Watanabe M, Nittoh T, Suzuki T, Kitoh A, Mue S, Ohuchi K
Department of Pathophysiological Biochemistry, Faculty of Pharmaceutical Sciences, Tohoku University, Miyagi, Japan.
Int Arch Allergy Immunol. 1995 Sep;108(1):11-8. doi: 10.1159/000237111.
Peritoneal eosinophilia was induced in rats using Ascaris suum extract as an antigen, and characteristics of granule proteins of eosinophils collected from the peritoneal cavity were investigated. Peritoneal eosinophilia was induced by injection of the antigen solution into the peritoneal cavity of the immunized rats that had been orally administered with cyclophosphamide. Peritoneal cells were collected 48 h after injection of the antigen solution, incubated in plastic dishes, and nonadherent cells were collected as an eosinophil-rich fraction, from which granule proteins were extracted. Granule proteins were then purified by cation exchange chromatography, gel filtration, copper chelate affinity chromatography, and reverse-phase HPLC. Two distinct basic proteins of which molecular weights are 18- and 17-kD were obtained. Partial N-terminal amino acid sequence analysis revealed that the 18-kD protein and the 17-kD protein were homologous to the human eosinophil cationic protein (ECP) and the human and guinea pig major basic protein (MBP), respectively. Both the two proteins showed strong bactericidal activity against Escherichia coli and Staphylococcus aureus. These results indicate that rat eosinophils also possess ECP and MBP in their granules as well as human and guinea pig eosinophils.
以猪蛔虫提取物作为抗原在大鼠中诱导产生腹腔嗜酸性粒细胞增多，并研究从腹腔收集的嗜酸性粒细胞颗粒蛋白的特性。通过将抗原溶液注射到经口服环磷酰胺免疫的大鼠腹腔中诱导产生腹腔嗜酸性粒细胞增多。在注射抗原溶液48小时后收集腹腔细胞，在塑料培养皿中孵育，收集非贴壁细胞作为富含嗜酸性粒细胞的部分，从中提取颗粒蛋白。然后通过阳离子交换色谱、凝胶过滤、铜螯合亲和色谱和反相高效液相色谱法纯化颗粒蛋白。获得了两种分子量分别为18 kD和17 kD的不同碱性蛋白。部分N端氨基酸序列分析表明，18 kD蛋白和17 kD蛋白分别与人嗜酸性粒细胞阳离子蛋白（ECP）以及人和豚鼠主要碱性蛋白（MBP）同源。这两种蛋白均对大肠杆菌和金黄色葡萄球菌表现出强大的杀菌活性。这些结果表明，大鼠嗜酸性粒细胞的颗粒中也像人和豚鼠嗜酸性粒细胞一样含有ECP和MBP。
