自然杀伤细胞对宫颈癌HeLa细胞裸鼠移植瘤的治疗效果  

Effects of Natural Killer Cells on the Treatment of HeLa Cancer Xenografts in Nude Mice

作　　者：杨柳 楼敏铭 冯郸 刘微微 马洁 YANG Liu;LOU Minming;FENG Dan;LIU Weiwei;MA Jie(Tianjin Kangting Biotechnology Group Co.,Ltd.,Tianjin,300385,China)

机构地区：[1]天津市康婷生物工程集团有限公司,天津300385

出　　处：《肿瘤药学》2020年第4期440-445,450,共7页Anti-Tumor Pharmacy

摘　　要：目的探讨自然杀伤(NK)细胞对人宫颈癌HeLa细胞的体外杀伤作用以及不同浓度NK细胞对宫颈癌HeLa细胞裸鼠移植瘤的治疗效果。方法采用含红色荧光蛋白(RFP)和荧光素酶报告基因(Luc)的慢病毒感染HeLa细胞,荧光倒置显微镜检测细胞中RFP基因的表达,体外成像技术检测Luc基因的表达。分别将不同浓度的NK细胞作用于HeLa-RFP/Luc细胞24、48、72 h,CCK-8法检测NK细胞对靶细胞的杀伤作用。将HeLa-RFP/Luc细胞接种于BALB/c-nu裸鼠皮下,建立宫颈癌裸鼠移植瘤模型,待肿瘤生长至100 mm3时,随机分为模型组、NK高剂量组、NK低剂量组、空白对照组,每组10只。给药4个疗程后,观察各组裸鼠的一般状况,每周测量体重及瘤体大小,体外活体成像观察裸鼠瘤体的荧光变化。治疗周期结束后,剥离瘤体,称重,取脾脏组织,采用ELISA法检测脾脏组织液中肿瘤相关基因的表达。结果NK细胞对HeLa-RFP/Luc细胞的体外杀伤率呈时间和剂量依赖性,效靶比越大则杀伤作用越强,且作用72 h后,各效靶比的杀伤率均达90%以上。成功筛选出共表达RFP和Luc基因的HeLa-RFP/Luc细胞株。NK细胞治疗对裸鼠的体重无明显影响(P>0.05),但随着治疗周期的延长,治疗组裸鼠瘤体体积和荧光值的增长速率均明显低于模型对照组(P<0.05)。与模型对照组相比,NK细胞治疗后各组裸鼠雌激素E2水平均无明显变化(P>0.05),IL-2、IFN-γ水平显著升高,IL-4水平显著降低(P<0.05)。结论NK细胞治疗可延缓人宫颈癌HeLa细胞的生长,下调促肿瘤因子的表达,且在治疗3个疗程后效果最佳。Objective To explore the effect of natural killer(NK)cells on apoptosis of cervical cancer HeLa cells,and the antitumor effect of NK cells at different doses on nude mice bearing cervical cancer xenografts.Methods HeLa cells were infected with lentivirus containing red fluorescent protein(RFP)and luciferase(Luc)genes for several days.The expression of RFP gene was detected by fluorescence inverted microscope,and the expression of Luc gene was detected by in vivo fluorescence imaging.HeLa-RFP/Luc cells were co-cultured with different concentrations of NK cells for 24 h,48 h and 72 h,and the killing effect was detected by CCK-8.BALB/c nude mice were adopted to establish a tumor-bearing model by inoculating subcutaneously HeLa-RFP/Luc cells.When the tumor grew up to 100 mm3,the xenografts nude mice were randomly divided into control group,model group,NK high-dose group and NK low-dose group,10 mice in each group,and got four courses of treatment.Then we measured body weight and tumor size weekly,and observed the changes of tumor fluorescence by in vivo fluorescence imaging system.After the treatment was finished,relevant tumors were striped and weighed,and spleen tissues were taken away.ELISA was applied to examine the expression of tumor-related factors in spleen tissues.Results The killing ratio of NK cells to HeLa-RFP/Luc cells in vitro was time-and dose-dependent.After 72 hours of co-culture,the killing ratio was more than 90%.The larger the effect-target ratio,the stronger the killing effect.HeLa-RFP/Luc cells co-expressing RFP and Luc genes were successfully screened out.No significant effects were observed on the body weight of nude mice after NK treatment(P>0.05).Along with the treatment courses continued,the growth rates of tumor volume and fluorescence in the NK treatment groups were significantly lower than that of the model control group(P<0.05).Compared with the model control group,there was no significant difference in estrogen level in NK treatment groups(P>0.05),but the levels of IL-2,IFN-γwere incr

关 键 词：自然杀伤细胞 宫颈癌HELA细胞 体外杀伤实验 裸鼠移植瘤 活体成像 肿瘤治疗 

分 类 号：R737.33[医药卫生—肿瘤]