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Biomedical subjects

H Asada

Publications and source records attributed to H Asada.

At least 55 records · Page 3Linked to original sources
PubMed2000

Impaired responses of peripheral blood mononuclear cells to staphylococcal superantigen in patients with severe atopic dermatitis: a role of T cell apoptosis.

Staphylococcus aureus colonization is an almost universal feature of atopic dermatitis. In order to investigate the role of staphylococcal enterotoxin B in the pathogenesis of atopic dermatitis, we assessed the correlation between clinical disease severity and proliferative response of peripheral blood mononuclear cells to staphylococcal enterotoxin B in patients with atopic dermatitis. Peripheral blood mononuclear cells from patients with mild atopic dermatitis showed significantly increased proliferative responses to staphylococcal enterotoxin B compared to controls. In contrast, peripheral blood mononuclear cells from patients with severe atopic dermatitis showed markedly suppressed proliferative responses. Additionally, longitudinal evaluation of peripheral blood mononuclear cell samples from the same patient demonstrated that proliferative responses were suppressed only at times of severe disease exacerbation. Mixing experiments, using autologous T cells and antigen presenting cells that were isolated at different time points from the same patient, demonstrated that T cells of severe atopic dermatitis patients were dysfunctional, but their antigen presenting cell function remained intact. We found no significant differences of interleukin-2 levels in the culture supernatants between healthy controls and atopic dermatitis groups. Fluorescence-activated cell sorter analysis for APO2.7 antigen, an early apoptosis cell marker, demonstrated that approximately 60% of staphylococcal-enterotoxin-B-stimulated T cells expressed APO2.7 antigen in severe atopic dermatitis cases. By contrast, 5%-20% of T cells expressed APO2.7 after staphylococcal enterotoxin B stimulation in cases of mild atopic dermatitis and in healthy controls. Nuclear staining with Hoechst 33258 also showed approximately 40% apoptotic cells in the CD19-CD16-PBMC of severe atopic dermatitis patients, compared with only 5%-10% in the mild atopic dermatitis group and in healthy controls. Blocking monoclonal antibody to Fas ligand partially prevented the staphylococcal-enterotoxin-B-induced apoptosis detected by APO2.7 expression and Hoechst 33258 staining. Suppressed proliferation of peripheral blood mononuclear cells in severe atopic dermatitis patients may be secondary to T cell death by apoptosis. These results suggest that an infection of S. aureus producing staphylococcal enterotoxin B may play a role in aggravation of atopic dermatitis by inducing apoptosis in T cells.

Adult↗
PubMed2000

Double involvement of extramammary Paget's disease in the genitalia and axilla.

We here reported a case of an 82-year-old man with double involvement of extramammary Paget's disease in the genitalia and axilla. Physical examination revealed erythema and reddish tumors on the pubic area and scrotum and irregular-shaped erythema on the left axilla. The skin biopsy sample from the genital area showed Paget's cells in the epidermis and upper dermis. The specimen from the left axilla showed Paget's cells scattered in the epidermis but not in the dermis.

Aged↗
PubMed2000

Annexin VII as a novel marker for invasive phenotype of malignant melanoma.

Both F10 and BL6 sublines of B16 mouse melanoma cells are metastatic after intravenous injection, but only BL6 cells are metastatic after subcutaneous injection. While examining the genetic difference between the two sublines, we found a marked reduction of annexin VII expression in BL6 cells. In addition, fusion cell clones of both sublines were as poorly metastatic as F10 cells after subcutaneous injection, and contained the annexin VII message as abundantly as F10 cells. Hence, we examined whether the annexin VII expression was correlated with the less malignant phenotype of clinical cases by immunohistochemistry. Immunoreactivities to anti-annexin VII antibody in melanoma cells were evaluated quantitatively by using skin mast cells as an internal positive control. Eighteen patients with malignant melanoma were divided into two groups: lymph node metastasis-negative and positive groups. The ratio of numbers of patients positive versus negative to the antibody was significantly larger in the former than in the latter group. These results not only indicated that annexin VII serves as a marker for less invasive phenotype of malignant melanoma, but also suggested a possible role of annexin VII in tumor suppression.

