Lin Sheng, Long Hai-xia, Xiang Tong, Zhu Bo, Xie Rong-kai
Department of Oncology, Xinqiao Hospital of the Third Military Medical University, Chongqin 400038, China.
Zhonghua Zhong Liu Za Zhi. 2011 Dec;33(12):896-9.
To isolate and identify the cancer stem cells from primary human ovarian cancer tissues.
Fresh tumor tissues from five cases of pathologically diagnosed ovarian cancers were taken, minced and then digested with collagenase and hyaluronidase to obtain single cell suspension. The erythrocytes were removed with ACK Lysis buffer. The suspensions were sorted by magnetic activated cell sorting (MACS) using CD133-binding microbeads. Then the sorted CD133(+) cells were verified by flow cytometry. The cells were cultured in serum-free medium supplemented with EGF, bFGF, insulin and BSA, and grew into spheroids. Immunofluorescence, differentiation and tumor formation tests of the cells were performed to characterize the properties of cancer stem cells.
The ovarian cancer stem cells were successfully isolated from primary human ovarian tumors, which formed typical spheroids in serum-free medium and had stronger ability of tumorigenesis. The results of related experiments verified that CD133 positive cells owned the properties of cancer stem cells.
The ovarian cancer stem cells presenting the characteristics of stemness in vitro and in vivo, have been successfully isolated from primary human ovarian tumor tissues by MACS. The isolated ovarian cancer stem cells could be used in future researches of their biological functions.
从原发性人类卵巢癌组织中分离并鉴定癌症干细胞。
取5例经病理诊断为卵巢癌的新鲜肿瘤组织，切碎后用胶原酶和透明质酸酶消化，以获得单细胞悬液。用ACK裂解缓冲液去除红细胞。使用结合CD133的微珠通过磁性激活细胞分选（MACS）对悬液进行分选。然后通过流式细胞术验证分选的CD133(+)细胞。将细胞在补充有表皮生长因子（EGF）、碱性成纤维细胞生长因子（bFGF）、胰岛素和牛血清白蛋白（BSA）的无血清培养基中培养，并长成球体。对细胞进行免疫荧光、分化和肿瘤形成试验，以表征癌症干细胞的特性。
成功从原发性人类卵巢肿瘤中分离出卵巢癌干细胞，其在无血清培养基中形成典型球体，且具有更强的肿瘤发生能力。相关实验结果证实CD133阳性细胞具有癌症干细胞的特性。
通过MACS已成功从原发性人类卵巢肿瘤组织中分离出在体外和体内均呈现干性特征的卵巢癌干细胞。分离出的卵巢癌干细胞可用于其生物学功能的未来研究。
