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Biomedical subjects

Naoki Yamamoto

Publications and source records attributed to Naoki Yamamoto.

At least 163 records · Page 9Linked to original sources
PubMed2003

An in vivo model for monitoring trans-differentiation of bone marrow cells into functional hepatocytes.

The plasticity of bone marrow cells (BMCs) remains controversial. The present study found that persistent injury induces efficient trans-differentiation of BMCs into functional hepatocytes. Mice with liver cirrhosis induced by carbon tetrachloride were injected with 1 x 10(5) non-treated green fluorescent protein (GFP)-positive BMCs via the tail vein. In these mice, transplanted GFP-positive BMCs efficiently migrated into the peri-portal area of liver lobules after one day, repopulating 25% of the recipient liver by 4 weeks. In contrast, no GFP-positive BMCs were detected following transplantation into control mice with undamaged livers. BMCs trans-differentiated into functional mature hepatocytes via immature hepatoblasts. Serum albumin levels were significantly elevated to compensate for chronic liver failure in BMC transplantation. These results reveal that recipient conditions and microenvironments represent key factors for successful cell therapy using BMCs.

Albumins↗
PubMed2003

Role of the zinc fingers of HIV-1 nucleocapsid protein in maturation of genomic RNA.

The nucleocapsid protein of HIV-1 consists of two basic amino acid regions and two zinc fingers. We investigated the requirement of these domains for the structural conversion of a 39mer RNA covering the dimerization initiation site by using three peptides; wild-type NCp7, a mutant in which the two zinc fingers are mutated, and another mutant in which the two zinc fingers are deleted. The two mutants exhibited similar conversion activities to each other, which were lower than that of the wild-type, indicating that the two basic regions exhibit some activity for RNA chaperone, as we suggested before, and the zinc fingers enhance the efficiency of this activity.

Amino Acid Sequence↗
PubMed2003

Helicobacter pylori induces RANTES through activation of NF-kappa B.

Helicobacter pylori-infected gastric mucosa displays a conspicuous infiltration of mononuclear cells and neutrophils. RANTES (short for "regulated upon activation, normal T cell expressed and secreted") is a chemoattractant cytokine (chemokine) important in the infiltration of T lymphocytes and monocytes. RANTES may therefore contribute to the cellular infiltrate in the H. pylori-infected gastric mucosa. The aim of this study was to analyze the molecular mechanism responsible for H. pylori-mediated RANTES expression. We observed that gastric epithelial cells produced RANTES upon coculture with H. pylori. In addition, H. pylori induced RANTES mRNA expression and an increase in luciferase activity in cells which were transfected with a luciferase reporter construct derived from the RANTES promoter, in gastric epithelial cells, indicating that the induction of RANTES production occurred at the transcriptional level. Induction of RANTES was dependent on an intact cag pathogenicity island. Activation of the RANTES promoter by H. pylori occurred through the action of NF-kappa B. Transfection of kinase-deficient mutants of I kappa B kinase (IKK) and NF-kappa B-inducing kinase (NIK) inhibited H. pylori-mediated RANTES activation. In contrast, tumor necrosis factor alpha- or interleukin-1/Toll-like receptor signaling molecules-such as mitogen-activated protein kinase/extracellular signal-regulated kinase kinase 1, MyD88, and interleukin-1 receptor-associated kinase-did not play a role in RANTES activation by H. pylori. Collectively, H. pylori induced NF-kappa B activation through an intracellular signaling pathway that involved IKK and NIK, leading to RANTES gene transcription. RANTES induction by H. pylori may play an important role in gastric inflammation.

Antineoplastic Agents↗
PubMed2003

Induction of protective immune responses against R5 human immunodeficiency virus type 1 (HIV-1) infection in hu-PBL-SCID mice by intrasplenic immunization with HIV-1-pulsed dendritic cells: possible involvement of a novel factor of human CD4(+) T-cell origin.

