A novel method for endothelial cell isolation
A novel method for endothelial cell isolation
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内皮细胞分离的新方法
DOI：
10.3892/or.2015.4490
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发表时间：
2016-03-01
期刊：
影响因子：
4.2
通讯作者：
Zhao, Yongxiang
中科院分区：
文献类型：
--
作者：
Mao, Qiqi;Huang, Xianing;Zhao, Yongxiang
本研究旨在建立一种快速、有效的肿瘤组织中新生血管内皮细胞的纯化方法。从携带源自小鼠肺癌刘易斯细胞的肿瘤的C57 BL/6小鼠分离新鲜组织，完全切碎并分成两部分。一部分用涡旋进行I型胶原酶消化以形成单细胞悬液，而另一部分消化但不进行涡旋。然后，使用抗CD 105抗体包被的Dynabeads分离CD 105细胞。将分离的CD 105（+）细胞在培养基中生长，并通过荧光激活细胞分选仪（FACS）检查CD 105的表面表达。Dil-Ac-LDL摄取实验和微管形成实验检测了CD 105（+）细胞对乙酰化LDL的摄取和维持毛细血管样结构形成的能力。免疫组化法检测刘易斯移植瘤新生血管内皮细胞（CD 105（+））的表达。有涡流的消化过程中获得的细胞数为5.70 ± 0.23 × 10 ~（4），远高于无涡流消化过程中获得的细胞数（0.32 ± 0.04 × 10 ~（4））（P
The present study aimed to develop a quick and efficient method for purification of newborn endothelial cells from tumor tissues. Fresh tissues were separated from C57BL/6 mice bearing tumors derived from mouse lung cancer Lewis cells, fully minced and divided into two parts. One part was subjected to collagenase type I digestion with a vortex to form a single-cell suspension, while another part was digested but without a vortex. Then, the CD105 cells were isolated using anti-CD105 antibody-coated Dynabeads. The isolated CD105(+) cells were grown in culture medium and examined for the surface expression of CD105 by a fluorescence-activated cell sorter (FACS). The uptake of acetylated LDL and the ability to maintain capillary tube-like structure formation in the CD105(+) cells were also examined by Dil-Ac-LDL uptake assay and tube formation assay. The expression of tumor newborn endothelial cells (CD105(+)) was tested in Lewis xenografts by immunohistochemistry. The number of cells which were obtained by the digestion process with a vortex was 5.70 +/- 0.23x10(4) much higher than the number without a vortex (0.32 +/- 0.04x10(4))(P