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目的:探讨甲状腺乳头状癌线粒体DNA(mt DNA)D环区基因突变和线粒体微卫星不稳定及其与临床病理参数之间的相关性。方法:选取96例甲状腺乳头状癌石蜡组织标本,提取DNA并对mt DNA D环区序列行巢式PCR扩增,通过正反向直接测序对扩增产物基因序列进行检测,利用Mitomaster软件将所测区的基因序列与剑桥标准系列(revised Cambridge reference sequence,r CRS)及相关数据库进行比对,分析D环区基因突变与临床病理参数之间的相关性。结果:采用巢式PCR成功扩增出91例甲状腺乳头状癌石蜡标本mt DNA D环区序列,扩增成功率达到94.8%;87例标本(90.6%)检测出突变位点35个,共255个突变,其中D310及D514区域存在微卫星不稳定;统计结果表明患者年龄、性别、肿瘤直径、TNM分期与mt DNA D环区突变个数无相关性(P>0.05)。结论:甲状腺乳头状癌mt DNA D环区基因存在高频突变,可能在甲状腺乳头状癌的发病中起重要作用。
Objective: To investigate the relationship between mt DNA D loop gene mutation and mitochondrial microsatellite instability and its clinicopathological parameters in papillary thyroid carcinoma. Methods: 96 specimens of papillary thyroid carcinoma were selected for DNA extraction. DNA was amplified and nested PCR was performed on the mtDNA D loop region. The sequence of the amplified product was detected by forward and reverse direct sequencing. Mitomaster software The gene sequences of the region were compared with those of the revised Cambridge reference sequence (rCRS) and related databases to analyze the relationship between mutations in the D-loop region and clinicopathological parameters. Results: The mtDNA D loop region of 91 cases of papillary thyroid carcinoma was successfully amplified by nested PCR. The success rate of amplification was 94.8%. A total of 255 mutations were detected in 87 cases (90.6%) There were some mutations in D310 and D514 regions. Microsatellite instability was observed in D310 and D514 regions. The statistical results showed no correlation between age, sex, tumor diameter and TNM stage and the number of mt DNA D loop mutations (P> 0.05). Conclusion: The mtDNA D loop gene of thyroid papillary carcinoma has high frequency mutation, which may play an important role in the pathogenesis of papillary thyroid carcinoma.