The potential of a dendritic cell (DC)-based vaccine against human immunodeficiency virus type 1 (HIV-1) infection in humans was explored with SCID mice reconstituted with human peripheral blood mononuclear cells (PBMC). HIV-1-negative normal human PBMC were transplanted directly into the spleens of SCID mice (hu-PBL-SCID-spl mice) together with autologous mature DCs pulsed with either inactivated HIV-1 (strain R5 or X4) or ovalbumin (OVA), followed by a booster injection 5 days later with autologous DCs pulsed with the same respective antigens. Five days later, these mice were challenged intraperitoneally with R5 HIV-1(JR-CSF). Analysis of infection at 7 days postinfection showed that the DC-HIV-1-immunized hu-PBL-SCID-spl mice, irrespective of the HIV-1 isolate used for immunization, were protected against HIV-1 infection. In contrast, none of the DC-OVA-immunized mice were protected. Sera from the DC-HIV-1- but not the DC-OVA-immunized mice inhibited the in vitro infection of activated PBMC and macrophages with R5, but not X4, HIV-1. Upon restimulation with HIV-1 in vitro, the human CD4(+) T cells derived from the DC-HIV-1-immunized mice produced a similar R5 HIV-1 suppressor factor. Neutralizing antibodies against human RANTES, MIP-1alpha, MIP-1beta, alpha interferon (IFN-alpha), IFN-beta, IFN-gamma, interleukin-4 (IL-4), IL-10, IL-13, IL-16, MCP-1, MCP-3, tumor necrosis factor alpha (TNF-alpha), or TNF-beta failed to reverse the HIV-1-suppressive activity. These results show that inactivated HIV-1-pulsed autologous DCs can stimulate splenic resident human CD4(+) T cells in hu-PBL-SCID-spl mice to produce a yet-to-be-defined, novel soluble factor(s) with protective properties against R5 HIV-1 infection.

AIDS Vaccines↗
PubMed2003

Intravenous inoculation of replication-deficient recombinant vaccinia virus DIs expressing simian immunodeficiency virus gag controls highly pathogenic simian-human immunodeficiency virus in monkeys.

To be effective, a vaccine against human immunodeficiency virus type 1 (HIV-1) must induce virus-specific T-cell responses and it must be safe for use in humans. To address these issues, we developed a recombinant vaccinia virus DIs vaccine (rDIsSIVGag), which is nonreplicative in mammalian cells and expresses the full-length gag gene of simian immunodeficiency virus (SIV). Intravenous inoculation of 10(6) PFU of rDIsSIVGag in cynomologus macaques induced significant levels of gamma interferon (IFN-gamma) spot-forming cells (SFC) specific for SIV Gag. Antigen-specific lymphocyte proliferative responses were also induced and were temporally associated with the peak of IFN-gamma SFC activity in each macaque. In contrast, macaques immunized with a vector control (rDIsLacZ) showed no significant induction of antigen-specific immune responses. After challenge with a highly pathogenic simian-human immunodeficiency virus (SHIV), CD4(+) T lymphocytes were maintained in the peripheral blood and lymphoid tissues of the immunized macaques. The viral set point in plasma was also reduced in these animals, which may be related to the enhancement of virus-specific intracellular IFN-gamma(+) CD8(+) cell numbers and increased antibody titers after SHIV challenge. These results demonstrate that recombinant DIs has potential for use as an HIV/AIDS vaccine.

AIDS Vaccines↗
PubMed2003

Rapid tumor formation of human T-cell leukemia virus type 1-infected cell lines in novel NOD-SCID/gammac(null) mice: suppression by an inhibitor against NF-kappaB.

We established a novel experimental model for human T-cell leukemia virus type 1 (HTLV-1)-induced tumor using NOD-SCID/gammac(null) (NOG) mice. This model is very useful for investigating the mechanism of tumorigenesis and malignant cell growth of adult T-cell leukemia (ATL)/lymphoma, which still remains unclear. Nine HTLV-1-infected cell lines were inoculated subcutaneously in the postauricular region of NOG mice. As early as 2 to 3 weeks after inoculation, seven cell lines produced a visible tumor while two transformed cell lines failed to do so. Five of seven lines produced a progressively growing large tumor with leukemic infiltration of the cells in various organs that eventually killed the animals. Leukemic cell lines formed soft tumors, whereas some transformed cell lines developed into hemorrhagic hard tumors in NOG mice. One of the leukemic cell lines, ED-40515(-), was unable to produce visible tumors in NOD-SCID mice with a common gamma-chain after 2 weeks. In vivo NF-kappaB DNA binding activity of the ED-40515(-) cell line was higher and the NF-kappaB components were changed compared to cells in vitro. Bay 11-7082, a specific and effective NF-kappaB inhibitor, prevented tumor growth at the sites of the primary region and leukemic infiltration in various organs of NOG mice. This in vivo model of ATL could provide a novel system for use in clarifying the mechanism of growth of HTLV-1-infected cells as well as for the development of new drugs against ATL.

