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

1.犬皮肤肥大细胞瘤来源（CM-MC）细胞被IgG和ige介导的机制激活。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以剂量依赖性方式抑制igg介导的[Ca^<2+>]i升高和组胺释放。采用免疫沉淀法和RT-PCR法检测CM-MC细胞中高亲和力IgG受体（fc γ - ri）蛋白和mRNA的表达。然后，鉴定了FcγRIα亚基的cDNA序列。犬FcγRIα cDNA全长1119bp （GenBank登录号AB 101519）。犬fc γ - ria cDNA与人和小鼠fc γ - ria cDNA的总体同源性分别为84%和78%。我们将fc γ - ri α cDNA转染到COS-7细胞中，检测单体IgG与细胞的结合情况。利用RT-PCR和5′/3′-RACE技术，从CM-MC细胞总RNA中克隆出犬fc γ - ri α的cDNA。将cDNA连接到哺乳动物表达载体上，转染到COS -7细胞中。用流式细胞术检测IgG与COS -7细胞的结合情况。物质P （10 ~ 30 μM）和C5a （0.5 ~ 1μM）对CM-MC具有剂量依赖性。[Ca^<2+>]i升高和脱颗粒被胰岛活化蛋白（IAP）抑制。因此，这些物质的活化似乎依赖于g蛋白偶联机制5。综上所述，CM-MC细胞可用于研究igg依赖机制引起的变应性炎症。影响P物质和C5a产生的环境化学物质可能影响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.