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Biomedical subjects

M Hasegawa

Publications and source records attributed to M Hasegawa.

At least 37 records · Page 2Linked to original sources

Self-energy correction to momentum-density distribution of positron-electron pairs.

Positron two-dimensional angular correlation of annihilation radiation (2D ACAR), i.e., the 2D projection of the electron momentum densities sampled by positron, in Si is employed to verify the prediction of the density functional theory within the local-density approximation (LDA). Carefully conducted test shows that the LDA introduces small but definite discrepancies to the 2D-ACAR anisotropies. Self-energy calculation using the GW method indicates that density-fluctuation contributes anisotropic momentum-density correction and thus improves the agreement between theory and experiment. These results provide valuable annotations to the arguments concerning the accuracy and validity of the LDA and GW schemes.

Journal Article↗

Evidence for muon neutrino oscillation in an accelerator-based experiment.

We present results for nu(mu) oscillation in the KEK to Kamioka (K2K) long-baseline neutrino oscillation experiment. K2K uses an accelerator-produced nu(mu) beam with a mean energy of 1.3 GeV directed at the Super-Kamiokande detector. We observed the energy-dependent disappearance of nu(mu), which we presume have oscillated to nu(tau). The probability that we would observe these results if there is no neutrino oscillation is 0.0050% (4.0 sigma).

Journal Article↗

Efficient and stable Sendai virus-mediated gene transfer into primate embryonic stem cells with pluripotency preserved.

Efficient gene transfer and regulated transgene expression in primate embryonic stem (ES) cells are highly desirable for future applications of the cells. In the present study, we have examined using the nonintegrating Sendai virus (SeV) vector to introduce the green fluorescent protein (GFP) gene into non-human primate cynomolgus ES cells. The GFP gene was vigorously and stably expressed in the cynomolgus ES cells for a year. The cells were able to form fluorescent teratomas when transplanted into immunodeficient mice. They were also able to differentiate into fluorescent embryoid bodies, neurons, and mature blood cells. In addition, the GFP expression levels were reduced dose-dependently by the addition of an anti-RNA virus drug, ribavirin, to the culture. Thus, SeV vector will be a useful tool for efficient gene transfer into primate ES cells and the method of using antiviral drugs should allow further investigation for regulated SeV-mediated gene expression.

Animals↗

Cysteinyl leukotrienes induce monocyte chemoattractant protein 1 in human monocytes/macrophages.

BACKGROUND: Monocytes/macrophages have a cysteinyl leukotriene 1 (CysLT1) receptor, but its function is poorly understood. Objective To elucidate the biological function of the CysLT1 receptor of human monocytes/macrophages. METHODS: We examined the production of TNF-alpha, IL-1beta, IL-2, IL-4, IL-6, IL-8, IL-10, monocyte chemoattractant protein 1 (MCP-1), macrophage colony-stimulating factor (M-CSF), and eotaxin induced by CysLTs (leukotriene (LT)C4, -D4, and -E4) in THP-1 cells, a human monocytic leukaemia cell line, and peripheral blood CD14+ monocytes/macrophages. Moreover, we examined the effect of CysLTs on the expression of beta-chemokine receptor 2B (CCR2B) as the receptor of MCP-1 by Western blot analysis. RESULTS: ELISA revealed that CysLTs induced MCP-1 in THP-1 cells and peripheral blood CD14+ monocytes/macrophages, but not other cytokines. PCR demonstrated that CysLTs increased MCP-1 mRNA expression in THP-1 cells, and Western blotting showed that CysLTs increased the expression of CCR2B in THP-1 cells. Moreover, we demonstrated that pranlukast, a CysLT1 receptor antagonist, blocked MCP-1 production by CysLTs in THP-1 cells almost completely, and partially inhibited MCP-1 release by CysLTs in peripheral blood CD14+ monocytes/macrophages and CCR2B expression by CysLTs in THP-1 cells. CONCLUSION: CysLTs induce MCP-1 and increase CCR2B expression in human monocytes/macrophages.

