腹水来源卵巢上皮性癌细胞系中侧群细胞的分离、鉴定及耐药性分析  

Identification and characterization of stem cells in an ovarian cancer cell line and examination their drug resistance

作　　者：魏振彤[1] 王医术[2] 于晓伟[1] 张松灵[1] 

机构地区：[1]吉林大学第一医院妇产科,长春130021 [2]病理生物学教育部重点实验室

出　　处：《中华妇产科杂志》2015年第6期452-457,共6页Chinese Journal of Obstetrics and Gynecology

基　　金：吉林省科技发展计划（20140520032JH）;吉林大学白求恩计划B（2012227）

摘　　要：目的由已建立的腹水来源卵巢上皮性癌（卵巢癌）细胞系中分离侧群（SP）细胞，鉴定SP细胞作为肿瘤干细胞样细胞的生物学特征，并对其进行耐药性分析。方法采用流式细胞仪分选该卵巢癌细胞系中的SP细胞；分别在体外分化条件下培养SP及非侧群（NSP）细胞，并分别测定其SP细胞比例用于比较其体外分化及自我更新能力；将SP及NSP细胞接种于非肥胖性糖尿病－严重免疫缺陷（NOD—SCID）小鼠的皮下及腹腔，比较SP及NSP细胞的成瘤率；采用活细胞计数法（CCK-8法）检测SP、NSP及未分选卵巢癌细胞（未分选细胞）对顺铂的耐药性，以50％抑制浓度（IC50）表示。结果分选前的腹水来源卵巢癌细胞中，SP细胞比例为（4．81±0．43）％，从该卵巢癌细胞系中可持续、稳定地分离出SP细胞。SP细胞经体外分化培养，3周后其中sP细胞比例为（4．89±0．33）％，与分选前SP细胞比例相近，两者比较，差异无统计学意义（P〉0．05）；而NSP细胞经体外分化培养，3周后其中sP细胞比例仅为（0．10±0．03）％，明显低于分选前SP细胞比例（P〈0．01）。皮下接种1．0×104个sP细胞6周后3只小鼠全部成瘤，皮下成瘤率为3／3，而接种1．0×104个NSP细胞12周后3只小鼠均未成瘤，皮下成瘤率为0，两者皮下成瘤率比较，差异有统计学意义（P〈0．01）；同时，腹腔接种相同数量的SP、NSP细胞后其成瘤能力与皮下接种后相似；原发肿瘤与移植瘤经HE染色和免疫组化染色显示，原发肿瘤与移植瘤均为卵巢低分化浆液性乳头状囊腺癌，均表达卵巢浆液性乳头状囊腺癌标志物CA125。SP、NSP、未分选细胞的顺铂IC50值分别为（2．33±0．14）、（1．60±0．04）、（1．81±0．03）μg／m]，SP细胞的顺铂IC50值明显高于NSP及未分选细胞（P〈0．05）；以上述未分选细胞的顺铂IC50进一步处理后，未分选细�Objective To isolate side population （SP） cells from an established ovarian cancer （OC） cell line, characterize these cells, and examine their drug resistance. Methods SP and non-SP （NSP） cells were isolated by fluorescence-activated cell sorting （FACS） , and cultured in differential conditions, then detected their SP ratio to compare their capability of differentiation and self-renewal. Moreover, SP and NSP cell tumorigenesis was examined by subcutaneous and intraperitoneal injection of these cells to nonobes ediabetic （NOD）-severe combined immundeficient（SCID） mice. Drug resistance to cisplatin was examined by cell counting kit-8 （CCK-8）.Results SP cells could be isolated stablly and insistently. There was （4.81±0.43）% of SP cells in the established OC cell line and （4.89±0.33）% of SP cells after cultured the isolated SP cells in differentiation condition, and there was no significant different between these two quantities （P〉0.05）. However, after cultured the NSP cells, there was only （0.10±0.03）% of SP cells which was significantly lower than that contained in the OC cell line （P〈0.01）. In the tumorigenesis assay 1.0×103 SP cells were injected subcutaneously and formed the xenografted tumors in 6 weeks （3/3）, and 1.0×104 NSP cells were injected subcutaneously and did not form xenografted tumors in 12 weeks （0）. The tumorigenic capability of SP cells was higher than that of NSP cells （P〈0.01）. Both the original and the xenografted tumors were low differentiated serous cystadenocarcinomas and expressed the ovarian serous cystadenocarcinomas CA125 marker after stained by HE and immunohistochemistry. Simultaneously, the SP ceils were also capable to form tumors as shown by intraperitoneal injection. In the drug resistance assay shown that the 50% inhibitory concentration （IC50） of the SP and NSP cells were respectively （2.33＋0.14）μg/ml and （1.60±0.04）μg/ml （P〈0.05）. After treated the unsorted OC cells with cisplatin, the

关 键 词：卵巢肿瘤 肿瘤干细胞 侧群细胞 抗药性 肿瘤 细胞系 肿瘤 ATP结合 匣式转运子 

分 类 号：R737.31[医药卫生—肿瘤]