Biomedical subjects

Chihiro Nakahara

Publications and source records attributed to Chihiro Nakahara.

5 recordsLinked to original sources

PubMed2005

Exosomes secreted from monocyte-derived dendritic cells support in vitro naive CD4+ T cell survival through NF-(kappa)B activation.

We investigated the effect of exosomes secreted from human monocyte-derived dendritic cells (Mo-DCs), which are generated from PBMCs in response to treatment with GM-CSF and IL-4, on naive CD4+ T cell survival in vitro. Exosomes isolated from culture supernatants of Mo-DCs (>90% purity) were purified with anti-HLA-DP, -DQ, -DR-coated paramagnetic beads. Purified exosomes prolonged the survival of naive CD4+ T cells (>98% purity) in vitro. Treatment with neutralizing mAb against HLA-DR significantly decreased the supportive effect of purified exosomes on CD4+ T cell survival. Exosomes increased nuclear translocation of NF-(kappa)B in naive CD4+ T cells, and NF-(kappa)B activation was significantly suppressed by anti-HLA-DR mAb or NF-(kappa)B inhibitor pyrrolidine dithiocarbamate (PDTC). In addition, PDTC inhibited the effect of exosomes on naive CD4+ T cell survival. Thus, exosomes secreted by Mo-DCs appear to support naive CD4+ T cell survival via NF-(kappa)B activation induced by interaction of HLA-DR and TCRs.

Antigens, CD↗

PubMed2004

Interleukin 1beta enhances invasive ability of gastric carcinoma through nuclear factor-kappaB activation.

PURPOSE: We examined the role of interleukin (IL)-1beta in activation of nuclear factor kappaB (NF-kappaB) and the biological function of activated NF-kappaB in gastric carcinoma cells. EXPERIMENTAL DESIGN: Human gastric carcinoma cell line GCTM-1 was used to examine NF-kappaB activation by immunostaining and electrophoretic mobility shift assay. Matrix metalloproteinase (MMP)-9 expression, which plays an important role in tumor invasion, was assessed by semiquantitative reverse transcription-PCR, Western blotting, and immunostaining. The invasive ability of GCTM-1 cells was measured by Matrigel invasion assay. In vivo expression of IL-1beta and MMP-9 and activation of NF-kappaB in 10 surgically resected gastric carcinoma specimens were examined immunohistochemically. RESULTS: IL-1beta enhanced NF-kappaB activation, MMP-9 expression, and the invasive ability of GCTM-1. A NF-kappaB inhibitor, pyrrolidine dithiocarbamate, suppressed both MMP-9 expression and invasiveness of IL-1beta-treated GCTM-1 cells. IL-1beta did not increase the invasive ability of GCTM-1 cells transfected with MMP-9 antisense oligonucleotide. Concomitant expression of IL-1beta and nuclear NF-kappaB was observed in 3 of 10 gastric carcinoma specimens. Cells producing IL-1beta were tumor-infiltrating macrophages in two specimens and gastric carcinoma cells in one specimen. CONCLUSIONS: One of the molecules that may play a role in NF-kappaB activation in some gastric carcinomas is IL-1beta. The present results suggest that IL-1beta increases the invasive ability of carcinoma cells through activation of NF-kappaB and the resulting MMP-9 expression.

Base Sequence↗

PubMed2004

Three-dimensional two-layer collagen matrix gel culture model for evaluating complex biological functions of monocyte-derived dendritic cells.

Dendritic cell-like cells (Mo-DCs) generated from peripheral blood monocytes with interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) have been used as tools to treat cancer patients (DC-vaccines). Because Mo-DCs have multiple antigen presentation-related functions, including phagocytosis, migration, cytokine production, and T cell stimulation, establishment of a method for simultaneously evaluating the various functions of Mo-DCs is important. We developed a new in vitro three-dimensional two-layer collagen matrix culture model that consists of a collagen gel containing Mo-DCs as the lower layer and a collagen gel containing necrotic GCTM-1 tumor cells and/or T cells as the upper layer. We used this system to observe simultaneously multiple functions of Mo-DCs by phase-contrast or fluorescence microscopy and to assess IL-12 secretion during more than 2 weeks of culture. We also observed interactions between Mo-DCs and necrotic GCTM-1 or T cells on an individual cell basis by time-lapse videomicroscopy. In addition, we collected Mo-DCs from the collagen gels by collagenase treatment and analyzed the expression of antigen presentation-related molecules such as HLA-DR, CD80, CD83, and CD86 on Mo-DCs. This model may be a useful tool for evaluation of the various functions of Mo-DCs used as DC vaccines and for studies of the complex behaviors of Mo-DCs in vivo.