Blotting, Western↗

Recognition and signal transduction mechanism of Escherichia coli heat-stable enterotoxin and its receptor, guanylate cyclase C.

Guanylate cyclase C (GC-C), a member of the membrane-bound GC family, consists of an extracellular domain (ECD) and an intracellular domain, which are connected by a single-transmembrane region. GC-C is a receptor protein, i.e. specifically stimulated by the endogenous peptides guanylin, uroguanylin, lymphoguanylin, and the exogenous peptide heat-stable enterotoxin (ST(a)), secreted by pathogenic Escherichia coli and acting on the intestinal brush border membranes. The binding of these peptide ligands to the ECD of GC-C results in the synthesis of cyclic GMP in cells, which, in turn, regulates a variety of intracellular physiologic processes. As the cloning of GC-C, its physiologic functions of each domain have been vigorously investigated. The structural characterization of the ligand-binding domain of the receptor promises to provide important clues for better understanding of the mechanisms of receptor recognition and activation. Recently, structural data for each domain of membrane-bound GCs and related proteins has become available. Coupling information obtained from such work and validation of structure-function relationships of GC-C and its ligands should allow for three-dimensional mapping of their interaction site in detail. Our approach to this issue involved designing photoaffinity-labeling ST(a) analogs, capable of binding covalently to the ligand-binding region of the ECD of GC-C. The photoaffinity-labeling ligand was used to covalently label a soluble form of the recombinant ECD protein. Mass spectrometric analyses of an endoproteinase digest of the ECD revealed that the ligand specifically bound to a narrow region contained in the membrane-proximal subdomain of the ECD of GC-C. These results will enable us to identify the possible binding motifs within the ligand-binding domain by computer modeling. In this review, we summarize the available data on the recognition mechanism between ST(a) and GC-C at the molecular level.

Amino Acid Sequence↗

Up regulated expression of fractalkine/CX3CL1 and CX3CR1 in patients with systemic sclerosis.

BACKGROUND: Fractalkine expressed on endothelial cells mediates activation and adhesion of leucocytes expressing its receptor, CX(3)CR1. Soluble fractalkine exhibits chemotactic activity for leucocytes expressing CX(3)CR1. OBJECTIVE: To determine the role of fractalkine and its receptor in systemic sclerosis (SSc) by assessing their expression levels in patients with this disease. METHODS: The expression of fractalkine and CX(3)CR1 in the skin and lung tissues was immunohistochemically examined. Circulating soluble fractalkine levels were examined by enzyme linked immunosorbent assay (ELISA). Blood samples from patients with SSc were stained for CX(3)CR1 with flow cytometric analysis. RESULTS: CX(3)CR1 levels on peripheral monocytes/macrophages and T cells were found to be raised in patients with diffuse cutaneous SSc. The numbers of cells expressing CX(3)CR1, including monocytes/macrophages, were increased in the lesional skin and lung tissues from patients with diffuse cutaneous SSc. Fractalkine was strongly expressed on endothelial cells in the affected skin and lung tissues. Soluble fractalkine levels were significantly raised in sera and were associated with raised erythrocyte sedimentation rates, digital ischaemia, and severity of pulmonary fibrosis. CONCLUSIONS: Up regulated expression of fractalkine and CX(3)CR1 cooperatively augments the recruitment of mononuclear cells expressing CX(3)CR1 into the affected tissue of SSc, leading to inflammation and vascular injury.

Adult↗

Increased E2F-1 expression via tumour cell proliferation and decreased apoptosis are correlated with adverse prognosis in patients with squamous cell carcinoma of the oesophagus.

