation.

3T3-L1 Cells↗
PubMed2005

Underrecognition of the severity of asthma and undertreatment of asthma in a rural area of Japan.

BACKGROUND AND AIM: Revised guidelines were released in Japan in 2003 for the assessment, treatment, and management of adult asthmatics, and similar guidelines for child asthmatics were released in 2002. We reassessed the severity and possible undertreatment of asthma according to these guidelines in stable asthmatics. METHODS: We reviewed medical records of 861 well-controlled asthmatic patients who, in April through June 2004 were cared for by 47 pulmonologists at 29 medical centers and 13 asthma clinics in a rural community in the San-in area of Japan. The physician obtained completed medical records about their symptoms and current treatment of the subjects, 726 adult and 135 children (aged 6 years or older) who were in stable condition and had had no exacerbations in the previous 3 months. The severity of asthma and current treatment for each patient were assessed according to the newly revised Japanese guidelines for the assessment, treatment, and management of adult and child asthmatics. RESULTS: In adult and child asthmatics, the percentage of predicted forced expiratory volume at 1 second (FEV1.0) was smaller and has a narrower distribution range than the percentage of predicted peak expiratory flow (PEF). When the severity of asthma was classified according to symptoms alone, 50% and 35% of those classified as mildly asthmatics patients with adults and children, respectively, had moderate to severe airflow limitation. Inhaled corticosteroids were prescribed to 90.6% of adult and 14.9% of child patients. When we compared the treatments that patients were actually receiving against the optimal treatments indexed according to a combined symptoms-FEV1.0 classification, we found that 49% of adult asthmatics were overtreated, 21% were properly treated, and 30% were undertreated. Among children, the respective percentages were 35%, 25%, and 40%. CONCLUSION: In well-controlled adult and child asthmatics, the severity of asthma is poorly judged when symptoms alone are considered. We suggest that the severity of asthma should be assessed through a combination of symptoms and the measurement of FEV1.0 during office visits. We also suggest that the proper dose of inhaled steroid needed to maintain stable conditions should be judged according to this combined symptoms-FEV1.0 classification.

Administration, Inhalation↗
PubMed2005

The Totton-Ken Seibu earthquake and exacerbation of asthma in adults.

The aim of the study was to characterize patients at risk for exacerbations of their asthma as a result of the Tottori-Ken Seibu earthquake and to identify factors that predict exacerbation of asthma after an earthquake. A retrospective cohort study-analysis was conducted of 156 asthmatic patients, aged 18 to 89 years, who were out-patients of Tottori University Hospital and who had completely recorded their asthmatic symptoms and measured their peak expiratory flow (PEF) rates for more than one year prior to the earthquake. Seventeen (11%) patients who experienced the earthquake were identified as having an exacerbation within one month after the earthquake. Diurnal variability of PEF during the month after the earthquake was compared to values during a matched month one year previously. When factors associated with exacerbation were identified by a review of the medical case notes and the contribution of these factors to the exacerbation was determined using multivariate analysis, airflow limitation was shown to be independently associated with exacerbation after the earthquake. Acute asthma attacks are more likely to occur within the first week after the earthquake event without diurnal PEF variability. Asthma is likely to worsen after an earthquake.

Adult↗
PubMed2004

ERK activation and subsequent RB phosphorylation are important determinants of the sensitivity to paclitaxel in lung adenocarcinoma cells.

Paclitaxel is used frequently in the treatment of advanced non-small cell lung cancer. This study was carried out in order to determine the role of extracellular signal-regulated kinases (ERK) and retinoblastoma protein (pRB) in the governing mechanism resistance to paclitaxel using two lung adenocarcinoma cell lines with differing sensitivities. In paclitaxel-sensitive Ma-10 cells, treatment with paclitaxel induced pRB phosphorylation at Ser795 and ERK activation. In contrast, in paclitaxel-resistant Ma-31 cells, paclitaxel dephosphorylated pRB at Ser795 without affecting ERK activity. A specific ERK inhibitor, PD98059, blocked paclitaxel-induced ERK activation and pRB phosphorylation at Ser795 in Ma-10 cells. Furthermore, PD98059 inhibited cell cycle progression during paclitaxel treatment, the accumulation of sub-G1 population, and the cytotoxic effect by paclitaxel in Ma-10 cells, suggesting that ERK activation by paclitaxel, subsequent pRB phosphorylation, and the cell cycle progression during paclitaxel treatment are important determinants of sensitivity to paclitaxel. These observations raise the possibility that the promotion of cell cycle during the exposure of lung cancer cells to paclitaxel may sensitize resistant cells to paclitaxel.