Antigen Presentation↗

PubMed2003

Cyclosporin-A enhances docetaxel-induced apoptosis through inhibition of nuclear factor-kappaB activation in human gastric carcinoma cells.

PURPOSE: We sought to determine whether cyclosporin-A (CsA) enhances docetaxel [Taxotere (TXT)]- induced apoptosis in human gastric carcinoma cells, and, if so, to determine the relation between this apoptosis and nuclear factor-kappaB (NF-kappaB) activation. EXPERIMENTAL DESIGN: Two human gastric carcinoma cell lines (GCTM-1 and MK-1), a human embryonic pulmonary fibroblast cell line, and human umbilical vein endothelial cells were used as drug targets. Apoptotic cell death was verified morphologically by nuclear fragmentation assay with Hoechst staining. Electrophoretic mobility shift assays were performed to check for nuclear translocation of NF-kappaB. The therapeutic effects of a combination of TXT and CsA were assessed in a mouse peritoneal dissemination model. RESULTS: A combination of CsA (5 micro M) and TXT (10 nM) significantly enhanced apoptotic cell death in both carcinoma cell lines but not in nonmalignant cell lines in comparison with the single-agent treatment alone. This effect was not related to drug uptake, efflux, or MDR1 expression. These effects were also observed in freshly obtained TXT-resistant gastric carcinoma cells isolated from a patient with malignant ascites. TXT alone induced NF-kappaB activation in both carcinoma cell types, and this activation was suppressed by CsA. A combination of TXT and NF-kappaB decoy, a well-known NF-kappaB inhibitor, also enhanced apoptotic cell death in the carcinoma cells. A combination of CsA and TXT significantly suppressed peritoneal dissemination in vivo relative to the single-agent effect. CONCLUSIONS: Treatment with CsA and TXT in combination may be an effective therapeutic strategy for patients with gastric carcinoma.

Adenocarcinoma↗

PubMed2003

PSK-mediated NF-kappaB inhibition augments docetaxel-induced apoptosis in human pancreatic cancer cells NOR-P1.

Docetaxel, a member of the taxane family, has been shown to induce apoptosis in a variety of cancer cells. However, toxicity at therapeutic doses has precluded the use of docetaxel alone for the management of cancer patients. PSK, a protein-bound polysaccharide, is widely used in Japan as an immunopotentiating biological response modifier for cancer patients. Our previous study showed that PSK induced downregulation of several invasion-related factors, suggesting an interaction of PSK with transcriptional factors. In this study, we showed that PSK dose dependently enhanced apoptosis induced by 1 nM of docetaxel in a human pancreatic cancer cell line NOR-P1. Furthermore, PSK inhibited docetaxel-induced nuclear factor kappa B (NF-kappaB) activation. Moreover, the expression of cellular inhibitor of apoptosis protein (cIAP-1), which is transcriptionally regulated by NF-kappaB and functions as an antiapoptotic molecule through interrupting the caspase pathway, was also inhibited by treatment with PSK plus docetaxel. As a result, PSK enhanced the docetaxel-induced caspase-3 activation. In addition, treatment by transfection of NF-kappaB decoy oligodeoxynucleotides (ODNs), but not scramble ones, inhibited the expression of cIAP-1 in NOR-P1 cells and induced a significant increase in docetaxel-induced apoptosis. Our data indicate that PSK suppresses the docetaxel-induced NF-kappaB activation pathway. Combination of PSK with a low dose of docetaxel may be a new therapeutic strategy to treat patients with pancreatic cancer.

Apoptosis↗