BACKGROUND: The retinoblastoma (Rb) pathway, which governs cell cycle progression, is frequently genetically altered in cancer, causing deregulated expression of the E2F-1 transcription factor, which promotes DNA synthesis and cell cycle progression. Recent studies show that E2F-1 also participates in apoptosis induction in a p53 dependent or independent manner. Despite its crucial role and paradoxical effects on cell turnover, the function of E2F-1 in human cancer is unclear. AIMS: To evaluate E2F-1 expression using immunohistochemistry in 43 surgically resected oesophageal squamous cell carcinoma (OSCC) specimens. METHODS: This study analysed the association of E2F-1 with tumour cell proliferation and apoptosis and the upstream regulators modulating these processes, and its impact on patient outcome. Tumour cell proliferation and apoptosis were assessed as percentage of MIB-1 positive or apoptotic cells (MIB-1 labelling index (MI) and apoptotic index (AI)), respectively. RESULTS: Entire specimens showed abnormal expression of one or more upstream regulators of pRb/E2F-1. Although E2F-1 positivity was not associated with the expression of upstream regulators, it showed a linear and positive correlation with MI but not AI. Patients with high MI, low AI, or high E2F-1 positivity had significantly shorter recurrence free survival. By multivariate analysis, high MI and low AI were independently associated with recurrence free survival, but E2F-1 was not. CONCLUSIONS: Increased cell proliferation and decreased apoptosis are associated with adverse prognosis in patients with OSCC. Although E2F-1 remains a controversial prognostic factor, its expression was closely associated with tumour cell proliferation and might influence clinical outcome, mainly via cell cycle progression.

Adult↗

Intravenous immunoglobulin does not increase FcgammaRIIB expression on monocytes/macrophages during acute Kawasaki disease.

OBJECTIVES: Intravenous immunoglobulin (IVIG) therapy has been reported to be effective for reducing the incidence of coronary artery lesions in Kawasaki disease (KD), an acute febrile vasculitis of unknown aetiology. Regarding the mechanism of IVIG in immune thrombocytopenic purpura (ITP), it has been reported that IVIG increases the expression of the inhibitory Fc receptor, FcgammaRIIB (CD32B), on splenic macrophages in a murine ITP model. Regarding the mechanism of IVIG during acute KD, we investigated whether or not IVIG increases the expression of FcgammaRIIB in peripheral blood CD14+ monocytes/macrophages. METHODS: The expression of FcgammaRIIB in peripheral blood CD14+ monocytes/macrophages was determined before and after IVIG therapy in 13 patients with acute KD by flow cytometry. RESULTS: The percentage of CD14+ CD32B+ monocytes/macrophages among peripheral blood mononuclear cells, the absolute number of CD14+ CD32B+ monocytes/macrophages and the percentage of CD14+ CD32B+ monocytes/macrophages among CD14+ monocytes/macrophages in patients with acute KD before IVIG therapy were significantly increased compared with those after IVIG therapy and in controls. CD14+ CD32B+ monocytes/macrophages decreased to within the normal range soon after IVIG therapy. CONCLUSIONS: IVIG therapy in patients with KD did not increase the expression of FcgammaRIIB in peripheral blood CD14+ monocytes/macrophages during the acute stage.

Acute Disease↗

Evidence for an oscillatory signature in atmospheric neutrino oscillations.

Muon neutrino disappearance probability as a function of neutrino flight length L over neutrino energy E was studied. A dip in the L/E distribution was observed in the data, as predicted from the sinusoidal flavor transition probability of neutrino oscillation. The observed L/E distribution constrained nu(micro)<-->nu(tau) neutrino oscillation parameters; 1.9x10(-3) 0.90 at 90% confidence level.

Journal Article↗

Search for electron neutrino appearance in a 250 km long-baseline experiment.

We present a search for electron neutrino appearance from accelerator-produced muon neutrinos in the K2K long-baseline neutrino experiment. One candidate event is found in the data corresponding to an exposure of 4.8 x 10(19) protons on target. The expected background in the absence of neutrino oscillations is estimated to be 2.4+/-0.6 events and is dominated by misidentification of events from neutral current pi(0) production. We exclude the nu(micro) to nu(e) oscillations at 90% C.L. for the effective mixing angle in the 2-flavor approximation of sin((2)2theta(microe)( approximately 1/2sin((2)2theta(13))>0.15 at Deltam(2)(microe)=2.8 x 10(-3) eV(2), the best-fit value of the nu(micro) disappearance analysis in K2K. The most stringent limit of sin((2)2theta(microe)<0.09 is obtained at Deltam(2)(microe)=6 x 10(-3) eV(2).

