Garrick R A
Department of Natural Sciences, Fordham College at Lincoln Center, Fordham University, 113 W. 60th Street, LL813, New York, New York 10023, USA.
Microvasc Res. 2000 May;59(3):377-85. doi: 10.1006/mvre.1999.2234.
There has been little development of endothelial cell cultures from vertebrates other than mammals. In this report the isolation of capillary endothelial cells from the rete mirabile of the eel, Anguilla rostrata, is described. The cells are isolated with collagenase H and dispase II. The cells are plated into fibronectin-hyaluronic acid coated flasks. The culture medium is M199 with Earle's salts supplemented with NaCl, HEPES, NaHCO(3), glutamine, pyruvate, heparin, antibiotics, endothelial cell growth supplement, and 20% serum. Cultures are incubated at 25 degrees C in humidified air. The rete mirabile contains pericytes in addition to endothelial cells. Variations in plating time, serum concentrations, and growth matrices were tried to separate the two cell types. The total number of endothelial cells and the ratio of endothelial cells to pericytes are the most important factors in obtaining pure cultures of capillary endothelial cells. Endothelial cells are isolated also from the endocardium, bulbus arteriosus, and large vessels. The initial isolates usually take 3-6 weeks to grow to confluence with subcultures taking about 2 weeks to confluence.
除哺乳动物外，脊椎动物内皮细胞培养的发展一直很少。在本报告中，描述了从美洲鳗鲡的奇静脉分离毛细血管内皮细胞的方法。用胶原酶H和Dispase II分离细胞。将细胞接种到纤连蛋白-透明质酸包被的培养瓶中。培养基是添加了NaCl、HEPES、NaHCO₃、谷氨酰胺、丙酮酸盐、肝素、抗生素、内皮细胞生长补充剂和20%血清的含Earle盐的M199。培养物在25℃的潮湿空气中孵育。奇静脉除了内皮细胞外还含有周细胞。尝试通过改变接种时间、血清浓度和生长基质来分离这两种细胞类型。内皮细胞总数以及内皮细胞与周细胞的比例是获得纯毛细血管内皮细胞培养物的最重要因素。内皮细胞也可从心内膜、动脉球和大血管中分离出来。最初的分离物通常需要3至6周才能生长至汇合，传代培养则需要约2周才能汇合。