Adolescent↗
PubMed2000

Accurate multiplex polymerase chain reaction assay for gender determination from a single cell.

For gender determination of preimplantation embryos or circulating fetal cells in maternal blood, we developed a multiplex polymerase chain reaction assay from a single cell. This assay which co-amplifies X (DXZ1)- and Y (DYZ1)-specific repeat sequences, yields a 308-bp band in females and two bands of 154 and 308 bp in males. In a randomized, blinded assay of 100 isolated single amniocytes, 99 (99%) were amplified successfully. All 50 of the XY cells were correctly diagnosed as male (100%), whereas 49 of the 50 XX cells were diagnosed as female (98%). This accurate and efficient assay may be applicable in these clinical settings.

Amniotic Fluid↗
PubMed2000

A role for heterologous gap junctions between melanoma and endothelial cells in metastasis.

F10 and BL6 sublines of B16 mouse melanoma cells are metastatic after intravenous injection, but only BL6 cells are metastatic after subcutaneous injection. We found that connexin (Cx) 26 is upregulated in BL6 cells. To examine gap junction formation, we devised a coculture system, in which an opened vein segment was placed at the bottom of a culture dish and then dye-labeled melanoma cells were seeded onto it. Immunohistochemistry indicated that the vein segment preserved the integrity of the endothelial monolayer. In this system, BL6 cells could transfer dye into endothelial cells but F10 cells could not. Transfection with wild-type Cx26 rendered F10 cells competent for coupling with endothelial cells and as spontaneously metastatic as BL6 cells. Conversely, transfection with a dominant-negative form of Cx26 rendered BL6 cells deficient in coupling and less metastatic. In human melanoma lesions, the level of Cx26 expression was low in melanoma cells residing in the basal layer, but significantly upregulated in melanoma cells invading the dermis. The results suggested that Cx26 plays a role in intravasation and extravasation of tumor cells through heterologous gap junction formation with endothelial cells.

Animals↗
PubMed1999

Frontal midline theta rhythms reflect alternative activation of prefrontal cortex and anterior cingulate cortex in humans.

Frontal midline theta rhythm (Fm theta) often appears on electroencephalogram (EEG) during consecutive mental tasks. To clarify the source of rhythmic activity, magnetoencephalogram (MEG) and EEG were simultaneously measured in six healthy volunteers during different mental tasks using whole head MEG system. MEG records were averaged every one cycle of Fm theta rhythms using individual positive peaks of Fm theta waves in Fz EEG as a trigger. Averaged theta components of MEG signals were analyzed with a multi-dipole model. Two sources were estimated to the regions both of the prefrontal-medial superficial cortex and anterior cingulate cortex (ACC). These regions were alternatively activated in about 40 to 120 degrees phase shift during one Fm theta cycle. From above results, we hypothesize that appearance of Fm theta during consecutive mental tasks reflects alternative activities of the medial prefrontal cortex and ACC.

Adult↗
PubMed1999

Grf40, A novel Grb2 family member, is involved in T cell signaling through interaction with SLP-76 and LAT.

We molecularly cloned a new Grb2 family member, named Grf40, containing the common SH3-SH2-SH3 motif. Expression of Grf40 is predominant in hematopoietic cells, particularly T cells. Grf40 binds to the SH2 domain-containing leukocyte protein of 76 kD (SLP-76) via its SH3 domain more tightly than Grb2. Incidentally, Grf40 binds to linker for activation of T cells (LAT) possibly via its SH2 domain. Overexpression of wild-type Grf40 in Jurkat cells induced a significant increase of SLP-76-dependent interleukin (IL)-2 promoter and nuclear factor of activated T cell (NF-AT) activation upon T cell receptor (TCR) stimulation, whereas the COOH-terminal SH3-deleted Grf40 mutant lacked any recognizable increase in IL-2 promoter activity. Furthermore, the SH2-deleted Grf40 mutant led to a marked inhibition of these regulatory activities, the effect of which is apparently stronger than that of the SH2-deleted Grb2 mutant. Our data suggest that Grf40 is an adaptor molecule involved in TCR-mediated signaling through a more efficient interaction than Grb2 with SLP-76 and LAT.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Medial prefrontal cortex generates frontal midline theta rhythm.

