Luo Shizhen, Truong Aaron H, Makino Ayako
Department of Physiology, University of Arizona, Tucson.
Department of Physiology, University of Arizona, Tucson;
J Vis Exp. 2016 Jul 3(113):53985. doi: 10.3791/53985.
Endothelial cells line the inner wall of blood vessels and play an important role in the regulation of vascular tone, vascular permeability, and new vascular formation. Endothelial cell dysfunction is implicated in the development and progression of many cardiovascular diseases including ischemic heart disease. To examine the function and characterization of coronary endothelial cells, cell isolation is the first step and it requires high purity and quantity to conduct subsequent experiments. This protocol describes an efficient method to isolate adult mouse coronary endothelial cells. The mouse heart is dissected and minced into small pieces. After the digestion of the heart using dispase and collagenase II, cells are washed and incubated with magnetic beads which are conjugated with anti-CD31 antibody. The beads with endothelial cells are washed several times and are ready to use in various applications, including imaging and molecular biological experiments. Efficient isolation yields approximately 10(4) cells per one heart with over 90% purity.
内皮细胞排列在血管内壁，在调节血管张力、血管通透性和新血管形成中发挥重要作用。内皮细胞功能障碍与包括缺血性心脏病在内的许多心血管疾病的发生和发展有关。为了研究冠状动脉内皮细胞的功能和特性，细胞分离是第一步，并且需要高纯度和数量才能进行后续实验。本方案描述了一种分离成年小鼠冠状动脉内皮细胞的有效方法。解剖小鼠心脏并切成小块。用分散酶和胶原酶II消化心脏后，洗涤细胞并与结合有抗CD31抗体的磁珠孵育。带有内皮细胞的磁珠洗涤几次后即可用于各种应用，包括成像和分子生物学实验。高效分离每颗心脏可产生约10(4)个细胞，纯度超过90%。
