Koyama M, Satoh K, Yoshida H, Suzuki S, Koie H, Takamatsu S
Department of Pathological Physiology, Hirosaki University School of Medicine, Japan.
Thromb Haemost. 1996 Oct;76(4):610-4.
Surface coverage with autogeneous endothelial cells is effective in reducing thrombogenicity of an artificial vascular graft, but procedure for obtaining the cells is invasive for patients. The purpose of this study was to establish cultures of human endothelial cells separated from a small piece of subcutaneous fat tissue. A piece of tissue weighing about 10 mg was obtained from subcutaneous fat using a biopsy needle, and treated with collagenase and dispase. Microvascular endothelial cells were selected and other types of cells contaminating the cultures were eliminated by scraping with a needle under a microscope. The yield of the cells was 8362 +/- 4264/10 mg of subcutaneous fat (n = 7). The cultures reached confluence in about 2 weeks. The cells were positive for von Willebrand factor, P-selectin, and uptake of acetylated low density lipoprotein. The cells produced 15.9 +/- 3.3 ng/mg cell protein/h of 6-ketoprostaglandin F1 alpha (n = 5) when stimulated with thrombin. Thrombin also stimulated the production of platelet-activating factor: 7653 +/- 4297 dpm/10(6) cells (n = 5). Endothelin-1 accumulation in the medium of unstimulated endothelial cells was 0.54 +/- 0.16 ng/mg cell protein/10 h (n = 8). As a preliminary experiment for graft seeding, the cells were also cultured on pieces of a gelatin-coated Dacron graft, and scanning electron microscopy revealed the surface coverage of the graft. We herein described about successful culture of human microvascular endothelial cells from subcutaneous fat tissue obtained using a biopsy needle. The cultured cells may be applicable to a seeded vascular graft.
自体内皮细胞覆盖可有效降低人工血管移植物的血栓形成性，但获取这些细胞的操作对患者具有侵入性。本研究的目的是建立从一小片皮下脂肪组织中分离的人内皮细胞培养物。使用活检针从皮下脂肪获取约10mg重的一块组织，并用胶原酶和分散酶处理。选择微血管内皮细胞，并通过在显微镜下用针刮除来消除污染培养物的其他类型细胞。细胞产量为8362±4264/10mg皮下脂肪（n = 7）。培养物在约2周内达到汇合。这些细胞对血管性血友病因子、P-选择素呈阳性，并能摄取乙酰化低密度脂蛋白。当用凝血酶刺激时，这些细胞产生15.9±3.3ng/mg细胞蛋白/小时的6-酮前列腺素F1α（n = 5）。凝血酶也刺激血小板活化因子的产生：7653±4297dpm/10⁶细胞（n = 5）。未刺激的内皮细胞培养基中内皮素-1的积累量为0.54±0.16ng/mg细胞蛋白/10小时（n = 8）。作为移植物接种的初步实验，这些细胞也在明胶包被的涤纶移植物片上培养，扫描电子显微镜显示了移植物的表面覆盖情况。我们在此描述了使用活检针从皮下脂肪组织成功培养人微血管内皮细胞的情况。培养的细胞可能适用于接种的血管移植物。