Frontal midline theta rhythm (Fm theta) is a distinct theta activity of EEG in the frontal midline area that appears during concentrated performance of mental tasks in normal subjects and reflects focused attentional processing. To tomographically visualize the source current density distributions of Fm theta, we recorded Fm theta by using a 64-channel whole-head MEG system from four healthy subjects, and applied a new analysis method, synthetic aperture magnetometry (SAM), an adaptive beam forming method. Fm theta was observed in the MEG signals over the bilateral frontal regions. SAM analysis showed bilateral medial prefrontal cortices, including anterior cingulate cortex, as the source of Fm theta. This result suggests that focused attention is mainly related to medial prefrontal cortex.

Adult↗
PubMed1999

Stability of brain natriuretic peptide (BNP) in human blood samples.

Stability of immunoreactivity of human brain natriuretic peptide (BNP) in blood samples was investigated. After storage of the whole blood samples in the blood collecting tubes made of glass or polyethylene terephthalate (PET), residual immunoreactivity of BNP in the plasma was measured by sandwich radioimmunoassay for human BNP. BNP in the blood samples collected in the PET tubes were kept more stable than that in the glass tubes. The results suggested that commercially available PET tubes would enable more accurate BNP values and this would also help to simplify the sample preparation.

Humans↗
PubMed1999

Growth-stimulating effects of tumor necrosis factor-alpha on simian virus 40-transformed human keratinocytes is linked to phosphorylation of retinoblastoma protein.

In this report, we show that tumor necrosis factor-alpha (TNF-alpha) has a strong growth-stimulating effect on simian virus 40 (SV40)-transformed keratinocytes, while it exerts a potent growth-inhibitory effect on normal human keratinocytes. Addition of TNF-alpha to SV40-transformed keratinocytes stimulated cell growth by 216, 246, or 248% at 25, 50, or 250 ng/ml after 4 days culture compared with untreated cells, while addition to normal human keratinocytes reduced growth to 65, 38, and 32% at the same concentrations, respectively. Cell cycle analysis by flow cytometry showed that treatment of SV40-transformed keratinocytes with 250 ng/ml TNF-alpha for 48 h increased the S-phase cells from 19.8 +/- 0.3 to 49.7 +/- 2.4%. The percentage of G1/G0 phase cells decreased from 62.6 +/- 1.6 to 32.4 +/- 3.3%. We also demonstrated by immunofluorescent staining that SV40 large T antigen was enhanced by TNF-alpha incubation. Additionally, we examined the hyper and hypo-phosphorylated state of retinoblastoma protein (pRB), the cell cycle regulatory protein. TNF-alpha induced hyperphosphorylated pRB in SV40-transformed keratinocytes. On the other hand, TNF-alpha suppressed it in normal keratinocytes. We hypothesize that the enhancement of SV40 large T antigen and hyperphosphorylated pRB is involved in the mechanism of growth stimulation of SV40-transformed keratinocytes by TNF-alpha.

Cell Division↗
PubMed1999

An accurate and rapid gender determination assay in single cells by the capillary polymerase chain reaction method.

PURPOSE: In preimplantation genetic diagnosis (PGD), a rapid and accurate assay has been required. We have therefore developed a capillary polymerase chain reaction (PCR) method using rapid thermal cycling programs to determine the gender of single amniocytes. METHODS: Single amniocytes from each amniotic fluid sample were isolated by micromanipulation and their gender was determined by a multiplex PCR assay in a capillary tube, using primers that amplify a 308-bp DXZ1 and a 154-bp DYZ1 repeat sequence on the X and Y chromosomes, respectively. RESULTS: All four thermal cycling programs, which took 180, 150, 120, and 90 min, were 100% accurate in diagnosing the gender of single amniocytes. No DNA contamination was observed in any samples. CONCLUSIONS: The multiplex PCR assay was rapid and accurate in diagnosing gender in single cells and may be clinically applicable in PGD.