Journal Article↗

Serum soluble CTLA-4 levels are increased in diffuse cutaneous systemic sclerosis.

OBJECTIVE: To determine serum levels of soluble cytotoxic T-lymphocyte associated molecule-4 (sCTLA-4) and clinical association in patients with systemic sclerosis (SSc). METHODS: Serum sCTLA-4 levels from 32 patients with diffuse cutaneous SSc (dSSc) and 27 patients with limited cutaneous SSc (lSSc) were examined by enzyme-linked immunosorbent assay (ELISA). For a longitudinal study, 211 sera from 30 SSc patients were analysed (follow-up 2.1-7.0 yr). RESULTS: Serum sCTLA-4 levels were elevated in dSSc patients compared with normal controls (n = 41), lSSc patients and patients with active systemic lupus erythematosus (SLE) (n = 23). By contrast, sCTLA-4 levels in patients with lSSc or SLE were normal. SSc patients with elevated sCTLA-4 levels had a shorter disease duration and more frequent presence of digital pitting scars, contracture of phalanges, diffuse pigmentation, pulmonary fibrosis and decreased percentage vital capacity (%VC) than those with normal sCTLA-4 levels. sCTLA-4 levels correlated positively with the extent of skin fibrosis, serum IgG levels and anti-topoisomerase I antibody levels. In a longitudinal study, sCTLA-4 levels decreased on a parallel with improvement of skin sclerosis in five dSSc patients. Skin sclerosis did not improve in two of six dSSc patients with high sCTLA-4 levels throughout the follow-up, while the remaining four patients showed improvement of skin sclerosis. CONCLUSION: These results suggest that sCTLA-4 correlates with disease severity and activity of SSc and that sCTLA-4 plays a role in immunological abnormalities of SSc, since sCTLA-4 may augment humoral immune responses as well as T-cell responses by interfering with B7-CTLA-4 interactions that induce negative signals in T and B cells.

Antigens, CD↗

Limits on the neutrino magnetic moment using 1496 days of Super-Kamiokande-I solar neutrino data.

A search for a nonzero neutrino magnetic moment has been conducted using 1496 live days of solar neutrino data from Super-Kamiokande-I. Specifically, we searched for distortions to the energy spectrum of recoil electrons arising from magnetic scattering due to a nonzero neutrino magnetic moment. In the absence of a clear signal, we found micro(nu)</=(3.6x10(-10))micro(B) at 90% C.L. by fitting to the Super-Kamiokande day-night spectra. The fitting took into account the effect of neutrino oscillation on the shapes of energy spectra. With additional information from other solar neutrino and KamLAND experiments constraining the oscillation region, a limit of micro(nu)</=(1.1x10(-10))micro(B) at 90% C.L. was obtained.

Journal Article↗

Frontotemporal dementia with Pick-type histology associated with Q336R mutation in the tau gene.

