树突细胞疫苗诱发抗肿瘤免疫特异性的体内研究  

Specificity of immunity induced by ovarian tumor extracts pulsed DC in vivo

作　　者：杨贺勤[1] 张震宇[1] 

机构地区：[1]首都医科大学附属北京朝阳医院妇产科,北京100020

出　　处：《现代妇产科进展》2009年第1期47-51,共5页Progress in Obstetrics and Gynecology

基　　金：国家自然科学基金资助项目(No:39970770)

摘　　要：目的:探讨卵巢肿瘤提取物负载的树突细胞(DC)体内诱发抗肿瘤免疫效应的特异性。方法:应用rrGM-CSF(终浓度40ng/ml)、rrIL-4(终浓度40ng/ml)及rrTNF-α(终浓度40ng/ml)体外培养Fischer344大鼠骨髓来源的单个核细胞,于培养第12天通过流式细胞仪检测细胞标志CD86、OX62抗原,用电镜观察细胞形态,证实获得树突细胞。分别利用NuTu-19卵巢肿瘤细胞和CBRH-7919大鼠肝癌细胞肿瘤提取物制备肿瘤抗原致敏Fischer344大鼠骨髓来源的树突细胞,得到两种肿瘤DC疫苗,即NuTu-DC和CBRH-DC。体内试验:Fischer344大鼠随机分为3组(1)对照组;(2)NuTu-DC治疗组;(3)CBRH-DC治疗组。于大鼠皮下接种NuTu-19细胞,5天后出瘤,开始于肿瘤接种对侧接种DC疫苗,每隔1周给予疫苗1次,共治疗2次。于第0天(未荷瘤时),第5天(出瘤后),第26天(治疗后1周),第40天(治疗后3周)天眼球取血分析各组动物外周血CD3/4/8表达水平;于荷瘤30,32,34,36,38,40天测量肿瘤体积,观察大鼠皮下肿瘤生长情况。于荷瘤40天处死各组实验动物,取肿瘤组织称重,评价治疗效果。结果:大鼠骨髓来源的骨髓单个核细胞(bone marrow mononuclearcells,BMMNC)在相关细胞因子的作用下可以培养出成熟的树突细胞,负载肿瘤提取物后,高表达表面抗原OX-62,CD-86。流式细胞仪检测外周血CD3/4/8水平显示:各组大鼠CD4/CD8比值升高,与未荷瘤时(第0天)相比,差异有统计学意义(P<0.05);CD3+CD4+T细胞表达百分率及CD4/CD8在治疗后3周时(第40天)NuTu-DC治疗组明显高于另两组(P<0.05),而对照组与CBRH-DC比较无差异。通过测量肿瘤生长曲线可见NuTu-DC治疗组各大鼠肿瘤体积较其它两组小,荷瘤第40天时,NuTu-DC治疗组为106.57±56.65mm3,对照组为299.78±118.17mm3,CBRH-DC治疗组为299.12±85.61mm3,与对照组和CBRH-DC组比较差异有统计学意义(P<0.01);而且NuTu-DC治疗组肿瘤平均瘤重为0.197±0.039g,与对照组和CBRH-DC治疗组有显著差异(P<0.01Objective：To study the specificity of immunity induced by ovarian tumor cells extracts pulsed dendrite cells in rats. Methods： The bone marrow mononuelear cells （BMMNC） were obtained and the mature DC were induced by culturing BMMNC in medium with granulocyte-macrophage colony stimulating factor （ GM-CSF）, interleukin4 （ IL-4 ） and tumor necrosis factor-α （TNF-α）. The shape of the DCs were observed, and the expression of the surface markers were analyzed by flow cytometry. After culturing for 5 days, the rats ovarian cancer cell line NuTu-19 extracts and cancer cell line CBRH-7919 extracts were added to pulse DC and acquire the tumor cells extracts pulsed DCs. After the tumor cells NuTu-19 were subcutaneously injected to the right armpit of the rats,rats vaccinated twice with NuTu-DC were compared to those vaccinated with CBRH-DC or PBS. Results ：The mature DCs were induced by BMMNC co-cuhuring with relative cytokines and cell surface phenotypes were highly expressed. CD/CD8 raised after tumor cells injection（P 〈 0.05 ）. On day 40, compared to the others, CD^＋ T cells and CD/ CD8 of the rats vaccinated with NuTu-DC raised remarkably （P 〈 0. 05 ）. Tumor volume and tumor weight were reduced in rats vaccinated with NuTu-DC, compared with rats vaccinated with CBRH-DC and PBS （P 〈0.01 ）,and its inhibition ratio was 64.45%. Tumor volume and weight did not reduced in rats vaccinated with CBRH-DC compared with those of PBs group （P 〉 0.05）. The inhibition ratio was O. 22%. Conclusion： NuTu-DC vaccine can induce active immunity against tumor cells in-vivo,and it is specific.

关 键 词：树突细胞 大鼠 近交F344 卵巢肿瘤 疾病模型 动物 肿瘤提取物 免疫疗法 

分 类 号：R737.33[医药卫生—肿瘤]