Study of the effect of environmental chemicals on the high-affinity IgG receptor-mediated signal transudation of mest cells.

1.犬皮肤肥大细胞瘤衍生（CM-MC）细胞激活IgG和IgE介导的机制。IgG介导的蛋白酪氨酸磷酸化和Ca^2+内流与IgE介导的相似。蛋白激酶抑制剂staurosporine呈剂量依赖性地抑制IgG介导的[Ca^<2+>]i升高和组胺释放。2.免疫沉淀法和RT-PCR法检测CM-MC细胞内高亲和力IgG受体（FcγRI）蛋白和mRNA的表达。对FcγRIα亚基的cDNA序列进行了鉴定。犬FcγRIα cDNA全长为1119 bp（GenBank登录号AB 101519）。犬Fcγ RI α cDNA与人和小鼠FcγRIα cDNA的同源性分别为84%和78%。以CM-MC细胞总RNA为模板，采用RT-PCR和5 '/3'-RACE技术克隆了犬FcγRIα的cDNA。将该cDNA与哺乳动物表达载体连接，转染COS-7细胞。4. P物质（10-30 μM）和C5 a（0.5-1μM）对CM-MC的激活作用呈剂量依赖性。[Ca胰岛活化蛋白（IAP）可抑制这些物质引起的[[2+]i升高和脱粒.因此，这些物质的激活可能依赖于G蛋白偶联机制。5.总之，CM-MC细胞可用于IgG依赖性机制引起的过敏性炎症的研究。影响P物质和C5 a产生的环境化学物质可能会影响IgG依赖性肥大细胞活化引起的过敏性炎症。

1.Canine cutaneous mastocytoma-derived (CM-MC) cells were activated both IgG- and IgE-mediated mechanisms. IgG-mediated protein tyrosine phosphorylation and Ca^<2+> influx were similar to those mediated by IgE. Protein kinase inhibitor staurosporine inhibited IgG-mediated [Ca^<2+>]i elevation and histamine release in a dose-dependent manner.2.The presence of high affinity IgG receptors (FcγRI) protein and mRNA in CM-MC cells was determined by immunoprecipitation and RT-PCR methods. Then, the cDNA sequence of FcγRIα subunit was identified. The entire canine FcγRIα cDNA was 1119bp long (GenBank accession number, AB 101519). The overall identity of the canine FcγRIa cDNA to its human and mouse counterparts was 84% and 78%, respectively.3.We transfected cDNA of FcγRIα to COS-7 cells to examine the binding of monomeric IgG to the cells. The cDNA of canine FcγRIα was cloned by RT-PCR and 5'/3'-RACE from total RNA of CM-MC cells. The cDNA was ligated to mammalian expression vector and transfected into COS -7 cells. The binding of IgG onto the COS -7 cells was detected by flow-cytometry.4.CM-MC was activated by Substance P (10-30 μM) and C5a (0.5-1μM) dose-dependently. [Ca^<2+>]i elevation and degranulation by these substances was inhibited by islet- activated protein (IAP). Therefore, the activation of these substances seemed to be dependent on G-protein-coupled mechanism5.In conclusion, CM-MC cells were useful for the study of allergic inflammation caused by IgG-dependent mechanisms. The environmental chemicals that affect Substance P and C5a production might influence allergic inflammation caused by IgG-dependent mast cell activation.