【摘要】 目的:评价应用99Tcm-rh-Annexin Ⅴ显像检测单次放疗后早期肿瘤细胞凋亡的可行性,并进一步探讨肿瘤组织内99Tcm-rh-Annexin Ⅴ的分布与凋亡调控蛋白bcl-2、bax表达的相关性。方法:采用直接标记法标记rh-Annexin Ⅴ作为核素凋亡显像示踪剂。将小鼠肝癌细胞(Hca-F25)接种于小鼠右腋下部位;随机分为A(n=9,对照组)、B(n=10,放疗组)两组,B组另分为3个亚组(分别为B1组,n=4;B2组n=3;B3组,n=3)。8d后肿瘤生长至直径约1cm时,B1、B2、B3组分别接受2Gy、5Gy、10Gy不同剂量直线加速器照射;1h后两组同时由鼠尾静脉注入99Tcm-rh-Annexin Ⅴ;4h后行SPECT显像并处死、取材,应用井型闪烁探测器检测荷瘤小鼠肿瘤组织内示踪剂分布,以%ID/g(每克组织百分注射剂量率)表示。采用TUNEL法检测肿瘤细胞凋亡;采用免疫组化SP法检测标本中bcl-2、bax蛋白表达。结果:B组肿瘤组织的%ID/g、TUNEL检测阳性细胞数及bax蛋白表达均明显多于A组(P<0.01);A组bcl-2蛋白表达及bcl-2/bax明显高于B组(P<0.05)。相关性研究表明,在A、B组中肿瘤组织的%ID/g与TUNEL阳性细胞数均呈明显正相关(r=0.801、0.846),而bcl-2蛋白表达与bax蛋白表达间无明确相关性;A组肿瘤组织内bcl-2、bax蛋白表达与肿瘤组织的%ID/g及TUNEL阳性细胞数均无明确相关性;B组肿瘤组织内,bcl-2蛋白表达与%ID/g及TUNEL阳性细胞数均无明确相关性,bax蛋白表达与%ID/g及TUNEL阳性细胞数均呈明显正相关(r=0.921、0.723),而bcl-2;bax比值与%ID/g及TUNEL阳性细胞数均呈明显负相关(r=-0.828、-0.687)。结论:肿瘤组织内99Tcm-rh-Annexin Ⅴ的分布可以反映放疗后早期肿瘤组织细胞凋亡的状况以及凋亡调控蛋白bcl-2、bax表达水平的变化。更多还原

【Abstract】 Objective:To determine the effectiveness of imaging with ～(99)Tc～m labelled recombinant human Annexin Ⅴ(～(99)Tc～m-rh-Annexin Ⅴ)as a reflection of the degree of apoptosis in tumor,and investigate the distribution of ～(99)Tc～m-rh-Annexin Ⅴ and its relationship with expression of bcl-2,bax early after radiotherapy.Methods:Eight days after being inoculated with allogenic hepatoma cells(Hca-F25)into the subcutaneous region the right axillary fossa,the mice(purebred 615)were divided into two groups randomly(group A was control group,n=9;group B was the study group,n=10).Group B was divided randomly into three groups and received different dose of radiotherapy(group B1,n=4,2Gy;group B2,n=3,5Gy;group B3,n=3,10Gy).One hour after radiotherapy,group A and group B received ～(99)Tc～m-rh-Annexin Ⅴ(3.7MBq 0.5μg per mouse)injection intravenously,and 4 hours later,the animals were imaged and sacrificed.The tumor samples were weighed and the radioactivity was determined with a well-type scintillation counter.The accumulation of ～(99)Tc～m-rh-Annexin Ⅴ in tumor was expressed as the %ID/g(percentage activity of injection dose per gram of tissue).Tumor cell apoptosis was examined by TUNEL methods,and the expression of bcl-2 and bax in tumor were determined with immunohistochemical methods.Results:Radiotherapy significantly increased the tumor uptake of ～(99)Tc～m-rh-Annexin Ⅴ and the positive number of TUNEL,as well as the expression of bax(P<0.01).The expression of bcl-2 and the bcl-2/bax was significantly higher in group A than in group B(P<0.01).The %ID/g in tumor correlated well with the positive number of TUNEL in group A and group B(r=0.801,0.846),but the expression of bcl-2 had no relationship with the expression of bax.In group A,the expression of bcl-2 and bax had no relationship with the %ID/g and the positive number of TUNEL in tumor.In group B,the expression of bax correlated well with the %ID/g and the positive number of TUNEL(r=0.921,0.723),and the bcl-2/bax correlated negatively with %ID/g and the positive number of TUNEL(r=-0.828,-0.687).Conclusion:The distribution of ～(99)Tc～m-rh-Annexin Ⅴ not only can truly reflected the degree of apoptosis in tumor,but also can reflect the change of bcl-2 and bax expression early after radiotherapy.更多还原