Animals↗
PubMed2002

Combined intrarectal/intradermal inoculation of recombinant Mycobacterium bovis bacillus Calmette-Guérin (BCG) induces enhanced immune responses against the inserted HIV-1 V3 antigen.

The development of a successful recombinant Mycobacterium bovis bacillus Calmette-Guérin (rBCG) vector-based vaccine for human immunodeficiency virus type 1 (HIV-1) requires the induction of high levels of HIV-1-specific immunity while at the same time maintaining immunity to tuberculosis. To examine a combined vaccination strategy for enhancement of immune responses specific for HIV-1, guinea pigs were inoculated with either a single or combination intradermal (i.d.), intrarectal (i.r.) and intranasal (i.n.) administration of rBCG-pSOV3J1 which secretes a chimeric protein of HIV-1 V3J1 peptide and alpha-antigen. Significant level of delayed-type hypersensitivity to both V3J1 peptide and tuberculin was induced in guinea pigs inoculated with human doses of rBCG-pSOV3J1 by a combination of intrarectal and intradermal routes. Guinea pigs inoculated by combined routes also had significantly higher titers of HIV-1-specific serum IgG and IgA compared with those animals immunized only intrarectally, which led to the enhanced neutralization activity against HIV-1(MN). In addition, the induction of high levels of IFNgamma and interleukin-2 (IL-2) mRNA in PBMC, splenocytes, and intraepithelial lymphocytes from the immunized animals was detected until at least 110 weeks post-inoculation. These results suggest that enhanced immune responses specific for HIV-1 are efficiently induced by combined intrarectal and intradermal immunization with rBCG-HIV, and antigen-specific Th1-type memory cells are maintained for more than 2 years in the immunized animals. Thus, inoculation with rBCG-HIV by combined routes represents an effective vaccination strategy to elicit high levels of HIV-1-specific immune responses.

AIDS Vaccines↗
PubMed2002

Lymphotoxin-beta receptor mediates NEMO-independent NF-kappaB activation.

Lymphotoxin-beta receptor (LTbetaR) is a member of the tumor necrosis factor receptor (TNFR) superfamily that activates nuclear factor-kappaB (NF-kappaB) through the IkappaB kinase (IKK) complex, the core of which is comprised of IKK1, IKK2 and NF-kappaB essential modulator (NEMO). We demonstrate here that the LTbetaR signaling to NF-kappaB activation does not necessarily require NEMO, which is essential for TNFR signaling. In the absence of NEMO, the p50 and RelB, but not RelA subunits of NF-kappaB are found in the nuclear DNA binding complexes induced by the LTbetaR signaling. Our results thus disclose NEMO-independent NF-kappaB activation by LTbetaR.

Active Transport, Cell Nucleus↗
PubMed2002

A recessive mutant cell line with a constitutive IkappaB kinase activity.

To search for negative regulatory components of the NF-kappaB activation pathways, we mutagenized Rat-1 fibroblasts and established a stable mutant cell line with a constitutive NF-kappaB activity. This mutant cell line, designated as TK26, showed permanently elevated I kappa B kinase (IKK) activity and a genetically recessive phenotype revealed by somatic cell hybridization between TK26 and Rat-1. Our results suggested that lack of a negative regulation of IKK could lead to permanent NF-kappaB activation. The TK26 cell line will be useful to genetically identify a component necessary for keeping the IKK complex under an inactive form in resting cells.

Animals↗
PubMed2002

A new ERK2 binding protein, Naf1, attenuates the EGF/ERK2 nuclear signaling.

Extracellular signal regulated kinase1/2 (ERK1/2), an important factor in signal transduction, controls cell growth, differentiation, and death. To elucidate the details of the mechanism of ERK1/2 signaling in human cells, we isolated Nef-associated factor 1 alpha (Naf1 alpha) by a yeast two-hybrid system, which bound to human ERK2. The binding was confirmed by a pull-down assay in vitro and immunoprecipitation in vivo. Upon EGF treatment, Naf1 alpha was phosphorylated by the EGF/MEK/ERK2 signal transduction pathway. To identify the role of Naf1 alpha in the ERK2 signaling, Naf1 alpha-expressing Saos-2 cells were analyzed for ERK2 nuclear translocation and activation of its downstream target. Overexpression of Naf1 alpha suppressed ERK2 entering into the nucleus and inhibited the ERK2-dependent Elk1-driven luciferase transcription, suggesting Naf1 alpha to be an attenuator of activated ERK2 signaling.

