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目的观察树突状细胞活化抗原B7-2基因联合自杀基因系统EC-CD/5-FC于实验性乳腺癌的体、内外治疗的协同作用。方法用重组腺病毒构建CD和B2-7载体,体外感染人乳腺癌MCF-7细胞,荧光显微镜观察其感染率。然后给与前药5-FC,通过MTT法检测MCF-7细胞的增殖情况;Annexi V-FTTC/PI双染流式细胞术检测MCF-7细胞的细胞周期。BALB/C小鼠腋窝下注射MCF-7细胞制备乳腺癌移植瘤模型,待肿瘤直径达0.5 cm大小,分别在肿瘤局部直接注射Ad CD/B7-2,然后连续10 d腹腔注射5-氟胞嘧啶(5-FC),免疫组化法检测肿瘤组织中CD8+T表达。结果 MTT法检测显示MCF-7细胞对前药有较高的敏感性,Ad CD/5-FC/Ad B7-2和Ad CD/5-FC组MCF-7细胞的增殖均受到明显抑制(P<0.05);而Ad B7-2治疗基因对MCF-7细胞的增殖没有任何影响。在感染复数为100时流式细胞仪检测,Ad CD/5-FC/Ad-B7-2组和Ad CD/5-FC组均出现典型的凋亡峰,细胞周期分析显示治疗后G0~G1期比率增多,G2~M及S期细胞减少。在MCF-7移植瘤模型中,Ad CD/5-FC/Ad-B7-2治疗后移植瘤的生长明显受到抑制;肿瘤瘤体内或瘤体周围CD8+T细胞浸润增加。结论 B7-2联合自杀基因系统对乳腺癌MCF-7细胞及其MCF-7细胞移植瘤均有明显的抑制作用,B7-2联合自杀基因Ad CD/5-FC系统时,共刺激分子B7-2可通过增强和诱导的特异抗肿瘤免疫,有效减少肿瘤负荷,增强Ad-CD/5-FC系统对肿瘤的杀伤作用。
Objective To observe the synergistic effect of dendritic cell activating antigen B7-2 gene and suicide gene system EC-CD / 5-FC in vivo and in vitro treatment of experimental breast cancer. Methods The CD and B2-7 vectors were constructed with recombinant adenovirus and in vitro infected with human breast cancer MCF-7 cells. The infection rate was observed by fluorescence microscopy. The proliferation of MCF-7 cells was detected by MTT assay. The cell cycle of MCF-7 cells was detected by Annexin V-FTTC / PI double staining flow cytometry. BALB / c mice were injected with MCF-7 cells under the armpit to establish a model of breast cancer xenografts. When the tumor diameter reached 0.5 cm in diameter, mice were injected with Ad CD / B7-2 directly in the tumor and then injected with 5-fluorouracil Pyrimidine (5-FC), and the expression of CD8 + T in tumor tissue was detected by immunohistochemistry. Results MTT assay showed that MCF-7 cells were highly sensitive to prodrugs and the proliferation of MCF-7 cells was significantly inhibited by Ad CD / 5-FC / Ad B7-2 and Ad CD / 5-FC (P <0.05), while Ad B7-2 treatment gene had no effect on the proliferation of MCF-7 cells. When the number of infected cells was 100, flow cytometry showed typical apoptosis peak in Ad CD / 5-FC / Ad-B7-2 group and Ad CD / 5-FC group. Cell cycle analysis showed that G0-G1 Period ratio increased, G2 ~ M and S phase cells decreased. In the MCF-7 xenograft model, the growth of transplanted tumor was significantly inhibited after Ad CD / 5-FC / Ad-B7-2 treatment. The infiltration of CD8 + T cells in or around the tumor increased. CONCLUSION: B7-2 combined with suicide gene system significantly inhibits breast cancer MCF-7 cells and MCF-7 cell xenografts. When B7-2 combined with suicide gene Ad CD / 5-FC system, costimulatory molecules B7- 2 can enhance the tumor-killing effect of Ad-CD / 5-FC system through enhanced and induced specific anti-tumor immunity, effectively reduce the tumor burden.