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-117- Immunology(6) 9/5 Room-A
CALCIUM INVOLVEMENT IN THE ACTIVATION OF CULTURED HUMAN MAST CELLS.
Hiroo Amano, Motohiro Kurosawa, Yasushi Igarashi, Kazunori Ohnishi, Osamu Ishikawa, Yoshiki Miyachi.
Department of Dermatology, Gunma University School of Medicine, Maebashi, Japan.
Mast cells release various mediators and play an essential role in the development of immediate type allergic reaction. Many investigations of mast cell functions have been conducted using rat peritoneal mast cells. The problem is that mast cells are heterogeneous and there may be functional differences in mast cells between rats and human beings. Human mononuclear cells were obtained from heparinized umbilical cord blood with Ficoll-paque, and cultured in a medium in the presence of SCF and IL-6. The purity of mast cells reached 100 % after 12 weeks. Mast cells incubated with fura-2/AM were activated, and the fluorescense of fura-2 was measured by a spectrofluorometer. The fluorescence images were also investigated microscopically with an image processor. When IgE-sensitized mast cells were incubated with anti-IgE, histamine was released in a dose-dependent and time-dependent manner. Calcium ionophore A23187 also induced histamine release. Fluorescence intensity increased obviously in fura-2-loaded mast cells exposed to the stimulators.
-118- Immunology(6) 9/5 Room-A
CHARACTERIZATION OF BONE MARROW-DERIVED MAST CELLS OBTAINED FROM NC MICE.
Takaaki Hiragun, Eishin Morita, Yoshikazu Kameyoshi, Toshihiko Tanaka, Shoso Yamamoto.
Department of Dermatology, Hiroshima University School of Medicine, Hiroshima, Japan.
NC mice spontaneously develop atopic eczema-like skin lesion associated with dermal mast cell hyperplasia. Skin histamine content of NC was 69.0mg/g wet tissue and significantly higher than those of C57BL/6 (16.1mg/g wet tissue) and BALB/c (19.4 mg/g wet tissue). To know a mechanism of this mast cell hyperplasia in NC mice, we investigated properties of bone marrow-derived mast cells (BMMC) obtained from NC mice. Histamine content of BMMC from NC was 0.67pg/cell and significantly higher than those of C57BL/6 (0.09pg/cell) and BALB/c (0.04pg/cell). Percent histamine release in BMMC was found to be similar in these three strains. Adhesion of BMMC to plastic dishes was higher in NC compared with C57BL/6 and BALB/c. These result indicate that NC mice has an abnormality in histamine content and adhesive property of mast cells.
-119- Immunology(6) 9/5 Room-A
FK506 AND CYCLOSPORIN A SELECTIVELY INHIBIT STEM CELL FACTOR-DEPENDENT SIGNAL TRANSDUCTION ANd UPREGULATE C-KIT EXPRESSION IN MOUSE MAST CELL LINE, MC/9 CELLS.
Fumihiko Ito, Noriaki Toyota, Hiruyuki Sakai, Hidetishi Takahashi, Hajime Iizuka.
Department of Dermatology, Asahikawa Medical College, Asahikawa, Japan.
Using mouse mast cell line, MC/9, we investigated the effects of immunosuppressants, FK506 and cyclosporin A (CsA), on IL-3-dependent or SCF-dependent proliferation and signal transduction. Neither FK506 nor CsA affected IL-3-dependent MC/9 cell proliferation, whereas they selectively inhibited SCF-dependent cell survival. MC/9 cells following these treatments could not be induced synergistic proliferation by both IL-3 and SCF and maintained their survival by SCF alone, resulting in apoptosis. Flow cytometry showed that they did not affect IL-3 receptor expression. However, immunoblot and RT-PCR analyses indicated increased c-kit protein and c-kit mRNA transcripts following FK506 and CsA treatments. In addition, adhesion assay revealed the increased adhesive MC/9 cells to NIH/3T3 cells following these treatments, consistent with that the increased c-kit protein is associated with membrane-bound SCF of NIH/3T3 cell. The increased c-kit receptor, however, seems to be not functional with regard to signal transduction. Neither FK506 nor CsA affected c-kit tyrosine phosphorylation and MAP kinase nuclear translocation following SCF stimulation.
-120- Immunology(6) 9/5 Room-A
INHIBITION OF HISTAMINE RELEASE BY MEDICINAL HERBAL EXTRACTS.
Akira Hatani, Mako Taniguchi, Hidenobu Okumura, Makoto Terai.
Shiga Central Research Laboratory, NOEVIR Co., Ltd., Shiga , Japan.
In general, immediate allergy reaction is one inducer for atopic dermatitis. This allergy is caused by chemical mediators released by basophiles and mast cells on deguranulation. Rat basophilic leukemia cells (RBL-2H3 cells) are known to release various chemical mediators induced by DNP-BSA antigen binding to anti-DNP IgE antibodies on the cell surface. We investigated the inhibitory effects of histamine release from RBL-2H3 cells for 200 medicinal herbal extracts.¡¡The histamine was measured by post-column HPLC method. The extract of Gambir, Rosae Fructus, Caryophlli Flos, Sanguisorba officinalls, Cinchonae Cortex, and Betula lenta inhibited the histamine release from th cells by more than 50%.¡¡Now we are investigating the main substance in each extracts.
-121- Immunology(6) 9/5 Room-A
SPONTANEOUS RELEASE OF SUBSTANCE-P IN SKIN OF CHRONIC URTICARIA.
Michihiro Hide, Osamu Koro, Sakae Kaneko, Eishin Morita, Kayo, Okahara, Naomasa Niimi, Shoso Yamamoto.
Department of Dermatology, Hiroshima University School of Medicine, Hiroshima, Japan.
Chronic Urticaria (CU) is often aggravated by psychological stress, suggesting involvement of neurological elements with this disease. In order to elucidate the involvement of neuropeptides with CU, we studied the release of substance-P (SP) in skin of patients with CU. Saline added in glass chambers with or without antigen on the abraded skin surface was collected every 30 min until 120 min. SP and histamine concentrations in collected samples were measured by EIA and RIA respectively. Periodical large release of SP was observed regardless of histamine concentration or addition of antigen, suggesting the involvement of spontaneous SP release with mast cell degranulation in CU.