Purification and Properties of a New Endotherium Growth Suppressing Factor from Macrophages

线性（1→3）-β-D-葡聚糖的羧甲基化衍生物（CMCD）直接作用于小鼠腹腔巨噬细胞和小鼠淋巴瘤P388 D1细胞，并在100μg/mL浓度下诱导其自身对牛动脉内皮细胞（BAE）的生长抑制活性。在100μg/mL CMCD存在下，从小鼠淋巴瘤P388 D1细胞培养物的条件培养基中纯化内皮细胞生长抑制因子（EGSF）。纯化步骤依次包括硫酸铵分级分离和六个阶段的柱层析。在还原条件下，纯化的EGSF在SDS-PAGE上显示两条带，用银染色，并且它们的分子量估计为约55和63 kDa，而纯化的EGSF的分子量通过使用Superdex 200 HR的凝胶过滤估计为约65 kDa。该因子强烈抑制来自牛动脉、人脐静脉和人真皮毛细血管的内皮细胞的增殖， ...更多信息 观察到抑制是可逆的。静脉注射纯化的EGSF给荷肉瘤180的小鼠，在16小时内引起肿瘤块中活肿瘤细胞数量的迅速减少。免疫组化结果显示，在此情况下，肿瘤灯内的组织发生出血性病变。因此，纯化的因子不仅表现出对内皮细胞的生长抑制活性，而且还表现出肿瘤消退活性。纯化的因子显着抑制体外牛动脉，人脐静脉，和成年人的真皮微血管内皮细胞在胶原凝胶上的小管形成。在胶原凝胶上完成内皮细胞的管形成后，纯化的因子在约5 ng/ml的浓度下在48 h内破坏管。这些发现表明EGSF是血管生成以及内皮细胞生长的有效抑制剂，并且可以通过抑制血管生成来实现实体瘤的消退。少

A carboxymethylated derivertives of a linear (1→3)-β-D-glucan (CMCD) acted directly on mouse peritoneal macrophages and mouse lymphoma P388D1 cells, and induced a growth suppressing activity for bovine artery endothelial cells (BAE) from themselves at a concentration of 100μg/mL. An endothelial cell growth-suppressing factor (EGSF) was purified from the conditioned medium of a mouse lymphoma P388D1 cell culture in the presence of 100μg/mL CMCD. The purification steps included, in order, ammonium sulfate fractionation and six stages of column chromatography. The purified EGSF showed two bands with silver staining on SDS-PAGE under reducing conditions and their molecular masses were estimated as approximately 55 and 63 kDa, while the molecular mass of the purified EGSF was estimated as about 65 kDa by gel filtration using Superdex 200HR. This factor strongly suppressed the proliferation of endothelial cells from bovine artery, human umbilical vein, and human dermal vas capillare and this … More suppression was observed to be reversible. The intravenous administration of the purified EGSF to sarcoma 180-bearing mouse caused a rapid decrease in the number of viable tumor cells in tumor lumps within 16h. Immunohistochemical study demonstrated that hemorrhagic disorder all over the tissue in the tumor lamp occurred under this circumstance. Thus, the purified factor exhibited not only growth suppressing activity for endothelial cells but also tumor regressing activity. The purified factor significantly inhibited in vitro tubulogenesis of bovine artery, human umbilical vein, and adult human darmal microvascular endothelial cells on collagen gel. After the tube formation of endothelial cells was completed on a collagen gel, the purified factor disrupted the tubes at a concentration of about 5 ng/ml within 48h. These findings demonstrate that EGSF is a potent inhibitor of angiogenesis as well as the growth of endothelial cells, and may bring about the regression of a solid tumor by inhibiting angiogenesis. Less