摘要
目的评价^99Tc^m-重组人膜联蛋白V(rh-AnnexinV)显像检测单次化疗后肿瘤细胞凋亡的可行性,探讨其在肿瘤组织的分布与凋亡调控蛋白Survivin、Caspase-3表达的相关性。方法采用^99Tc^m直接标记法标记rh-AnnexinV,作为核素凋亡显像剂。将小鼠肝癌细胞Hca-F25接种于小鼠右腋窝皮下;区组随机化分为A(对照组,9只)和B(化疗组,10只)组,肿瘤生长至直径约1cm时,B组一次性接受环磷酰胺腹腔内注射,剂量为按体质量150mg/kg,A组注射同体积生理盐水;20h后2组同时由鼠尾静脉注入^99Tc^m-rh-Annexin V;4h后行SPECT显像并处死、取肿瘤组织,用井型γ闪烁计数器检测荷瘤小鼠肿瘤组织内放射性分布,以每克组织百分注射剂量率(%ID/g)表示:采用原位末端标记法(TUNEL)检测肿瘤细胞凋亡;采用免疫组织化学SP法检测标本中Survivin、Caspase-3蛋白表达。采用SPSS10.0软件对数据进行统计学处理。结果单次化疗后B组肿瘤组织内^99Tc^m-rh-Annexin V分布[(0.478±0.123)%ID/g]、TUNEL检测阳性细胞数[(18.030±5.600)个/视野]及Caspase-3蛋白表达[(3.266±0.482)%]均明显高于A组[分别为(0.332±0.061)%ID/g,(6.744±2.325)个/视野,(2.387±0.387)%],A组Survivin蛋白表达[(4.227±0.636)%]明显高于B组[(3.407±0.519)%],差异有统计学意义(F值依次为10.502,31.507,18.971,9.560,P均〈0.01)。相关性研究表明,肿瘤组织的放射性摄取(%ID/g)与TUNEL阳性细胞数呈明显正相关(r=0.858,P=0.000);肿瘤组织内Survivin蛋白表达与Caspase-3蛋白表达呈明显负相关(r=-0.818,P=0.000);肿瘤组织内^99Tc^m-rh—AnnexinV分布与Caspase-3蛋白表达呈明显正相关,与Survivin蛋白表达呈明显负相关(P〈0.01)。结论肿瘤组织内^99Tc^m-rh—AnnexinV的分布可以反映化疗后早期肿瘤组织细胞凋亡的状况以及Survivin、Caspase-3表达水平的变化。
Objective Recently, molecular imaging for detecting cellular apoptosis is developing rapidly. The aim of the study was to determine the effectiveness of imaging with^99Tc^m labelled recombinant human Annexin V (^99Tc^m-rh-Annexin V ) as a reflection of apoptosis in tumor, and related its distribution with expression of Survivin and Caspase-3 after a single dose of chemotherapy. Methods Eight days after being inoculated with allogenic hepatoma cells( Hca-F25 ) into right axillary fossa, the mice ( purebred 615 ) were randomly divided into two groups (control group A, n =9;and treated group B, n = 10). Group B was received a single dose of chemotherapy intraperitoneally (cyclophosphamide, 150 mg/kg). Groups A and B were given^99Tc^m-rh-Annexin V (3.7 MBq ·0.5 μg^-1 per mouse) intravenously 20 h later. Four hours aften^99Tc^m-rh-Annexin V injection, the animals were imaged and sacrificed, and the tumor samples were weighed and the radioactivity was determined in a well-counter. The accumulation of^99Tc^m-rh-Annexin V in tumor was expressed as the percentage activity of injection dose per gram of tissue ( % ID/g). Tumor cell apoptosis was examined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) method, and the expression of Survivin and Caspase-3 in tumor were determined with immunohistochemical method. SPSS 10.0 was used for data analysis. Results Single dose chemotherapy tsignificantly increased the tumor uptake of^99Tc^m-rh-Annexin V [ (0.478 ± 0. 123 ) % ID/g vs (0. 332 ±0. 061) % ID/g] and the positive number of TUNEL [ ( 18. 030 ±5. 600) cells/field vs (6. 744 ±2. 325) cells/field], as well as the expression of Caspase-3 [ (3.266±0.482)% vs (2. 387±0. 387)%, F was 10.502, 31. 507, 18. 971, respectively, all P 〈 0. 01 ], but the expression of Survivin was significantly higher in group A than in group B [(4.227±0.636)% vs (3.407±0.519)%, F =9.560, P〈0.01]. The % ID/g in tumor correlated well with the positive number of TUNEL( n = 19, r = 0. 858, P = 0.000), and he expression of Survivin correlated negatively with the expression of Caspase-3 ( r = - 0. 818, P = 0. 000). In group A and B, the distribution of^99Tc^m-rh-Annexin V correlated positively well with the expression of Caspase-3 and negatively with the expression of Survivin ( P 〈 0.01 ). Conclusion The distribution of^99Tc^m-rh-Annexin V can not only reflect the extent of apoptosis, but also the change of Survivin and Caspase-3 expression in tumor after a single dose of chemotherapy.
出处
《中华核医学杂志》
CAS
CSCD
北大核心
2008年第2期92-96,共5页
Chinese Journal of Nuclear Medicine
基金
辽宁省教育厅2006-2007高等学校科研项目计划(20060204)