Cell Nucleus↗
PubMed2002

Bay 11-7082 inhibits transcription factor NF-kappaB and induces apoptosis of HTLV-I-infected T-cell lines and primary adult T-cell leukemia cells.

Human T-cell leukemia virus type I (HTLV-I) is the causative agent of an aggressive form of leukemia designated adult T-cell leukemia (ATL). We have previously demonstrated that all T-cell lines infected with HTLV-I and primary leukemic cells from ATL patients display constitutively high activity of transcription factor NF-kappaB. In this study we showed that Bay 11-7082, an inhibitor of NF-kappaB, induced apoptosis of HTLV-I-infected T-cell lines but only negligible apoptosis of HTLV-I-negative T cells. Bay 11-7082 rapidly and efficiently reduced the DNA binding of NF-kappaB in HTLV-I-infected T-cell lines and down-regulated the expression of the antiapoptotic gene, Bcl-x(L), regulated by NF-kappaB, whereas it had little effect on the DNA binding of another transcription factor, AP-1. Although the viral protein Tax is an activator of NF-kappaB, Bay 11-7082-induced apoptosis of HTLV-I-infected cells was not associated with reduced expression of Tax. Furthermore, Bay 11-7082- induced apoptosis of primary ATL cells was more prominent than that of normal peripheral blood mononuclear cells, and apoptosis of these cells was also associated with down-regulation of NF-kappaB activity. Our results indicate that NF-kappaB plays a crucial role in the pathogenesis and survival of HTLV-I-infected leukemic cells and that it is a suitable target for the prevention and treatment of ATL.

Adult↗
PubMed2002

Differential regulation of aquaporin-5 and -9 expression in astrocytes by protein kinase A.

Aquaporins (AQPs) transport water through the membranes of numerous tissues, but the molecular mechanisms for regulating water balance in brain are unknown. In this study, we investigated the effects of a protein kinase A (PKA) activator on the expression of AQP4, 5 and 9 in cultured rat astrocytes. Treatment of the cells with dbcAMP caused decreases in AQP5 mRNA and protein and increases in AQP9 mRNA and protein in time- and concentration-dependent manners. However, AQP4 mRNA and protein were not changed by treatment with dbcAMP. The dbcAMP-induced effects on AQP5 and AQP9 mRNAs were inhibited by PKA inhibitors. In addition, pretreating the cells with an inhibitor of protein synthesis, cycloheximide, inhibited the increase in AQP9 mRNA induced by dbcAMP, but not the decrease in AQP5 mRNA. These results suggest that signal transduction via PKA may play important roles in regulating the expression of AQP5 and AQP9, and the effect on AQP9 may be mediated by some factors induced by dbcAMP.

Animals↗
PubMed2002

Activation of cyclin D1 and D2 promoters by human T-cell leukemia virus type I tax protein is associated with IL-2-independent growth of T cells.

Our aim was to examine the involvement of G(1) cell-cycle regulators in cell growth dysregulation induced by HTLV-I. Compared to uninfected cells, higher expression levels of cyclin D1 and D2 mRNA were detected in HTLV-I-infected T-cell lines, which were at least in part mediated by the viral transforming protein Tax since Tax activated both cyclin D1 and D2 promoters in the human T-cell line Jurkat. A Tax mutant that did not activate NF-kappaB failed to activate cyclin D1 and D2 promoters. Inhibitors of NF-kappaB (dominant negative IkappaBs mutants) suppressed Tax-dependent activation of cyclin D1 and D2 promoters, indicating that Tax-induced activation was mediated by NF-kappaB. Wild-type and mutant Tax capable of activating NF-kappaB, but not Tax mutant incapable of activating NF-kappaB, converted cell growth of a T-cell line from being IL-2-dependent to being IL-2-independent; and this conversion was associated with IL-2-independent induction of cyclins D1 and D2. Our data suggest that induction of cyclins D1 and D2 by Tax is involved in IL-2-independent cell-cycle progression as well as IL-2-independent transformation of primary human T cells by HTLV-I. High expression levels of cyclin D1 and D2 mRNAs were also detected in some patients with ATL. Our findings link HTLV-I infection to changes in cellular D-type cyclin gene expression, transformation of T cells and subsequent development of T-cell leukemia.

Animals↗
PubMed2002

Polyamine depletion induces apoptosis through mitochondria-mediated pathway.

