口腔医学研究 ›› 2015, Vol. 31 ›› Issue (4): 361-364.

• 临床研究论著 • 上一篇    下一篇

人淋巴管内皮细胞的分离和鉴定

魏军成1, 张壁2, 陈琦2, 王贻宁2, 赵艳2*   

  1. 1. 华中科技大学同济医学院附属同济医院妇产科 湖北 武汉 430030;
    2. 武汉大学口腔医院修复科
  • 收稿日期:2014-12-15 出版日期:2015-04-28 发布日期:2016-04-29
  • 通讯作者: 赵艳,Email: zhao.yan@whu.edu.cn
  • 作者简介:魏军成(1975~ ),男,湖北人,博士,主治医师,主要从事妇科肿瘤研究。
  • 基金资助:
    国家自然科学基金青年项目(编号:81400544 )

Isolation and Characterization of Human Lymphatic Endothelial Cells.

WEI Jun-cheng, ZHANG Bi, CHEN Qi, et al   

  1. Department of Obstetrics and Gynecology, Tongji Hospital, Huazhong University of Science and Technology, Wuhan 430030
  • Received:2014-12-15 Online:2015-04-28 Published:2016-04-29

摘要: 目的:分离培养人淋巴管内皮细胞(LECs),为研究淋巴管新生在淋巴管瘤复发及肿瘤淋巴结转移中的作用奠定基础。方法:HE染色和免疫组织化学法了解人淋巴管瘤的组织病理特征;胶原酶消化法从淋巴管瘤组织中分离淋巴管内皮细胞;光镜和透射电镜观察淋巴管内皮细胞的形态和超微结构;MTT法检测细胞体外增殖。结果:HE染色显示舌淋巴管瘤中具有大量扩张的管腔,免疫组织化学染色显示管腔内皮细胞表达FLT-4、和LYVE-1;体外培养的淋巴管内皮细胞呈"铺路石"样外观;在透射电子显微镜下,观察到淋巴管内皮细胞特征性细胞器如weibel-palade小体、胞质小突和质膜小泡;MTT结果显示细胞于第3天开始进入增殖活跃期,第5天进入高峰期,第6、7天进入平台期;在Ⅰ型胶原凝胶中细胞形成管腔样结构。结论:应用胶原酶消化法可以有效的从淋巴管瘤组织中分离得到淋巴管内皮细胞。

关键词: 淋巴管瘤, 淋巴管内皮细胞, 细胞培养

Abstract: Objective: To assist further investigation of the function of lymphangiogenesis during cancer metastasis. Methods: Hematoxylin-eosine staining and immunohistochemistry were used to detect histopathological features of lymphangioma. Human lymphatic endothelial cells were isolated by collagenase digestion, and were cultured in M199 medium added with vascular growth factor and heparin. Morphology and ultrastructure of cells were observed under contras phase light microscope and transmission electron microscope. MTT was applied to survey proliferation of lymphatic endothelial cells. Results: Hematoxylin-eosine staining sections revealed that numerous dilated vessels were generally presence in glossal lymphanigoma specimen. FLT-4 and LYVE-1 were positive in lymphatic endothelial cells. LECs showed a typical cobblestone appearance observed under contrast phase light microphage. The typical granule, weibel-palade body, that was unique to the endothelial cells, is exhibited in the ultrastructure of lymphatic endothelial cells. Gap intercellular junctions in adjacent lymphatic endothelial cells were overviewed in the ultrathin sections of lymphatic endothelial cells. Lymphatic endothelial cells formed tube-like structures and communicated with each other in the collagenⅠgel to form a complex vascular plexus similar to the vessel system in vivo. Conclusion: We have successfully isolated and expanded LECs in vitro, and provided a favorable model for researching the phenotypic and functional properties of LECs.

Key words: Lymphangioma , Lymphatic endothelial cell, Cell culture

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