人卵巢癌微血管内皮细胞的培养及体外管腔样结构形成的观察.docVIP

人卵巢癌微血管内皮细胞的培养及体外管腔样结构形成的观察 徐 燕，梁志清，王 丹，谢 尧（第三军医大学西南医院妇产科，重庆 400038) 摘要：目的 建立人卵巢癌微血管内皮细胞(ovarian carcinoma-derived microvascular endothelial cellsODMECs)免疫磁珠分选体系并观察ODMEC体外二维管腔样结构（tubule-like structure，TLS)的形成。利用结合CD31抗体的MACS MicroBeads 免疫磁珠分选系统纯化ODMEC。对所获得的ODMEC进行鉴定DIL-acLDL）和结合凝集素Ⅰ（UEA-Ⅰ）的能力，观察其在体外培养体系中二维管腔样结构的形成。结果 所获ODMEC流式分析细胞纯度可达99.6%表达CD31、FⅧ-Rag；具有结合UEA-Ⅰ和摄取DIL-acLDL的能力；培养过程中观测到ODMECs可形成二维管腔样结构。本研究成功建立了人ODMEC的免疫磁珠；微血管内皮细胞；卵巢癌；细胞培养uman ovarian cancer microvascular endothelial cells and formation of lumen-like structure in vitro XU Yan，LIANG Zhi-qing，WANG Dan, XIE Yao（Department of Obstetrics and Gynecology, Southwest Hospital, Third Military Medical University, Chongqing 400038, China）Abstract: Objective To establish the magnetic activated cell sorting system (MACS)for isolation and incubation of human ovarian carcinoma-derived microvascular endothelial cells (ODMEC) and to observe their ability to form tubule-like structure (TLS) in vitro. Methods MACS was used to purify ODMEC with antiCD31 antibody and ODMEC were identified according to their morphological，biochemical and phenotypic characteristics.ODMEC to intake acetylated-low-density lipoprotein (DIL-acLDL) and Ulex europeaus agglutinin -1 (UEA-Ⅰ) was assayed to observe the formation of TLS in vitro. Results Flow cytometry showed that the purity of ODMEC was up to 99.6 %. The ODMEC were characterized by cobble stones, contact inhibition, expressed CD31，CD105FⅧ-Rag, and exhibited the ability to bind to UEA-1,intake DIL-acLDL and form TLS during incubation. Conclusion The MACS we established can be used to isolate and incubate human ODMEC and study the formation characteristics of TLS. Key Words:magnetic beads; microvascular endothelium cell; ovarian carcinoma；cell culture 由于肿瘤的生长和转移离不开血管的生成，因此抗肿瘤血管生成一直是研究的热点。多项研究表明，受微环境的影响，肿瘤血管内皮细胞在形态结构、基因的表达、以及遗传学水平具有异质性[1]，这一特点推动了来源于不同瘤体组织的微血管内皮细胞分离和培养。目前体外培养血管内皮细胞已成为实验室常规方法，其广泛应用已有30多年的历史，在此期间分离来源于不同组织的血管内皮细胞特别是微血管的方法得以不断改进和提高。在前期实验中[2]我们利用卵巢癌微血管内皮细胞(ovarian