Wang Yong, Liu Ya-hui, Jiang Jian-shuai, Cui Han-bin
Department of Hepatobiliary & Pancreatic Surgery, Ningbo First Hospital, Ningbo, China.
Zhonghua Yi Xue Za Zhi. 2012 Dec 25;92(48):3434-7.
To explore a new efficient purification method of hepatocellular carcinoma (HCC) stem cells and identify their features.
Human hepatocellular carcinoma cell line SMMC-7721 was cultured in sphere-culture system of polyhedra-treated dish and tumor stem cell specific medium. Upon the formation of cellular sphere, the cells were inoculated subcutaneously into immunocompromised mice and received the interventions of different concentrations of cisplatin. Then the drug-resistant cells were purified and re-cultured in TSC medium. Finally the stem cell markers and tumor stem cell markers were determined through real-time polymerase chain reaction (PCR), immunofluorescence method and flow cytometry.
Through the double filter of TSC medium and cisplatin-resistance, SMMC-7721 stem cells could be grown in a suspended form and formed spheres in TSC medium. The stem cell markers (NANOG, OCT-4, SOX-2 and Notch) and tumor stem cell markers (CD24, 90.0%; CD133, 6.1%; CD90, 4.8%) were all over-expressed in purified cancer stem cells as compared with ordinary cells. And the over-expression of CD24 was the most obvious.
The combination of in vitro cell culture with TSC medium, in vivo proliferation and cisplatin resistance test is a new efficient method of purifying hepatocellular carcinoma stem cells. Tumor stem cell with stem cell characteristics and an over-expression of CD24 may be cloned from SMMC-7721.
探索一种新的高效纯化肝癌干细胞的方法并鉴定其特征。
将人肝癌细胞系SMMC - 7721培养于经多面体处理的培养皿的球状体培养系统及肿瘤干细胞特异性培养基中。细胞球形成后，将细胞皮下接种于免疫缺陷小鼠，并给予不同浓度顺铂干预。然后纯化耐药细胞并在TSC培养基中重新培养。最后通过实时聚合酶链反应（PCR）、免疫荧光法和流式细胞术检测干细胞标志物和肿瘤干细胞标志物。
通过TSC培养基和顺铂耐药双重筛选，SMMC - 7721干细胞能够以悬浮形式生长并在TSC培养基中形成球体。与普通细胞相比，纯化后的癌干细胞中干细胞标志物（NANOG、OCT - 4、SOX - 2和Notch）和肿瘤干细胞标志物（CD24，90.0%；CD133，6.1%；CD90，4.8%）均呈过表达。且CD24的过表达最为明显。
体外细胞培养与TSC培养基、体内增殖和顺铂耐药试验相结合是一种新的高效纯化肝癌干细胞的方法。具有干细胞特征且CD24过表达的肿瘤干细胞可能可从SMMC - 7721中克隆获得。