Amnion↗
PubMed1999

Human herpesvirus 6 infects dendritic cells and suppresses human immunodeficiency virus type 1 replication in coinfected cultures.

Human herpesvirus 6 (HHV-6) has been implicated as a cofactor in the progressive loss of CD4(+) T cells observed in AIDS patients. Because dendritic cells (DC) play an important role in the immunopathogenesis of human immunodeficiency virus (HIV) disease, we studied the infection of DC by HHV-6 and coinfection of DC by HHV-6 and HIV. Purified immature DC (derived from adherent peripheral blood mononuclear cells in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4) could be infected with HHV-6, as determined by PCR analyses, intracellular monoclonal antibody staining, and presence of virus in culture supernatants. However, HHV-6-infected DC demonstrated neither cytopathic changes nor functional defects. Interestingly, HHV-6 markedly suppressed HIV replication and syncytium formation in coinfected DC cultures. This HHV-6-mediated anti-HIV effect was DC specific, occurred when HHV-6 was added either before or after HIV, and was not due to decreased surface expression or function of CD4, CXCR4, or CCR5. Conversely, HIV had no demonstrable effect on HHV-6 replication. These findings suggest that HHV-6 may protect DC from HIV-induced cytopathicity in AIDS patients. We also demonstrate that interactions between HIV and herpesviruses are complex and that the observable outcome of dual infection is dependent on the target cell type.

Cell Survival↗
PubMed1998

Changes in extracellular glutamate and GABA levels in the hippocampal CA3 and CA1 areas and the induction of glutamic acid decarboxylase-67 in dentate granule cells of rats treated with kainic acid.

For the evaluation of glutamatergic and GABAergic transmission during seizures, rat hippocampal CA1 and CA3 areas were separately assessed by brain microdialysis, and extracelluar glutamate and GABA were measured through the course of the seizures after a systemic administration of kainic acid (KA). The generalized convulsion started at about 1.5 h and was suppressed by diazepam at 2 h after the KA treatment. In the CA3 area, extracellular glutamate started to increase soon after the KA injection and returned to the control level at about 1.5 h. A decrease and then slight increase of the extracellular glutamate level in CA3 followed the diazepam injection. In the CA1 area, in contrast, a long-lasting decrease of extracellular glutamate was observed. The extracellular GABA concentration in the CA3 area increased immediately after the systemic administration of KA and returned to the normal level at about 3.5 h. A second increase in the extracellular GABA in this area began at about 4.5 h after the KA treatment. In the CA1 area, an increase of extracellular GABA began at about 3.5 h after KA administration (much later than that observed in the CA3 area) and was maintained throughout the observation. In situ hybridization showed a transient expression of glutamic acid decarboxylase (GAD)-67 mRNA in the granule cell layer of the dentate gyrus at 4 and 6 h, whereas GAD65 mRNA was unaffected. GABA immunoreactivity in the same area and mossy fibers in the CA3 were increased most significantly at 8 h after administration of KA. The possible relation of GABA induction in mossy fibers with the delayed increase in extracellular GABA in CA3 was discussed.

Analysis of Variance↗
PubMed1998

Natural killer cell-derived large granular lymphocyte lymphoma of lung developed in a patient with hypersensitivity to mosquito bites and reactivated Epstein-Barr virus infection.

