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放射性核素标记的Ku70 反义寡核苷酸对甲状腺癌荷瘤小鼠疗效研究
王奇金,邹大进*
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(第二军医大学长海医院内分泌科, 上海 200433)

摘要:

目的研究放射性核素标记的Ku70反义寡核苷酸(ASODNs)对甲状腺癌荷瘤小鼠的放疗疗效及其作用机制。方法采用131I标记Ku70 ASODNs,建立甲状腺癌TT细胞荷瘤小鼠模型,通过Ku70 ASODNs和(或)放射处理,观察小鼠成瘤率、死亡率和肿瘤生长情况。利用Annexin Ⅴ/PI染色后流式细胞术(FCM)检测肿瘤细胞凋亡水平。蛋白质印迹分析检测不同处理后小鼠肿瘤组织Ku70蛋白的表达以及细胞增殖(PCNA)和凋亡指标(Bcl2)的表达。结果经131I标记Ku70 ASODNs处理后,肿瘤组织Ku70蛋白表达下调；肿瘤细胞生长抑制,与对照组相比肿瘤体积降低,而且小鼠成瘤率下降,死亡率亦降低(P<0. 01)；并且131IASODNs组肿瘤体积与单纯131INa处理组相比也有降低(P<0. 05)。细胞凋亡检测发现131IASODNs组肿瘤细胞的凋亡率(35.6%)高于ASODNs组(10.4%)和生理盐水组(9.2%),与131INa组(26.6%)相比也有统计学差异(P<0. 05)。进一步检测发现131IASODNs组肿瘤组织细胞中的PCNA和Bcl2表达与对照组和ASODNs组相比都降低。结论131I标记Ku70 ASODNs能显著抑制甲状腺癌肿瘤细胞的生长,促进肿瘤细胞凋亡,其作用机制与Ku70表达抑制和PCNA及Bcl2信号通路相关。

关键词: 甲状腺肿瘤；放射性核素；反义寡核苷酸类 Ku70；增殖细胞核抗原； bcl-2

DOI：10.3724/SP.J.1008.2011.033

投稿时间：2010-07-07修订日期：2010-12-08

基金项目:

Therapeutic effects of radioisotope labeled Ku70 antisense oligodeoxynuclecotide on thyroid carcinoma implanted in nude mice

WANG Qi-jin,ZOU Da-jin*

(Department of Endocrinology, Changhai Hospital, Second Military Medical University, Shanghai 200433, China)

Abstract:

ObjectiveTo investigate the therapeutic effects of radioisotope labeled Ku70 antisense oligodeoxynuclecotide (ASODNs) on thyroid carcinoma implanted in nude mice and the related mechanism. MethodsThe Ku70 ASODNs labeled with 131I was used to treat the tumorbearing mouse model derived from thyroid carcinoma TT cells. The tumorforming rate,mortality rate and tumor growth were observed and calculated after treatment with 131IASODNs, 131INa, ASODNs or NS(normal saline). AnnexinⅤ/PI assay was used to examine the apoptosis of tumor cells by flow cytometry. Western blotting analysis was performed to determine the protein expressions of Ku70, proliferating cell nuclear antigen (PCNA, a cell proliferation marker) and Bcl2(a cell apoptosis marker). ResultsAfter treatment with 131IASODNs, the Ku70 protein level was downregulated in the tumor tissues and the growth of tumor was inhibited. The tumor volume, tumorforming rate and mortality rate were significantly decreased in 131IASODNs group than in the NS control group (P<0. 01). The tumor volume of 131IASODNs group was also significantly smaller than that in the 131INa group (P<0. 05); the apoptosis rate of 131IASODNs group (35.6%) was significantly higher than that of the ASODNs group (10.4%), NS group (9.2%) and 131INa group (26.6%) (P< 0.05). Further investigation found that PCNA and Bcl2 protein levels in 131IASODNs group were lower than those in NS and ASODNs groups. ConclusionThe Ku70 131IASODNs can effectively inhibit the growth of thyroid carcinoma and promote apoptosis of TT tumor cells, which might be related to the downregulation of Ku70 and the changes of PCNA and Bcl2 signal pathway.

Key words: thyroid neoplasms radionuclides antisense oligonucleotides Ku70 proliferating cell nuclear antigen bcl-2