Adenocarcinoma↗
PubMed2003

Insulin-stimulated phosphorylation of a Rab GTPase-activating protein regulates GLUT4 translocation.

Insulin stimulates the rapid translocation of intracellular glucose transporters of the GLUT4 isotype to the plasma membrane in fat and muscle cells. The connections between known insulin signaling pathways and the protein machinery of this membrane-trafficking process have not been fully defined. Recently, we identified a 160-kDa protein in adipocytes, designated AS160, that is phosphorylated by the insulin-activated kinase Akt. This protein contains a GTPase-activating domain (GAP) for Rabs, which are small G proteins required for membrane trafficking. In the present study we have identified six sites of in vivo phosphorylation on AS160. These sites lie in the motif characteristic of Akt phosphorylation, and insulin treatment increased phosphorylation at five of the sites. Expression of AS160 with two or more of these sites mutated to alanine markedly inhibited insulin-stimulated GLUT4 translocation in 3T3-L1 adipocytes. Moreover, this inhibition did not occur when the GAP function in the phosphorylation site mutant was inactivated by a point mutation. These findings strongly indicate that insulin-stimulated phosphorylation of AS160 is required for GLUT4 translocation and that this phosphorylation signals translocation through inactivation of the Rab GAP function.

3T3 Cells↗
PubMed2003

Elevated urinary 8-hydroxydeoxyguanosine, a biomarker of oxidative stress, and lack of association with antioxidant vitamins in chronic obstructive pulmonary disease.

OBJECTIVE: The purpose of this study was to investigate whether patients with COPD are under oxidative stress and to elucidate the relationship between the level of oxidative stress and antioxidant vitamins. METHODOLOGY: Nineteen male patients with COPD and 13 age- matched male control subjects were studied. Urinary 8-hydroxydeoxyguanosine (8-OHdG) concentrations were determined using an enzyme-linked immunosorbent assay kit and corrected for creatinine concentrations. Serum levels of vitamin C, alpha-tocopherol, and beta-carotene were determined by high performance liquid chromatography. RESULTS: The median (interquartile range) 8-OHdG excretion was 8.1 ng/mg (5.3-10.9 ng/mg) in control subjects and 12.2 ng/mg (9.8-15.5 ng/mg) in COPD patients (P < 0.01). Urinary 8-OHdG levels were significantly elevated in ex-smokers in the COPD group compared with ex-smokers in the control group. Urinary 8-OHdG level was negatively correlated with FVC (r = -0.42, P = 0.016), FEV1 (r = -0.49, P = 0.0048), and oxygen tension in arterial blood (r = -0.41, P = 0.0005). No significant differences in antioxidant levels were demonstrated between the two groups. There were no significant correlations between urinary 8-OHdG excretion and the serum concentrations of antioxidant vitamins. CONCLUSION: The burden of oxidative stress was observed to increase in COPD patients as judged by urinary 8-OHdG. A depletion of antioxidant vitamins in serum was not essential for this phenomenon. Elevated urinary 8-OHdG level may not be attributable to smoking status or to antioxidant vitamins in COPD.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed2003

Sequence variability and candidate gene analysis in two cancer patients with complex clinical outcomes during morphine therapy.

In this case report, we present genetic differences in two morphine-related gene sequences, UDP-glucuronosyltransferase 2B7 (UGT2B7) and mu opioid receptors (MOR1), in two cancer patients whose clinical responses to morphine were very different [i.e., sensitive (patient 1) and low responder (patient 2)]. In addition, allelic variants in the UGT2B7 gene were analyzed in 46 Japanese individuals. Amplified DNA fragments for the two genes of interest were screened using single strand conformation polymorphism and then sequenced. In the UGT2B7 gene, 12 single nucleotide polymorphisms (SNPs) were newly identified with an allelic frequency ranging from 0.022 to 0.978. Six SNPs in the promoter region (A-1302G, T-1295C, T-1111C, G-899A, A-327G, and T-125C) and two coding SNPs (UGT2B7*2 in exon 2 and C1059G in exon 4) appeared to be consistently linked. Remarkable differences in the nucleotide sequence of UGT2B7 were observed between the two patients; in contrast to patient 1 who had "reference" alleles at almost SNP positions, but a rare ATTGAT*2(AT)C haplotype as homozygosity, patient 2 was a homozygous carrier for the predominant GCCAGC*1(TC)G sequence. Serum morphine and two glucuronide concentrations in patient 2 suggest that the predominant GCCAGC*1G sequence was not associated with a "poor metabolizer" phenotype. In the MOR1 gene, patient 1 had no SNPs, whereas patient 2 was a heterozygous carrier for both the G-1784A and A118G alleles. The present study describes substantial differences in genotype patterns of two genes of interest between the two patients. The results necessitate larger trials to confirm these observations in larger case control studies.