A 17-year-old female developed natural killer (NK) cell-derived large granular lymphocyte (LGL) lymphoma of the lung. She had a past history of hypersensitivity to mosquito bites (HMB). After an eight-year chronic, active Epstein-Barr virus (EBV) infection, she developed multiple lung lesions and pleural effusion. In the effusion, 60% of the cells were LGL. They were CD2+, 3-, 16+, 56+, 57+, 45RO+/RA + weak, and possessed strong NK activity. No rearrangement of T-cell-receptor genes was detected. From all these results, a diagnosis of NK-LGL lymphoma of the lung was made. EB virus DNA was detected in cells infiltrating the pleural effusion. The clonality of the LGLs was determined by Southern blot hybridization with the terminal repeat sequence of EB virus as a probe, and by chromosomal abnormalities. The patient died from respiratory failure. Necropsy of the lung revealed diffuse lymphoma composed of polymorphic cells with typical angiocentric lesions. Reportedly, lymphomas of NK lineage show predominantly extranodal involvement, and primary lung lesions are rare. In the pleural effusion of the present case, abnormally high levels of soluble Fas ligand, interleukin-10 and interferon gamma were detected. This hypercytokinemia, reflecting the microenvironment of lymphoma cells, may play a role in the progression of the lymphoma and organ injury in the lung.

Adolescent↗
PubMed1998

Tube device for facilitating distal anastomosis in aortic arch replacement.

The technique of placing an inverted graft into the descending thoracic aorta facilitates and secures the distal anastomosis in aortic arch replacement, especially in the anastomosis beyond the transverse arch. We developed a simple technique using a pair of thin-walled tubes to enable the arch graft, with its four branches, to be smoothly inserted into the flaccid, normal-caliber descending aorta. The use of these tubes simplified the procedure, resulting in time saving.

Anastomosis, Surgical↗
PubMed1998

Synchronization of normal human keratinocyte in culture: its application to the analysis of 1,25-dihydroxyvitamin D3 effects on cell cycle.

In this report, we applied the isoleucine-deprivation method to normal human keratinocytes in cultivation with serum-free MCDB153 medium. The growing cells were synchronized by transfer to MCDB153 medium prepared without isoleucine, and the degree of synchronization was analyzed by the cell cycle distribution and phosphorylation of retinoblastoma protein (pRB), one of the tumor suppressor gene products. 1,25-dihidroxyvitamin D3 (1,25(OH)2D3), a biologically active form of vitamin D3, is supposed to induce the differentiation and growth inhibition of human keratinocytes caused by cell cycle arrest. We examined its effect on cell cycle kinetics, especially progression from G1/G0 to S-phase, by using this synchronized system. We showed that 10(-6) M 1,25(OH)2D3 inhibited the progression from G1/G0 to S-phase strikingly in synchronized normal human keratinocytes.

Blotting, Northern↗
PubMed1998

Endogenous EGF-family growth factors are necessary for the progression from the G1 to S phase in human keratinocytes.

Recently several endogenous epidermal growth factor (EGF)-family growth factors (transforming growth factor-alpha, amphiregulin, and heparin-binding EGF-like growth factor) have been identified in human keratinocytes. These factors are known to play an important role in the regulation of cell proliferation. Here we show that the interaction between these factors and EGF receptor are key factors in the progression from the G1 phase to the S phase (the G1/S progression) in human keratinocytes. In this study, human keratinocytes were cultured in serum-free MCDB153 medium and then partially synchronized by isoleucine deprivation. After synchronization, the number of S phase cells increased and reached a maximum after 18-24 h. The immediate addition of anti-EGF receptor blocking antibody (1 microg per ml) to synchronized cells decreased S phase cells by 42.5% compared with untreated keratinocytes at 18 h. By contrast, the addition of anti-EGF receptor antibodies at 12 h or later did not alter the percentage of S phase cells. Northern blot analysis of synchronized cells demonstrated that mRNA expression of transforming growth factor-alpha, amphiregulin, heparin-binding EGF-like growth factor, and EGF receptor reached a maximum within 0.5-3 h after synchronization, when many cells initiated progression from the G1 to the S phase. The results show that anti-EGF receptor antibodies block the G1/S progression and the rapid increase of mRNA expression of endogenous EGF-family growth factors and EGF receptor during G1/S progression. These findings indicate that growth factor binding and EGF receptor activation are involved in the G1/S cell cycle progression of human keratinocytes.

Amphiregulin↗