结外NK/T细胞淋巴瘤敏感药物筛选  

Drug sensitivities of extranodal NK/T-cell Lymphoma cells cultured in different culture systems

作　　者：刘蕾[1,2] 郭宏强[1,2] 杨树军[1,2] 

机构地区：[1]河南省肿瘤医院 [2]郑州大学附属肿瘤医院内科,河南郑州450008

出　　处：《中华肿瘤防治杂志》2015年第2期86-90,共5页Chinese Journal of Cancer Prevention and Treatment

基　　金：国家自然科学基金(81101797);河南省卫生厅项目(2011020157)

摘　　要：目的应用无血清培养基(serum free medium,SFM)初步富集肿瘤耐药细胞,筛选出结外NK/T细胞淋巴瘤(extranodal NK/T-celllymphoma,ENKTCL)的敏感药物。方法应用加入10%人血清和700U/mL人白细胞介素-2(interlukin-2,IL-2)的1640培养基和加入700U/mL人IL-2的SFM VIVO-15TM,分别培养SNK-6细胞;MTT法分别检测2种培养体系中表柔比星(adriamycin,ADM)、奥沙利铂(oxaliplatin,L-OHP)、吉西他滨(gemcitabine,GEM)、左旋门冬酰胺酶(L-Asparaginase,L-ASP)、阿糖胞苷(cytosine arabinoside,Ara-C)和甲氨蝶呤(methotrexate,MTX)等药物作用的IC50;2种培养体系中分别加入上述各种药物作用32h后,7AAD/PE-AnnexinⅤ染色法流式细胞术检测各处理组细胞的凋亡率,并分析比较。结果无血清培养体系中细胞部分悬浮生长,每个悬浮球由>50个数目不等的细胞组成,悬浮球体积较有血清中明显增大。有血清和SFM中IC50含量,ADM分别为(0.12±0.018)和(0.751±0.14)μg/mL,P<0.001;Ara-C分别为(0.217±0.002)和(0.822±0.028)μg/mL,P<0.001;MTX分别为(0.023±0.001)和(0.032±0.002)μg/mL,P=0.002。药物作用后,有血清和SFM中SNK-6细胞的凋亡率,ADM分别为(45.23±2.58)%和(30.91±2.13)%,P=0.041;Ara-C分别为(56.12±2.32)%和(41.47±2.46)%,P=0.034;MTX分别为(24.42±1.92)%和(13.01±2.28)%,P=0.045;GEM分别为(15.05±2.05)%和(41.45±1.74)%,P<0.001。结论SFM VIVO-15TM可用于ENKTCL SNK-6细胞培养;无血清培养后的SNK-6细胞对ADM、Ara-C和MTX耐药性增强,对L-OHP、GEM和L-ASP敏感。OBJECTIVE To enrich tumor-initiating cells from serum free medium （SFM） supplemented with human interleukin-2 （IL-2）. To screen the chemosensitivity of drugs for extranodal NK/T-cell lymphoma （ENKTCL） in vitro. METHODS SNK-6 cells were cultured in 10% human serum culture medium and SFM VIVO-15TM supplemented with human IL-2. The growth status of cells cultured in two kinds of system were closely observed and recorded. Drug sensitiv- ities of SNK-6 cells to Adriamycin （ADM）, Oxaliplatin （L-OHP）, Gemcitabine （GEM）, L-Asparaginase （L-ASP）, cyto- sine arabinoside （Ara-C） and Methotrexate （MTX） were analyzed by MTT assays. PE Annexin V was adopted by Flow cytometry to detect the apoptosis of SNK-6 cells dealed with ADM,L-OHP,GEM,L-ASP,Ara-C and MTX for 32 hours. RESULTS SNK-6 cells of ENKTCL grow in a suspention state in the two different culture systems,especially in SFM VIVO-15TM supplemented with human IL-2. Tumorspheres were initially observed at the fifth day of serum-free culture. Each tumorsphere composed of more than fifty cells. The IC50 of ADM, Ara-C and MTX in serum and serum free medium were respectively （0.12±0. 018） and （0. 751±0.14） μg/mL（P〈0. 001）,（0. 217±0. 002） and （0. 822±0. 028） μg/mL （P〈0. 001）,（0. 023±0. 001） and （0. 032±0. 002） μg/mL（P=0. 002）. The apoptosis rates of ADM,Ara-C,MTX and L-ASP in serum and serum-free medium were respectively （45.23 ± 2.58）% and （30. 91 ± 2.13）% （P = 0. 041）, （56.12±2.32）% and （41.47±2.46）% （P=0.034）,（24.42±1.92）% and （13.01±2.28）% （P=0.045）, （15.05± 2. 05）% and （41.45±1.74）% （P〈0. 001）. CONCLUSIONS The SFM VIVO-15TM supplemented with human IL-2 can be used to culture ENKTCL cells. SNK-6 cells cultured in SFM show drug resistance to ADM,Ara-C and MTX, but are sensitive to L-OHP,GEM, L-ASP.

关 键 词：MTT 细胞凋亡 无血清培养基 SFM 肿瘤起源细胞 

分 类 号：R733.1[医药卫生—肿瘤]