活体细胞凋亡检测筛选肝癌敏感化疗药物的实验研究

Screening sensitive chemotherapeutic drug for liver cancer by apoptosis detecting in vivo

目的:通过动物实验探讨99 Tcm-HYNIC-Annexin V活体细胞凋亡检测筛选肝癌敏感化疗药物的可行性。方法:20只VX-2荷瘤免随机分为4组,通过治疗剂量的紫杉醇(PAC,n=6),5-氟尿嘧啶(5-FU,n=5),表柔比星(EPI,n=5)分组干预治疗,同时建立对照组(C,n=4)。分别予化疗前2h、化疗后46h静脉注射99 Tcm-HYN-IC-Annexin V,2h后再以单光子发射计算体层摄影(SPECT)成像,选取肿瘤部位放射性计数(T)及腿部非肿瘤区部位放射性计数(NT),计算其比值(T/NT),进行半定量分析,确定敏感化疗药物。取肿瘤组织,分别予免疫组化(Tunel)和流式细胞仪(FCM)分析组织细胞凋亡率,放射性核素聚集显像与肿瘤细胞凋亡进行相关性分析。实验数据以SPSS 17.0软件分析。结果:99 Tcm-HY-NIC-Annexin V放射显像显示,化疗前各组肿瘤部位均无明显显像(F=0.025,P=0.980),放射性核素分布主要聚集在肾脏及膀胱区。药物干预48h后,对照组仍无显像,而药物干预组显像清楚,放射性计数显著高于对照组(F=16.52,P<0.001)及化疗前(P<0.01)。药物干预组中,PAC组肿瘤组织放射性显像显著高于5-FU组和EPI组(P值分别为0.042和0.003),而5-FU组与EPI组间放射性显像差异无统计学意义(P=0.224)9。9 Tcm-HYNIC-Annexin V活体细胞凋亡检测结果与免疫组化及流式细胞检测结果一致(R2=0.537 7,P=0.002;R2=0.695 4,P<0.001)。结论9:9 Tcm-HYNIC-Annexin V核素显像检测能够反映活体组织细胞凋亡情况,该方法能够在活体状态下筛选敏感化疗药物。

OBJECTIVE：To investigate the feasibility of screening sensitive chemotherapeutic drugs for liver cancer by 99 Tcm-HYNIC-Annexin V detecting apoptosis in vivo. METHODS： New Zealand rabbits implanted VX-2 in liver were divided into control （C,n=4） ,paclitaxel （PAC, n=6）, 5-Fluorouracil （5-FU, n=5）, and Epirubicin （EPI, n=5） group. Imagings of the liver tumor were detected by SPECT through intravenous injection 99Tem-HYNIC-Annexin V before and after treatment for 48 h, Tumor radioactive count proportion to non-tumor sites （T/NT） was calculated. The tumor was taken out and divided into two pieces for Tunel immunohistochemical analysis and flow eytometry （FCM）. Data were analyzed with variance analysis and correlation analysis through soft SPSS 17.0. RESULIS： The SPECT showed that all groups had no significant tumor imaging before the treatment （F= 0. 025, P= 0. 980）. Radioactivity distributed mainly in kidney and bladder area. It is no significant tumor imaging in control group instead of the PAC, 5-FU and Epi group after treatment for 48 h. The Tumor/non-Tumor （T/NT） of PAC, 5-FU, Epi group after treatment for 48 h was significant distinguished from that of the control group （F=16.52, P〈0. 001） and all groups prior to treatment （P〈0.01）. The T/NT of PAC was significant distinguished from that of 5-FU and EPI group（P= 0. 042; P= 0. 003）. There was no different between 5-FU and EPI group（P= 0. 224）. The TINT was significant correlate with Tunel positive cells and apoptotic rate of the tumor （R2 = 0. 537 7,P=0. 002;R2：0. 695 4,P〈0. 001）. CONCLUSION： Apoptotic cell can be detected by SPECT through 99 Tcm-HYNIC-Annexin V which can screen chemotherapy sensitive drug in vivo for liver cancer.