In this report, we describe the clinical and neuropathological features of a case of familial frontotemporal dementia (FTD), with onset at 58 years of age and disease duration of 10 years, associated with a novel mutation, Q336R, in the tau gene (tau). In vitro studies concerning the properties of tau proteins bearing this mutation, with respect to microtubule assembly and tau filament aggregation, are reported. Clinically, the patient showed alterations in memory, language and executive functions and marked behavioural change consistent with FTD, although the extent of memory impairment was more than is characteristic of FTD. At autopsy, there was degeneration of the frontal and temporal lobes associated with the presence of hyperphosphorylated tau proteins in swollen (Pick) cells and intraneuronal inclusions (Pick bodies). By immunohistochemistry, the Pick bodies contained both 3-repeat and 4-repeat tau proteins although, because no fresh tissues were available for analysis, the exact isoform composition of the aggregated tau proteins could not be determined. Neurons within frontal cortex contained neurofibrillary tangle-like structures, comprising both straight and twisted tubules, or Pick bodies in which the filaments were short and randomly orientated. In vitro, and in common with other tau missense mutations, Q336R caused an increase in tau fibrillogenesis. However, in contrast to most other tau missense mutations, Q336R increased, not decreased, the ability of mutant tau to promote microtubule assembly. Nonetheless, this latter functional change may likewise be detrimental to neuronal function by inducing a compensatory phosphorylation that may yield increased intracellular hyperphosphorylated tau species that are also liable to fibrillize. We believe the mutation is indeed pathogenic and disease causing and not simply a coincidental rare and benign polymorphism. Since this mutation is segregating with the FTD clinical and neuropathological phenotype, it has not been found in unaffected individuals and it has novel functional properties in vitro which are likely to be detrimental to neuronal function in vivo.

Aged↗

Expression of human coagulation factor VIII in adipocytes transduced with the simian immunodeficiency virus agmTYO1-based vector for hemophilia A gene therapy.

We demonstrate that transduction of adipocytes with a simian immunodeficiency virus agm TYO1 (SIVagm)-based lentiviral vector carrying the human coagulation factor VIII gene (SIVhFVIII) resulted in expression of the human FVIII transgene in vitro and in db/db mice in vivo. Cultured human adipocytes were transduced with the SIVagm vector carrying the GFP gene in a dose-dependent manner and transduction of adipocytes with SIVhFVIII resulted in efficient expression of human coagulation factor VIII (hFVIII; 320 +/- 39.8 ng/10(6) adipocytes/24 h) in vitro. Based upon successful transduction of adipocytes by SIV vectors carrying the lacZ gene in vivo in mice, the adipose tissue of db/db mice was transduced with SIVhFVIII. There was a transient appearance of human FVIII in mouse plasma (maximum 1.8 ng/ml) on day 11 after the injection. Transcripts of human FVIII transgene and human FVIII antigen also were detected in the adipose tissue by RT-PCR and immunofluorescence, respectively, on day 14. Emergence of anti-human FVIII antibodies 14 days after the injection of SIVhFVIII may explain the disappearance of human FVIII from the circulation. These results suggest that transduction of the adipocytes with vectors carrying the human FVIII gene may be potentially applicable for gene therapy of hemophilia A.

Adipocytes↗

Postischemic administration of Sendai virus vector carrying neurotrophic factor genes prevents delayed neuronal death in gerbils.

Sendai virus (SeV) vector-mediated gene delivery of glial cell line-derived neurotrophic factor (GDNF) and nerve growth factor (NGF) prevented the delayed neuronal death induced by transient global ischemia in gerbils, even when the vector was administered several hours after ischemia. Intraventricular administration of SeV vector directed high-level expression of the vector-encoded neurotrophic factor genes, which are potent candidates for the treatment of neurodegenerative diseases. After occlusion of the bilateral carotid arteries of gerbils, SeV vector carrying GDNF (SeV/GDNF), NGF (SeV/NGF), brain-derived neurotrophic factor (SeV/BDNF), insulin-like growth factor-1 (SeV/IGF-1) or vascular endothelial growth factor (SeV/VEGF) was injected into the lateral ventricle. Administration of SeV/GDNF, SeV/NGF or SeV/BDNF 30 min after the ischemic insult effectively prevented the delayed neuronal death of the hippocampal CA1 pyramidal neurons. Furthermore, the administration of SeV/GDNF or SeV/NGF as late as 4 or 6 h after the ischemic insult also prevented the death of these neurons. These results indicate that SeV vector-mediated gene transfer of neurotrophic factors has high therapeutic potency for preventing the delayed neuronal death induced by transient global ischemia, and provides an approach for gene therapy of stroke.