Aged↗
PubMed2003

[Cavitary pulmonary metastasis from bladder cancer: a case report].

A 69 year-old [correction of 63] man who had had a radical cystectomy for bladder cancer was admitted to our hospital because of hemosputum and right femoral pain. His chest radiograph and computed tomogram showed a mass shadow with a cavity in the left upper lung field. Sputum cytology showed class V squamous cell carcinoma and a bone scintigram showed right femoral metastasis. Despite radiotherapy to the left upper lung and the right femur, the patient's condition worsened, and he died of respiratory failure after hospitalization for about 1 month. At autopsy, pathologic studies of lung cancer revealed mixed-type transitional cell carcinoma, squamous cell carcinoma and adenocarcinoma. A diagnosis of metastatic lung cancer from bladder cancer was made. Cavitating pulmonary metastasis is uncommon. We report a rare case of pulmonary metastasis from bladder cancer, with mixed-type histopathology at both primary and metastatic sites.

Adenocarcinoma↗
PubMed2002

Dissociation of DNA damage and mitochondrial injury caused by hydrogen peroxide in SV-40 transformed lung epithelial cells.

BACKGROUND: Since lung epithelial cells are constantly being exposed to reactive oxygen intermediates (ROIs), the alveolar surface is a major site of oxidative stress, and each cell type may respond differently to oxidative stress. We compared the extent of oxidative DNA damage with that of mitochondrial injury in lung epithelial cells at the single cell level. RESULT: DNA damage and mitochondrial injury were measured after oxidative stress in the SV-40 transformed lung epithelial cell line challenged with hydrogen peroxide (H2O2). Single cell analysis of DNA damage was determined by assessing the number of 8-oxo-2-deoxyguanosine (8-oxo-dG) positive cells, a marker of DNA modification, and the length of a comet tail. Mitochondrial membrane potential, DeltaPsim, was determined using JC-1. A 1 h pulse of H2O2 induced small amounts of apoptosis (3%). 8-oxo-dG-positive cells and the length of the comet tail increased within 1 h of exposure to H2O2. The number of cells with reduced DeltaPsim increased after the addition of H2O2 in a concentration-dependent manner. In spite of a continual loss of DeltaPsim, DNA fragmentation was reduced 2 h after exposure to H2O2. CONCLUSION: The data suggest that SV-40 transformed lung epithelial cells are resistant to oxidative stress, showing that DNA damage can be dissociated from mitochondrial injury.

Journal Article↗
PubMed2002

Group V phospholipase A2 induces leukotriene biosynthesis in human neutrophils through the activation of group IVA phospholipase A2.

We reported previously that exogenously added human group V phospholipase A(2) (hVPLA(2)) could elicit leukotriene B(4) (LTB(4)) biosynthesis in human neutrophils (Han, S. K., Kim, K. P., Koduri, R., Bittova, L., Munoz, N. M., Leff, A. R., Wilton, D. C., Gelb, M. H., and Cho, W. (1999) J. Biol. Chem. 274, 11881-11888). To determine the mechanism of the hVPLA(2)-induced LTB(4) biosynthesis in neutrophils, we thoroughly examined the effects of hVPLA(2) and their lipid products on the activity of group IVA cytosolic PLA(2) (cPLA(2)) and LTB(4) biosynthesis under different conditions. As low as 1 nm exogenous hVPLA(2) was able to induce the release of arachidonic acid (AA) and LTB(4). Typically, AA and LTB(4) were released in two phases, which were synchronized with a rise in intracellular calcium concentration ([Ca(2+)](i)) near the perinuclear region and cPLA(2) phosphorylation. A cellular PLA(2) assay showed that hVPLA(2) acted primarily on the outer plasma membrane, liberating fatty acids and lysophosphatidylcholine (lyso-PC), whereas cPLA(2) acted on the perinuclear membrane. Lyso-PC and polyunsaturated fatty acids including AA activated cPLA(2) and 5-lipoxygenase by increasing [Ca(2+)](i) and inducing cPLA(2) phosphorylation, which then led to LTB(4) biosynthesis. The delayed phase was triggered by the binding of secreted LTB(4) to the cell surface LTB(4) receptor, which resulted in a rise in [Ca(2+)](i) and cPLA(2) phosphorylation through the activation of mitogen-activated protein kinase, extracellular signal-regulated kinase 1/2. These results indicate that a main role of exogenous hVPLA(2) in neutrophil activation and LTB(4) biosynthesis is to activate cPLA(2) and 5-lipoxygenase primarily by liberating from the outer plasma membrane lyso-PC that induces [Ca(2+)](i) increase and cPLA(2) phosphorylation and that hVPLA(2)-induced LTB(4) production is augmented by the positive feedback activation of cPLA(2) by LTB(4).