Polyamines, namely putrescine, spermidine, and spermine, are essential for cell survival and proliferation. A decrease in intracellular polyamine levels is associated with apoptosis. In this study, we used inhibitors of polyamine biosynthesis to examine the effect of polyamine depletion. A combination of inhibitors of ornithine decarboxylase, S-adenosylmethionine decarboxylase, or spermidine synthase decreased intracellular polyamine levels and induced cell death in a WEHI231 murine B cell line. These cells exhibited apoptotic features including chromatin condensation and oligonucleosomal DNA fragmentation. Addition of exogenous polyamines reversed the observed features of apoptotic cell death. Similar effects were also observed in other cell lines: a human B cell line Ramos and a human T cell line Jurkat. Depletion of polyamines induced activation of caspase-3 and disruption of the mitochondrial membrane potential (Delta psi m). Inhibition of caspase activities by an inhibitor prevented the apoptotic nuclear changes but not Delta psi m disruption induced by polyamine depletion. Overexpression of Bcl-xl, an anti-apoptotic Bcl-2 family protein, completely inhibited Delta psi m disruption, caspase activation, and cell death. These results indicate that the depletion of intracellular polyamines triggers the mitochondria-mediated pathway for apoptosis, resulting in caspase activation and apoptotic cell death.

Adenosylmethionine Decarboxylase↗
PubMed2002

Developmental changes and localization of mouse brain serine proteinase mRNA and protein in mouse brain.

Serine proteases are known to be involved in neural development and various functions in the central nervous system. Mouse brain serine proteinase (mBSP) is expressed almost exclusively in the mouse brain and it has been characterized at the molecular and biochemical levels. In this study, we analyzed the developmental changes and localization of mBSP mRNA and protein in the mouse brain, using reverse transcription-polymerase chain reaction, Western blotting, and immunohistochemistry. Expression of mBSP was strong in the white matter and the nerve tracts after postnatal day 30, especially in the cerebellum and the medulla oblongata. These results suggest that mBSP contributes to development and sustaining the functions in the mouse brain.

Animals↗
PubMed2002

Synthesis and evaluation of pseudopeptide analogues of a specific CXCR4 inhibitor, T140: the insertion of an (E)-alkene dipeptide isostere into the betaII'-turn moiety.

A 14-residue peptide, T140, strongly inhibits the T-cell line-tropic HIV-1 (X4-HIV-1) infection, since this peptide functions as a specific antagonist against a chemokine receptor, CXCR4. T140 takes an antiparallel beta-sheet structure with a type II' beta-turn. In the present paper, we have designed and synthesized several T140 analogues, in which an (E)-alkene dipeptide isostere was inserted into the type II' beta-turn moiety, as a bridging study to develop nonpeptidic CXCR4 inhibitors. It has been proven that the turn region of T140 can be replaced by the above surrogate with the maintenance of strong anti-HIV activity.

Anti-HIV Agents↗
PubMed2002

Human T-cell leukemia virus type I Tax transactivates the matrix metalloproteinase-9 gene: potential role in mediating adult T-cell leukemia invasiveness.

Human T-cell leukemia virus type I (HTLV-I) is the etiologic agent of adult T-cell leukemia (ATL) and of tropical spastic paraparesis/HTLV-I-associated myelopathy. Infiltration of various tissues by circulating leukemic cells is a characteristic of ATL. Matrix metalloproteinases (MMPs), which mediate the degradation of the basement membrane and extracellular matrix, play an important role in metastasis and tumor cell dissemination. The aim of this study was to explore whether expression of MMP-2 and MMP-9 was deregulated by HTLV-I infection. The data showed that HTLV-I-infected T-cell lines expressed high levels of MMP-9 compared with uninfected T-cell lines. In contrast, the levels of the related MMP-2 were not significantly altered by HTLV-I infection. In addition, the elevated expression of MMP-9 in HTLV-I-infected cells was attributable to the action of the viral transactivator protein Tax. The results show that Tax can activate the MMP-9 promoter and induce MMP-9 expression in T cells, indicating that the constitutive expression of MMP-9 in virus-infected cell lines is at least in part mediated by Tax. Activation of the MMP-9 promoter by Tax occurs mainly through the action of NF-kappaB and SP-1. The biologic significance of these observations was validated by the following 2 findings: MMP-9 expression was increased in primary ATL cells, and plasma MMP-9 levels were elevated in ATL patients. In addition, plasma levels of MMP-9 correlated with organ involvement in ATL patients. Together these data suggest that overexpression of MMP-9 in HTLV-I- infected cells may be in part responsible for the invasiveness of ATL cells.

Enzyme Induction↗