Animals↗

Generation of a recombinant Sendai virus that is selectively activated and lyses human tumor cells expressing matrix metalloproteinases.

Malignant tumor cells often express matrix metalloproteinases (MMPs) at a high level to enable their dissemination and metastasis. Sendai virus (SeV), a nonsegmented negative strand RNA virus, spreads in the target tissues in vivo via cleavage activation of the viral fusion glycoprotein by a tissue-specific, trypsin-like enzyme. By deleting the viral matrix protein, we previously generated a recombinant SeV that does not bud to mature virions, but is highly fusogenic and spreads extensively from cell to cell in a trypsin-dependent manner. Here, we changed the tryptic cleavage site of the fusion glycoprotein of this virus to a site susceptible to MMPs. The resulting recombinant virus was no longer activated by trypsin but spread efficiently in cultured cells supplemented with MMP2 or MMP9 and in human tumor cell lines expressing these MMPs. Furthermore, the virus spread extensively in tumor cells xenotrasplanted to nude mice without disseminating to the surrounding normal cells, leading to the inhibition of the tumor growth in the mice. These results demonstrate the selective targeting and killing of human tumor cells by recombinant SeV technology and greatly advance the reemerging concept of oncolytic virotherapy, which currently appears to rely largely upon a natural preference of certain viruses for cancer cells.

Animals↗

Expansion of genetically corrected neutrophils in chronic granulomatous disease mice by cotransferring a therapeutic gene and a selective amplifier gene.

Hematopoietic stem cell gene therapy has not provided clinical success in disorders such as chronic granulomatous disease (CGD), where genetically corrected cells do not show a selective advantage in vivo. To facilitate selective expansion of transduced cells, we have developed a fusion receptor system that confers drug-induced proliferation. Here, a 'selective amplifier gene (SAG)' encodes a chimeric receptor (GcRER) that generates a mitotic signal in response to estrogen. We evaluated the in vivo efficacy of SAG-mediated cell expansion in a mouse disease model of X-linked CGD (X-CGD) that is deficient in the NADPH oxidase gp91phox subunit. Bone marrow cells from X-CGD mice were transduced with a bicistronic retrovirus encoding GcRER and gp91phox, and transplanted to lethally irradiated X-CGD recipients. Estrogen was administered to a cohort of the transplants, and neutrophil superoxide production was monitored. A significant increase in oxidase-positive cells was observed in the estrogen-treated mice, and repeated estrogen administration maintained the elevation of transduced cells for 20 weeks. In addition, oxidase-positive neutrophils were increased in the X-CGD transplants given the first estrogen even at 9 months post-transplantation. These results showed that the SAG system would enhance the therapeutic effects by boosting genetically modified, functionally corrected cells in vivo.

Animals↗

A defective nontransmissible recombinant Sendai virus mediates efficient gene transfer to airway epithelium in vivo.

Recombinant Sendai virus (SeV)-mediated gene transfer to differentiated airway epithelial cells has shown to be very efficient, because of its ability to overcome the intra- and extracellular barriers known to limit gene delivery. However, this virus is transmission competent and therefore unlikely to be suitable for use in clinical trials. A nontransmissible, replication-competent recombinant SeV has recently been developed by deleting the envelope Fusion (F) protein gene (SeV/DeltaF). Here we show that SeV/DeltaF is able to mediate beta-galactosidase reporter gene transfer to the respiratory tract of mice in vivo, as well as to human nasal epithelial cells in vitro. Further, in an ex vivo model of differentiated airway epithelium, SeV/DeltaF gene transfer was not importantly inhibited by native mucus. When compared to the transmission-competent SeV in vivo, no difference in gene expression was observed at the time of peak expression. The development of an F-defective nontransmissible SeV, which can still efficiently mediate gene transfer to the airway epithelium, represents the first important step towards the use of a cytoplasmic RNA viral vector in clinical trials of gene therapy.

Adult↗