Calcium↗
PubMed2002

A method to identify serine kinase substrates. Akt phosphorylates a novel adipocyte protein with a Rab GTPase-activating protein (GAP) domain.

This study describes a method for the identification of the substrates of specific serine kinases. An antibody specific for the phosphomotif generated by the kinase is used to isolate phosphorylated substrates by immunoprecipitation, and the isolated proteins are identified by tandem mass spectrometry of peptides. This method was applied to the identification of substrates for the protein kinase Akt, which specifically phosphorylates the RXRXXS/T motif. 3T3-L1 adipocytes were treated with insulin to activate Akt, and the putative Akt substrate proteins were isolated by immunoprecipitation with an antibody against the phospho form of this motif. This led to the identification of a novel 160-kDa substrate for Akt. The 160-kDa substrate for Akt, which was designated AS160, has a Rab GAP domain. Recombinant AS160 was shown to be a substrate for Akt, and two sites of phosphorylation, both in RXRXXS/T motifs, were identified by mass spectrometry and mutation. Insulin treatment of adipocytes caused AS160 to redistribute from the low density microsomes to the cytosol.

3T3 Cells↗
PubMed2002

Insulin receptor substrate 4 associates with the protein IRAS.

The insulin receptor substrates (IRSs) are key components in signaling from the insulin receptor, and consequently any proteins that interact with them are expected to participate in insulin signaling. In this study we have searched for proteins that interact with IRS-4 by identifying the proteins that coimmunoprecipitated with IRS-4 from human embryonic kidney 293 cells by microsequencing through mass spectrometry. A group of proteins was found. These included phosphatidylinositol 3-kinase, a protein previously identified as an IRS-4 interactor, and several proteins for which there was no previous evidence of IRS-4 association. One of these proteins, named IRAS, that had been found earlier in another context was examined in detail. The results from the overexpression of IRAS, where its amount was about the same as that of IRS-4, indicated that IRAS associated directly with IRS-4 and showed that the increased complexation of IRS-4 with IRAS did not alter the insulin-stimulated tyrosine phosphorylation of IRS-4 or the association of IRS-4 with phosphatidylinositol 3-kinase or Grb2. On the other hand, overexpression of IRAS enhanced IRS-4-dependent insulin stimulation of the extracellularly regulated kinase. The domains of IRAS and IRS-4 responsible for the association of these two proteins were identified, and it was shown that IRAS also associates with IRS-1, IRS-2, and IRS-3.

Adaptor Proteins, Signal Transducing↗
PubMed2002

IL-5-induced integrin adhesion of human eosinophils caused by ERK1/2-mediated activation of cPLA2.

We examined the mechanism by which interleukin (IL)-5 causes beta(2)-integrin adhesion of human eosinophils. IL-5 caused time-dependent activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38alpha in eosinophils as detected by their phosphorylation. Preincubation of eosinophils with U0126, a mitogen-activated protein kinase/ERK kinase inhibitor, suppressed IL-5-induced activation of cytosolic phospholipase A(2) (cPLA(2)) and eosinophil adhesion, and p38 inhibition by SB203580 had neither effect. ERK1/2 phosphorylation and eosinophil adhesion were blocked by inhibition of the src-family tyrosine kinase, Janus tyrosine kinase (JAK)2, or phosphoinositide-3 kinase (PI3K). Coimmunoprecipitation assay demonstrated that Lyn, a src-family tyrosine kinase, was constitutively associated with PI3K. Inhibition of src-tyrosine kinase but not JAK2 suppressed PI3K activation. Our data suggest that IL-5 induces beta(2)-integrin adhesion of human eosinophils by regulation of cPLA(2) activation caused by ERK1/2 phosphorylation. This phosphorylation results from activation of PI3K and protein tyrosine kinases. We also find that src-family tyrosine kinase, possibly Lyn, is the upstream kinase causing PI3K activation.

Cell Adhesion↗