p14ARF基因遗传变异与喉癌相关性及其介导线粒体途径致细胞凋亡作用机制研究

The morbidity of Laryngeal Squamous Cell Carcinoma (LSCC) is second in Head and Neck Squamous Cell Carcinoma (HNSCC).Five years survival rate is only 30-40% in advanced patients. It is one of the most serious malignant tumors for human health hazards.. The domestic and abroad researches in recent years show that p14ARF gene plays an important role in the process and incidence of HNSCC. Our preliminary works suggest that the mitochondrial function, p14ARF polymorphisms are associated with the risk and prognosis of HNSCC (including LSCC) and Second Primary Malignancy(SPM) . And successfully built an eukaryotic expression vector model of mitochondrial membrane protein p32 interaction with p14ARF..Hypothesis: The polymorphisms in the second exon of the p14ARF gene are associated with the risk and prognosis of LSCC. The mechanism may be that the p14ARF genetic variants effect the interaction with P32, causing the abnormalities of the mitochondrial membrane potential, cell cycle and apoptosis, eventually leading to carcinogenesis.. For the first time, we investigate the association between the single nucleotide polymorphisms (SNPs) of p14ARF gene and the risk and prognosis of LSCC. And using molecular epidemiology and molecular biology experimental methods, we explore the function mechanism of p14ARF polymorphisms which leading to cell carcinogenesis.Therefore, this research will provide a new train of thought for the early diagnosis or prognosis and individualized treatment of Laryngeal Squamous Cell Carcinoma.

喉癌发病率居头颈鳞癌第二位，晚期患者5年生存率仅为30-40%，是严重危害人类健康的恶性肿瘤之一。近年国内外研究发现p14ARF基因在头颈鳞癌的发病过程中起到重要作用，我们前期工作也表明线粒体功能改变、p14ARF多态性与包括喉癌在内的头颈鳞癌的二次原发肿瘤相关，并成功构建了证明线粒体膜蛋白p32与p14ARF的相互作用的真核表达载体。据此推测：p14ARF基因第二外显子多态性改变与喉癌的发病风险及预后相关，其调控机制可能是发生遗传变异的p14ARF影响与p32相互作用后，引起线粒体膜电位改变，进而细胞生长周期及凋亡发生异常，最终导致细胞癌变。本课题首次在明确p14ARF遗传变异与中国人群喉癌发病风险及预后相关性的基础上，利用分子流行病学及生物学实验研究手段，从功能学上探索p14ARF多态性导致细胞癌变的作用机制。为喉癌的早期诊断、预后判断以及个体化治疗提供了新思路。

本实验通过使用荧光定量PCR定量检测了P14ARF基因与P32基因的在喉癌中的表达水平，探讨了此二者基因在喉癌发生发展过程中可能起到的作用。实验发现P14ARF基因在喉癌早期表达水平上升，可能是由于肿瘤抑制基因P14ARF于肿瘤早期过表达试图抑制肿瘤的发展有关，在肿瘤晚期表达水平下降甚至缺失。P32基因在喉癌中的表达水平与癌旁正常喉粘膜组织相比未见统计学差异，可能暗示P32基因并不能作为喉癌发生发展以及诊断的一个分子指标。另外，通过免疫组化染色检测了P14ARF蛋白以及P32蛋白在喉癌组织中的表达水平，结果与荧光定量PCR结果基本一致。综上所述，P14ARF在喉癌中早期表达水平上升，晚期下降，可以作为喉癌发生发展以及分子诊断的指标，P32基因在喉癌的发生发展中并没有起到关键作用。